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Comparison of Different Gonadotropin-Releasing Hormone Plus Prostaglandin F2alpha Synchronization Protocols in Bos taurus, Bos indicus, and Bos indicus x Bos taurus Postpartum Cows and Heifers

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Comparison of Different Gonadotropin-Releasing Hormone Plus Prostaglandin F2alpha Synchronization Protocols in Bos taurus, Bos indicus, and Bos indicus x Bos taurus Postpartum Cows and Heifers
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ESTERMAN, REINA D. ( Author, Primary )
Copyright Date:
2008

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Subjects / Keywords:
Breeding ( jstor )
Cattle ( jstor )
Conception rate ( jstor )
Estrus ( jstor )
Estrus cycle ( jstor )
Heifers ( jstor )
Ovulation ( jstor )
Pregnancy rate ( jstor )
Prostaglandins ( jstor )
Zebu ( jstor )

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University of Florida
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University of Florida
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Copyright Reina D. Esterman. Permission granted to the University of Florida to digitize, archive and distribute this item for non-profit research and educational purposes. Any reuse of this item in excess of fair use or other copyright exemptions requires permission of the copyright holder.
Embargo Date:
5/31/2008
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659815228 ( OCLC )

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1 COMPARISON OF DIFFERENT GONADOT ROPIN-RELEASING HORMONE PLUS PROSTAGLANDIN F2 SYNCRHONIZATION PROTOCOLS IN Bos taurus , Bos indicus and Bos indicus Bos taurus POSTPARTUM COWS AND HEIFERS By REGINA D. ESTERMAN A THESIS PRESENTED TO THE GRADUATE SCHOOL OF THE UNIVERSITY OF FLOR IDA IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE UNIVERSITY OF FLORIDA 2007

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2 Copyright 2007 By Regina D. Esterman

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3 To my family for providing support and encour agement all my life and to my friends for providing insight and laughter.

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4 ACKNOWLEDGMENTS First and foremost, I would like to thank my mentor, Dr. Joel Yelich for his guidance, patience, and intellectual support throughout my graduate program. I would also like to thank the members of my supervisory committee, Drs. Pe ter J. Hansen and Owen Rae, for their insight and contributions to my growth as a scientist. Special thanks are extended to the member s of my lab, Brad Austin, Steaven Woodall, and Erin McKinniss. Their time, assistance, a nd willingness to help w ith these projects were invaluable. My lab partners have made my gra duate experience more enjoyable. I would also like to thank the staff of the Santa Fe Beef Research Unit a nd Beef Research Unit for their cooperation and assistance during the experiments. Additional grat itude is expressed to the Bar L Ranch and Laramore family in Mariana, Florid a for cattle used in this research and their assistance and hospitality during th e course of the experiments. Finally, I would like to thank my friends a nd fellow graduate students for the good times and laughs that have been had. Also, thanks fo r challenging my scientific ideas and trying to make me a more humorous person. I truly appr eciate everyone’s encouragement and support.

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5 TABLE OF CONTENTS Page ACKNOWLEDGMENTS...................................................................................................4 LIST OF TABLES...............................................................................................................7 LIST OF FIGURES...........................................................................................................10 LIST OF ABBREVIATIONS............................................................................................11 ABSTRACT....................................................................................................................... 12 CHAPTER 1 INTRODUCTION......................................................................................................14 2 LITERATURE REVIEW...........................................................................................16 Hypothalamic-Pituitary-Ovarian Axis........................................................................16 Estrous Cycle..............................................................................................................19 Overview.............................................................................................................19 Estrous Detection.................................................................................................22 Follicular Development..............................................................................................26 Corpus Luteum Function and Luteolysis....................................................................30 Overview.............................................................................................................30 Luteolysis............................................................................................................32 Estrous Synchronization.............................................................................................36 Overview.............................................................................................................36 Prostaglandin F2 .................................................................................................37 Progestins............................................................................................................41 Gonadotropin Releasing Hormone (GnRH)........................................................45 GnRH + CIDR + PGF2 Systems........................................................................48 3 EFFECTIVENESS OF NEW VS. ONCE-USED CIDR AND CLOPROSTENOL SODIUM VS. DINOPROST TROMETHAMINE IN A GnRH/CIDR + PGF2 PROTOCOL IN LACTATING Bos indicus Bos taurus BEEF COWS..................52 Introduction.................................................................................................................52 Materials and Methods...............................................................................................53 Results........................................................................................................................ .57 Discussion...................................................................................................................61 Implications................................................................................................................69

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6 4 EFFECTIVENESS OF CLOPROSTENOL SODIUM VS. DINOPROST TROMETHAMINE IN A GnRH/CIDR + PGF2 SYNCHRONIZATION PROTOCOL IN ANGUS, BRAHMAN, and BRAHMAN ANGUS COWS AND HEIFERS....77 Introduction.................................................................................................................77 Materials and Methods...............................................................................................78 Results........................................................................................................................ .81 Discussion...................................................................................................................85 Implications................................................................................................................92 5 FOLLICLE DEVELOPMENT, ESTROUS CHARACTERISTICS, AND EFFECTIVENESS OF A GnRH/CIDR + PGF2 SYNCHRONIZATION PROTOCOL IN POSTPARTUM LACTATING ANGUS ( Bos taurus ) AND BRANGUS ( Bos indicus Bos taurus ) BEEF COWS.....................................................................................100 Introduction...............................................................................................................100 Materials and Methods.............................................................................................101 Results.......................................................................................................................1 07 Discussion.................................................................................................................113 Implications..............................................................................................................119 6 CONCLUSIONS AND IMPLICATIONS...............................................................128 LIST OF REFERENCES.................................................................................................136 BIOGRAPHICAL SKETCH...........................................................................................157

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7 LIST OF TABLES Table Page 3-1 Main effects for estrous, c onception and pregnancy rates of Bos indicus Bos taurus cows synchronized with cont rolled intravaginal progesterone-releasing device (CIDR: New vs. Used) treatments and prostaglandin F2 (Cloprostenol sodium(Cloprostenol) vs. Dinoprost tromethamine-(Di noprost)) treatments administered at CIDR removal. ............................................................................................................................... ...70 3-2 Simple treatment effects for estr ous, conception and pregnancy rates of Bos indicus Bos taurus cows synchronized with controlled intravaginal proge sterone-releasing device (CIDR: New vs. Used) tr eatments and prostaglandin F2 (Cloprostenol sodium(Cloprostenol) vs. Dinoprost tromethamine -(Dinoprost)) treatments administered at CIDR removal. .......................................................................................................71 3-3 Days postpartum (DPP) effects on estr ous, conception and pregnancy rates of Bos indicus Bos taurus cows synchronized with two cont rolled intravaginal progesteronereleasing device (CID R) treatments and two prostaglandin F2 treatments administered at CIDR removal. ...................................................................................................72 3-4 Main effects for estrous, c onception and pregnancy rates of Bos indicus Bos taurus cows synchronized with cont rolled intravaginal progesterone-releasing device (CIDR: New vs. Used) treatments and prostaglandin F2 (Cloprostenol sodium(Cloprostenol) vs. Dinoprost tromethamine-(Dinoprost)) treatm ents administered at CIDR removal.. ............................................................................................................................... ...73 3-5 Simple treatment effects for estr ous, conception and pregnancy rates of Bos indicus Bos taurus cows synchronized with controlled intravaginal proge sterone-releasing device (CIDR: New vs. Used) tr eatments and prostaglandin F2 (Cloprostenol sodium(Cloprostenol) vs. Dinoprost tromethamine -(Dinoprost)) treatments administered at CIDR removal..........................................................................................................74 3-6 Days postpartum (DPP) effects on estr ous, conception and pregnancy rates of Bos indicus Bos taurus cows synchronized with two cont rolled intravaginal progesteronereleasing device (CID R) treatments and two prostaglandin F2 treatments administered at CIDR removal. ...................................................................................................75 4-1 Treatment (TRT), year, and treatment year (TRT YEAR) effects for estrous, conception and pregnancy rates of Angus, Brahman, and Brahman Angus cows synchronized with a controlled intravaginal progesterone-releasing device (CIDR) and two prostaglandin F2 (cloprostenol sodium: cloproste nol vs. dinoprost tromethamine: dinoprost) treatments administ ered at CIDR removal..............................................94

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84-2 Estrous, conception and pregnancy rates for Angus (AN), Brahman (BR), and Brahman Angus cows synchronized with a contro lled intravaginal pr ogesterone-releasing device (CIDR) and two prostaglandin F2 (cloprostenol sodium and dinoprost tromethamine) treatments administered at CIDR removal......................................95 4-3 Cow age effects for estrous, conception and pregnancy rates of Angus, Brahman, and Brahman Angus cows synchronized with a cont rolled intravaginal progesteronereleasing device (CIDR) and two prostaglandin F2 (cloprostenol sodium and dinoprost tromethamine) treatments administered at CIDR removal......................................96 4-4 The effect of days postpartum (DPP) on estrous, conception and pregnancy rates of Angus, Brahman, and Brahman Angus cows synchronized with a controlled intravaginal progesterone-releasing de vice (CIDR) and two prostaglandin F2 (cloprostenol sodium and dinoprost trometha mine) treatments administered at CIDR removal.....................................................................................................................97 4-5 Treatment (TRT), year, and treatment year (TRT YEAR) effects for estrous, conception and pregnancy rates of Angus, Brahman, and Brahman Angus heifers synchronized with a controlled intravaginal progesterone-releasing device (CIDR) and two prostaglandin F2 (cloprostenol sodium: cloproste nol vs. dinoprost tromethamine: dinoprost) treatments administ ered at CIDR removal..............................................98 4-6 Breed effects for estrous, conception a nd pregnancy rates of Angus (AN), Brahman (BR), and Brahman Angus heifers synchronized with a controlled intravaginal progesterone-releasing device (C IDR) and two prostaglandin F2 (cloprostenol sodium and dinoprost tromethamine) treatments admi nistered at CIDR removal by breed. 99 5-1 Effect of breed and estrous cycling st atus on ovulation rates to GnRH and ovulatory follicle size (LS mean SE) in Angus and Brangus cows synchronized with a 7 d CIDR treatment with prostaglandin F2 (PGF2 ) administered at CIDR removal...........122 5-2 Effect of breed and ovula tion status at GnRH (Ovulated – OV vs. No ovulation No-OV) on follicle sizes (LS mean SE) and estrogen concentrations (LSmean SE) for follicles ovulating after PGF2 in Angus and Brangus cows synchronized with a 7 d CIDR treatment with prostaglandin F2 (PGF2 ) administered at CIDR removal..123 5-3 Effect of breed and ovulati on status at GnRH on progester one concentrations (LS means SE) at CIDR removal, CL regression rate, an d incidence of silent estrus of Angus and Brangus cows synchronized with a 7 d CIDR treatment with prostaglandin F2 (PGF2 ) administered at CIDR removal...............................................................................124 5-4 Estrous characteristics as determined by HeatWatch of Angus and Brangus cows synchronized with a 7 d CIDR tr eatment with prostaglandin F2 (PGF2 ) administered at CIDR removal. .....................................................................................................125

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95-5 Effect of breed and estrous cycling status on estrous, conception and pregnancy rates of Angus and Brangus cows synchronized with a 7 d CIDR treatment with prostaglandin F2 (PGF2 ) administered at CIDR removal..........................................................126 5-6 Effect of breed and ovulation status at GnRH on estrous, conception and pregnancy rates of Angus and Brangus cows synchr onized with a 7 d CIDR treatment with prostaglandin F2 (PGF2 ) administered at CIDR removal....................................127

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10 LIST OF FIGURES Figure Page 3-1 Effect of interval from PGF to the onset of estrus in Bos indicus Bos taurus cows synchronized with two controlled intravag inal progesterone-rel easing device (CIDR) treatments and two prostaglandin F2 treatments administered at CIDR removal. .76 5-1 Effect of breed (Angus vs. Br angus on the interval from PGF2 to the onset of HeatWatch estrus. Estrous response is reported as the percentage of cows in estrus during each time interval within a breed divide d by the total in estrus within a breed. Cows received GnRH concurrent with a CIDR followed by PGF2 7 d later. ......120 5-2 Effect of ovulation status to GnRH on interval from PGF2 to onset of estrus. Estrous response is reported as the percen tage of cows in estrus duri ng each time interval within ovulation status divided by the total in estrus within ovulation stat us. Cows received GnRH concurrent with a CIDR followed by PGF2 7 d later in Angus and Brangus cows. Estrous response by ovulation status to GnRH tended to be different P = 0.07 as determined by survival analysis...............................................................................121

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11 LIST OF ABBREVIATIONS AI artificial insemination BCS body condition score BW body weight CIDR controlled-intravaginal progesterone-releasing device CL corpus luteum DPP days postpartum FSH follicle stimulating hormone GnRH gonadotropin-releasing hormone LH luteinizing hormone PGF2 prostaglandin F2 MGA melengestrol acetate

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12 Abstract of Thesis Presen ted to the Graduate School of the University of Florida in Partial Fulfillment of the Requirements for the Degree of Master of Science COMPARISON OF DIFFERENT GONADOT ROPIN-RELEASING HORMONE PLUS PROSTAGLANDIN F2 SYNCRHONIZATION PROTOCOLS IN Bos taurus , Bos indicus and Bos indicus Bos taurus POSTPARTUM COWS AND HEIFERS By Regina D. Esterman May 2007 Chair: Joel V. Yelich Major: Animal Sciences A series of experiments were conducte d to compare different Select Synch synchronization protocols, which consist of gonadotropin-releasing hormone (GnRH) administration followed 7 d later with prostaglandin F2 (PGF2 ) combined with a vaginal progesterone insert (CIDR), in Bos taurus , Bos indicus , and Bos indicus Bos taurus postpartum cows and heifers. In Exp. 1, postpartum cows (n = 284) were synchronized with the Select Synch protocol in a 2 2 factorial design comparing a ne w CIDR vs. once-used CIDR and comparing the prostaglandins cloprostenol sodium vs. dinoprost tromethamine. Following PGF2 , estrus was detected for 5 d and cows were ar tificially inseminated (AI) 8 to 14 h after an observed estrus. In Exp. 2, postpartum cows (n = 259) received the same treatments as Exp. 1, but estrus was detected for 3 d followed by AI and all cows not exhibiting estrus by 72 h were timed-AI concomitant with GnRH 76 to 80 h after PGF2 . In both experiments, estrous response, conception, and synchronized pregnancy rates we re similar (P > 0.05) between new CIDR vs. once-used CIDR and cloprostenol sodium vs. di noprost tromethamine. Across treatments, mean synchronized pregnancy rates were 32.6% for Exp. 1 and 51.6% for Exp. 2. As days from calving increased, response to treatment increased in both experiments. Conception rate was

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13 greater (P < 0.05) for cows detected in estrus 72 h (54.1%) compared to cows > 72 h (23.4%) after PGF2 . In Exp. 3, postpartum cows (n = 335) and 2 yr old heifers (n = 163) of Bos taurus and varying degrees of Bos indicus breeding received a similar protocol as Exp. 2, with a new CIDR and either cloprostenol sodium or dinopros t tromethamine at CIDR removal. In cows, estrous responses and conception rates were sim ilar between treatments. Timed-AI pregnancy rates tended (P = 0.06) to be grea ter, and synchronized pregnanc y rates were greater (P < 0.05) for cloprostenol sodium (57.2%) compared to di noprost tromethamine (46.2%). Breed had no effect on synchronized pregnancy rates but as days from calving increased, response to treatment increased. In heifers, estrous response, con ception, and synchronized pregnancy rates were similar (P > 0.05) across tr eatments and breeds. In Exp. 4, postpartum Angus ( Bos taurus ; n = 31) and Brangus ( Bos indicus Bos taurus ; n = 22) cows were synchronized with Select Sync h plus a new CIDR. Ovar ian follicle response to GnRH was monitored using ultrasonography. Breed did not influence ovulation rate to GnRH. Cows which ovulated to GnRH had gr eater (P < 0.05) luteal regression, estrous response, and synchronized pregnancy rate compared to cows that failed to ovulate, regardless of breed. Intervals from PGF2 to estrus and distribution of estr us were similar (P < 0.05) between breeds. Angus had a longer (P < 0.05) estrus (63 h 46 min) with more (P < 0.05) mounts (49.0) compared to Brangus (7 h 30 mi n; 21.5 mounts; respectively). Es trous response, conception rate, and pregnancy rates were similar across breeds. In conclusion, CIDR (new vs. once-used) and PGF2 type (cloprostenol sodium vs. dinoprost tromet hamine) used in the Select Synch protocols resulted in similar synchronized pregnancy rates in three of four studies Furthermore, response to the Select Synch protocols improved as the interval from calving lengthened in cattle of Bos indicus breeding.

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14 CHAPTER 1 INTRODUCTION Artificial insemination (AI) allows beef producer s to use superior genetics within their cow herd by using bulls that they would normally not be able to purchase on their own. In order to effectively and efficiently utilize AI, estrous synchronization is comm only used to decrease the amount of estrous detection required for th e AI breeding period. In an unsynchronized group of cows, estrous detection would normally span at least 21 d, the average length of the estrous cycle, to allow all cows an opportunity to di splay estrus and be inseminated. In estrous synchronized cows, estrous detection can be decreas ed to < 7 d, shortened to 3 d with the use of a timed-AI for cows not exhibiting estrus, or eliminated completely by performing a timed-AI where all cows are inseminated at a single, predetermined time. Use of estrous synchronization protocols also increases the number of cows pre gnant early in the breeding season, resulting in a heavier and more uniform calf crop the following year. Estrous synchronization is achieved by admi nistering hormone treatments to cows on designated days. Common treatments includ e gonadotropin-releasing hormone (GnRH), progestogens, and prostaglandin F2 (PGF2 ). The primary function of GnRH is to induce ovulation. When administered at the start of a synchroniza tion protocol, it allows for synchronization of follicle waves and when admi nistered after the end of a synchronization protocol induces ovulation for a timed-AI. Proges togens prevent animals from displaying estrus during the duration of the treatment period, genera lly 7 to 14 d, and can induce estrous cyclicity in some anestrous cows and prepube rtal heifers. Prostaglandin F2 functions to regress the corpus luteum (CL), which causes the cow to exhi bit estrus 2 to 7 d later. The aforementioned hormone treatments can be utilized either separately or in comb ination to synchronize estrus in both postpartum beef cows and (or) yearling heifers.

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15 One of the most common synchronization pr otocols involves administration of GnRH followed 7 d later by PGF2 resulting in a synchronized estrus during the 5 d following PGF2 . This protocol is termed the Select Synch prot ocol. A common problem with the Select Synch protocol is that some cattle exhibit estrus 1 to 2 d before the PGF2 treatment. Another commonly used protocol is to combine the Se lect Synch protocol with an intravaginal progesterone-releasing devi ce (CIDR) at GnRH with CIDR removal at PGF2 . The addition of the CIDR allows for a highly synchronous estrus in the 2 to 5 d following PGF2 and it can induce estrus in some anestrous cows and prepuberta l heifers. This is termed the Select Synch + CIDR protocol. In subtropical regions, such as Florida, cattle containing some Bos indicus breeding are commonly raised due to their superior tolerance to high temperatures, humidity, parasites, and low quality forages compared to Bos taurus cattle. However, subtle differences in the reproductive physiology and hormona l profiles in cattle of Bos indicus breeding can modify how they respond to some estrous synchronization pr otocols. These modifications, in combination with the fact that estrus is difficult to detect in cattle of Bos indicus breeding, often results in decreased pregnancy rates to synchr onization protocols compared to Bos taurus cattle. Therefore the objectives of this re search are to evaluate the effec tiveness of the Select Synch and the Select Synch + CIDR protocol for sync hronizing postpartum cows and heifers of Bos taurus , Bos indicus , and Bos indicus Bos taurus breeding.

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16 CHAPTER 2 REVIEW OF LITERATURE Hypothalamic-Pituitary-Ovarian Axis Overview Reproductive function in cattle is largely in fluenced by hormones produced within the brain. The pulsatile release of gonadotropin releasing hormone (GnRH) from the hypothalamus regulates the release of hormones from the ante rior pituitary, including follicle stimulating hormone (FSH) and luteinizing hormone (LH). The latter two hormones are under the positive control of GnRH, which regulates their pulsatile release thro ughout the estrous cycle. Gonadotropin releasing hormone, a decapepti de (Senger, 1999), is produced in the hypothalamus. Release of GnRH is mediated by tonic and surge centers in different nervous centers of the hypothalamus. The ventromedial and arcuate nuclei comprise the tonic center, which mediates the basal rel ease of GnRH. The tonic cent er yields high frequency, low amplitude pulses observed during th e luteal phase of the estrous cycle. In contrast, the suprachiasmatic and preoptic nuclei comprise the surge center, which mediates the low frequency, high amplitude pulses of GnRH associat ed with ovulation. The surge is preceded by estradiol release from the dominant follicle, en hancing the LH surge necessary for ovulation (Wettemann et al., 1972). In a study by Vizcarra et al. (1999), GnRH intravenously infused every hour resulted in more fre quent LH pulses compared to in fusion every 4 h. However, FSH secretion tended to be increased with infusion of GnRH every 4 h. In addition, Molter-Gerard et al. (1999) used GnRH immunizations and exogen ous GnRH injections in set frequencies to demonstrate that low GnRH pulse frequency enhanced the stor age of FSH such that basal concentrations are consistently available for follicul ar recruitment. They also demonstrated that higher frequency pulses of GnRH increased stor age of LH in preparation for the LH surge

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17 preceding ovulation. Findings from Vizcarra et al. (1999) and Molter-G erard et al. (1999) indicate that LH and FSH secr etion are differentially regulate d based upon the frequency and duration of GnRH stimulation. The transfer of GnRH from the hypothalamus to the anterior pituitary, where it acts to release LH and FSH, utilizes a unique system called the hypothalamo-hypophyseal portal system. Axons from nerve cells in the tonic and surge centers of the hypothalamus extend into the pituitary stalk and terminate on a capillary plexus where GnRH is released. This allows for transfer of minute amounts of GnRH to the ante rior pituitary. Without this system present, GnRH cannot effectively stimulate release of LH or FSH (Anderson et al., 1981). Follicle stimulating hormone f unctions to stimulate follicular maturation and production of the steroid hormone estradio l, through aromatization of andr ogens. Receptors for FSH are only located on granulosa cells of growing follicle s, acting to promote follicular recruitment, growth, and continued maturation. Follicles can grow to a diameter of 4 mm without the influence of gonadotropins. However, for con tinued growth to 9 mm , FSH stimulation is required, and to reach ovulatory size, LH pulses are required (Gong et al., 1996). Both FSH and LH are secreted together during th e mid-luteal phase of the estrous cycle. However, FSH is also secreted in separate pulses, and the pulse freque ncy declines from the earlyto mid-luteal phase of the estrous cycle (Walters et al., 1984). Luteinizing hormone is the dominant luteotr ophic factor in the bovine, and without LH, CL function ceases (Simmons and Hansel, 1964; Hoffmann et al., 1974). Additionally, LH is required for progesterone produc tion by the CL (Armstrong a nd Black, 1966). Luteinizing hormone has a very short half-life in circulation, and therefore it mu st be secreted in a pulsatile manner in order to effectively reach its target . Luteinizing hormone release is dependent upon

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18 pulsatile release of GnRH from the hypothalamus (Anderson et al., 1981). The release of GnRH and subsequent release of LH varies throughout th e estrous cycle (Walters et al., 1984). Pulse concentration and amplitude do not change from th e earlyto mid-luteal phases of the estrous cycle; however there is a 2 fold increase in freq uency of pulses in the ear ly-luteal phase (Walters et al., 1984). Rahe et al. (1980) reported a pulse frequency of 20 to 30 pulses/24 h in the early luteal phase and 6 to 8 pulses/ 24 h in the mid-luteal phase. A reduced LH secretion has been reported in ovariectomized Brahman cows compared to Hereford cows in response to a GnRH challe nge (Griffen and Randel, 1978). Similarly, Rhodes et al. (1978) observed a decrease in peak concentration of the LH surge afte r estradiol treatment in Brahman cows when compared to Hereford cows. In addition, Brahman cows exhibited a longer interval from exogenous estradiol treatm ent to LH peak when compared to Hereford cows. This may be a result of the Brahman pituitary having a decreased responsiveness to gonadotropin stimulation compared to Bos taurus breeds. Hormones produced by the ovary, including estr adiol and progesterone, also play key regulatory roles throughout the estrou s cycle. The pattern of estradio l release is similar to that of LH, and pulses of estradiol may be caused by pulses of LH (Walters et al., 1984). As progesterone concentrations dec line during the late-luteal phase of the estrous cycle, estrogen secretion begins to increase (Wettemann et al., 1 972). This rise in estrogen first reduces, but then increases the ability of the pituitary to re lease LH, thus causing the LH surge that leads to ovulation (Kesner et al., 1981).

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19 Estrous Cycle Overview The bovine estrous cycle consists of a seri es of events which includes the growth and development of a follicle, its subsequent ovulati on, and preparation of the uterus for conception, all to allow repeated attempts for pregnancy. The estrous cycle can be divided into two major parts, the follicular phase and luteal phase. The follicular phase includes the time from regression of the corpus luteum (CL) through ovulation, and can be furt her sub-divided into proestrus and estrus. The luteal phase includes the time from ovulation through regression of the CL and can be further sub-divided into metestrus and diestrus. Estrus is the most easily distinguished pha se in the bovine, as it is the time when the female is sexually receptive to the male. This behavior is caused by high concentrations of estradiol, the predominate hormone of the follicular phase. Estradiol increases about 3 d prior to estrus from approximately 3.6 pg/ml during the lut eal phase, to a peak concentration of 9.7 pg/ml the day before estrus (Wettemann et al., 1972). Estradiol concentr ations have been reported to be greater in Bos taurus (Angus) cows than Bos indicus (Brahman) cows for days 7 to 17 of the estrous cycle (Segerson et al., 1984 ). However, Alvarez et al. ( 2000) reported no differences in basal or maximum estradiol concentrations for Angus and Brahman cows. Elevated estradiol concentrations also regulate the surge of LH that leads to ovulation (Walters and Schallenberger, 1984). Ovulation occurs approximately 26 h after th e onset of behavioral estrus (Plasse et al., 1970; Pinheiro et al., 1998). The expression of estrus allows for copul ation to occur, and provides the opportunity for pregnancy. The primary and most reliable indicator of estrus is for a cow to allow herself to be mounted by another cow (Glencross et al., 1981). Secondary si gns of estrus include mounting

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20 other cows, clear mucous discharge from the vul va, swelling of the vulva, restlessness, following other cows, chin resting, and lip curling. These signs can occur before, during, or after estrus and do not relate to the time of ovulation, but shou ld instead be used as clues that a cow should be observed more closely (Nebel, 2003). The duration of estrus varies between cattle of Bos taurus and Bos indicus breeding, and may also vary for cows undergoing a spontan eous naturally occurring estrus versus a synchronized estrus. Richardson et al. (2002) observed that Bos taurus heifers which underwent a synchronized and a spontaneous estrus did not differ in duration. Rae et al. (1999) reported that the duration of a synchronized estrus to be 8.5 h for Angus ( Bos taurus ) and 6.7 h for Brahman ( Bos indicus ) heifers, while crossbred ( Bos indicus Bos taurus ) heifers had a duration of 11.9 h. Many factors, includi ng breed (Rae et al., 1999), envir onment (Landaeta-Hernandez et al., 2002), number of animals in estrus at a time (Hurnick et al., 1975; La ndaeta-Hernandez et al., 2002), and estrous detection method (Rae et al., 1 999) influence the duration of estrus; however, multiple studies indicate that Bos indicus heifers (Plasse et al., 1970) and cows (Randel, 1984; Pinheiro et al., 1998) have a shorter duration of estrus compared to Bos taurus heifers (Richardson et al., 2002) and cows (Schams et al., 1977). Metestrus is the stage following estrus from ovulation until the CL is fully functional, and generally lasts 3 to 5 d. Ovulation occurs during metestrus in cattle, approximately 24 h after the onset of estrus. In Bos taurus beef cows, ovulation occurs 24 to 36 h from the onset of estrus (Looper et al., 1998; Rorie et al., 1999). Plasse et al. (1970) reported that ovulation occurred 25.6 h after the beginning of estrus and 18.9 h afte r the end of estrus in Brahman heifers. Likewise, Pinheiro et al. (1998) re ported that ovulation occurred 26.6 h after the onset of estrus in Nelore ( Bos indicus ) cows. After the LH surge induces ovulation, the ovulated follicle

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21 undergoes cellular and tissue remodeling changes to form the CL. During metestrus, the predominate hormone switches from estradiol to progesterone produced by the CL. However, progesterone concentrations remain low as the CL be gins to develop. As the CL develops in size and progesterone output increases, the cow enters diestrus. Diestrus is the longest phase of the estrous cycle, lasting 10 to 14 d, and is characterized by increased progesterone secre tion. Progesterone concentrati ons from 1 to 13 ng/mL are observed when a CL is present in Bos indicus cows (Ruiz-Cortes and Olivera-Angel, 1998). In Bos taurus cows, progesterone concentrations from 4.7 to 12.0 ng/mL have been reported for cows with a CL (Henricks et al ., 1971). Regardless of whether or not a pregnancy occurs, the CL forms into a fully functional, progesterone-pr oducing structure. If an embryo is present in the uterus, the CL and high progesterone concentr ations are maintained throughout gestation. If pregnancy does not occur, the CL remains active until d 17 to 18 of the estrous cycle, at which time it undergoes luteolysis, which is stimulated by prostaglandin F2 secretion from the uterus. Progesterone concentrations rapidly dec line allowing for anot her estrous cycle. Proestrus occurs 3 to 4 d before estrus and is characterized by increased follicular growth. Following regression of the CL and a decline in progesterone, the predominate hormone becomes estradiol (Schams et al., 1998). During proestrus, a single dominate follicle emerges from a pool and is selected to grow into an ovulatory follicle which produces estradiol. The length of the estrous cycle is approximately 21 d in Bos taurus breeds (Hansel et al., 1973). Alvarez et al. (2000) re ported no difference in estrous cy cle length of Senepol (tropical Bos taurus ; 20.4 d), Angus (temperate Bos taurus ; 19.5 d), or Brahman (tropical Bos indicus ; 19.7 d) cows. However, Plasse et al. (1970) observed an estrous cycle length of 27.7 d in Brahman heifers evaluated over a full year, indicating that Bos indicus animals may have a

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22 longer estrous cycle. Length of the estrous cycl e may also differ between heifers and cows, as Plasse et al. (1970) reported an average length of 27.7 d in Brahman heifers, compared to Llewelyn et al. (1987), who obser ved a 23 d estrous cycle in Boran cows. However, in Bos taurus beef cows (Zollers et al., 1989) and heifer s (Mihm et al., 2000), estrous cycle length has little variation from 21 d. Estrous Detection Estrous detection is the process of determining if an animal is in estrus and is primarily conducted by visual observation. Cows display es trus following proestrus, when they have a large follicle producing adequate estradiol concentrations to induce estrous behavior. Estrus can be spontaneous and naturally occurring, or resu lt from initiation of luteolysis by administration of prostaglandins. Detection of estrus can be one of the mo st challenging aspects of an artificial insemination (AI) program. The primary underlyi ng changes in the cow that these methods of detection rely on include increased physical activity and locomotion (Farris et al., 1954), swelling of the vulva (Foote et al., 1979), a nd most importantly standing to be mounted by another cow. Pedometry is a method of estrous detecti on which monitors the physical activity and locomotion based upon the number of steps a cow ta kes over a given period of time. Cows may be four times as active during estrus than any other phase of the estrous cycle (Kiddy, 1977). This is monitored by a mechanically activated leg band that collects the information, which is later evaluated by a reader (Senger, 1994). Using this information, the optimal time for insemination can be predicted (Maa tje et al, 1997). Pedometry is predominately used in dairies,

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23 as its application to beef operati ons is not practical due to the frequency of which the pedometer would need to be read. Another method of estrous dete ction is electrical resistance of reproductive tissues. This method monitors changes in the el ectrical resistance of fluids us ing a probe inserted into the vagina. Electrical resistance of vaginal fluids is indicative of th e stage of the estrous cycle the cow is in (Foote et al., 1979). These change s are notable due to vulvar mesenchymal tissue being 74% heavier during estrus than during dies trus because of tissue hydration (Ezov et al., 1990). This method of estrus detection is not co mmonly utilized in beef operations due to the frequency of which cows would have to be worked through a handling facility. Therefore, estrous detection in beef operati ons relies primarily on the behavior of cows during estrus by allowing themselves to be mo unted by other cows, which is also the most reliable indicator of estr us (Glencross et al., 1981). To dete rmine whether or not a cow in estrus has been mounted by another cow, a variety of products are available to producers including paints, pressure sensitive patches, scratch-o ff patches, and computer transponders. These products are affixed to the area just anterior to the cow’s tail hea d, where they will be activated by the mounting cow. Painting of the tail head, used in conj unction with visual obs ervation, is the least expensive of these products, and has been indicat ed to yield similar conception rates to more expensive alternatives (Xu et al., 1998). Another product, the Kamar Heatmount Detector (Steamboat Springs, CO), is a pressure sensitive device, which releases red ink when activated and is visible from a distance on the cow. The device contains a timing mechanism which requires approximately three seconds of mounting to change from white to red. However, the

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24 Kamar has been reported to yield false pos itive readings about 13.2 % of the time (Gwazdauskas et al., 1990). A newer product on the market is the Estrus Alert Heat Detector (Western Point, Inc., Merrifield, MN), a scratch-off patch in the form of a sticker. This product features a silver, scratch-off surface, similar to that of a lottery ticket, which scratc hes away to reveal a brightly colored signal layer. Unlike the Kamar, the Estrus Alert allows a producer to determine if a cow has received only a few mounts compared to a large number of mount s based on the degree to which the patch is scratched off. The HeatWatch (DDx, Inc., Denver, CO) system was the first remote sensing system to allow 24 hour monitoring of standing events asso ciated with estrus (Neb el, 2003). This system utilizes a computer tran sponder tucked into a mesh patch, which is affixed to the tailhead of the cow. The transponder records data for each individual cow, includ ing date, time, and duration of each standing event. This information is transmitted to a computer where the HeatWatch software develops management and i ndividual cow reports (Nebel, 2003). Teaser animals can also be used to aid in th e detection of estrus. These include peniledeviated, epididymectomized, vasectomized, or other bulls incapable of impregnating a female, as well as androgenized steers and cows, all of wh ich can be outfitted with chin ball markers to aid in the detection of cows in estrus (Burns and Spitzer, 1992). The chin ball marker works like a ball point pen, leaving ink marks on th e dorsal surface of the cow in estrus. Breed differences are also apparent in the onset of estrus, ability to detect estrus, duration of estrus, time of day estrus occu rs, and the intensity of estrus. Rae et al. (1999) observed estrus durations for Angus ( Bos taurus ) heifers to be approximately 8.5 hours, while Brahman ( Bos indicus ) heifers were only 6.7 h. They also observed crossbred ( Bos indicus Bos taurus )

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25 heifers had an average estrus dur ation of 11.9 h. Other studies have observed estrus durations of 11 to 12 h in synchronized Bos taurus heifers (Richardson et al., 2002). Pinheiro et al. (1998) observed a 10.9 h estrus duration in Nelore ( Bos indicus ) cows. Rae et al. (1999) reported that breed had a significant effect on mounting activity during a s ynchronized estrus in Angus, Brahman, and Angus Brahman, receiving an average of 19, 25, and 37 mounts, respectively. Additionally, estrus may be mo re difficult to detect in Bos indicus cattle. Behavi oral estrus in Bos indicus breeds may be expressed more by secondary signs of estrus, such as head butting (Galina et al., 1982; Orih uela et al., 1983). Bos indicus cattle may also have an increased occurrence of silent heats and ab sence of visual estrus (Dawuda et al., 1989; Lamothe-Zavaleta et al., 1991), but comparisons to Bos taurus cows were not made in these studies. Synchronization of estrus promotes the form ation of sexually-active groups, which in turn increases the moun ting activities of both Bos taurus and Bos indicus cows (Hurnick et al., 1975; Landaeta-Hernandez et al., 2002) compared to a spontaneous, naturally occurring estrus. Social status in the herd can also effect behavioral estrus. Landaeta-He rnandez et al. (2004) observed that dominate cows tended to receive le ss mounts than subordinate and intermediate cows, due to the dominate cows avoiding being mounted by subordinate cows. For this reason, dominate cows may take longer to be detected in estrus. The time of day which cows display estrus can also be influenced by breed. It has been indicated that the onset of estrus occurs more frequently during the nighttime hours than daytime hours in both Bos taurus (Stevenson et al., 1996) and Bos indicus (Pinheiro et al., 1998) cows. However, while Landaeta-Hernandez et al. (2002) observed more cows initiate estrus during night hours (60%) than day hours (40%), they observed more Senepol ( Bos taurus ) cows initiate estrus during the daytime hours than Brahman cows.

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26 The environment the cow is in can contribute to the number of mounts the cow receives. Cows that are consuming feed, in a crowded pe n or aisle, or housed on poor footing will demonstrate less mounting activity (Britt et al., 1986). Cows which have foot problems will display less mounting activity rega rdless of whether the problem is structural, subclinical, or clinical (Leonard et al ., 1994). During a synchronized estr us, mounting activity is not effected by whether another cow or teaser bull is moun ting the estrus cow (Pinheiro et al., 1998). However, the same study indicated that cows un dergoing a natural estrus received more mounts from a bull than another cow. Follicular Development Overview Like many mammalian species, cattle are born with a finite amount of oogonia available during their lifetime. Oogonia ar e developed during mid-gestati on when the primordial germ cells migrate to the gonadal ridge and are arre sted in prophase I of meiosis and stored in primordial follicles (Hirshfield et al., 1991). It has been hyp othesized that granulosa cells modulate the transcriptional activity of the ooc yte genome (De La Fuente and Eppig, 2001). Around the time of puberty, these inhibitory f actors are removed by LH stimulation, and just prior to ovulation meiotic divisions resume (Senger, 1999). Oogonia stored in the ovary awaiting signals to grow are maintained in a quiescent stage and are known as primordial follicles. The primordial follicle is characterized by a single layer of granulosa cells surrounding the oocyte, and from this stage follicles gradually and continually leave the resting pool to begin growth (Fortune, 1994). These follicles can be in a quiescent state, with a single layer of flattened granulosa cells, or in an active state, with a single layer of flattened, and some cuboidal cells (Fair et al., 1997b). A quiescent, primordial follicle will have

PAGE 27

27 5 to 8 flattened granulosa cells; however, when the follicle is activated, it matures to greater than 8 cells and the number of flattened cells declines rapidly as they become increasingly cuboidal (Braw-Tal, 2002). At this stage of follicular growth, th e cells are still tran scriptionally inactive (Fair et al., 1997b). The mechanism by which these follicles are gradually released and signaled to grow is unknown (Fortune, 1994). Once the oocyte reaches an 18-cell stage, it is completely surrounded by cuboidal granulosa cells and is termed a primary follicle (Braw-Tal, 2002). As the number of cuboidal granulosa cells grow, they begi n producing mRNA for the protein follistatin (Braw-Tal, 1994). Follistatin acts to inhibit activin A, thereby protecting the growing follicle from the growthinhibiting effects of activin A (Braw-Tal, 1994) . Additionally, the ooc yte begins to produce growth differentiation factor 9 (GDF9) a nd bone morphogenic protei n 15 (BMP15), both of which are structurally similar to transforming growth factor(TGF ) and expressed exclusively in the oocyte (McGrath et al., 1995; Dube et al., 1998) to increas e granulosa proliferation (BrawTal, 2002). At about the same time, the surround ing granulosa cells begin to produce kit ligand to promote oocyte growth (Braw-Tal, 2002). The oocyte continues to grow a second layer of cuboidal gran ulosa cells, and is termed a secondary follicle (Fair et al ., 1997b; Braw-Tal, 2002). At th is stage, the oocyte becomes transcriptionally active (Fair et al., 1997b), develops the zona pellucida, and forms gap junctions between the oocyte and granulosa ce lls (Fair et al., 1997a). Hence, the follicle moves from being dependent on intraovarian signals to being an autonomous unit wher e its growth is regulated only by follicle-produced factors (Braw-Tal, 2002). More layers of granulosa cells are added and a cavity or antrum begins to surround the oocyte, at wh ich point it is termed a tertiary follicle. The antrum begins to accumulate follicular fluid a nd the size of the follicle begins to increase.

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28 Granulosa cells differentiate into inner, cu mulus cells and outer, mural cells (Vanderhyden, 2002). The tertiary follicle continues to grow into a mature Graafian follicle capable of ovulation. From this stage, the follicle will gr ow continuously until it reaches one of two fates, atresia or ovulation (Fortune, 1994). Once follicles have developed to a graafian st age, they begin to progress through stages of recruitment, selection, and dominance. In cattle, follicles grow in a wave-like pattern to ensure that at any given time there is an e ligible follicle available for ovulation (Sirois and Fortune, 1988). Typically, there ar e two to three follicular waves per estrous cycle, with some studies showing more two-wave cycles (Ginther et al., 1989), and others showing more threewave cycles (Savio et al., 1988; Sirois and Fo rtune, 1988). In two-wave cycles, the waves begin around days 2 and 11 of the estrous cycle, whereas in cows with three-wave cycles, the waves begin around days 2, 9, and 16 of the estrous cy cle (Sirois and Fortune, 1988). The number of follicular waves a cow has may be related to the level of dominance, or progesterone output, of the CL, with a three-wave cow having a more do minant CL than a two-wave cow. Townson et al. (2002) observed that cattle experiencing tw o follicular waves ovulated larger, older, and potentially less fertile follicles than cattle ovul ating after three follicular waves, as shown by their subsequent pregnancy rates. This study co nfirmed the trend of higher pregnancy rates in three-wave cows over two-wave cows seen by Ahmad et al. (1997), as well as the theory by Mihm et al. (1994) that a few days longer gr owth prior to ovulation compromises pregnancy rates. Atresia is a form of apoptosis, or programm ed cell death. Atresia is the most common fate of follicles, as only a select number will actually achieve dominance and ovulate. While atresia can occur at any stage of follicular growth, it is not evenly distributed across follicular

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29 development. In cattle, atresia is greatest just before the final stages of follicular dominance (Fortune, 1994). For a follicle to grow from a 300 m antral foll icle to a 3 to 5 mm follicle detectable via ultrasound, it is calculated to take more than 30 d (Lussier et al., 1987). Up to this point in development, follicles can grow independently of gonadotropin stimulation (Scaramuzzi et al., 1993). Recruitment begins with a rise in FSH, wh ich stimulates the growth of a cohort of small follicles beyond 4 mm in diameter (Adams et al ., 1992; Ginther et al., 1997). As FSH declines, fewer follicles continue to grow, while others undergo atresia (Austin et al., 2001). In a study by Austin et al. (2001), they obser ved that by about 33 h post-FSH peak, there were two follicles still growing, and by 53 h a dominant follicle coul d be identified and measured at 8.5 mm. This dominant follicle grows between 12 to 20 mm an d has enhanced estradio l and inhibin production to prevent growth of another cohort (Ginther et al., 1999). The dominant follicle continues to grow for an additional 3 to 4 d before it regres ses from the negative feedback of progesterone on LH (Sunderland et al., 1994). The decline in estradiol from the fo rmer dominant follicle causes another FSH rise and another wave begins (Sunderland et al., 1994). If luteolysis occurs during the time of dominance of the second dominant follicl e, it will ovulate; however, if luteolysis does not occur during dominance, the follicle regresses and another follicular wave is initiated (Cooke et al., 1997). Breed may also effect follicular growth and deve lopment in cattle. It has been shown that non-lactating Brahman cows had significantly greater numbers of small (2 to 5 mm), medium (6 to 8 mm) and large ( 9 mm) follicles when compared to non-lactating Angus and Senepol cows (Alvarez et al., 2000). However, there was no difference in the occurrence of two vs. three follicular waves or estrous cycle length among the breeds. Bos indicus commonly have estrous

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30 cycles with up to four follicular waves in both heifers (Rhodes et al., 1995) and cows (Zeitoun et al., 1996; Martinez et al., 2003). A study by Viana et al. (2000) showed Gir ( Bos indicus ) cattle experienced mostly three (60%) and four (26.7% ) follicular waves. However, the number of follicular waves did not affect the estrous cycl e length. Numerous studies have also noted a smaller dominant follicle size of 10 to 12 mm in Bos indicus cows on the last follicular wave (Bo et al., 1993a,b; Figueiredo et al., 1997; Rhodes et al., 1995a), compared to 14 to 20 mm in Bos taurus cows (Ginther et al., 1989; Kastel ic et al., 1990; Bo et al., 1993b). Corpus Luteum Function and Luteolysis Overview The corpus luteum is a heterogeneous struct ure on the ovary with a unique population of cells with different morphological, endocrinol ogical, and biochemical properties (Niswender and Nett, 1988). It is comprised of endothelial cells, small luteal cells , large luteal cells, fibroblasts, smooth muscle cells, immune cells , and pericytes (O’Shea et al., 1989; Farin et al., 1986; Hansel et al., 1991). There are two types of steroidogeni c cells in the CL, known as small luteal cells and large luteal cells. These cells produce proge sterone when the animal is both cycling and pregnant (O’Shea, 1987). The CL is formed from the granulosa and theca cells of the antral follicle (O’Shea, 1987). Preparation for the transformation of these cells to progesterone producing luteal cells is initiated prior to ovulation (McNatty and Sawers, 1975), bu t can occur independently of ovulation (Kesler et al., 1981). This transformation is termed lute inization and is characterized by the change from production of estradiol to progest erone (Juengel and Niswender, 1999). The granulosa cells of the follicle differentiate into large luteal cells (O ’Shea, 1987), and are stimulated to produce progesterone primarily by growth hormone (Lie bermann and Schams, 1994). Growth hormone

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31 receptors on the CL are located primarily on large luteal cells (Lucy et al., 1993). The theca cells of the follicle differentiate into small lute al cells (Donaldson a nd Hansel, 1965), and are stimulated to produce progesterone primarily by LH (Niswender and Nett, 1988). Luteinizing hormone receptors on the CL are primarily locat ed in small luteal cells. The mechanism by which LH stimulates progesterone production in volves formation of cAMP and activation of protein kinase A (Schams and Beri sha, 2004). It has also been s uggested that large luteal cells may differentiate into small lu teal cells (Fisch et al., 1989) and small luteal cells may differentiate into large luteal cells (Cran, 1983). Luteal structur es that appear to be in an intermediate stage between the tw o have been identified (Priedkaln s et al., 1968). Small luteal cells are primarily responsible for higher ma gnitude, LH-stimulated progesterone production, whereas large luteal cells are responsible for ba sal progesterone secretion (Hansel et al., 1991). The actions of prostaglandin F2 (PGF2 ), are primarily mediated by the large luteal cells, as they contain most of the PGF2 receptors (Pate, 1994). After the LH surge and ovulation, the walls of the follicle fold in (O’Shea et al., 1980), granulosa cells hypertrop hy (Fawcett et al., 1969), and gap junc tions between granulosa cells are reduced (Murdoch, 1985) to form the CL. Puls atile secretion of LH is required for proper formation of the CL from days 2 to 7 of the es trous cycle and for full fu nctionality of the CL from days 7 to 12 of the estrous cycle; however, it is not necessary for the CL to be maintained and produce normal circulating progesterone concen trations past d 12 (Pet ers et al., 1994). The CL is one of the few tissues capable of angiogenesis, or the generation of new blood vessels from existing blood vessels by migration and proliferation of endothelial cells (Schams and Berisha, 2004). The development of this micro-circulatory system involves the breakdown of the follicular basement membrane, endothelial cell proliferation, and development of capillary

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32 lumina (Smith et al., 1994). The precise contro l of angiogenesis is essential for normal luteal function, and once complete, the CL becomes one of the most highly vascularized organs in the body, and has the greatest rate of blo od flow (Wiltbank et al., 1988). Differences between breed types have been observed for CL function, CL weight, and progesterone output. Rhodes et al . (1982) reported that Hereford Holstein heifers displayed heavier CL weights and had grea ter progesterone output than Brah man heifers. However in the same study, systemic progesterone concentrations did not differ between the breeds. In contrast, Segerson et al. (1984) reported great er mean progesterone concentra tions from d 7 to d 17 of the estrous cycle for Angus (5.3 ng/mL) than Brahma n (4.1 ng/mL) cows. Corpus luteum size has been reported to be 17 to 21 mm in diameter in Bos indicus cows (Bo et al., 1993a,b; Figueiredo et al., 1997; Rhodes et al., 1995a). In Bos taurus cows, CL size has been reported to be 20 to 30 mm (Ginther et al., 1989; Kastelic et al., 1990; Bo et al., 1993b) . Rhodes et al. (1982) also indicated that Brahman heifers ma y show a more seasonal effect of CL progesterone content than Hereford Holstein heifers, with summer CL progesterone content being lower than in the winter; however, systemic progesterone concentrat ions did not differ for season or breed. In contrast, Zeitoun et al. (1996) reported higher ci rculating progesterone concentrations for Brahman cows in the spring (6.3 ng/mL) than in th e fall (4.8 ng/mL). It has also been reported that CL of Brahman cattle are more deeply im bedded in the ovarian stroma and can be more difficult to detect by rectal palpation (Rhodes et al., 1982; Plasse et al., 1968). Luteolysis Prostaglandin F2 is the main luteolytic agent in the bovine and initiates CL regression (Gooding et al., 1972; Inskeep, 1973). At the end of the estrous cycle, the CL is regressed by episodic release of PGF2 from the uterus. The PGF2 is transferred from the uterus to the ovary

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33 where it acts on the CL by a unique counter-current exchange system between the uterine vein and ovarian artery (McCracken et al., 1972). This subsequently causes a marked decrease in progesterone concentrations between d 16 and 19 of the estrous cycle (Hansel et al., 1973) and leads to behavioral estrus and ovulation (Thatcher and Chenault, 1976). Prostaglandin F2 has a very short half-life and is rapidly inactivated by oxidation in the bloods tream from a single pass through the lungs (Kindahl, 1980). For this reaso n, a counter-current exchange system evolved to transport PGF2 from the uterus, by the uterine arter y, to the ovary and return by the ovarian vein, thus preventing the effective amount of PGF2 from being lost in systemic circulation (McCracken et al., 1972). McCracken et al. (1972) also discovered that infusion of as little as 25 g/hr of PGF2 into the artery leading to the ovary resu lted in a rapid decrease in progesterone secretion; however, the sm all concentrations of PGF2 were ineffective at inducing luteolysis when administered into pe ripheral circulation. It has been shown that exposur e to, or inhibition of, progest erone regulates the timing of release of PGF2 , and subsequent CL regression (Ottobr e et al., 1980; Schams et al., 1998). In vivo studies have shown that pr ior to d 12 of the estrous cy cle progesterone inhibits PGF2 release (Schams and Berisha, 2004). Additiona lly, progesterone exposure to the endometrium enhances synthesis of PGF2 and has a priming effect on luteolys is (Boshier et al., 1981). After d 12 the luteal tissue begins to lose its sensitivity for progesterone and PGF2 is no longer inhibited. Progesterone down-re gulates its own receptors within the endometrium, thereby decreasing its own actions, and causing an increase in the actions of estradiol (Robinson et al., 2001). The decrease in progesterone receptors, a nd increase in estrogen receptors leads to an increase in oxytocin receptor s (Vallet et al., 1990). The in crease in estradiol causes high frequency, low amplitude oxytocin release fr om the hypothalamic oxytocin pulse generator

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34 (McCracken et al., 1999) and enhances oxytocin r eceptors in the uterus (Meyer et al., 1988). Oxytocin produced from both the posterior pituitary and endometrium activates sub-luteolytic PGF2 concentrations from the uterus (Hooper et al., 1986). The release of uterine PGF2 acts on PGF2 receptors in luteal cells to further stimul ate the release of oxytoc in (Hooper et al., 1986). These events culminate to cause secretion of larg e amounts of PGF2 from the uterus and increases in oxytocin to induce luteolysis. All of these events c ontinue until the PGF2 receptor response system is desensitized and luteal oxytoci n release ceases (McCra cken et al., 1999). Regression of the CL involves both functi onal and structural degradation. The progesterone producing capability of the CL is decreased rapidly at luteolysis by several mechanisms including the decrease in gene e xpression for steroidogenic acute regulatory protein (StAR), 3 -hydroxysteroid dehydrogenase (3 -HSD), and LH-receptor, all of which are important in maintaining lute al function (Tsai et al., 2001). Steroidogenic acute regulatory protein is an essential co mponent in the regulation of steroid biosynthesis, 3 -HSD is a key enzyme in the production of steroid hormones, and reduction in LH-receptor inhibits steroidogenesis. Other stimulat ory factors for functional degrad ation include the up-regulation of gene expression for c-fos , prostaglandin G/H synthase -2 (PGHS-2), and monocyte chemoattractant protein-1 (MCP-1), all of which are induced by PGF2 (Tsai et al., 2001). The protein cfos, is a primary response gene which regulat es protein synthesis and release, PGHS-2 is an enzyme which metabolizes arachidonic acid to form important intermediates to prostaglandin production, and MCP-1 is an infl ammatory mediator which increases macrophages within the CL during luteal regr ession. Physical degr adation includes a decrease in cytoplasmic granulation, rounding of the cell out line, peripheral vacuolation of the large luteal cells, condensation of the cytoplasm, loss of prominent nuclei, and an increase in prominence of

PAGE 35

35 connective tissue (Hansel et al., 1973). As the CL ages, artery walls thicken and begin to deteriorate, which contributes to declining progesterone concen trations (Donaldson and Hansel, 1965). The vasoconstrictive properties of PGF2 may also cause decreased blood flow through the CL to contribute to lut eal regression (Schams and Beri sha, 2004). Collectively, these processes cause the sharp declin e of progesterone concentrations, allowing for an increase LH pulse amplitude and frequency in response to ri sing estradiol concentrat ions (Chenault et al., 1975) The actions of exogenous PGF2 are not effective for luteolys is during all stages of the estrous cycle (Lauderdale, 1972; Henricks et al., 1974). Early (d 7) in the estrous cycle, PGF2 is less effective at inducing luteol ysis than later in the cycle (d 15) (Tanabe and Hann, 1984). It was hypothesized that this difference was due to a lack of luteal PGF2 receptors, however this was disproved by Wiltbank et al. (1995) who reported that PGF2 receptor concentration and affinity for both developing (days 2 to 4 of the es trous cycle) and active CL (days 6 to 10 of the estrous cycle) were similar. Therefore, furthe r studies by Silva et al. (2000) investigated the ability of PGF2 to be converted to its inac tive form, 13,14-dihydro-15-keto PGF2 (PGFM), by the enzyme 15-hydroxyprostaglandin dehydrogenase (PGDH). The amount of PGDH increased during the estrous cycle and pregnancy when PGF2 was not an effective luteolysin, but did not increase during the times when PGF2 was an effective luteolysin. Furthermore, Silva et al. (2000) reported that the amount of mRNA for PGDH was greater on days PGF2 was not effective and significantly lower when PGF2 was effective. They also observed that concentrations of cyclooxygenase (COX-2) incr eased on similar days, but were undetectable when PGF2 was effective.

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36 Another proposed reason fo r the ineffectiveness of PGF2 to induce luteolysis early in the estrous cycle is the lack of endot helin-1 (ET-1), a promoter of lu teolysis that appears in large quantities in the CL just after PGF2 exposure (Wright et al., 2001). Levy et al. (2000) reported low concentrations of ETconverting enzyme (E TE-1), the enzyme which converts ET-1 to its active state, in the luteal tissue early in the es trous cycle. They subsequently observed a fourfold increase in ETE-1 during the transition from the early to mid-luteal phase of the estrous cycle, indicating the lack of ac tive ET-1 early in the estrous cy cle may contribute to why the early developing CL is not susceptible to PGF2 -induced luteolysis. Estrous Synchronization Overview Estrous synchronization is a powerful manageme nt tool that allows producers to have a large number of cattle in estrus during a short pe riod of time to make artificial insemination more practical. Through the use of hormone treatmen ts, the estrous cycle can be manipulated in several ways. Prostaglandin F2 can be used to shorten the lu teal phase and induce estrus in cows with a functional CL. Progestins can le ngthen the luteal phase and prevent ovulation during the treatment period, which is generally 7 to 14 d. Finally, GnRH can induce ovulation, either to synchronize follicular waves, or to induce ovulation for a timed-AI protocol. Synchronization of the estrous cycle can be achieved in several ways including synchronization of estrus, synchronization of folli cle wave growth, and (or) a combination of both. The combination of these two synchroni zation schemes can yield a tight synchrony of estrus that allows producers to e ither inseminate after an observed estrus or a timed-AI protocol. In order to synchronize estrus or follicle development, exogenous administration of GnRH, PGF2 , and progestins can be used either alone or in conjunction in development of effective

PAGE 37

37 estrous synchronization protocols. The different protocols must be matched to an individual producer’s needs, facilities, and labor resources. Prostaglandin F2 Prostaglandin F2 functions to regress the CL causi ng a sharp decline in progesterone production, which allows for estradio l to increase and the cow to s ubsequently exhib it estrus 3 to 5 d later. In order for PGF2 to exert its luteolytic effects, the cow must have a functional CL (Rowson et al., 1972). Prostaglandin F2 and its analogs are less eff ective at causing luteolysis during the early luteal phase of the cycle (Rowso n et al., 1972; Lauderdale et al., 1974). In a study by Chenault et al. (1976), progesterone co ncentrations were d ecreased to < 0.5 ng/mL within 24 h after PGF2 administration. Exogenous PGF2 administration can be used to initiate luteolysis without compromising the fertility of the subsequent estrus (Lauderdale et al., 1974). Stage of the estrous cycle also has a significan t effect on the both the expression of estrus and the interval from PGF2 administration to the onset of estrus . Kiracofe et al. (1985) observed that when PGF2 was administered on days 1 to 4 of th e estrous cycle very few displayed estrus and when inseminated had significantly decreased fertility. Tanabe and Hann (1984) observed estrus in 86.0, 90.0, and 98.0% of da iry heifers administered PGF2 on d 7, 11, and 15 of the estrous cycle, respectively. They also identified a significant difference in the interval to onset of estrus, with d 7 heifers displa ying estrus earliest (43.9 h), d 11 he ifers the latest (71.5 h), and d 15 heifers intermediately (53.0 h). However, the synchrony of estrus over the 24 h interval from 32 to 56 h was 88.4, 13.3, and 73.5% for d 7, 11, and 15, respectively. Similarly, Watts and Fuquay (1985) reported estrous responses of 43.0, 83.6 and 100% for cows administered PGF2 on days 5-7, 8-11 and 12-15, respectively. Within the same treatment groups, they also observed an average interval to estrus of 59, 70, and 72 h. Additionally, King et al. (1982) observed that

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38 the stage of cycle when PGF2 is given influences the interval to estrus in both heifers and cows, with cows early in the estrous cycle (d 5 to 9) showing estrus earlier following PGF2 than cows late in the estrous cycle (d 10 to 15). They also indicated that the interval to estrus after PGF2 is shorter for heifers than cows. In a large New Zealand study of dair y herds using a single dose of PGF2 , cows treated on either d 7 or d 15/16 of the estrous cycle dem onstrated the most preci se synchrony of estrus (Macmillan and Henderson, 1984). In the same study, cows which received PGF2 on days 12 or 13 showed the most variable estrous re sponse, and cows which received PGF2 on days 15 or 16 were the most likely to be detected in estr us within 10 d. This, along with other studies, demonstrates the significant effects that stage of the estrous cycle has on the subsequent interval to estrus in both Bos taurus (Macmillan and Henderson, 1984; Stevenson et al., 1984) and Bos indicus cattle (Cornwell et al., 1985). Differences in the synchrony of estrus following PGF2 administered at different stages of the estrous cycle result from different stages of follicular development. If the cow has a dominant folli cle in its growing phase at the time of CL regression by PGF2 , the interval to estrus will be shorter compared to a cow which has a regressing follicle and will, therefore, have to recruit and mature a new follicle before ovulation can occur. Conception rates appear to be effected by stage of the estrous cycle, although the data are somewhat conflicting. Watts and Fuquay ( 1985) observed concepti on rates of 56.8, 62.1, and 78.3% for heifers administered PGF2 on d 5 to 7, d 8 to 11, and d 12 to 15 of the estrous cycle, respectively. In contrast, other studies have observed no difference in pregnancy rates when PGF2 was administered duri ng the early or late luteal phase of the estrous cycle (Stevenson et al., 1984). In a study by Diskin et al. (2001), it was indicated that stage of cycle effects may be

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39 less pronounced in dairy heifers because they have a shorter interval to estr us with less variation than cows. There is some evidence to suggest that breed type may also have an effect on the efficacy of PGF2 . Initial work reported a considerable difference in estrous response after a PGF2 induced estrus. In Bos taurus cattle, estrous responses of 70 to 95% have been reported after a single dose of PGF2 (Tanabe and Hann, 1984; Watts and Fuquay, 1985). However, estrous responses of 46 to 62% (Landivar et al., 1985; Ori huela et al., 1983) were observed after a single dose of PGF2 in Bos indicus cows. In a study by Pinheiro et al . (1998) with a group of Nelore ( Bos indicus ) cows and heifers, they reported an estrous response of only 46.4 and 33.3% respectively, following two doses of PGF2 11 d apart. Additional work suggests that the decrease in estrous response maybe due to a decreased effectiveness of PGF2 in cattle of Bos indicus breeding. In Bos indicus cattle, the decreased estrous re sponse may be due to inhibition of estrus expression caused by incomplete re gression of the CL, and therefore elevated progesterone concentrations (Pi nheiro et al., 1998; Rekwot et al., 1999). Cornwell et al. (1985) reported estrous responses of only 50 and 67% for Brahman heifers treated with PGF2 on days 7 and 10 of the estrous cycle. However, in he ifers that did not disp lay estrus, progesterone concentrations decreased initially, but began to increase to pre-treatment concentrations within 48 h after PGF2 . This effect has also been observed in Bos taurus cattle (Chenault et al., 1976; Copelin et al., 1988). To prevent the partial luteolysis in Bos indicus cows, the dose of PGF2 has been split into two consecutive doses, administered 24 h ap art (Cornwell et al., 1995). Administration of the second dose sustains the luteolytic actions of PGF2 and prevents the rebounding of progesterone concentrations caused by incomplete CL regression. In two recent studies in Bos

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40 indicus females, the dose of PGF2 was split into two, half doses , administered 24 h apart in heifers (Bridges et al., 2005) and cows (Por tillo et al., 2006). Th e split-dose of PGF2 increased estrous response, timed-AI pregnancy rate, a nd synchronized pregnanc y rates for heifers (Bridges et al., 2005) as compared to a single dose, by increasing CL regression. While CL regression was enhanced in mature cows, th ere was no difference in estrous response or synchronized pregnancy rate s (Portillo et al., 2006) . A split-dose of PGF2 does not appear to be necessary in Bos taurus cows, as high CL regression rates (85 to 100%) are already achieved in these cows (King et al., 1982; Tanabe and Hann, 1984; Kirakofe et al., 1985). Two major chemical classes of PGF2 are commercially available, dinoprost tromethamine and cloprostenol sodium. Dinopr ost tromethamine is a naturally occurring prostaglandin marketed under the names of Lutalyse, Prostamate, and In-Synch, and is administered as a 5 mL (25 mg dinoprost) intramuscular injecti on. Cloprostenol sodium is a synthetic, structurally similar analogue of PGF2 marketed under the names of Estrumate and Estroplan, and is administered as a 2 mL (500 g cl oprostenol) intramuscular or subcutaneous injection. It has been used to synchronize es trus in cows for many years and has resulted in acceptable pregnancy rates (Johnson, 1978; Seguin et al., 1983; Jackson et al., 1979). The comparison of cloprostenol sodium and dinoprost tromethamine to synchronize estrus has been well documented (Young and Anderson, 1986; Salverson et al., 2002) in Bos taurus cattle, but few comparisons of their effectiveness in cattle of Bos indicus breeding have been conducted (Hiers et al., 2003). Hiers et al. (2003) obser ved a 5% numerically greater timed-AI pregnancy rate with cloprostenol sodium over dinoprost tromethamine in Bos indicus Bos taurus cows synchronized with GnRH + PGF2 system combined with melengestrol acetate (MGA). However, in Bos taurus heifers, Salverson et al. (2002) obser ved no difference in the efficacy of

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41 cloprostenol sodium and dinoprost tromethamine when used 19 d after a 14 d MGA treatment. More consistent conception rates are achieved with PGF2 when exogenous progesterone is administered in the immediate pre-inj ection period (Macmillian et al., 2003). Progestins Exogenous progestins mimic the luteal phas e of the estrous cy cle and prevent the expression of estrus. Progestin s are commonly used to artific ially extend the estrous cycle during an estrous synchronization protocol. Se veral types of progestins are used including natural progesterone and synthetic progestins including norgestomet and MGA. The two most commonly used progestins for estrous synchroni zation include the controlled intravaginal progesterone-release de vice (CIDR), which contains proge sterone, and MGA, a feed through progesterone. The CIDR is a T-shaped intravaginal insert which is made up of a nylon spine covered by an elastic, silicone coating. The CIDR available for commercial use in the United States contains 1.38 g of progesterone and has been successfully used to synchronize estrus in both heifers and postpartum beef cows (Lucy et al., 2001). The typical dur ation of use of the CIDR is 7 days with an injection of PGF2 either the day before CIDR re moval (Lucy et al., 2001) or the day of CIDR removal to induce lute olysis (Larsen et al., 2006). It is estimated that approximately 0.7 g of progesterone are used durin g its initial use (Savio et al., 1993) suggesting that there is a potential for reuse of the CIDR an additional time. Melengestrol acetate is an orally-administered progestin, and when admi nistered at a rate of 0.5 mg/head/d it effectively suppresses estrus (Zimbelman a nd Smith, 1966). Treatments of progestins generally range from 7 to 14 d in leng th, and when administered for periods greater than 9 d, they lead to a reduction in fertility of the subsequent ovulation (Hill et al., 1971). This

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42 reduction in fertility is caused by the developm ent of what is termed a persistent dominant follicle (Guthrie et al., 1970). Melengestrol aceta te is most commonly used in heifers, and is administered for 14 d, followed by an injection of PGF2 17 (Brown et al., 1988) to 19 d (Lamb et al., 2000) after M GA withdrawal. Ireland and Roche (1982) observed that the c oncentrations of exogenously administered progesterone were negatively correlated with an an imal’s ability to secrete LH. Cows receiving subluteal concentrations of progesterone have an increased frequency of LH pulses during the course of treatment (Roberson et al., 1989). Th is insufficient concentr ation of circulating progesterone allows LH frequency to increase, es tradiol to rise, and follicles to experience a prolonged period of dominance (Kojima et al., 1992). This pattern of LH s ecretion prevents the wave-like growth seen in a norma lly cycling cow, resulting in a single follicle that grows and remains dominant with no further follicle recrui tment for the duration of progestin treatment (Sirois and Fortune, 1990). Depe nding on the length of progestin treatment, the follicle may be aged and less fertile upon ovulation, with the pe riod of dominance being directly correlated to the reduction in fertility (Mihm et al., 1994). When low concentrations of progesterone are administered to inhibi t estrus over extended (> 9 d) periods of time, it has been observed that follicles grow to a larger maximum size (Zimbelman and Smith, 1966), persist on the ovary (Trimberger and Hansel, 1955), and lead to a temporary reduction in fertility (Hill et al., 1971). Additionall y, it was observed that low doses of progesterone increased LH pulse frequency an d prolonged pre-ovulatory s ecretion of estradiol (Sirois and Fortune, 1990). Hill et al. (1971) observed decreased pregnancy rates caused by ovulation failure, fertilization failure, and a bnormal oocytes following a 14 d MGA treatment. Chow et al. (1982) also observe d greater LH and estradiol con centrations during the estrus

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43 following exogenous progesterone treatment compared to non-treated heifers. Guthrie et al. (1970) and Kinder et al. (1996) concluded that the development of a persistent dominant follicle was the cause of reduced fertility following a lo ng term exogenous progesterone treatment. In a study by Ahmad et al. (1995), in which cows were set up to ovulate e ither normal, growing follicles or persistent follicles, they observed la rger follicles and greater estradiol concentrations in cows with persistent follicles. In the same study following ovulati on, less oocytes/embryos were recovered from cows with persistent follicles than growing follicles, and fewer embryos from persistent follicles reached a 16 cell stage at 14%, compared to those from growing follicles at 86%. Uterine secretions may also be altered in animals that undergo a long-term progestin treatment (Wordinger et al., 1972; Gibbons et al., 1973). Episodic release of LH is c ontrolled by the circulating con centrations of progesterone (Roberson et al., 1989). To more closely mimic a cow’s natural luteal phase, Stock and Fortune (1993) did a study in heifers with three treatme nts including a blank CI DR, one CIDR, or two CIDRs, over a period of 14 d. The blank CIDR served as a control, one CIDR mimicked a synchronization program, and two CI DRs mimicked the greater proge sterone concentrations in a normal luteal phase. The single CIDR treatment result ed in ovulatory follicles that were larger in size and remained dominant for three times as lo ng compared to controls. The two CIDR group had more follicular waves per cycle than the control and one CIDR treatments. This study demonstrated that the progester one provided in a normal, one CI DR protocol was not sufficient to fully suppress LH pulses resulting in a single fo llicle that maintained dominance, and led to a decrease in fertility (S tock and Fortune, 1993). In a study by Wehrman et al. (1996), embryos we re transferred into recipient cows that had ovulated a persistent dominant follicle 7 d prev iously. Pregnancy rates were similar between

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44 recipients that ovulated normal follicles and thos e that ovulated a persistent dominant follicle. These results, in conjunction with the findings of Mihm et al. (1994) that progesterone concentrations following ovulation of a persistent dominant follicle did not vary from those ovulating a normal follicle, indicate that the reduction in fertility is a result of problems with the follicle and not other factors, such as the uterine environment. It should also be noted that these effects do not carry over to the second estrus following withdrawal of progesterone treatment (Roberson et al., 1989). Follicles which undergo a prolonged period of dominance, even over a short period of time, can also have reduced fertility. Austin et al. (1999) observed no differences in the subsequent fertility of heifers which ovulated follicles after a period of 2, 4, 6, or 8 d of dominance, with pregnancy rates of 89, 68, 78 and 71% respectively. However, in heifers with dominant follicle durations of 10 or 12 d ha d decreased pregnancy rates of 52 and 12%, respectively. This indicates th at follicles on the ovary only one to two days longer can result significantly less fe rtile ovulations. One of the advantages of using progestins is th at it has been shown to initiate estrus and ovulation in some anestrous cows (Fike et al., 1997; Lucy et al., 2001) and prepubertal heifers (Imwalle et al., 1998; Fike et al., 1999; Hall et al., 1997). This e ffect is achieved by stimulating LH secretion both during and after progesterone tr eatment, thereby accelerating follicle growth in both anestrous cows (Garcia-Winder et al., 1987) and prepubertal heifers (Anderson et al., 1996). After a 7 d CIDR treatment, Fike et al. (1997) initiated estrus or CL formation in 70% of anestrous cows whereas Lucy et al. (2001) observed approximately 40% of anestrous cows to resume cycling. Similarly, Fike et al. (1997) observed 66.1 to 81.4% of anestrous cows and prepubertal heifers in estrus within 180 h following MGA treatment. Hall et al. (1997)

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45 implanted heifers with norgestomet implants for 10 d, and observed that while puberty was induced, it was dependent on age, with a greater number of olde r heifers achieving puberty than younger heifers. Within 10 d after a 7 d MGA tr eatment, Imwalle et al. (1998) induced puberty in 8/8 MGA treated heifers, compared to only 4/9 control heifers. Gonadotropin Releasing Hormone (GnRH) In a normally cycling animal, when progesteron e begins to decrease and estradiol from the dominant follicle rises, GnRH is triggered an d LH is released, leading to ovulation of the eligible follicle (Fortune et al., 1988). Administration of an exogenous gonadotropin releasing hormone agonist (GnRH) works indi rectly through the ante rior pituitary to in itiate ovulation. Gonadotropin releasing hormone can be admi nistered exogenously as a 2 mL (100 g) intravenous or intramuscular injection a nd is marketed under the names of Fertagyl, Factrel, and Cystorelin. The effects of GnRH are observed with in 2 to 4 h of in jection through an increase in peripheral concentr ations of both LH and FSH (Che nault et al., 1990; Stevenson et al., 1993). This induced LH surge causes ovulation w ithin 24 to 32 h after treatment (Thatcher et al., 1989; Macmillan and Thatcher, 1991; Pursley et al., 1994) of large (> 9 mm; Sirois and Fortune, 1988), dominant (Guilbault et al., 1993) ov arian follicles in th eir growth phase of development. However, GnRH will not initiate ovulation of all large follicles, particularly follicles which are regressing or in a state of at resia (Guilbault et al., 1993) and have a declining number of LH receptors (Rollosson et al., 1994). Following ovulation, estradiol concentrations are dramatically decr eased in peripheral circulation both after a GnRH treatment induced LH surge (Twagiramunga et al., 1994b) and LH surge caused by removal of a large follicle (Hughes et al., 1987). If a follicle is, in fact, ovulated to GnRH, the rapid decrease in estrogen output w ill result in an FSH surge, causing recruitment

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46 of a new wave of follicular growth 1 to 2 d later (Ko et al., 1991; Adams et al., 1992). Additionally, the FSH peak provided by the GnRH injection 2 to 4 h after treatment will stimulate the recruitment of a new follicular wa ve (Chenault et al., 1990; Rettmer et al., 1992). One of the reasons that GnRH does not initiate ovulation of all large follicles is due to stage of cycle effects. In a study by Moreira (2000) in dairy heifers, an Ovsynch protocol initiated on days 2, 5, 10, 15, and 18 of the estrou s cycle resulted in ovu lation rates of 0, 100, 25, 60, and 100%, respectively, to the initial GnRH in jection. These results are expected, as on d 2 of the estrous cycle a dominant follicle is not ye t present to ovulate to GnRH. On d 5, a first wave dominant follicle is presen t to ovulate to GnRH. By d 10 most of the first wave dominant follicles have already become atretic, and thus will not ovulate to GnRH. However, some animals may already have a second wave dominant follicle eligible for o vulation. On d 15, most cows will have a second wave domi nant follicle capable of ovulati on, but this is more variable than the first wave due to differences in dura tion of dominance of th e first wave dominant follicle. On d 18, cows are in the proestrus phas e and thus yield high ovula tion rates. A similar study by Vasconcelos (1999) in l actating dairy cows observed lo west ovulation rates for cows administered GnRH on days 1 to 4 (23%), modera te ovulation rates on days 10 to 16 (54%), and greatest ovulation rates on days 5 to 9 and 17 to 21 (96 and 77%, respectively). The use of GnRH agonists have been shown to synchronize follicular growth and yield precise ovulation times without compromising fert ility (Twagiramungu et al., 1995). It has been suggested with mixed results that treatment w ith GnRH agonists may alter the subsequent CL progesterone output. There have been reports of both increased (Macmillan et al., 1985a,b; Stevenson et al., 1993) and decreased (Ford and Stormshak, 1978, Rodger and Stormshak, 1986) progesterone production following treatment with a GnRH agonist, as well as numerous studies

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47 which showed no changes (Macmillan and Thatch er, 1991; Prescott et al., 1992). Studies by Twagiramungu et al. (1992b; 1994a) demonstrated that both the CL existing at the time of GnRH treatment, and that which was formed as a result of a GnRH-induced ovulation, respond to luteolysis by exogenous PGF2 . Therefore, GnRH-induced CL are similar to a naturally formed CL, at least in regard to luteolysis. Since GnRH can initiate ovulation and s ynchronization of follicle waves, it was combined with PGF2 which was administered 7 d after GnRH in the initial development of the GnRH + PGF2 estrous synchronization program (Thatc her et al., 1989; Macmillan and Thatcher, 1991; Pursley et al., 1994). This system allows for the turnover of any large follicle on the ovary, the growth of a new follicular wave cont aining the pre-ovulatory follicle, and subsequent luteolysis to eliminate the CL to bring the cow into estrus. This results in a gr oup of cattle with highly synchronized follicular growt h, and a precise estrus following PGF2 –induced luteolysis. Estrous rates of 70 to 83% (Thatcher et al., 1989; Coleman et al., 1991; Twagiramungu et al., 1992b,c) and pregnancy rates of 65 to 85% (C oleman et al., 1991; Twagiramungu et al., 1992b,c), both over a 4 d period, have been re ported in beef and dairy cattle. Another use of GnRH is to induce ovulation during the period after PGF2 following a synchronization protocol to reduce or eliminate the need for es trous detection. This is possible, because a great degree of synchrony is achieved, with 56 to 76% of cows displaying estrus between 24 and 72 h after PGF2 (Twagiramungu et al., 1992b,c). With the addition of a second GnRH injection, given 48 hrs after PGF2 , Pursley et al. (1994) obs erved that dairy cows and heifers ovulated between 24 and 32 h after GnRH. It also appears that breed may have an effect on respons e to GnRH. In a study by Griffin and Randel (1978), ovariectomized Brahman and He reford cows were treated with GnRH and

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48 the resulting LH response was significantly lower for Brahman cows, indica ting that they may be less responsive to GnRH-induced LH release. GnRH + CIDR + PGF2 Systems The combination of GnRH and PGF2 has been extensively st udied as a synchronization program in Bos taurus cattle but less thoroughly researched in cattle of Bos indicus breeding. The base estrous synchronization system is a co mbination of GnRH followed 7 d later with PGF2 , followed by a multitude of different AI pr ograms. The different programs are termed Select Synch, Co-synch, Ovsynch, and Hybrid-Sync h. The Select Synch protocol involves estrous detection and subsequent AI over a set number of days , typically 5 d (Thompson et al., 1999; Stevenson et al., 2000). The Co-Synch system is a timed-AI protocol, in which cows are administered GnRH and timed-AI 48 h after PGF2 (Stevenson et al., 2000; Geary et al., 2001). The OvSynch system consists of a timed-AI conducted approximately 16 h after the second GnRH, which is typically given 48 h after PGF2 . The Hybrid-Synch system uses a combination of estrous detection and timed-AI to allow all cows to be inseminated. This system consists of 3 d of estrous detection and AI th rough 72 h and all cows not exhi biting estrus are administered GnRH and timed-AI 72 to 80 h after PGF2 . The effectiveness of GnRH + PGF2 systems in Bos taurus cattle have been well established. Conception rates of 46 to 55% have been achieved in lactating dairy cows (Pursley et al., 1995) and 26 to 63% in dairy heifer s (Schmitt et al., 1996) in a GnRH + PGF2 protocol followed by detected estrus or timed-AI. Concep tion rates of 33 to 69% have also been achieved in Bos taurus beef cows (Geary et al., 1998; Stevenson et al., 2000). Lemaster et al. (2001) compared the Select Synch, Co-synch, and Hybrid-Synch protocols in cows of Bos indicus breeding. Similar pregnancy ra tes were achieved for Co-synch

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49 and Hybrid-Synch with 31.0% and 35.5%, respectivel y, but there was a decr ease in the Select Synch system with pregnancy rates of 20.8%. This decrease was attributed to the difficultly of estrus detection in Bos indicus cows, as well as the lower number of Bos indicus cows inseminated. One of the problems with the Select Synch sy stem is the premature expression of estrus several days before PGF2 (DeJarnette et al., 2001) resulting in the need for additional estrous detection. The addition of a pr ogestin between GnRH and PGF2 treatments prevents the need for the extra estrous detection (Thompson et al., 1 999; DeJarnette et al., 2003). This progestin can come in the source of either MGA or a CIDR. Another benefit of the progestin is that it can induce cyclicity in some anestrous cows (Fik e et al., 1997; Lucy et al., 2001), and may be particularly beneficial in co ws that calve late in the br eeding season or are in poor body condition (Stevenson et al., 2000; Lamb et al., 2001) Although limited, the data would suggest that the CIDR may be more effective in inducing estrus than the MGA treatment. Perry et al. (2004) induced ovulation and estrous cycles in more early postpartum Bos taurus beef cows using a 7 d CIDR treatment vs. normal (0.55 mg/kg) or high (4.41 mg/kg) levels of MGA over the same treatment period. Similarly, Stevenson et al. (2003) achieved pregnancy rates of 46 and 55% when MGA or a CIDR were added to the Co-Synch protocol , respectively, for postpartum Bos taurus beef cows. In a group of Bos indicus and Bos indicus Bos taurus heifers, Kerr et al. (1991) saw pregnancy rates of 48.8% using a 7 d CIDR treatment with pre gnant mare serum gonadotropin (PMSG) and PGF2 on d 7, compared to only 18.6% pregna ncy rates with two doses of PGF2 12 d apart.

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50 Martinez et al. (2002b) observe d that the addition of a CIDR to a Co-Synch timed-AI treatment protocol in lactating Bos taurus beef cows yielded similar results to that without a CIDR, with pregnancy rates of 42.9 and 45.1%, resp ectively. The results of this study are in agreement with those of Johnson et al. (2000), who observed similar pregnancy rates in CoSynch + CIDR (45%) and Co-Synch (47%) treatments for postpartum Bos taurus beef cows. However, Martinez et al. (2002a), reported an increase in pregnancy rates from 39.1 to 68.0% with the addition of a CIDR to the Co-Synch system in Bos taurus beef heifers. In contrast, Kawate et al. (2004) reported significantly greater (72.5 vs. 47.7%) conception rates for postpartum Bos taurus beef cows given an Ovsynch + CIDR treatment compared with Ovsynch alone. Similarly, Lamb et al. (2001) reported increased pregnancy rates for Co-synch + CIDR (58.6%) over Co-synch (48.1%) treatments with an even greater increase in pregnancy rate among anestrous Bos taurus beef cows , with 59% for Co-synch + CI DR compared with 39% for Co-synch treated cows. Using an Ovsynch system without a CIDR , Barros et al. (2000) achieved similar pregnancy rates with Bos indicus cows as those from Bos taurus cows, ranging from 42 to 48% (Fernandes et al, 2001; Williams et al., 2002). Most of the CIDR synchronization programs di scussed above were conducted with the 7 d CIDR containing 1.38 g of progesterone. Macmillia n et al. (1991) reported that a 1.9 g CIDR releases progesterone for at least 15 d. Ta king that into account, producers often question whether a CIDR can be reused. In a series of experiments by Col azo et al. (2004), the effectiveness of previously used CIDRs was e xplored using treatments of a new CIDR, one once-used CIDR, one twice-used CI DR, and two twice-used CIDRs in Bos taurus beef heifers undergoing a timed-AI protocol. Similar pregna ncy rates were observed with a new and once-

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51 used CIDR, but pregnancy rate s were significantly decreased with a twice-used CIDR. However, when two, twice-used CIDRs were used concurrently, there was no change in pregnancy rates compared to the ne w and once-used CIDR treatments. Limited work has been done using the Hybr id-Synch protocol to synchronize estrus in postpartum beef cows, particularly those of Bos indicus breeding.

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52 CHAPTER 3 EFFECTIVENESS OF NEW VS. ONCE-USED CIDR AND CLOPROSTENOL SODIUM VS. DINOPROST TROMETHAMINE IN A GnRH/CIDR + PGF2 PROTOCOL IN LACTATING Bos indicus Bos taurus BEEF COWS Introduction Estrous synchronization allows for a large num ber of cows to display estrus and ovulate over a short period of time. This allows for e ither the reduction of daily estrous detection over the entire estrous cycle to 3 to 5 d or its elimin ation with incorporation of a timed-AI, where all cows are inseminated at a predetermined time. A frequently used and effective synchronization protocol in Bos taurus cattle is administration of Gn RH followed 7 d later with PGF2 (Thatcher et al., 1989; Pursley et al., 1995). However, a common problem with the GnRH + PGF2 protocol is expression of estr us several days before PGF2 (DeJarnette et al., 2001). This problem can be eliminated with addition of a progestogen concomitant with GnRH and removed at PGF2 (DeJarnette et al., 2004). A ddition of progestogen can also increase the percentage of anestrous cows that exhibit estr us (Stevenson et al., 2000; Larsen et al., 2006). Limited research has employed the GnRH + PGF2 protocol either with (Hie rs et al., 2003) or without a progestogen (Lemaster et al., 2001) in Bos indicus Bos taurus cattle with limited success. The reason(s) for the less than acceptable results ar e unclear, but may be due to a decreased responsiveness of the CL to th e luteolytic actions of PGF2 (Lemaster et al., 2001). The ability of prostaglandins like cloprost enol sodium and dinoprost tromethamine to synchronize estrus has been documented (Young and Anderson, 1986; Salverson et al., 2002) in Bos taurus cattle, but only one study in cattle of Bos indicus breeding has been conducted (Hiers et al., 2003). Hiers et al. (2003) reported a similar timed-AI pregnancy rate between cloprostenol sodium and dinoprost tromethamine in non-lactating Bos indicus Bos taurus cows synchronized with GnRH + PGF2 protocol combined with melengestrol acetate.

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53 The intravaginal progesterone-releasing insert (CIDR) is an effective synchronization product that also induces estrus in some anestrous cattle (Lucy et al., 2001; Larsen et al. 2006). A new CIDR (1.9 g of progesterone) maintains circulating progesteron e concentrations > 1 ng/mL for at least 15 d after insertion (Macmillan et al., 1991) suggesting that a CIDR could be used for two consecutive 7 d treatments and stil l suppress the expression of estrus. Reuse of a CIDR has been investigated in Bos taurus beef cows with no difference in pregnancy rates compared to a new CIDR when used with estradiol cypionate at CIDR insertion and PGF2 at CIDR removal (Colazo et al., 2004); although, si milar studies have not been conducted in Bos indicus Bos taurus cattle. Data are inconclusive as to wh ether or not cloprostenol sodium is more effective in cattle of Bos indicus breeding. Therefore, the objectives of these experime nts were to evaluate the effectiveness of a new CIDR compared to a onc e-used CIDR and cloprostenol sodium compared to dinoprost tromethamine in two GnRH + PGF2 synchronization protocols in postpartum lactating Bos indicus Bos taurus cows. Materials and Methods Two experiments were conducted from January to March of 2005 and 2006 at the Bar L Ranch, Marianna, Florida. In Experime nt 1, multiparous postpartum lactating Bos indicus Bos taurus cows (n = 284) were used. Mean ( SD) cow age was 5.7 1.9 yr, days postpartum (DPP) was 58.0 12.5 d, body weight (BW) was 499 48 kg, and body condition score was 5.2 0.5 (BCS: 1 = emaciated, 9 = obese; Ri chards et al., 1986). The degree of Bos indicus breeding ranged from approximately 10 to 38% with the remainder being Bos taurus breeding. The experiment was a 2 2 factorial design. At the start of the experiment (d 0), cows were equally distributed by DPP and cow age to one of two progesterone treatm ents, which included a

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54 new CIDR (1.38 g; Eazi-Breed CIDR, Pfizer Animal Health, New York, NY) and a onceused CIDR (new CIDR used once and autoclaved before re-use). All cows received GnRH (100 g i.m.; Fertagyl, Intervet, Boxmeer, The Netherlands) at CIDR insertion and BCS were collected. On d 7, CIDR were removed and cows within each CIDR treatment were equally distributed by DPP, BCS, and cow age to receive either of two PGF2 treatments, which included cloprostenol sodium (500 g i.m.; Estrumate, Schering-Plough Veterinary Corp., Kenilworth, NJ) or dinoprost tromethamine (25 mg i.m.; Prostamate, Agrilabs, St. Joseph, MO). All cows received an Estrus Alert estrous detection patch (Estrus Alert, Western Point, Inc., Merrifield, MN) at CIDR removal to aid in estrous detection. Estrus was visually detected three time s daily at 0700, 1200, and 1700 h for 5 d following PGF2 . Estrus was defined as a co w standing to be mounted by anot her cow and/or a half to full red Estrus Alert patch. Cows were artifi cially inseminated (AI) 8 to 12 h after observed in estrus. Frozen-thawed semen from a single sire was used and cows were inseminated by two AI technicians. Seven days after the last cow was inseminated, bulls were placed with cows. Pregnancy was diagnosed approximately 55 d afte r AI using a real-time B-mode ultrasonography machine (Aloka 500V, Corometrics Medical Systems, Wallingford, CT) with a 5.0 MHz transducer. Due to the 7 d period in which no cows were inseminated or bred by the clean-up bull, differences in fetal size (Curran et al., 1986 ) were used to determine whether a pregnancy resulted from the synchronized breeding or clean-up bull. Estrous response was defined as the number of cows displaying estrus for 5 d after PGF2 and inseminated divided by the total number of co ws treated. Conception rate was defined as the number of cows that became pre gnant to AI divided by the number of cows that displayed estrus and were AI. Synchronized pregnancy rate was the number of cows pregnant to AI divided by

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55 the total number of cows treated. Thirty-day pr egnancy rate was the number of cows pregnant during the first 30 d of the breed ing season divided by the total number of cows treated. The GENMOD procedure of SAS (SAS Inst. Inc. , Cary, NC) was used for the statistical analysis for Experiment 1. The main effects of CIDR and PGF2 treatments, and CIDR PGF2 were evaluated for estrous response, concep tion, synchronized pregna ncy, and thirty-day pregnancy rates. The aforementioned response va riables were also evaluated for the simple treatment effects of new CIDR + cloprosteno l, new CIDR + dinoprost, once-used CIDR + cloprostenol, and once-used CIDR + dinoprost. Cow age, DPP, BCS, and interval from PGF2 to the onset of estrus were include d as covariates. When covariat es were significant (P < 0.05), they were treated as independent variables. The effects of DPP were divided into three categories ( 50 d, 51 to 69 d, 70 d), BCS divided into two categories ( 5, > 5), and interval from PGF2 to the onset of estrus divided into five categories (48, 60, 72, 84, 96 hr). The effect of DPP and BCS along with the appropriate tw o-way interactions with main and simple treatment effects were evaluated for estrous response, conception, synchronized pregnancy, and thirty-day pregnancy rates. Th e effect of interval from PGF2 to the onset of estrus and appropriate two-way interactions with main and simple treatme nt effects were evaluated for conception rate. The effect of AI technician a nd all appropriate interactions with the main and simple effects were tested for conception rates. The distribution of estr us after CIDR removal for the main and simple treatment effects were analyzed using the LIFETEST procedure (survival analysis) of SAS. Two animals were not present at the pregnancy diagnosis, and therefore, were removed from all statistical analyses. In Experiment 2, multiparous postpartum lactating Bos indicus Bos taurus cows (n = 259) were utilized. Mean ( SD) cow age was 6.9 1.9 yr, DPP was 48.5 12.8 d, and BCS

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56 was 5.1 0.5. The degree of Bos indicus breeding ranged from approximately 10 to 38% with the remainder being Bos taurus breeding. The experiment was a 2 2 factorial design and animals were assigned to the same synchr onization treatments as Experiment 1. Estrus was detected as described in Experime nt 1. Cows were AI 8 to 12 h after observed in estrus. All cows that had not displayed estrus by 0800 h, 73 h after PGF2 were timed-AI and administered GnRH between 76 and 80 h after PGF2 . Cows in estrus at the 72 h observation were inseminated 8 to 12 h later, an d not included in the timed-AI. Cows were inseminated with frozen-thawe d semen from five pre-assigned sires and inseminated by a single AI technician. Seven days after the last cow was inseminated, clean-up bulls were placed with cows. Pregnancy was di agnosed approximately 56 d after AI using a real-time B-mode ultrasound (Aloka 500V, Coro metrics Medical Systems, Wallingford, CT) with a 5.0 MHz transducer. Due to the 7 d period in which no cows were inseminated or bred by the clean-up bull, differences in fetal size (Curran et al., 1986) were used to determine whether a pregnancy resulted from the synchroni zed breeding or by the clean-up bull. Estrous response was defined as the number of cows displaying estrus for 3 d after PGF2 and inseminated divided by the total number of co ws treated. Conception ra te was the number of cows that displayed estrus, were AI and became pregnant, divided by the number of cows that displayed estrus and were AI. Timed-AI pregna ncy rate was the number of cows that failed to display estrus, were timed-AI and became pregnant , divided by the total number of cows that were timed-AI. Synchronized pregnancy rate was the total number of cows that were inseminated and became pregnant, divided by the total number of cows treated. Thirty-day pregnancy rate was the number of cows pregnant during the fi rst 30 d of the breeding season divided by the total number of cows treated.

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57 The GENMOD procedure of SAS (SAS Inst. In c., Cary, NC) was used for the statistical analysis of Experiment 2. The main effects of CIDR and PGF2 treatments, and CIDR PGF2 were evaluated for estrous response, concepti on, timed-AI pregnancy, synchronized pregnancy, and thirty-day pregnancy rates. The aforementi oned response variables were also evaluated for the simple treatment effects of new CIDR + cloprostenol, new CIDR + dinoprost, once-used CIDR + cloprostenol, and once-used CIDR + dinopros t. Cow age, DPP, BCS, and interval from PGF2 to onset of estrus on conception rate were in cluded as covariates. When covariates were significant (P < 0.05) they were tr eated as independent variables. Days postpartum were divided into three categories (< 40 d, 40 to 59 d, 60 d) and BCS divided into two categories ( 5, > 5). The effect of DPP and BCS cat egories along with main and simple treatment effects and appropriate two-way interactions were evaluated for estrous response, conception, synchronized pregnancy, and thirty-day pregnancy rates. Since AI sires were pre-assign ed to cows before AI and not equally distributed across tr eatments, AI sire effects were not included in the statistical analysis. One cow lost a CIDR between d 0 and d 7, and another cow was not present for pregnancy diagnosis. Both cows were removed from all statistical analysis. Results In Experiment 1, estrous response, concep tion, and synchronized pregnancy rates were similar (P > 0.05) for main effects of CIDR and PGF2 treatments (Table 3-1). Thirty-day pregnancy rates were similar (P > 0.05) for new (78.0%) vs. once-used (79.4%) CIDR and for cloprostenol (77.3%) vs. dinoprost (80.1%). There were no (P > 0.05) CIDR PGF2 treatment effects on estrous response, conception, synchronized pregnancy, or thirty-day pregnancy rates. Additionally, simple treatment e ffects were similar (P > 0.05) for estrous response, conception, and synchronized pregnancy rates (Table 3-2). Thirty-day pregnancy rates were similar (P >

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58 0.05) for new CIDR + cloprostenol (74.7%), new CIDR + dinoprost (81.3%), once-used CIDR + cloprostenol (80.0%), and once-used CIDR + dinopr ost (78.9%). The over all estrous response, conception, synchronized pregnancy, and thir ty-day pregnancy rates were 68.4, 47.7, 32.6, and 78.7%, respectively. When included as a covariate, DPP influenced (P < 0.05) estrous response, conception, synchronized pregnancy, and thirty -day pregnancy rates for main and simple treatment effects; however, when BCS was included as a covariate it did not (P > 0.05) effect estrous response, conception, sync hronized pregnancy, and thirty-d ay pregnancy rates for main and simple treatment effects. As previously indicated, DPP effected (P < 0.05) estrous response, conception, and synchronized pregnancy rates (T able 3-3). However, there were no (P > 0.05) treatment DPP effects for either main or simple treatment effects. Cows that were long ( 70 d) postpartum had a greater (P < 0.05) estrous response co mpared to cows that were short ( 50 d) and medium (50 to 69 d) postpartum, which were similar (P > 0.05 ). Conception rates were similar (P > 0.05) for cows that were medium and long postpartum, bot h of which were greater (P < 0.05) than for cows that were short postpartum. Synchronize d pregnancy rates were greater (P < 0.05) for cows that were long postpartum compared to short and medium postpartum cows, which were different (P < 0.05). Thirty-day pregnancy rate s were greater (P < 0.05) for cows that were medium (79.1%) and long (87.8%) postpartum co mpared to cows that were short (71.6%) postpartum. Cows that were medium and shor t postpartum had similar (P > 0.05) thirty-day pregnancy rates. The BCS between DPP categories were similar (P > 0.05) and had no effect (P > 0.05) on estrous response, conception, synchroni zed pregnancy, and thirty -day pregnancy rates when included in the DPP model as a covariate.

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59 The distribution of estrus after PGF2 treatment was similar (P > 0.05) for the main and simple treatment effects when evaluated with the survival analysis (data not shown). Figure 3-1 shows the distribution of estrus for all treatments combined. No cows were detected in estrus until 48 h after PGF2 . The mean interval from PGF2 to onset of estrus (64.4 16.0 h) was not influenced (P > 0.05) by main or simple treatment e ffects. Interestingly, th ere was an effect (P < 0.01) of interval from PGF2 to onset of estrus on conception rate (Figure 3-1). Cows which displayed estrus 48, 60, 72, and 84 h after PGF2 had similar (P > 0.05) conception rates, but cows which displayed estrus at 48, 60, and 72 h had greater (P < 0.05) conception rates compared to cows that displayed estrus 96 h after PGF2 . Cows that displaye d estrus at 84 h had similar (P > 0.05) conception rate s compared to cows which disp layed estrus at 96 h. When included as a covariate, DPP in fluenced (P < 0.05) conception rate, but there was no (P > 0.05) interval from PGF2 to onset of estrus DPP effect on conception rate . Conception rate was not influenced (P > 0.05) by AI technician or its inter actions with main and simple treatment effects. Cow age and BCS did not affect (P > 0.05) estrous response, conception rate, synchronized pregnancy, or thirty -day pregnancy rates when include d as covariates for main and simple treatment effects. For Experiment 2, estrous response, concep tion, timed-AI pregna ncy, and synchronized pregnancy rates were similar (P > 0.05) for main effects of CIDR and PGF2 treatments (Table 34) and simple treatment effects (Table 3-5). Th irty-day pregnancy rates were similar (P > 0.05) for new (83.2%) vs. once-used (76.9%) CIDR and for cloprostenol (76.7%) vs. dinoprost (83.3%). There were no (P > 0.05) CIDR PGF2 treatment effects on estrous response, conception, timed-AI pregnancy, synchronized pregnanc y, or thirty-day pregna ncy rates. Thirtyday pregnancy rates were similar (P > 0.05) fo r new CIDR + cloprostenol (82.5%), new CIDR +

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60 dinoprost (83.7%), once-used + CIDR cloproste nol (71.2%), and once-used + CIDR dinoprost (82.8%). The pooled estrous response, c onception, timed-AI pregnancy, synchronized pregnancy, and thirty-day pregnancy rates were 50.2, 60.2, 32.3, 46.3, and 80.0%, respectively. The CIDR treatment tended (P = 0.10) to in fluence conception rate as 14.1% more new CIDR cows became pregnant compared to once-us ed CIDR cows. (Table 3-4). The interval from PGF2 to onset of estrus was not (P > 0.05) aff ected by main or simple treatment effects, with a mean interval of 58.6 10.3 h pooled across treatments. Interval from PGF2 to onset of estrus did not (P > 0.05) e ffect conception rates. Days postpartum affected (P < 0.05) estrous response, conception, timed-AI pregnancy, synchronized pregnancy, and thir ty-day pregnancy rates when in cluded as a covariate for main and simple treatment effects (Table 3-6). There were no (P > 0.05) interactions of DPP with the main effect of CIDR treatment or simple treatm ent effects. Estrous response was greater (P < 0.05) for long ( 60 d) compared to short (< 40 d) a nd medium (40 to 59 d) postpartum cows, which were similar (P > 0.05) to each other. Conception and timed-AI pregnancy rates were greater (P < 0.05) for long compared to short po stpartum cows, but were similar (P > 0.05) to medium postpartum cows. Short postpartum cows had similar (P > 0.05) conception and timedAI pregnancy rates compared to medium postpar tum cows. Synchronized pregnancy rates were greater (P < 0.05) for long compared to medium and short (P < 0.05), whic h were different (P < 0.05). Thirty-day pregnancy rate was greater (P < 0.05) for long (94.5%) postpartum cows compared to short (70.3%) or medium (76.9%) post partum cows, which were similar (P > 0.05). There was a PGF2 treatment DPP effect (P < 0.05) on estrous response. Cloprostenol treated cows that were short (34.2%) and medium postpartum (49.1%) had similar (P > 0.05) estrous responses compared to dinoprost treated cows that were short (44.4%) and medium postpartum

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61 (41.8%). However, long postpartum cows treated with cloprostenol (79.0%) had a greater (P < 0.05) estrous response compared to dinoprost treated cows (5 7.2%). The BCS between DPP categories were similar (P > 0.05). Body condition score did not (P > 0.05) affect estrous response, timed-AI pregnancy, or synchronized pregnancy rates, but did affect c onception and thirty-day pregnancy rates when included as a covariate for the main and simp le treatment effects. Cows with a BCS 5 (64.5%) tended (P = 0.10) to have a greate r conception rate compared to cows with a BCS < 5 (48.6%). Cows with a BCS 5 (84.3%) had a greater (P < 0.05) thir ty-day pregnancy rate compared to cows with a BCS < 5 (68.6%). There were no (P > 0.05) interactions of BCS with the main or simple treatment effects. Cow age did not affect (P > 0.05) estrous re sponse, conception rate, timed-AI pregnancy, synchronized pregnancy, or thirty -day pregnancy rates when include d as a covariate for main and simple treatment effects. Discussion There were no CIDR (new vs. once-used) or PGF2 (cloprostenol sodium vs. dinoprost tromethamine) main effects on estrous response, conception, synchronized pregnancy, or thirtyday pregnancy rates for postpartum lactating Bos indicus Bos taurus cows in Experiment 1. The 5 d estrous response was 23.2% greater and the synchronized pregnancy rate was 11.8% greater than reported by Lemaster et al. (2001) in postpartum lactating Bos indicus Bos taurus beef cows synchronized with the Select Synch prot ocol without a CIDR treatment. The ability of a new CIDR to induce estrous cycles in anestrous cows has been previously reported (Lucy et al., 2001; Larsen et al. 2006). However, it is not kno wn if a once-used CIDR would induce estrus in anestrous cows to the same extent as a new CI DR. It is also unclear what influence CIDR

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62 treatments had on inducing estrus in anestrous cows in the presen t study since cycling status was not evaluated. The estrous response a nd synchronized pregnancy rates of Bos indicus Bos taurus cows are similar (Kojima et al., 2000; Stevens on et al., 2000), but also less (Geary et al., 1999; Larson et al., 2006) compared to studies of Bos taurus cows synchronized with the Select Synch protocol. It appears that the key factor resu lting in reduced synchronized pregnancy rates to the Select Synch protocol in Bos indicus Bos taurus cows compared to Bos taurus cows is the decreased estrous response, which results in significantly fewer cows being inseminated and lower synchronized pregnancy rates. Estrus is difficult to detect in cattle of Bos indicus breeding (Randel, 1984; Galina et al., 1994). Cows of Bos indicus breeding exhibit a shorte r and less intense estr us (Plasse et al., 1970; Randel, 1984; Pinheiro et al., 1998), may show more subtle signs of estrus (Galina et al., 1982; Orihuela et al., 1983), ha ve fewer mounts during estrus (Rae et al., 1999), a nd often initiate estrus during the evening hours (Pinheiro et al., 1998; Landaeta-He rnandez et al., 2002). It does not appear that estrous detection was a major problem in Experiment 1, because cows were intensely monitored by visual detection during the morning, mid-day, and early evening hours. Furthermore, Estrus Alert patches aided in detecting any es trus, which either began in, or occurred entirely during the evening hours. Even in pos tpartum lactating cows of Bos indicus breeding known to be cycling at the start of a Select Synch program without a CIDR, the 5 d estrous response was only of 55.7% (Lemaster et al., 2001). Because of the low estrus response in cattle of Bos indicus breeding synchronized with the Sel ect Synch protocol, utilization of some type of timed-AI program may be necessary. As previously indicated, conception rate was not influenced by the main effects of CIDR or PGF2 treatments. The conception rates were si milar to those reported by Lemaster et al.

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63 (2001) in Bos indicus Bos taurus cows synchronized with Sele ct Synch without a CIDR but substantially less than reports in Bos taurus cows synchronized with Select Synch either with (Larson et al., 2006) or without (Geary et al., 1999; Larson et al., 2006) a CIDR. Additionally, interval from PGF2 to the onset of estrus had a signi ficant effect on c onception rates in Experiment 1. For cows displaying estrus 84 and 96 h after PGF2 , conception rates were reduced by > 30% compared to cows di splaying estrus 48 to 72 h after PGF2 . This finding is of even greater significance since a single AI sire was used to inseminate all cows and there were no AI technician, BCS, or cow age effects on con ception rate. These resu lts suggest that there are differences in ovarian follicle development at PGF2 , which compromised fertility of the subsequent estrus. The reduced conception rates are probably due to differences in duration of dominance of the ovulatory follicle at PGF2 . Follicles with longer durations of dominance have decreased fertility when inseminated after an obs erved estrus (Mihm et al., 1994; Austin et al., 1999; Townson et al., 2002). Several factors coul d be affecting ovarian follicle dominance. The interval to estrus following PGF2 is dependent on the stage of ovarian follicular growth when PGF2 is administered (Geary et al., 1999; Hittinger et al., 2004). Cows with a mature dominant follicle present at PGF2 display estrus earlier after PGF2 compared to cows with a growing and developing follicle at PGF2 (Kastelic et al., 1990), which takes longer to exhibit estrus. Incorporation of a CIDR into the Select Synch protocol in Bos indicus Bos taurus cows appears to have modified the ovarian follicular development during the CIDR treatment. In cows synchronized with the Select Synch protoc ol without a CIDR, some cows display estrus either at PGF2 or from 0 to 36 h after PGF2 (Lemaster et al., 2001). However, with the addition of the CIDR in the present experiment, no cows exhibited estrus until 48 h after PGF2 ,

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64 indicating that presence of the CIDR delayed the expression of estrus and probably affected ovarian follicle development. This could be mani fested in decreased fertility of cows displaying estrus 84 and 96 h after PGF2 . At CIDR removal, progesterone profiles va ry depending on the cycling status, stage of estrous cycle, and ovulation stat us to GnRH. Progesterone profiles could include: 1) high progesterone resulting from ovulation to GnRH on d 0, 2) high progesterone from follicle turnover without ovulation to GnRH, 3) high prog esterone from an existing CL, with no follicle turnover, and 4) low progesterone from a noncyc ling cow with no follicular turnover. During periods of high progesterone treatm ent, pulsatile LH secretion is enhanced, resulting in increased ovarian follicle development (Imwalle et al., 1998) . Therefore, it could be hypothesized that the CIDR treatment may enhance pulsatile LH secreti on, particularly in the absence of a functional corpus luteum (CL), which could enhance and possibly advance dominant follicle development. This could result in follicles that have longer durations of dominance and decreased fertility when inseminated after an observed estrus (Mih m et al., 1994; Austin et al., 1999; Townson et al., 2002). In contrast to the present study, Bridges et al. (2005) observed decreased conception rates in yearling Bos indicus Bos taurus heifers which displayed estr us 0 to 48 h (32.1%) after PGF2 compared to heifers that displayed es trus at 60 (48.8%) and 72 h (59.1%) when synchronized with a 14 d MGA treatment followed 19 d later by PGF2 . Estrous detection was not conducted after 72 h as heifers that did not exhi bit estrus were timed-AI. In the Bridges et al., (2005) study, heifers would not have been exposed to a proge stogen during the 7 d prior to PGF2 treatment.

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65 In Experiment 2, there were no CIDR, PGF2 , or simple treatment effects on estrous response, conception, timed-AI, or synchronized pregnancy rates. The 3 d estrous response was 13.0% greater, timed-AI pregnanc y rate was 7.7% greater, and th e synchronized pregnancy rate was 10.8% greater compared to a report by Lema ster et al. (2001) in postpartum lactating Bos indicus Bos taurus cows synchronized with the Select Synch + timed-AI protocol without a CIDR. The addition of a timed-AI to the Select Synch protocol in Experiment 2 increased synchronized pregnancy rates by 13.7% compared to Experiment 1 where only estrous detection was utilized. Lemaster et al. (2001) also obser ved a similar increase in synchronized pregnancy rate when a timed-AI was added to the Select S ynch protocol without a CI DR. Furthermore, the ability of the CIDR to induce cyclicity in some anestrous cows (Lucy et al., 2001; Larson et al., 2006) could have also aided in increasing the synchronized pregna ncy rates in the present study compared to Lemaster et al. (2001) using th e Select Synch plus timed-AI without a CIDR. The greater synchronized pregna ncy rates for Experiment 2 compared to Experiment 1 can be attributed to an increased conception rate as well as the additiona l cows inseminated at timed-AI. Incorporation of the timed-AI allows a ll cows an opportunity to be inseminated. This appears to be very important in cattle of Bos indicus breeding because of the difficulty in detecting estrus (Galina et al., 1994) and increased inci dence of “silent estrus” (Dawuda et al., 1989; Lamothe-Zavaleta et al., 1991). The timed -AI plus GnRH may en hance conception rates of cows that exhibit estrus from 84 to 120 h after PGF2 by ovulating follicles before the oocytes are aged and fertility begins decline (Mihm et al., 1994). This is supported by greater timed-AI pregnancy rates (32.3%) in Experiment 2 co mpared to conception rates (24.4%) for cows observed in estrus 84 and 96 h after PGF2 in Experiment 1. Addition of a timed-AI to the Select

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66 Synch protocol appears to be imperative wh en trying to maximize pregnancy rates to a synchronized breeding in postpartum lactating Bos indicus Bos taurus cows. Results from both experiments indicate that PGF2 treatments, cloprostenol sodium and dinoprost tromethamine, are e qually effective luteolysins in postpartum lactating Bos indicus Bos taurus cows as estrous response, conception, tim ed-AI, and synchronized pregnancy rates were similar between treatments. This is in agreement with a study by Salverson et al. (2002), who reported similar estrous response, concep tion, and synchronized pregnancy rates in Bos taurus heifers synchronized with 14 d MGA treatment followed 19 d later with PGF2 treatments of cloprostenol sodium and dinopros t tromethamine. In non-lactating Bos indicus Bos taurus cows synchronized with a modified Co-synch prot ocol, Hiers et al. (2003) reported a similar synchronized pregnancy rate be tween cloprostenol sodium and dinoprost tromethamine treated cows. Cows treated with a new CIDR tended to ha ve a 14.1% greater conception rate compared to once-used CIDR in Experiment 2. In cont rast, CIDR treatment (n ew CIDR vs. once-used CIDR) had no effect on conception rates in Expe riment 1. Differences in the quantity of progesterone released by the once-used CIDR may have affected the underlying ovarian follicular dynamics (Stock and Fortune, 1993), whic h could have affected conception rates. Low blood progesterone concentrations in the absence of a CL re sults in increased LH pulse frequency, which promotes prolonged follicle grow th and ovarian follicle dominance (Stock and Fortune, 1993), which leads to ovulation of follicles that have decreased fert ility at fertilization (Mihm et al., 1994; Austin et al., 1999; Towns on et al., 2002). The aforementioned scenario could have occurred in some cows tr eated with a once-used CIDR treatment.

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67 It is unclear what proportion of follicles ovulated to GnRH at CIDR insertion but previous research in Bos indicus Bos taurus cows indicates that acro ss several stages of the estrous cycle was only 27% (Hiers et al., 2006) of follicles ovulate to GnRH. However, in Bos taurus cows, > 60% of cows ovulate follicles to GnRH at random stages of the estrous cycle (Vasconcelos et al., 1999; Moreira et al., 2000). Further research ha s shown that as the interval from PGF2 to estrus lengthens (Mihm et al., 1994; Austin et al., 1999; Townson et al., 2002) aged follicles, that are potentially less fertile, are ovulated. Therefore, if fewer cows failed to ovulate to GnRH at CIDR insertion and had lo w progesterone concentr ation during the CIDR treatment, it could have had a negative effect on fertility of the subsequent dominant follicle ovulating after CIDR removal. In contrast, rece nt research in our lab with lactating Brangus cows has shown that interval from PGF2 to the onset of estrus was not influenced by whether a follicle ovulated or did not ovulate to GnRH at CI DR insertion in the Sele ct Synch protocol (See Chapter 5). This further suggests that there are considerable effect s on ovarian follicle development in Bos indicus Bos taurus cows during a Select Synch pr otocol with a CIDR that need to be characterized. Nutritional status has a dire ct influence on reproductive f unction and cycling status of lactating postpartum beef cows (Smith et al., 1976; Oyedipe et al., 1982). Body condition score (BCS) can be used as an indirect measure of nutritional status (Ric hards et al., 1986). Body condition score affected reproductive functi on in Experiment 2. Cows with a BCS 5 had greater conception and thirty-d ay pregnancy rates, and tende d (P = 0.08) to have greater synchronized pregnancy rates compared to cows with a BCS < 5. These findings are consistent with other reports stressing the importance of BCS on response to a synchronized AI breeding and subsequent pregnancy rates (Stevenson et al ., 2000; Dejarnette et al., 2004; Larson et al.,

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68 2006). Cows with a BCS 5 had a 15.7% greater th irty-day pregnancy rate compared to cows with a BCS < 5, indicating that importance of body condition at br eeding on subsequent pregnancy rates early in the breeding season. Postpartum interval, measured as days from calving to the start of the synchronization protocol, often has a strong influence on the number of cows cycling within a herd (Stevenson et al., 2000; Lamb et al., 2001). Days postpar tum had significant and similar effects on reproductive performance in Experiments 1 and 2 so the results will be discussed together. In Experiment 1, estrous response was increas ed by 28.8%, conception rate by 37.8%, and synchronized pregnancy rate by 40.5% for long ( 70 d) compared to short ( 50 d) postpartum cows. Similarly, in Experiment 2, estrous resp onse was increased by 29.3%, conception rate by 27.2%, timed-AI pregnancy rate by 32.2%, and s ynchronized pregnancy rate by 36.1% for long ( 60 d) compared to short ( 40 d) postpartum cows. These data are similar to that of Stevenson et al. (2000) who observed great er reproductive performance for cows with longer postpartum intervals in lactating Bos taurus beef cows. They reported a 5.7% increase in cycling status for each 10 d increase in postpartum length. Postpa rtum data from both experiments stress the importance of knowing where cows are in relatio n to calving when starti ng a synchronization program in postpartum lactating cows of Bos indicus Bos taurus breeding. In summary, estrous response, conception, timed-AI, synchron ized pregnancy, and thirtyday pregnancy rates were similar for new CI DR vs. once-used CIDR and cloprostenol vs. dinoprost treatments when evaluated in Bos indicus Bos taurus cows in both experiments. As interval from PGF2 to the onset of estrus increased, con ception rate decreased significantly in Experiment 1. Synchronized pregnancy rates were greater with the addition of a timed-AI in Experiment 2 compared to no timed-AI in Experiment 1. For both experiments, estrous

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69 response, conception, synchronized pregnancy, and thirty-day pregnancy rates increased as the days from calving to the start of the synchronization pr otocol increased. Implications Synchronized pregnancy rates were similar between cloprostenol sodium and dinoprost tromethamine treatments in Select Synch + CIDR protocols. The decreased estrous response of the Select Synch + CIDR protocol compromises the protocols overall effectiveness, but is improved with addition of a timed-AI 3 d after CIDR removal. Additional research needs to be conducted to determine if a once-used CIDR co nsistently results in decreased reproductive performance in postpartum cows compared to a new CIDR in a Select Synch + CIDR timed-AI protocol. The overall effectiven ess of Select Synch protocols are significantly influenced by days postpartum at the start of treatment and prod ucers need to pay partic ular attention to when synchronization programs are implemented in relation to calving in postpartum Bos indicus Bos taurus cattle.

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70 Table 3-1 Main effects for estrous , conception and pregnancy rates of Bos indicus Bos taurus cows synchronized with controlled intravag inal progesterone-releasing device (CIDR: New vs. Used) treatments and prostaglandin F2 (Cloprostenol sodium(Cloprostenol) vs. Dinoprost tromethamine-(Dinoprost)) treatments ad ministered at CIDR removal. Number in parenthesis is the number of cows in each category (Experiment 1).a Variable n Estrous response (%)b Conception rate (%)c Synchronized pregnancy rate (%)d New CIDR 141 70.9 (141) 45.0 (100) 31.9 (141) Used CIDR 141 66.0 (141) 50.5 (93) 33.3 (141) P-value P > 0.05 P > 0.05 P > 0.05 Cloprostenol 141 68.8 (141) 48.5 (97) 33.3 (141) Dinoprost 141 68.1 (141) 46.9 (96) 31.9 (141) P-value P > 0.05 P > 0.05 P > 0.05 a All cows received GnRH (100 g) at the initiation of either a 7 d new CIDR or once-used CIDR. Cows received eith er cloprostenol sodium (500 g) or dinoprost tromethamine (25 mg) at CIDR removal. Estrus was detected for 5 d, and cows that exhibited estrus were AI approximately 8 to 12 hours later. b Percentage of cows disp laying estrus 5 d after PGF2 of total treated. c Percentage of cows pregnant to AI of th e total that exhibited estrus and were AI. d Percentage of cows pregnant during the synchronized breeding of the total treated.

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71 Table 3-2 Simple treatment effects for estrous , conception and pregnancy rates of Bos indicus Bos taurus cows synchronized with controlled intr avaginal progesterone-releasing device (CIDR: New vs. Used) treatments and prostaglandin F2 (Cloprostenol sodi um(Cloprostenol) vs. Dinoprost tromethamine-(Dinoprost)) treatments administered at CIDR removal. Number in parenthesis is the number of co ws in each category (Experiment 1).a Treatments N Estrous response (%)b Conception rate (%)c Synchronized pregnancy rate (%)d New CIDR Cloprostenol 71 71.8 (71) 47.1 (51) 33.8 (71) New CIDR Dinoprost 70 70.0 (70) 42.9 (49) 30.0 (70) Used CIDR Cloprostenol 70 65.7 (70) 50.0 (46) 32.9 (70) Used CIDR Dinoprost 71 66.2 (71) 51.1 (47) 33.8 (71) a All cows received GnRH (100 g) at the initiation of either a 7 d new CIDR or once-used CIDR. Cows received eith er cloprostenol sodium (500 g) or dinoprost (25 mg) at CIDR removal. Estrus was detected for 5 d, and cows that exhibited estrus were AI approximately 8 to 12 hours later. b Percentage of cows disp laying estrus 5 d after PGF2 of total treated. c Percentage of cows pregnant to AI of th e total that exhibited estrus and were AI. d Percentage of cows pregnant during the synchronized breeding of the total treated.

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72 Table 3-3 Days postpartum (DPP) effects on estrous , conception and pregnancy rates of Bos indicus Bos taurus cows synchronized with two cont rolled intravaginal progesteronereleasing device (CID R) treatments and two prostaglandin F2 treatments administered at CIDR removal. Number in parenthesis is th e number of cows in each category (Experiment 1).a DPP n Estrous response (%)b Conception rate (%)c Synchronized pregnancy rate (%)d 50 109 57.8e (109) 27.0e (63) 15.6e (109) 51-69 91 64.8e (91) 49.2f (59) 31.9f (91) 70 82 86.6f (82) 64.8f (71) 56.1g (82) a All cows received GnRH (100 g) at the initiation of either a 7 d new CIDR or once-used CIDR. Cows received eith er cloprostenol sodium (500 g) or dinoprost tromethamine (25 mg) at CIDR removal. Estrus was detected for 5 d, and cows that exhibited estrus were AI approximately 8 to 12 hours later. b Percentage of cows disp laying estrus 5 d after PGF2 of total treated. c Percentage of cows pregnant to AI of th e total that exhibited estrus and were AI. d Percentage of cows pregnant during the synchronized breeding of the total treated. e,f,g Means without a common superscript w ithin a column differ (P < 0.05)

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73 Table 3-4 Main effects for estrous, con ception and pregnancy rates of Bos indicus Bos taurus cows synchronized with controlled intravag inal progesterone-releasing device (CIDR: New vs. Used) treatments and prostaglandin F2 (Cloprostenol sodium(Cloprostenol) vs. Dinoprost tromethamine-(Dinoprost)) treatments ad ministered at CIDR removal. Number in parenthesis is the number of cows in each category (Experiment 2).a Treatments N Estrous response (%)b Conception rate (%)c Timed-AI pregnancy rate (%)d Synchronized pregnancy rate (%)e New CIDR 125 51.2 (125) 67.2 (64) 32.8 (61) 50.4 (125) Used CIDR 130 49.2 (130) 53.1 (64) 31.8 (66) 42.3 (130) P-value P > 0.05 P = 0.10 P > 0.05 P > 0.05 Cloprostenol 129 53.5 (129) 59.4 (69) 28.3 (60) 45.0 (129) Dinoprost 126 46.8 (126) 61.0 (59) 35.8 (67) 47.6 (126) P-value P > 0.05 P > 0.05 P > 0.05 P > 0.05 a All cows received GnRH (100 g) at the initiation of either a 7 d new CIDR or once-used CIDR. Cows received eith er cloprostenol sodium (500 g) or dinoprost tromethamine (25 mg) at CIDR removal. Estrus was detected for 3 days, and cows that exhibited estrus were AI approximately 8 to 12 hours later. Cows which ha d not displayed estrus were timed-AI at 76 to 80 hours and given GnRH. b Percentage of cows disp laying estrus 3 d after PGF2 of total treated. c Percentage of cows pregnant to AI of th e total that exhibited estrus and were AI. d Percentage of cows pregnant to time d-AI of the total that were timed-AI. e Percentage of cows pregnant during the synchronized breeding of the total treated.

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74 Table 3-5 Simple treatment effects for estrous , conception and pregnancy rates of Bos indicus Bos taurus cows synchronized with controlled intr avaginal progesterone-releasing device (CIDR: New vs. Used) treatments and prostaglandin F2 (Cloprostenol sodi um(Cloprostenol) vs. Dinoprost tromethamine-(Dinoprost)) treatments administered at CIDR removal. Number in parenthesis is the number of co ws in each category (Experiment 2).a Treatments n Estrous response (%)b Conception rate (%)c Timed-AI pregnancy rate (%)d Synchronized pregnancy rate (%)e New CIDR Cloprostenol 63 60.3 (63) 68.4 (38) 20.0 (25) 49.2 (63) New CIDR Dinoprost 62 41.9 (62) 65.4 (26) 41.7 (36) 51.6 (62) Used CIDR Cloprostenol 66 47.0 (66) 48.4 (31) 34.3 (35) 40.9 (66) Used CIDR Dinoprost 64 51.6 (64) 57.6 (33) 29.0 (31) 43.8 (64) a All cows received GnRH (100 g) at the initiation of either a 7 d new CIDR or once-used CIDR. Cows received eith er cloprostenol sodium (500 g) or dinoprost tromethamine (25 mg) at CIDR removal. Estrus was detected for 3 days, and cows that exhibited estrus were AI approximately 8 to 12 hours later. Cows which ha d not displayed estrus were timed-AI at 76 to 80 hours and given GnRH. b Percentage of cows disp laying estrus 3 d after PGF2 of total treated. c Percentage of cows pregnant to AI of th e total that exhibited estrus and were AI. d Percentage of cows pregnant to time d-AI of the total that were timed-AI. e Percentage of cows pregnant during the synchronized breeding of the total treated.

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75 Table 3-6 Days postpartum (DPP) effects on estrous , conception and pregnancy rates of Bos indicus Bos taurus cows synchronized with two cont rolled intravaginal progesteronereleasing device (CID R) treatments and two prostaglandin F2 treatments administered at CIDR removal. Number in parenthesis is th e number of cows in each category (Experiment 2).a DPP N Estrous response (%)b Conception rate (%)c Timed-AI pregnancy rate (%)d Synchronized pregnancy rate (%)e < 40 74 39.2f (74) 44.8f (29) 20.0f (45) 29.7f (74) 40-59 108 45.4f (108) 57.1f,g (49) 33.9f,g (59) 44.4g (108) 60 73 68.5g (73) 72.0g (50) 52.2g (23) 65.8h (73) a All cows received GnRH (100 g) at the initiation of either a 7 d new CIDR or once-used CIDR. Cows received eith er cloprostenol sodium (500 g) or dinoprost tromethamine (25 mg) at CIDR removal. Estrus was detected for 3 days, and cows that exhibited estrus were AI approximately 8 to 12 hours later. Cows which ha d not displayed estrus were timed-AI at 76 to 80 hours and given GnRH. b Percentage of cows disp laying estrus 3 d after PGF2 of total treated. c Percentage of cows pregnant to AI of th e total that exhibited estrus and were AI. d Percentage of cows pregnant to time d-AI of the total that were timed-AI. e Percentage of cows pregnant during the synchronized breeding of the total treated. f,g,h Means without a common superscript within a co lumn differ (P < 0.05).

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76 50.0 63.6 48.9 28.6 18.2 0 10 20 30 40 50 60 70 80 90 100 4860728496Conception rate, % Figure 3-1: Effect of interval fr om PGF to the onset of estrus in Bos indicus Bos taurus cows synchronized with two controlled intravaginal progesterone-releasing de vice (CIDR) treatments and two prostaglandin F2 treatments administered at CIDR removal. Means without a common letter between columns differ (P < 0.05). Numbers in parenthesis indicate the number of cows inseminated for each category (Experiment 1). (68) (44) (45) (14) (22) Interval from PGF2 to onset of estrus, h a a a a, b b

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77 CHAPTER 4 EFFECTIVENESS OF CLOPROSTENOL SODIUM VS. DINOPROST TROMETHOMINE IN A GnRH/CIDR + PGF2 SYNCHRONIZATION PROTOCOL IN ANGUS, BRAHMAN, and BRAHMAN ANGUS COWS AND HEIFERS Introduction Cattle of Bos indicus breeding are commonly used by producers in tropical and subtropical regions of the world due to their superior tolerance to high temperatures, humidity, parasites, and utiliz ation of low quality forages compared to Bos taurus cattle. Slight differences in the reproduc tive physiology of Bos indicus compared to Bos taurus cattle include a reduced capacity for LH secretion (Randel, 1984), an earlier LH surg e and ovulation relative to the onset of estrus (Munro, 1988), and a greater sensitiv ity to exogenous gonadotrophins (Randel, 1984). Behavioral differences are also apparent in Bos indicus cattle, including a shorter, less evident estrus (Galina et al., 1982) and increased occu rrence of ‘silent estrus’ (Galina et al., 1996). Utilization of the estrous synchronization pr otocol of GnRH followed 7 d later by PGF2 is commonly used in Bos taurus cows (Thatcher et al., 1989; Purs ley et al., 1995; Stevenson et al., 2000). A common problem with the GnRH + PGF2 system is expression of estrus several days prior to PGF2 (DeJarnette et al., 2001), which can be prevented with the addition of a progestogen between the GnRH and PGF2 treatments (DeJarnette et a., 2004). Addition of a progestogen like melengestrol acetate (Stevenson et al., 2000) or the intravaginal progesterone releasing device (Lucy et al., 2001; Larsen et al., 2006) to the GnRH + PGF2 system can also have a beneficial effect by increasing the number of anestrous cows that exhibit estrous cycles. Studies using the GnRH + PGF2 systems with (Lemaster et al., 2001) or without (Barros et al., 2000; Hiers et al., 2003) a progest ogen have been conducted in Bos indicus Bos taurus cattle with limited success.

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78 The effectiveness of prostaglandins like cl oprostenol sodium and dinoprost tromethamine to synchronize estrus has been well documented in Bos taurus cattle (Young and Anderson, 1986; Salverson et al., 2002), but only one comparison in cattle of Bos indicus breeding has been conducted (Hiers et al., 2003). Hiers et al. (2003) reporte d only a numerically greater synchronized pregnancy rate for cloprostenol sodium compared to dinoprost tromethamine in Bos indicus Bos taurus cows synchronized with GnRH + PGF2 system combined with melengestrol acetate. Therefore, the first objective of these experi ments was to evaluate the effectiveness of two PGF2 treatments, cloprostenol sodium and di noprost tromethamine, in a GnRH + PGF2 synchronization program combined with a CIDR for synchronizing heifers and postpartum lactating cows. A second objective was to ev aluate breed effects for cows of Angus ( Bos taurus ), Brahman ( Bos indicus ), and Brahman Angus breeding for resp onses to the GnRH + PGF2 synchronization program. Materials and Methods Two experiments were conducted over a two year period from February to May of 2005 and 2006 at the University of Florida, Department of Animal Sciences Beef Research Unit. In Experiment 1, multiparous postpartum lactati ng cows of varying degrees of Brahman ( Bos indicus ) and Angus ( Bos taurus ) breeding (n = 157, Year 1; n = 178, Year 2) were used. For year 1, cows had a mean ( SD) age of 5.3 2.5 years, days postpartum (DPP) of 69.1 14.3, body weight (BW) of 543 65 kg, and body conditi on score of 5.1 0.5 (BCS: 1 = emaciated, 9 = obese; Richards et al., 1986). For year 2, cows had an averag e age of 5.0 2.4 years, DPP of 67.0 16.7, BW of 551 64 kg, and BCS of 5.1 0.7. Breed types represented included Angus, Brahman, and different pe rcentages of Brahman Angus breeding. The Brahman Angus cows

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79 were Angus Brahman, Angus Brahman (Brangus), Angus Brahman, and Angus Brahman. On d 0, all cows rece ived GnRH (100 g i.m.; Fertagyl, Intervet, Boxmeer, The Netherlands), and a new CIDR ( 1.38 g progesterone; Eazi-Breed CIDR, Pfizer Animal Health, New York, NY) and BW and BCS were recorded. On d 7, CIDR was removed and cows were equally distributed by breed and DPP to receive either of two PGF2 treatments, which included cloprostenol sodium (500 g i.m.; Estrumate, Schering-Plough Veterinary Corp., Kenilworth, NJ) or dinoprost trom ethamine (25 mg i.m.; Lutalyse, Pfizer Animal Health, New York, NY). All cows also received an Estrus Alert patch (Estrus Alert, Western Point, Inc., Merrifield, MN) to aid in estrous detection. Estrus was visually detected three time s daily at 0700, 1200, and 1700 h for 3 d following PGF2 . Estrus was defined as a co w standing to be mounted by anot her cow and/or a half to full red Estrus Alert patch. Cows were artifi cially inseminated (AI) 8 to 12 h after an observed estrus. All cows that had not displayed estrus by 0800 h, 73 h after PGF2 were timed-AI and administered GnRH between 76 and 80 h after PGF2 . Cows in estrus at the 72 h observation were inseminated 8 to 12 h later, an d not included in the timed-AI. Cows were inseminated using frozen-thawed semen from multiple pre-assigned sires and were inseminated by three AI technicians. Pr egnancy was diagnosed approximately 29 d after insemination using a real-time B-mode ultrasoun d (Aloka 500V, Corometrics Medical Systems, Wallingford, CT) with a 5.0 MHz tr ansducer during both years. Estrous response was defined as the number of cows displaying estrus and inseminated for 3 d after PGF2 divided by the total number of cows tr eated. Conception rate was the number of cows that displayed estrus, were inseminate d and became pregnant, divided by the number of cows that displayed estrus and were inseminated. Timed-AI pregnancy ra te was the number of

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80 cows that failed to display estrus, were tim ed-AI, and became pregnant divided by the total number of cows that were timed-AI. Synchroni zed pregnancy rate was the total number of cows pregnant to the AI divided by th e total number of cows treated. The GENMOD procedure of SAS (SAS Inst. In c., Cary, NC) was used for the statistical analysis. The effects of PGF2 treatment, breed, year, and all appropriate inte ractions were evaluated for estrous response, conception, timed-AI pregnancy, and synchronized pregnancy rates. Days postpartum, BCS and cow age were included as covariates for the evaluation of estrous response, conception, ti med-AI pregnancy, and synchroni zed pregnancy rates. When covariates were significant (P < 0.05) they we re treated as independent variables. Days postpartum was divided into three categories ( 55 d, 56 to 74 d, 75 d) and cow age was divided into three categories (3 yr, 4 to 5 yrs, > 5 yrs). When significant (P < 0.05), the effects of DPP and cow age were also analyzed with treatment, year, breed, and all appropriate interactions for estrous response, conception, timed-AI pregnancy, and synchronized pregnancy rates. The effect of interval from PGF2 to the onset of estrus, PGF2 treatment, year, breed, and appropriate interactions were ev aluated for conception rate. In year 1, one cow lost a CIDR between d 0 and d 7, and incomplete data was av ailable on another cow; therefore, both cows were eliminated from all statistical analyses. In year 2, two cows lost their CIDR between d 0 and d 7 and were eliminated from all statistical analyses. In Experiment 2, two-year old virgin heifer s of varying degrees of Angus and Brahman breeding (n = 89, Year 1; n = 74, Y ear 2) were used. For year 1, heifers had a mean ( SD) BW of 433 38 kg and BCS of 5.6 0.7. For year 2, heifers had a mean BW of 465 43 kg and BCS of 5.2 0.5. Breed types represented the same six breed types as Experiment 1. Heifers were synchronized using the same protocol and estrus detection methods described in

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81 Experiment 1. Heifers were inseminated using frozen-thawed semen from multiple pre-assigned sires and were inseminated by the same three AI technicians as Experiment 1. Estrous response, conception, timed-AI pregnancy, and synchronized pregnancy rates are as described for Experiment 1. Estrous response was defined as the number of heifers displaying estrus and inseminated for 3 d after PGF2 divided by the total number of heifer s treated. Conception rate was the number of heifers that displayed estrus, were inseminated and became pregnant, divided by the number of heifers that displayed estrus and were inseminated. Timed-AI pregnancy rate was the number of heifers that failed to display estrus, were timedAI, and became pregnant divided by the total number of heifers that were timed-A I. Synchronized pregnancy rate was the total number of heifers pregnant to the AI di vided by the total numbe r of heifers treated. For statistical analysis, the GENMOD proce dure of SAS (SAS Inst. Inc., Cary, NC) was used. The effects of PGF2 treatment, breed, year, and all appropriate interactions were evaluated for estrous response, conception, timed-AI pregnancy, and synchronized pregnancy rates. Body condition score was included as a co variate. The effect of interval from PGF2 to the onset of estrus, PGF2 treatment, year, breed, and appropria te interactions were evaluated for conception rate. In year 1, two heifers lost their CIDR between d 0 and d 7, and were eliminated from all statistical analyses. In year 2, no anim als were removed from the statistical analyses. Results Estrous response was similar (P > 0.05) between PGF2 treatments and years (Table 4-1), but estrous response was affected (P < 0.05) by br eed in Experiment 1 (Table 4-2). Angus and Brangus cows had a greater (P < 0.05) estrous response compared to Brahman and other breed categories, which were similar (P > 0.05) to each other. There were no (P > 0.05) PGF2

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82 treatment breed, PGF2 treatment year, breed year, or PGF2 treatment year breed effects on estrous response. Cow age affected es trous response (P < 0.05) when included as a covariate and the age categories are presented in Table 4-3. Threeyear old cows had a decreased (P < 0.05) estrous response compared to cows that were 4 and 5 and > 5 years of age, which had similar (P > 0.05) estrous responses. Days postpartum also a ffected estrous response when included as a covariate and the resulting DPP categories are presente d in Table 4-4. Short ( 55 d) postpartum cows had a lower (P < 0.05) estrous response than medium (56 to 74 d) postpartum cows, but were similar (P > 0.05) to long ( 75 d) postpartum cows. Medium postpartum cows had a similar (P > 0.05) estrous response compared to long postpartum cows. There was an interaction (P < 0.05) of PGF2 treatment by DPP on estrous response. Cows that were short and medium postpartum had simila r (P > 0.05) estrous responses between PGF2 treatments. However, long postpartum cows treat ed with cloprostenol had a greater (P < 0.05) estrous response (61.0%) compared to dinoprost tr eated cows (45.2%). Body condition score did not (P > 0.05) influence estrous response when in cluded as a covariate. The average interval from PGF2 to the onset of estrus was not effected (P > 0.05) by PGF2 treatment or breed of cow, nor did it effect (P > 0.05) conception rates. For cows that exhibited estrus, the average interval from PGF2 to the onset of estrus was 52.8 8.6 h, Conception rate was simila r (P > 0.05) between PGF2 treatments (Table 4-1) and breed of cow (Table 4-2). There were no (P > 0.05) PGF2 treatment breed, PGF2 treatment year, breed year, or PGF2 treatment year breed effects on conception ra te. However, greater (P < 0.01) conception rates were observe d for year 1 compared to year 2 (Table 4-1). Cow age also affected conception rate (P < 0.05) when included as a covariate (Table 4-3). Three-year old cows had a similar (P > 0.05) conception rate comp ared to 4 and 5 year old cows, but greater (P

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83 < 0.05) conception rates compared to cows > 5 year s of age. Cows age 4 and 5 had similar (P > 0.05) conception rates compared to cows > 5 years old. Days postpartum influenced (P < 0.05) conception rate when included as a covariate (T able 4-4). Long postpartum cows had a greater (P < 0.05) conception rate compared to short po stpartum cows, but were similar (P > 0.05) compared to medium postpartum cows. S hort postpartum cows had a similar (P > 0.05) conception rate compared to medium postpartu m cows. Body condition score did not (P > 0.05) influence conception rate when included as a covariate. Timed-AI pregnancy rate was influenced (P < 0.05) by treatment, but was not affected (P > 0.05) by breed or year. Time d-AI pregnancy rates tended to be greater (P = 0.06) for cloprostenol treated cows compared to dinoprost tr eated cows (Table 4-1). There were no (P > 0.05) PGF2 treatment breed, PGF2 treatment year, breed year, or PGF2 treatment year breed effects on timed-AI pregnancy rate. Cow age and DPP did not (P > 0.05) influence timed-AI pregnancy rates when included as covariates. Synchronized pregnancy rate wa s effected (P < 0.05) by trea tment and year (Table 4-1), but was not (P > 0.05) influenced by breed (Table 4-2). Cloprostenol treated cows had greater (P < 0.05) synchronized pregnancy rates compared to dinoprost treated cows (Table 4-1). There were no (P > 0.05) PGF2 treatment breed, PGF2 treatment year, breed year, or PGF2 treatment year breed effects on synchronized pregnanc y rate. Synchronize d pregnancy rates were greater (P < 0.01) for year 1 compared to year 2 (Table 4-1). Age did not (P > 0.05) influence synchronized pregnancy ra te when included as a covariat e. Days postpartum tended (P = 0.06) to influence synchronized pregnancy rate when included as a covariate (Table 4-4). Synchronized pregnancy rate te nded (P = 0.07) to be greater for medium postpartum cows compared to short postpartum cows, but medium postpartum cows were similar (P > 0.05) to

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84 long postpartum cows. Long postpartum cows ha d greater (P < 0.05) synchronized pregnancy rates compared to short postpartum cows. The overall estrous response, conception rate, timed-AI pre gnancy rate, and synchronized pregnancy rate pooled across treatments were 50.5, 62.1, 41.0, and 51.6%, respectively. For Experiment 2, estrous response wa s not influenced (P > 0.05) by PGF2 treatment or year (Table 4-5), but was affected (P < 0.05) by breed (Table 4-6). Although estrous response was different among breeds, there was no specific tr end for estrous response across the different degrees of Brahman breeding (Table 4-6). There were no (P > 0.05) PGF2 treatment breed or PGF2 treatment year effects on estrous response . However, there was a breed year effect (P < 0.05) on estrous response. Estrous responses for the breed groups by year were Angus 57.9 and 50.0%, Angus Brahman 18.8 and 38.5%, Angus Brahman (Brangus) 71.4 and 18.2%, Angus Brahman 44.0 and 70.0%, Angus Brahman 44.4 and 36.4%, and Brahman 61.5 and 72.7%, for years 1 and 2, respectiv ely. When included as a covariate, BCS did not (P > 0.05) influence estrous response. The average interval from PGF2 to onset of estrus for cows which displayed estrus prior to timed-AI were not affected (P > 0.05) by PGF2 treatment or breed. The average interval from PGF2 to onset of estrus was 59.1 10.5 h. The interval from PGF2 to onset of estrus did not (P > 0.05) effect conception rates. Conception rate was not in fluenced (P > 0.05) by PGF2 treatment (Table 4-5), year, breed, PGF2 treatment breed or PGF2 treatment year. Body condition score did not (P > 0.05) affect conception rate wh en included as a covariate. Timed-AI pregnancy rate was influenced (P < 0.05) by PGF2 treatment (Table 4-5), but was not affected (P > 0.05) by breed, year, PGF2 treatment breed or PGF2 treatment year.

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85 Timed-AI pregnancy rate was greater (P < 0.05) for dinoprost treated heifers compared to cloprostenol treated heifers. Body condition score had no (P > 0.05) effect on timed-AI pregnancy rate when incl uded as a covariate. Synchronized pregnancy rate was not influenced (P > 0.05) by PGF2 treatment (Table 45), year, breed, PGF2 treatment breed, and PGF2 treatment year. There was a breed year effect (P < 0.05) on synchronized pregnancy rate . Synchronized pregnancy rates for the breed groups across each year were Angus 47.4 and 75.0%, Angus Brahman 18.8 and 61.5%, Angus Brahman (Brangus) 57.1 and 18.2%, Angus Brahman 48.0 and 45.0%, Angus Brahman 44.4 and 27.3%, and Brahman 53.9 and 63.6% , for years 1 and 2, respectively. Body condition score did not (P > 0.05) effect synchr onized pregnancy rate when included as a covariate. The overall estrous response, concepti on, timed-AI pregnancy, and synchronized pregnancy rates pooled across treatmen ts were 48.5, 60.8, 31.0, 45.5%, respectively. Discussion There were no PGF2 treatment (cloprostenol sodium vs. dinoprost tromethamine) effects on estrous response or conception rates for pos tpartum lactating Angus, Brahman, and Brahman Angus cows in Experiment 1. Timed-AI pregnancy rate tended to be greater for cows that received cloprostenol compared to those rece iving dinoprost. Synchronized pregnancy rates were greater for cloprostenol cows compared to dinoprost cows. Larson et al. (2006) observed a similar 3 d estrous response in Bos taurus cows synchronized with the Select Synch + timed-AI with a CIDR protocol compared to the Angus co ws in the current study. In contrast, the 3 d estrous response for Brahman and Brahman Angus cows was 21.1% less compared to the Larson et al. (2006) study. Additionally, the 3 d estrous response observed in Experiment 1 was

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86 17.5% greater compared to a previous report by Le master et al. (2001) in postpartum lactating Bos indicus Bos taurus beef cows synchronized with the Select Synch + timed-AI protocol without a CIDR. Addition of a CIDR is lik ely the primary reason for the considerable differences between the Lemaster et al. (2001) and the present study. Incorporation of a CIDR into a synchronization protocol i nduces estrous cycles in some an estrous cows (Lucy et al., 2001; Larsen et al., 2006). However, it is unclear what effect the CIDR had on inducing estrous cycles in anestrous cows in the current study, as estrous cycling status was not evaluated. The estrous response and synchronized pregnancy rate observed in Angus, Brahman, and Brahman Angus cows in Experiment 1 are greater than some repo rts (Stevenson et al., 2000), but less than others (Larson et al., 2006) in Bos taurus cows synchronized with the Select Synch + timed-AI protocol with CIDR. Estrous response and synchronized pr egnancy rates for Experiment 1 were similar to results obtained in a recent study in our lab in postpartum lactating Bos indicus Bos taurus cows synchronized with the Select Synch + tim ed-AI protocol with a CIDR (See Chapter 3, Experiment 2). There was a breed effect on es trous response with Angus co ws having a greater estrous response compared to all breed compositions except the Angus Brahman cows. While estrous response for the Brahman Angus cows in this study were gr eater than previous studies in Bos indicus Bos taurus cows (Lemaster et al., 2001), it is less than observed in Bos taurus cows (Larson et al., 2006). Estrus is difficult to detect in cattle of Bos indicus breeding (Randel, 1984; Galina et al., 1994). Decreased durations of estrus, combined with an increased incidence of “silent estrus” in cattle of Bos indicus breeding (Galina et al., 1996) make it difficult to detect, even during a synchronized estrus. Estrous dete ction was probably adequate, as cows were intensely monitored by visual detection during the morning, mid-day, and early evening hours.

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87 Furthermore, use of Estrus Alert patches should have aided in detecting any missed estruses, which began in, or occurred entire ly during the evening hours. It is unclear what effect estrous cycling status had on estrus res ponse since it was not determined. Even in cows known to have had estrous cycles at the start of a synchronization protocol, Lema ster et al. (2001) reported a 3 d estrous response of only 45.7% in Bos indicus Bos taurus cows synchronized with the Select Synch + timed-AI protocol. Conception rate was not influenced by PGF2 treatment. Similar conception rates have been reported in Bos taurus cows administered cloprostenol or dinoprost on d 8 of the estrous cycle (Seguin et al., 1985). Larson et al. (2006) observed similar conception rates in Bos taurus cows synchronized with the Select Synch + timed -AI protocol with a CIDR compared to the current study with all breeds pooled and Angus ( Bos taurus ) alone. Lemaster et al. (2001) observed similar conception rates to the current study in Bos indicus Bos taurus synchronized with the Select Synch + timed-AI protocol without a CIDR. Theref ore, it appears that if cows exhibit estrus, breed composition does not appear to have a signific ant effect on conception rates. Timed-AI pregnancy rate tended to be grea ter (14.3%) for cloprostenol compared to dinoprost PGF2 treatments. This result is in contrast to a previous study in our lab (See Chapter 3) that reported no difference in timed-AI pre gnancy rates between cloprostenol and dinoprost in Bos indicus Bos taurus cows synchronized with the same Select Synch + timed-AI protocol with a CIDR. The reason for the difference is unclear. Additional research will have to be conducted to determine if cloprostenol is a more effective luteolysin than dinoprost, which could result in more cows undergoing luteolysis a nd thereby ovulating to GnRH at timed-AI. Whatever the reason for the increase, the fact that timed-AI pregnancy rates ranged from 30 to 50% across all breed types and allowed all cows an opportunity to be inseminated, reflects the

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88 importance of this type of timed-AI program in Brahman and Brahman Angus cows. Larson et al. (2006) reported a 15.0% lower timed-AI pregna ncy rate compared to all breeds when pooled and 12.5% lower than Angus cows in the present study. This re sult is likely due to a decreased estrous response in the current study compared to that of the Larson et al. (2006) study. Differences in follicle development probably delay the expression of estrus in the Brahman and Brahman Angus cows to around the time the timed-AI is performed. Therefore, although only about 50% of these cows display es trus over the 3 d period following PGF2 , many of the remaining cows are ready to exhibit estrus aroun d the timed-AI. In comparison, the Larson et al. (2006) study observed 69.2% of cows in estrus over the 3 d period following PGF2 , as Bos taurus cows likely display estrus sooner following PGF2 compared to Bos indicus or Bos indicus Bos taurus cows. Timed-AI pregnancy rates are then reduced for these cows, as a majority of synchronized cows have already displa yed estrus and the timed-AI includes a greater proportion of non-responders compared to a timed-AI in Bos indicus type cattle. Synchronized pregnancy rate was greater for cloprostenol compared to dinoprost treated cows. The increase was due to small and nonsignificant increases in estrous response, conception rate and timed-AI pregnancy rates for cloprostenol treated cows, which had an additive effect and resulted in a significant increa se in synchronized pregna ncy rates compared to dinoprost treated cows. In contrast, a similar increase in pregnancy rate for cloprostenol compared to dinoprost treated cows was not observed in Bos taurus cows when administered on d 8 of the estrous cycle (Seguin et al., 1985) . Although breed had no significant effect on synchronized pregnancy rate, Angus cows had numerically greater synchronized pregnancy rates compared to all other breed combinati ons with the exception of the Angus Brahman cows. Synchronized pregnancy rates for the Brahman Angus cows were 16.1% greater than a report

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89 by Lemaster et al. (2001) in Bos indicus Bos taurus cows synchronized with the Select Synch + timed-AI protocol without a CIDR. Larson et al. (2006) reported a 6.4 % greater synchronized pregnancy rate in Bos taurus cows synchronized with the Select Synch + timed-AI protocol with a CIDR, compared to the current study with al l breeds pooled. Larson et al. (2006) observed similar synchronized pregnancy rates compared to the Angus ( Bos taurus ) cows in the current study. Additional research with th e same breed types will need to be conducted to determine if synchronized pregnancy rates cont inue to be greater for clopro stenol compared to dinoprost treated cows, and if they conti nue to be similar between the breed types used in the present study. Cow age had a significant effect on estrous response and conception rate. Estrous response was less for 3-year old cows compared to cows 4 years of age, but conception rates were greater for 3 to 5 year old cows compared to cows > 5 years of age. It should be noted that the 3-year old cows were primiparous cows having their first calf. Hence, the decreased estrous response could have resulted from a decreased pe rcentage of 3-year olds going through estrous cycles at the start of the treatment. However, estrous cycling status was not determined in the present study. With that said, th e 3-year olds still had the grea test conception rate of all age groups. In a multi-year study by Renquist et al. (2006), cow age had no effect on pregnancy rate in multiparous Bos taurus cows. However, the Renquist et al. (2006) study did not include primiparous cows, as the current study did. Postpartum interval, measured as days from calving to breeding, can have a strong influence on the number of cows cycling within a herd at the start of the breeding season (Stevenson et al., 2000; Lamb et al., 2001). Such a phenomenon appears to be the case in the current study. Estrous response was incr eased by 11.4%, conception rate by 26.4%, and

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90 synchronized pregnancy rate by 14.4% for long ( 75 d) compared to short ( 55 d) postpartum cows. Timed-AI pregnancy rate was not infl uenced by DPP. These data are supported by Stevenson et al. (2000) who observed greater reproductive performance with longer postpartum intervals in lactating Bos taurus beef cows. They reported a 5.7% increase in cycling status for each 10 d increase in postpartum length. Data from the present study and others (Stevenson et al., 2000; Lamb et al., 2001) indicates the importa nce of knowing where cows are in relation to calving when starting a synchronization progr am in postpartum lactating cows of Bos taurus , Bos indicus, and Bos indicus Bos taurus breeding. In Experiment 2, 3 d estrous response was not affected by PGF2 treatment for Bos taurus, Bos indicus, and Bos indicus Bos taurus heifers. Breed had a significant effect on estrous response as the Angus, Angus Brahman, and Brahman heifers had similar but greater estrous responses than the other breed combinations of Angus Brahman breeding. In Bos taurus heifers synchronized with the Select Sync h + timed-AI with a CIDR, Lamb et al. (2006) observed a 25.5% greater 3 d estrous resp onse compared to the current study across all breeds. Additionally, Lamb et al. (2006) obser ved 18.4% greater estrous response compared to the Angus ( Bos taurus ) heifers in the current st udy. It should be noted that heifers in the current study were two years old and most heifers shoul d have been cycling at the start of the experiment. Furthermore, animal numbers used in the experiment are small and additional numbers are needed to adequately study the breed effects. Finally, heifers were managed in a single group throughout the experime nt and this should have added to an increased ability to detect estrus in the Brahman, and Brahman Angus heifers. Several st udies have reported that estrus is easier to detect when multiple animals are in estrus at the same time and there is an

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91 increase in the mounting activity of heifers (H urnick et al., 1975; La ndaeta-Hernandez et al., 2002) Conception rates were similar between PGF2 treatments, which is in agreement with Salverson et al. (2002) who observed similar c onception rates for clopr ostenol and dinoprost treatment administered 19 d after a 14 d MGA treatment in Bos taurus heifers. Conception rates were also similar across all breed combinations, similar to Experiment 1. Lamb et al. (2006) observed a similar conception rate in Bos taurus heifers synchronized with the Select Synch + timed-AI with a CIDR compared to the Angus and across all breeds in the current study. Rae et al. (1999) also observed similar first service conception rates in A ngus, Brahman, and Angus Brahman synchronized with an estradiol/proge stin injection followed by a 9 d norgestomet implant. These data suggest that as long as a he ifer exhibits estrus, conception rates will likely be similar across several breed ty pes of Angus, Brahman, and Brahman Angus heifers when synchronized with a Select Synch + timed-AI with a CIDR or similar type protocol. Timed-AI pregnancy rate wa s greater for dinoprost trea ted heifers compared to cloprostenol treated heifers, whic h is opposite to the results from Experiment 1. Salverson et al. (2002) observed a similar timed-AI pregnancy rate for cloprostenol and dinoprost administered 19 d after a 14 d MGA treatment in a small group of Bos taurus cows. It is unclear why timedAI pregnancy rate was decrease d for cloprostenol compared to dinoprost treated heifers in the current study, as all previous repo rts all indicate either similar or numerically greater pregnancy rates for cloprostenol compared to dinoprost. Timed-AI pregnancy rate was similar across all breeds in the current study. Larson et al. (2006) observed similar timed-AI pregnancy rates to the current study in Bos taurus heifers synchronized with the Se lect Synch + timed-AI with a CIDR protocol.

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92 Synchronized pregnancy rate was similar betw een cloprostenol and di noprost treatments. This observation is in agreement with Salverso n et al. (2002) who repor ted similar synchronized pregnancy rates for cloprostenol and dinoprost treatments administered 19 d after a 14 d MGA treatment in Bos taurus heifers. The synchronized pregna ncy rates for Angus and Brahman heifers were similar but greater th an the other breed combinations. Larson et al. (2006) observed 11.6% greater synchronized pregnancy rates in Bos taurus heifers synchronized with the Select Synch + timed-AI with a CIDR compared to all breeds pooled in the current study. In addition, the synchronized pregnancy rates for the Angus and Brahman were similar to Bos taurus heifers synchronized with a 14 d MGA treatment followe d by GnRH 17 or 19 d later (Salverson et al., 2002). No comparison has been investigated between Bos taurus, Bos indicus, and Bos indicus Bos taurus heifers synchronized with the Se lect Synch + timed-AI protocol. In summary, PGF2 treatments of cloprostenol an d dinoprost yield similar estrous responses and conception rates, but may increas e timed-AI and synchronized pregnancy rates in postpartum lactating cows of Bos indicus breeding. Days postpartum has a major effect on response of Bos indicus Bos taurus cows to a synchronization pr otocol. Breed differences were apparent only for estrous response. Implications The Select Synch + timed-AI protocol combined with a CIDR appears to be an equally effective synchronization protocol in Angus, Brahman, and Brahman Angus postpartum cows and heifers. Additionally, clopros tenol sodium resulted in great er synchronized pregnancy rates compared to dinoprost tromethamine across all breeds of postpartum cows but not 2 year old heifers. The overall effectiveness of the Sele ct Synch + timed-AI protocol combined with a CIDR was significantly influenced by days from calving to when the s ynchronization treatment

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93 was initiated. Cows with longer calving inte rvals had greater synchr onized pregnancy rates, regardless of breed. Hence, producers need to pa y particular attention to when they start a synchronization protocol in re lation to days from calving in Angus, Brahman, and Brahman Angus postpartum cows.

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94 Table 4-1 Treatment (TRT), year, and treatment year (TRT YEAR) effects for estrous, conception and pregnancy ra tes of Angus, Brahman, and Brahman Angus cows synchronized with a controlled intr avaginal progesterone-releasing device (CIDR) and two prostaglandin F2 (cloprostenol sodium: cloprostenol vs. dinoprost tromethamine: dinoprost) treatments admini stered at CIDR removal. Number in parenthesis is the number of cows in each category (Experiment 1).a Treatments N Estrous response (%)b Conception rate (%)c Timed-AI pregnancy rate (%)d Synchronized pregnancy rate (%)e Cloprostenol 166 54.2 (166) 64.4 (90) 48.7 (76) 57.2 (166) YEAR 1 78 57.7 (78) 82.2 (45) 54.6 (33) 70.5 (78) YEAR 2 88 51.1 (88) 46.7 (45) 44.2 (43) 45.5 (88) Dinoprost 169 46.8 (169) 59.5 (79) 34.4 (90) 46.2 (169) YEAR 1 79 48.1 (79) 71.1 (38) 39.0 (41) 54.4 (79) YEAR 2 90 45.6 (90) 48.8 (41) 30.6 (49) 38.9 (90) P value TRT ns ns 0.06 < 0.05 YEAR ns < 0.01 ns < 0.05 TRT YEAR ns ns ns ns a All cows received GnRH (100 g) at the initiation of a 7 d CIDR treatment. Cows received either cloprostenol sodium (500 g) or dinoprost tromethamine (25 mg) at CIDR removal. Estrus was detected for 3 d, and cows that exhibite d estrus were AI approximately 8 to 12 h later. Cows which had not displayed estrus were timed-AI at 76 to 80 h and received GnRH. b Percentage of cows disp laying estrus 3 d after PGF2 of total treated. c Percentage of cows pregnant to AI of th e total that exhibited estrus and were AI. d Percentage of cows pregnant to time d-AI of the total that were timed-AI. e Percentage of cows pregnant during the synchronized breeding of the total treated. f No (P > 0.05) TRT BR, TRT YEAR, BR YEAR, TRT BR YEAR effects.

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95 Table 4-2 Estrous, conception and pregnancy ra tes for Angus (AN), Brahman (BR), and Brahman Angus cows synchronized with a controlled intravaginal progesteronereleasing device (CIDR) and two prostaglandin F2 (cloprostenol sodium and dinoprost tromethamine) treatments administered at CIDR removal. Number in parenthesis is the number of cows in each category (Experiment 1).a Breed Typef Variable AN 3/4 AN 5/8 AN 1/2 AN 1/4 AN BR n 70 70 35 97 32 31 Estrous response (%)b 62.9g (70) 44.3h (70) 68.6g (35) 45.4h (97) 37.5h (32) 45.2h (31) Conception rate (%)c 68.2 (44) 54.8 (31) 50.0 (24) 72.7 (44) 50.0 (12) 57.1 (14) Timed-AI pregnancy rate (%)d 38.5 (26) 48.7 (39) 36.4 (11) 45.3 (53) 35.0 (20) 23.5 (17) Synchronized pregnancy rate (%)e 57.1 (70) 51.4 (70) 45.7 (35) 57.7 (97) 40.6 (32) 38.7 (31) a All cows received GnRH (100 g) at the initiation of a 7 d CIDR treatment. Cows received either cloprostenol sodium (500 g) or dinoprost tromethamine (25 mg) at CIDR removal. Estrus was detected for 3 d, and cows that exhibite d estrus were AI approximately 8 to 12 h later. Cows which had not displayed estrus were timed-AI at 76 to 80 h and received GnRH. b Percentage of cows disp laying estrus 3 d after PGF2 of total treated. c Percentage of cows pregnant to AI of th e total that exhibited estrus and were AI. d Percentage of cows pregnant to time d-AI of the total that were timed-AI. e Percentage of cows pregnant during the synchronized breeding of the total treated. f Breed type by fraction of Angus breeding with remainder being Brahman breeding. g,h Means without a common subscript within a row differ (P < 0.05).

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96 Table 4-3 Cow age effects for estrous, conception and pregnancy rates of Angus, Brahman, and Brahman Angus cows synchronized with a controlled intravaginal progesterone-releasing device (C IDR) and two prostaglandin F2 (cloprostenol sodium and dinoprost tromethamine) treatments admini stered at CIDR removal. Number in parenthesis is the number of cows in each category (Experiment 1).a Cow age, yr Variable 3 4 5 > 5 Estrous response, (%)b 37.3f (110) 53.8g (119) 60.4g (106) Conception rate, (%)c 73.2f (41) 65.6f,g (64) 51.6g (64) Timed-AI pregnancy rate, (%)d 39.1 (69) 43.6 (55) 40.5 (42) Synchronized pregnancy rate (%)e 51.8 (110) 55.5 (119) 47.2 (106) a All cows received GnRH (100 g) at the initiation of a 7 d CIDR treatment. Cows received either cloprostenol sodium (500 g) or dinoprost tromethamine (25 mg) at CIDR removal. Estrus was detected for 3 d, and cows that exhibite d estrus were AI approximately 8 to 12 h later. Cows that did not displayed estrus were timed-AI at 76 to 80 h and received GnRH. b Percentage of cows disp laying estrus 3 d after PGF2 of total treated. c Percentage of cows pregnant to AI of th e total that exhibited estrus and were AI. d Percentage of cows pregnant to time d-AI of the total that were timed-AI. e Percentage of cows pregnant during the synchronized breeding of the total treated. f,g Means without a common subscrip t within a row differ (P < 0.05).

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97 Table 4-4 The effect of days postpartu m (DPP) on estrous, conc eption and pregnancy rates of Angus, Brahman, and Brahman Angus cows synchronized with a controlled intravaginal progesterone-releasing de vice (CIDR) and two prostaglandin F2 (cloprostenol sodium and dinoprost trometha mine) treatments administered at CIDR removal. Number in parenthesis is the num ber of heifers in each category (Experiment 1).a DPP, days Variable 55 56 74 75 Estrous response (%)b 40.9f (71) 53.8g,h (132) 52.3f,h (132) Conception rate (%)c 44.8f (29) 60.6f,g (71) 71.0g (69) Timed-AI pregnancy rate (%)d 38.1 (42) 45.9 (61) 38.1 (63) Synchronized pregnancy rate (%)e 40.9f,i (71) 53.8g,j (132) 55.3g (132) a All cows received GnRH (100 g) at the initiation of a 7 d CIDR treatment. Cows received either cloprostenol sodium (500 g) or dinoprost tromethamine (25 mg) at CIDR removal. Estrus was detected for 3 days, and cows that ex hibited estrus were AI approximately 8 to 12 h later. Cows which had not displayed estrus we re timed-AI at 76 to 80 h and received GnRH. b Percentage of cows disp laying estrus 3 d after PGF2 of total treated. c Percentage of cows pregnant to AI of th e total that exhibited estrus and were AI. d Percentage of cows pregnant to time d-AI of the total that were timed-AI. e Percentage of cows pregnant during the synchronized breeding of the total treated. f,g,h Means without a common subscript within a row differ (P < 0.05). i, j Means without a common subscript w ithin a row tend to differ (P < 0.07).

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98 Table 4-5 Treatment (TRT), year, and treatment year (TRT YEAR) effects for estrous, conception and pregnancy ra tes of Angus, Brahman, and Brahman Angus heifers synchronized with a controlled in travaginal progester one-releasing device (CIDR) and two prostaglandin F2 (cloprostenol sodium: cloprostenol vs. dinoprost tromethamine: dinoprost) treatments admini stered at CIDR removal. Number in parenthesis is the number of heifer s in each category (Experiment 2).a Treatments N Estrous response (%)b Conception rate (%)c Timed-AI pregnancy rate (%)d Synchronized pregnancy rate (%)e Cloprostenol 80 48.8 (80) 66.7 (39) 19.5 (41) 42.5 (80) YEAR 1 43 44.2 (43) 79.0 (19) 16.7 (24) 44.2 (43) YEAR 2 37 54.1 (37) 55.0 (20) 23.5 (17) 40.5 (37) Dinoprost 83 48.2 (83) 55.0 (40) 41.9 (43) 48.2 (83) YEAR 1 46 50.0 (46) 52.2 (23) 34.8 (23) 43.5 (46) YEAR 2 37 46.0 (37) 58.8 (17) 50.0 (20) 54.1 (37) TRT ns ns < 0.05 ns YEAR ns ns ns ns TRT YEAR ns ns ns ns a All heifers received GnRH (100 g) at the initiation of a 7 d CIDR treatment. Heifers received either cloprostenol sodium (500 g) or dinoprost tromethamine (25 mg) at CIDR removal. Estrus was detected for 3 d, and heifers that e xhibited estrus were AI approximately 8 to 12 h later. Heifers which had not displayed estrus were timed-AI at 76 to 80 hours and given GnRH. b Percentage of heifers displayi ng estrus during the 3 d after PGF2 of the total treated. c Percentage of heifers pregnant to AI of the total that exhibited estrus and were AI. d Percentage of heifers pre gnant to timed-AI of the total that were timed-AI. e Percentage of heifers pregnant during the 3 day synchronized breeding of the total treated. f No (P > 0.05) TRT BR, TRT YEAR, TRT BR YEAR interactions.

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99 Table 4-6 Breed effects for estrous, conception and pregnancy rates of Angus (AN), Brahman (BR), and Brahman Angus heifers synchroni zed with a controlled intravaginal progesterone-releasing de vice (CIDR) and two prostaglandin F2 (cloprostenol sodium and dinoprost trometha mine) treatments administered at CIDR removal by breed. Number in parenthesis is the number of heif ers in each category (Experiment 2).a Breed Typef Variable AN 3/4 AN 5/8 AN 1/2 AN 1/4 AN BR n 27 29 18 45 20 24 Estrous response (%)b 55.6g,j (27) 27.6h,i (29) 38.9g,i (18) 55.6g,j (45) 40.0g,h (20) 66.7j (24) Conception rate (%)c 53.3 (15) 62.5 (8) 71.4 (7) 56.0 (25) 50.0 (8) 75.0 (16) Timed-AI pregnancy rate (%)d 58.3 (12) 28.6 (21) 9.1 (11) 35.0 (20) 25.0 (12) 25.0 (8) Synchronized pregnancy rate (%)e 55.6 (27) 37.9 (29) 33.3 (18) 46.7 (45) 35.0 (20) 58.3 (24) a All heifers received GnRH (100 g) at the initiation of a 7 d CIDR treatment. Heifers received either cloprostenol sodium (500 g) or dinoprost tromethamine (25 mg) at CIDR removal. Estrus was detected for 3 days, and heifers that exhibited estrus were AI approximately 8 to 12 h later. Heifers which had not displayed estrus were timed-AI at 76 to 80 h and received GnRH. b Percentage of heifers displayi ng estrus during the 3 d after PGF2 of the total treated. c Percentage of heifers pregnant to AI of the total that exhibited estrus and were AI. d Percentage of heifers pre gnant to timed-AI of the total that were timed-AI. e Percentage of heifers pregnant during the 3 d synchronized breeding of the total treated. f Breed type by fraction of Angus breeding with remainder being Brahman breeding. g,h,i,j Means without a common subscrip t within a row differ (P < 0.05).

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100 CHAPTER 5 FOLLICLE DEVELOPMENT, ESTROUS CHARACTERISTICS, AND EFFECTIVENESS OF A GnRH/CIDR + PGF2 SYNCHRONIZATION PROTOCOL IN POSTPARTUM LACTATING ANGUS ( Bos taurus ) AND BRANGUS ( Bos indicus Bos taurus ) BEEF COWS Introduction Historically, estrous synchroni zation systems for postpartum cows have been developed in Bos taurus cattle. One of the most common synchronization system utilizes GnRH with PGF2 7 d later (Macmillan and Thatcher, 1991; Purs ley et al., 1995), followed by 5 d of estrous detection, which is also known as the “Select Synch” protocol. A common problem with the Select Synch protocol is premature expre ssion of estrus several days prior to PGF2 , resulting in additional estrous detection (DeJarnette et al ., 2001). Addition of an exogenous progestogen between GnRH and PGF2 eliminates the need for additiona l estrous detection (Thompson et al., 1999; DeJarnette et al., 2003) with the added benefit of inducing cyclicity in some anestrous cows (Fike et al., 1997; Lucy et al., 2001). Howe ver, the resulting synchronized pregnancy rates of the GnRH + PGF2 protocols in cattle of Bos indicus breeding either with (Yelich, 2002) or without (Lemaster et al., 2001) a progestogen are less compared to Bos taurus cattle. The exact reason(s) for the compromised res ponse are unclear but it could be due to the subtle differences in the re productive physiology of cattle of Bos indicus breeding compared to Bos taurus cattle. Differences in concentratio ns of reproductive hormones and altered sensitivities of their release have been noted for LH (Griffen and Randel, 1978), estradiol (Segerson et al., 1984), and progesterone (Rhode s et al., 1982; Segerson et al., 1984) between Bos indicus and Bos taurus cattle. Cattle of Bos indicus breeding also have an increased percentage of three-wave follicle growth patterns (Rhodes et al., 1995; Zeitoun et al., 1996; Martinez et al., 2003) than Bos taurus cattle, which could be effecting the ability of GnRH to

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101 initiate ovulation and synchronize follicle development. Characteris tics associated with estrus are also different between Bos indicus and Bos taurus cattle. Estrus is more difficult to detect (Galina et al., 1982; Orihuela et al., 1983), shorter in duration, (R ae et al., 1998), and there is a greater incidence of “silent estrus” (Dawuda et al ., 1989; Lamothe-Zavaleta et al., 1991) in cattle of Bos indicus breeding. Therefore, objectives of this experiment were to compare the response of postpartum lactating Angus ( Bos taurus ) and Brangus ( Bos indicus Bos taurus ) cows to administration of GnRH concomitant with a 7 d progesterone insert with PGF2 administered at insert removal for the following variables: 1) size of follicles ovul ating to GnRH, 2) follicle size and corresponding hormone concentrations at and after insert removal, 3) estrous characteristics after PGF2 , and 4) AI and breeding season pregnancy rates. Materials and Methods The experiment was conducted from April to June of 2005 at the University of Florida, Department of Animal Sciences Santa Fe Beef Re search Unit. Postpartum lactating two-year old Angus (n = 31) and Brangus (n = 22) cows we re utilized. Throughout the experiment, cows were housed in similar sized pastures by breed, and fed stored Bermuda grass hay ad libitum and an energy supplement to meet nutrient requirem ents. For two weeks prior to the start of experiment, cows were moved through the worki ng facilities two to thr ee times per week to acclimate them to frequent handling. Day 0 was de signated as the start of the experiment. On days -12 and -2, blood samples were collected by jugular venipuncture into evacuated tubes with an anticoagulant (EDTA; Becton, Dickinson and Company, Franklin Lakes, NJ) for the determination of progesterone concentrations to determine estrous cycle status. After collection, blood samples were immediately placed on ice, centrifuged (3000 g for 15 min), and plasma

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102 was separated and stored at -20C until further analysis. A cow was determined to have estrous cycles (cycling) if either sample had progesterone concentrations 1 ng/mL, and no estrous cycles (noncycling) if progesteron e concentrations were < 1 ng/mL at both samples. At the start of synchronization, body weight (BW) was 402 35 kg for Angus and 421 52 kg for Brangus, body condition score (BCS: 1 = emaciated, 9 = obes e; Richards et al., 1986) was 4.8 0.3 for Angus and 5.2 0.4 for Brangus, hip height ( HH) was 128 3 cm for Angus and 131 4 cm Brangus, and Angus cows were 94.9 23.5 days postpartum (DPP) while Brangus cows were 91.8 13.7 DPP. To facilitate ul trasound examinations, cows were evenly distributed by estrous cycling status (cycling vs. noncycling), DPP, a nd BCS into four groups, two each of Angus and Brangus. Groups were treated identically thr oughout the experiment and each group was housed in a 4 acre pasture. One Angus group and one Brangus group began the experiment on the same day, while the remaining Angus and Brangus cows started the next day. The day that each group started the experiment was designated as d 0. On d 0 of the experiment, cows within each group received GnRH (100 g; i.m., Fertagyl, Intervet, Boxmeer, The Netherlands) and a CIDR (1.38 g progesterone; Eazi-Breed CIDR, Pfizer Animal Health, New York, NY). On days 0, 2, 7, artificial insemination (AI), and eight days after an observed synchronized estr us, plasma and serum samples were collected by jugular venipuncture for evaluati on of progesterone and estradiol, respectively. Plasma samples were treated as previously described, and serum samples were collected into sterile evacuated tubes (Becton, Dickinson and Company, Franklin Lakes, NJ), allowe d to clot at room temperature, centrifuged (3000 g for 15 min), and serum was separated and stored at -20C until further analysis.

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103 Additionally on days 0, 2, 7, AI, and eight da ys after an observed estrus, cows were evaluated by transrectal u ltrasonography using a real-time B-mode ultrasonography machine (Aloka 500V, Corometrics Medica l Systems, Wallingford, CT) with a 7.5 MHz transducer. At each ultrasonography evaluation, height and width of all luteal structures, luteal cavities, and follicles 5 mm in diameter were measured with th e internal calipers of the ultrasonography machine and their locations on the ovaries were recorded. Volume of the CL was calculated using the formula for volume of a sphere ( d3/6). When a luteal cavity was present, its volume was subtracted from the volume of the outer sphe re resulting in net lu teal volume (CL volume) represented by luteal tissue. The CL resulting from the GnRH induced ovulation was termed the accessory CL and when two CL were present, total CL volume was calculated as the sum of the volume of each CL. On d 2 of the experime nt, ultrasonography was conducted to determine if GnRH initiated ovulation, with ovulati on defined as disappearance of the largest follicle from the previous ultrasonography examination. On d 7 of the experiment, CIDR were removed and cows received PGF2 (25 mg, i.m., Prostamate, Agrilabs, St. Joseph, MO). Cows were also evaluated by ultrasonography to confirm the site of ovulation by the presence of a newly formed CL in the location of the previously ovulated follicle and all follicles 5 mm in diameter were recorded. The CIDR retention rate was 100% fo r the 7 d treatment period. Estrus was monitored using electronic heat detection monitors (HeatWatch, DDx, Boulder, CO), which were applied on either d 2 or 3 of the experiment. Estrus was defined as three or more mounts within a 4 h period with the end of estrus defined as the last mount recorded prior to a period of extended inactivity of at least 8 h (Landaet a et al., 1999). Mounts with a duration 1 sec were recorded. Duration of estrus was calculated by subtracting the time

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104 of the initial mount from the time of the last mount. Interval from PGF2 (d 7) to the onset of estrus was calculated as the interval from PGF2 to first mount as detected by HeatWatch. Total number of mounts received during estrus was also determined. Additionally, a visual, daily estrous detection was conducted at 0700, 1200, and 1700 h for 5 d following PGF2 . Visual estrus was defined as a cow standing to be mounted by another cow and (or) presence of a clear vaginal mucous discharge. Individuals c onducting visual estrous detection did not have access to any HeatWatch data during the 5 d estrous detection period. Cows were AI by a single AI technician 8 to 12 h after declared in estrus by the HeatWatch system. Cows were inseminated using frozen-thawed semen from multiple sires, which were preassigned to cows before the start of the experiment. An ultrasonography exam was performed at AI to determine follicle size and presence of any luteal structures. At both 72 and 120 h after PGF2 , any cow not displaying estrus was ultrasounded and a plasma sample was taken via jugular veinipuncture fo r evaluation of progesterone con centrations. Eight days after the expression of estrus, cows were evaluated by ultrasonography to confirm the site of ovulation of a dominant follicle at AI as indicated by the pr esence of a CL in the location of the previously ovulated follicle. A plasma sample was also coll ected to evaluate CL function as determined by progesterone concentrations. Cows that did not exhibit estrus during the synchronized breeding were evaluated by ultrasonography approximately 8 d after the end of the synchronized breeding period to determine if ovulati on occurred by the presence of a CL. Estrous detection as determined by HeatWatch and AI continued for an additional 25 d after the synchronized breeding. Angus and Brangus cows were placed in individual breeding pa stures with a single bull of good fertility for an additional 40 d breeding period. Pregnancy was diagnosed

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105 approximately 30d after AI, at the end of the breeding season, and ag ain 30 d after the end of the breeding season using a real-time B-mode ultrasound with a 5.0 MHz transducer. Corpus luteum regression was defined as the number of cows which had a CL as determined by ultrasonography evaluati on and progesterone concentrations 1 ng/mL on d 7 of the experiment and either exhibited estrus or had progesterone concentrations of < 1.0 ng/mL by 72 h following PGF2 . Cows that had progesterone < 1 ng/mL without a CL present on d 7 of the experiment as determined by ultrasonography eval uation were not included in the CL regression analysis. Follicle growth from d 7 to AI was determined by the difference in size of the eventual ovulatory follicle on d 7 from the size of the folli cle present at AI which subsequently ovulated. Days of follicle growth were the number of days from d 7 to the day of AI, and were determined in half day increments. Follicle growth rate was defined as the follicle growth from d 7 to AI, divided by the days of follicle growth. Estrous response was defined as the number of cows displaying estrus as determined by HeatWatch during the 5 d after PGF2 divided by the total numb er of cows treated. An additional visual estrous response was defined as the number of co ws displaying estrus by visual observation, divided by the total number of cows treated. Concep tion rate was defined as the number of cows that displayed estrus, were inseminated, and became pregnant, divided by the number of cows that displayed estrus and were inseminated. Synchronized pregnancy rate was the number of cows pregnant to the AI divided by the total number of cows treated. Breeding season pregnancy rate is the number of cows pr egnant at the end of 70 d breeding season, which included the 30 d AI breeding a nd a 40 d clean-up bull breeding divided by the total number treated.

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106 Progesterone concentrations were done in mu ltiple assays using RIA (Seals et al., 1998) with intraand interassay CV of 11.6 and 12.8%, respectively. Se nsitivity of the assay was 0.01 ng/mL. Estrogen concentrations were done in multiple assays using RIA (Yelich et al., 1997) with intraand interassay CV of 37.8 and 67.5%, respectively. Se nsitivity of the assay was 1.3 pg/mL. The GENMOD procedure of SAS (SAS Inst. Inc. , Cary, NC) was used for the statistical analysis of categorical data. The effect of br eed, cycling status, and in teraction were evaluated for ovulation rate to GnRH, estrous response, CL regression to PGF2 , ovulation rate after AI, conception rate, and synchronized pregnancy ra te, while DPP, BCS, and interval from PGF2 to the onset of estrus were include d as covariates. The effect of ovulation or no ovulation to GnRH, breed, and interaction were evaluated for estrous response, CL regression, ovulation rate for AI, conception rate, and synchronized pregnancy ra te, while DPP, BCS, and interval from PGF2 to the onset of estrus were included as covariates. When covariates were significant (P < 0.05) they were treated as independent variables. Days postpartum was divided into three categories, 60 d (short), 61 to 99 d (medium), and 100 d (long). When significan t, the effects of DPP and interaction with breed were evaluated for ovulat ion rate to GnRH, estrous response, ovulation rate for AI, conception rate, and synchronized pre gnancy rate. The effect of interval from PGF2 to the onset of estrus along w ith the two-way interaction with breed was evaluated for conception rate. The effect of breed, cycling status, and th e interaction on follicle diameters at GnRH, CIDR removal, and AI as well as luteal tissue vol ume and progesterone concentrations at CIDR removal and 8 d after estrus were analyzed usi ng GLM procedure of SAS. The effect of breed, cycling status, and the interacti on on estrogen concentrations at CI DR removal and AI were also analyzed using GLM procedure of SAS. The effect of breed, ovulation or no ovulation to

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107 GnRH, and the interaction on follicle growth rate s from CIDR removal to AI were analyzed using GLM procedure of SAS. The distribution of estrus after CIDR removal for breed and ovulation to GnRH status were analyzed using the LIFETEST pro cedure (survival analysis) of SAS. Results At the start of the experiment, 66.0% of the cows were cycling and cycling status was similar (P > 0.05) between Angus (74.2%; 23/31) and Brangus (54.6%; 12/22) cows. Cycling status was effected (P < 0.05) by DPP with fewe r short postpartum cycling (28.6%) compared to long postpartum cows (77.1%), while medium postpartum cows (54.6%) had a similar (P > 0.05) percent cycling compared to short and long groups. The average size of follicle that ovulated to GnRH was similar (P > 0.05) between Angus and Brangus cows (Table 5-1). Size of the fo llicle that ovulated to GnRH was not (P > 0.05) affected by cycling status, and ther e was no (P > 0.05) effect of breed cycling status. Likewise, ovulation to GnRH was similar (P > 0.05) between Angus and Brangus (Table 5-1). Ovulation rate to GnRH was not (P > 0.05) influenced by BCS or DPP when the vari ables were included as covariates. Cycling status effected (P < 0.05) ovulation to GnRH, with 54.3% of cycling and 27.8% of noncycling cows ovulating, but there was no (P > 0.05) breed cycling status effect on ovulation rate. At PGF2 , size of the follicle that eventually ovulated was similar (P > 0.05) between Angus and Brangus (Table 5-2). There were no (P > 0.05) ovulation status to GnRH, or breed ovulation status to GnRH effects on follicle si ze on d 7. Estrogen concentrations on d 7 were greater (P < 0.05) for Angus compared to Brangus cows, but were similar (P > 0.05) for cows

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108 that ovulated and failed to ovulate (Table 5-2). There was no (P > 0.05) breed cycling status effect on estrogen concentrati on on d 7 (Data not shown) Progesterone concentrations on d 7 for cows with a CL were similar (P > 0.05) between Angus and Brangus (Table 5-3) and were also similar (P > 0.05) for cows that ovulated and failed to ovulate to GnRH (Table 53). There was no (P > 0.05) breed ovulation status to GnRH effect on progesterone concentrations on d 7. Cycling status prior to GnRH influenced (P < 0.01) whether cows had high progesterone concentrations (> 2 ng/mL) on d 7, with 44.4% of noncycling and 82.9% of cycling cows with high progesterone. There was no (P > 0.05) breed cycling status effect on progesterone concentrations on d 7 (Data not shown). Cycling status affected (P < 0.01 ) the likelihood that a cow would have a CL at CIDR removal, with 50.0% of noncycling and 100.0% of cycling cows having a CL. Number of cows with a functional CL at PGF2 was similar (P > 0.05) between Angus (87.1%; 27/31) and Brangus (77.2%; 17/22) cows. In contrast, CL regression was influenced (P < 0.05) by breed, with more (P < 0.05) Angus havi ng CL regression compared to Brangus (Table 5-3). Cows that ovulated to GnRH (100.0%; 24/24) tended (P = 0.07) to have greater CL regression than cows not ovulating to GnRH (9 0.0%; 18/20). There was (P < 0.05) a breed ovulation status to GnRH effect on CL regression (Table 5-3). Corpus luteum regression was not (P > 0.05) influenced by BCS or DPP when included as covariates. The mean intervals from PGF2 to the onset of HeatWatch estrus were similar (P > 0.05) between Angus and Brangus (Table 5-4). The dist ribution of estrus was also similar (P > 0.05) between Angus and Brangus (Figure 5-1) as de termined by survival analysis. The mean intervals from PGF2 to the onset of HeatWatch estrus were not influenced (P > 0.05) by

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109 ovulation status to GnRH. Cows that ovulated to GnRH had an interval of 63 h 43 m 4 h 8 m and cows that did not ovulate ha d an interval of 65 h 14 m 4 h 35 m. There was also no (P > 0.05) breed ovulation status effect on interval from PGF2 to the onset of HeatWatch estrus. The distribution of estrus by ovulation status to GnRH is presented in Figure 5-2. When analyzed by survival analysis, distribution of estr us tended (P = 0.07) to differ between cows that ovulated compared to cows that failed to ovulate to GnRH. Cycling status and breed cycling status did not (P > 0.05) in fluence interval from PGF2 to the onset of HeatWatch estrus (Data not shown). Estrous response wa s not influenced by breed (Table 5-5). However, the mean duration of estrus was greater (P < 0.05) fo r Angus compared to Brangus (Table 5-4). Furthermore, Angus received more (P < 0.05) total mounts compared to Brangus (Table 5-4). HeatWatch estrous response was not (P > 0.05) infl uenced by cycling status prior to GnRH (Table 5-5). However, there tended (P = 0.10) to be cycling status and breed cycling status effects (P = 0.06) on HeatWatch estrous response (Table 55). A greater (P < 0.05) number of cows that ovulated to GnRH displayed estrus compared to cows that did not ovulate to GnRH (Table 5-6). However, ther e were no (P < 0.05) breed or breed ovulation to GnRH effects on HeatWatch estrous response. When included as covariates, BCS and DPP did not (P > 0.05) influence HeatWatch estrous response for either cyc ling status or ovulation to GnRH effects. It is interesting to note that four Brangus cows had “silent estruses,” and all four of those cows failed to ovulate to GnRH, but three out of the four cows had high progesterone at PGF2 . In contrast, in the one Angus cow that exhibite d a “silent estrus,” ovula tion occurred to GnRH and progesterone was high at PGF2 . The HeatWatch system detected 75.5% of cows in estrus during the 5 d after PGF2 (Angus = 80.7%; Brangus = 68.2%), which was simila r (P > 0.05) to visual estrous detection

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110 with 66.0% of cows were observed in estrus during the 5 d after PGF2 (Angus = 67.7%; Brangus = 63.6%). Size of the eventual ovulatory follicle on d 7, size of the eventual ovulatory follicle at AI, estrogen concentrations at AI, and follicle growth rate between day 7 and AI were similar (P > 0.05) between breeds and were not ( P > 0.05) in fluenced by ovulation to GnRH nor breed ovulation to GnRH (Table 5-2). There we re no (P > 0.05) cycling status or breed cycling status effects on any of the afor ementioned variables (Data not s hown). Estrogen concentrations at AI were greater (P < 0.05) for Brangus compar ed to Angus (Table 5-2). However, there were no effects (P > 0.05) of cycling status, ovulation st atus to GnRH, or the respective interactions with breed on estrogen concentrations at AI. Ovulation rate after PGF2 was similar (P > 0.05) for Angus (83.8%; 26/31) and Brangus (86.4%; 19/22). Cycling status tended (P = 0.07) to effect th e number of cows ovulating after PGF2 , with 91.4% of cycling and 72.2% of noncyc ling cows ovulating. There was (P < 0.05) a breed cycling status effect on ovulation rate after PGF2 . Cycling Angus (95.7%; 22/23) and Brangus (83.3%; 10/12) had similar (P > 0.05) ovulation rates after PGF2 , as did noncycling Angus (50.0%; 4/8) and Brangus (90% ; 9/10). In contrast, cycli ng Angus cows had a greater (P < 0.05) ovulation rate after PGF2 compared to noncycling Angus cows, while cycling and noncycling Brangus cows had similar (P > 0.05) ovulation rates after PGF2 . Ovulation rate after PGF2 was influenced (P < 0.05) by whether cows ovul ated or did not ovul ate to GnRH. Cows that failed to ovulate to GnRH had an ovulation rate after PGF2 of 75.9% (n = 22/29), while cows that ovulated to GnRH had a 95.8% (n = 23/24) ovulatio n rate after PGF2 . There was no (P > 0.05) breed ovulation to GnRH or ovulation to GnRH cycling status effects on ovulation

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111 status after PGF2 . When included as covariates, BCS and DPP did not (P > 0.05) influence ovulation rate after PGF2 . Eight days after the synchronized estrus, CL area and progesterone concentrations for cows which displayed estrus were similar (P > 0.05) between Angus (6004 446 mm2; 3.5 0.3 ng/mL) and Brangus (5921 576 mm2; 4.1 0.3 ng/mL), respectively. There were no (P > 0.05) cycling status, ovulati on status to GnRH, breed cycling status, or breed ovulation status to GnRH effects on CL area or progesterone concentrations 8 d after the synchronized estrus. The largest follicle present on the ovary was similar (P > 0.05) between Angus (13.8 0.4 mm) and Brangus (13.5 0.5 mm). Conception rate was not (P > 0.05) affected by breed or ovula tion status to GnRH (Table 5-6). There were no (P > 0.05) effects of breed ovulation status to GnRH or ovulation to GnRH cycling status on conception rate. Inte restingly, cycling status affected (P < 0.01) conception rates, as 100% of noncycling cows that displayed estrus conceived compared to only 72.4% of cycling cows. Ther e were no (P > 0.05) breed cycling status effects on conception rate. The interval from PGF2 to onset of estrus did not (P > 0.05) effect conception rate. The number of mounts a cow received as measured by HeatWatch tended to effect (P = 0.07) conception rate. Cows receiving 20, 21 to 59, and 60 mounts had conception rates of 63.6 (24), 81.0 (21), and 100.0% (8), re spectively. Conception rates were similar (P > 0.05) for cows with 20 and 21 to 59 mounts, as well as for cows with 21 to 59 and 60 mounts, but were greater (P < 0.05) for cows with 60 mounts compared to cows with 20 mounts. There were no (P > 0.05) effects of breed number of mounts received cate gories on conception rate. When included as covariates, DPP and BCS did not (P > 0.05) in fluence conception rate.

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112 Synchronized pregnancy rate was not (P > 0.05) influenced by breed or cycling status (Table 5-5). There tended (P = 0.09) to be a breed cycling status eff ect on synchronized pregnancy rate (Table 5-5). Ovulation status to GnRH ef fected (P < 0.05) synchronized pregnancy rate with 75.0% of cows that ovulated to GnRH and 48.3% of cows that did not ovulate to GnRH becoming pregnant. There tended (P = 0.08) to be an ovulation status to GnRH cycling status effect on sync hronized pregnancy rate. Noncycli ng cows that ovulated to GnRH (100.0%) had greater (P < 0.05) s ynchronized pregnancy rates compared to noncycling cows that did not ovulate to GnRH (46.2%), and cycling co ws that did (68.4%) or did not (50.0%) ovulate to GnRH. There was no (P > 0.05) of breed ovulation status to Gn RH effect on synchronized pregnancy rate. When included as a covariate, BCS did not e ffect (P > 0.05) synchronized pregnancy rate. Days postpartum influenced (P < 0.05) synchronized pregnancy rate when included as a covariate, and there was an intera ction (P < 0.05) of breed and DPP. Angus cows that were short, medium, and long postpartum had similar (P > 0.05) synchronized pregnancy rates of 50.0 (2/4), 80.0 (4/5) and 63.6% (16/22 ), respectively. Howe ver, short postpartum (100.0%; 3/3) Brangus cows had greater (P < 0.05) synchronized pregnancy rates compared to medium postpartum (83.3%; 5/6) Brangus cows . Additionally, medium postpartum Brangus cows had a greater synchronized pregnancy ra te compared to long postpartum (30.8%; 4/13) Brangus cows. During the 30 d following the synchronized br eeding period, 28.3% of cows returned to estrus. The number of cows that returned to estrus was not (P > 0.05) influenced by breed, cycling status, ovulation status to GnRH, or AI sire. Thirty-day pregnancy rate was similar (P > 0.05) between Angus (71.0%; 22/31) and Brangus ( 81.8%; 18/22). When included as covariates, DPP and BCS did not (P > 0.05) effect thirty-day pregnancy rate. Overall breeding season

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113 pregnancy rates tended (P = 0.06) to be influenced by breed with more Brangus (100%) than Angus (90.3%) being pregnant. Days postpartum affected breeding season pregnancy rate, with short, medium, and long postpartum cows at the start of the sync hronized breeding having pregnancy rates of 71.4, 100.0, and 97.1%, respectively. When included as a covariate, BCS did not (P > 0.05) effect breedi ng season pregnancy rate. Discussion Ovulation rate to GnRH was similar betw een Angus (45.2%) and Brangus (45.5%) cows. Previous work in our lab with Bos indicus Bos taurus cows indicated an ovul ation rate of 27% in cows treated with GnRH across several stages of the estrous cycle (Hie rs et al., 2006). In contrast, Geary et al. (2000) reported a 66% ovulation rate to GnRH in cycling and noncycling Bos taurus beef cows in a Select Synch protocol w ithout a CIDR. Additionally, Moreira et al., (2000) reported a 58% ovulation rate and Vasconcelos et al. (1999) a 64% ovulation rate in cycling Bos taurus cows across several stages of the estrou s cycle. The majority of the data in Bos taurus cows suggest that GnRH induces ovulati on in approximately 60% of cows across random stages of the estrous cycle, whic h was similar to previous reports in Bos taurus cows (Vasconcelos et al., 1999; Moreir a et al., 2000). The below averag e ovulation response to GnRH in Angus cows was due to the very low response to GnRH in noncycling Angus cows (12.5%) compared to the high ovulation ra te GnRH in cycling cows (56.5%). In contrast, ovulation to GnRH in Brangus cows was less than 50% fo r both cycling and noncycling cows. Although, the Brangus cows had a greater response than that reported by Hier s et al. (2006) in Bos indicus Bos taurus cows, it is still considerably less than reports in Bos taurus cows (Geary et al., 2000; Vasconcelos et al., 1999; Moreira et al., 2000). Although, the ability of GnRH to initiate ovulation can be influenced by stage of the estrous cycle in cattle of Bos indicus (Hiers et al.,

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114 2006) and Bos taurus (Vasconcelos et al., 1999; Moreira et al., 2000) breeding, stage of estrous cycle was not known in the current study. What is even less clear is why Brangus cows had GnRH induced ovulation rates less than 50% for cycling and noncycling cows. Cattle of Bos indicus breeding have an increased incidence of three and four wave ovarian follicular growth patterns (Rhodes et al., 1995; Bo et al., 2003), which could increase th e likelihood that fewer cows will respond to GnRH across random stages of the estrous cycle. With that said, size of follicle ovulating to GnRH was similar for Angus and Brangus cows, which agrees with studies in both Bos taurus (Crowe et al., 1993; Thompson et al., 1999) and cattle of Bos indicus breeding (Hiers et al., 2006). Therefore, it appears that follicl es need to be at least 10 mm in diameter when a sufficient number of LH receptors are present (Lucy et al., 1992) to ovulate to an exogenous GnRH, regardless of breed. The PGF2 induced luteolysis was greater for Angus compared to Brangus cows. The rate of luteolysis of Angus co ws is similar to a report in Bos taurus cows (Vasconcelos et al., 1999) but slightly less than another report in Bos taurus cows (Geary et al . (2000). In both studies, cattle were synchr onized with a GnRH + PGF2 protocol. The rate of luteolysis in the Brangus cows is similar to a report by Hiers et al. (2006) in cattle of Bos indicus breeding. In a recent study with Bos indicus Bos taurus heifers synchronized with 14 d MGA treatment with PGF2 19 d later, Bridges et al. (2005) reported a CL regression rate that was approximately 79.1%. However, in the same study, yearling Bos taurus heifers and two year old Bos indicus Bos taurus heifers had similar CL regression rates. Therefore, data from the present study and the literature are inconclusive as to whether cattle of Bos indicus breeding consistently have decreased luteolysis to a single PGF2 treatment compared to Bos taurus cattle. It is interesting to note that luteolysis in the Brangus cows was 100% for cows that ovulated to GnRH, but only

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115 71.4% for cows that did not ovulate to GnRH. The increased CL regression in cows that ovulated to GnRH could be due to increased progesterone output from the newly formed CL, making it and possibly any original CL present more responsive to PGF2 . Bridges et al. (2005) who observed greater CL regression in Bos indicus Bos taurus heifers with progesterone concentrations 3 ng/mL compared to heifers with pr ogesterone concentrations < 3 ng/mL. Howard and Britt (1990) as well as Moreira et al . (2000) reported that increased progesterone concentrations, particularly when an accessory CL formed from a GnRH induced ovulation was present, lead to enhanced luteolysis to PGF2 . HeatWatch estrous response was similar between Angus and Brangus cows, although, numerically (12.5%) more Angus co ws exhibited estrus compared to Brangus cows. Estrous response for Angus cows was similar to previous studies of Bos taurus cows synchronized with GnRH 6 or 7 d prior to PGF2 (Twagiramungu et al., 1992c; Gear y et al., 2000), whereas, the Brangus cows had a considerably reduced estr us response compared wi th Twagiramungu et al. (1992c) and Geary et al. (2000). Brangus cows had a 23% greater estrous response than a report by Lemaster et al. (2001) in Bos indicus Bos taurus cows synchronized with the Select Synch protocol without a CIDR. Cycling cows, regard less of breed, tended to have a greater estrous response compared to noncycling cows, which is similar to reports in both Bos taurus (Stevenson et al. 2000) and Bos indicus (Lemaster et al., 2001) cows sy nchronized with the Select Synch protocol without a CIDR. Additionally, estrous re sponse was greater for cows that ovulated to GnRH on d 0, compared to cows that did not ovulate to GnRH on d 0 regardless of breed. Therefore, the increased estrous response of cows that ovulated to GnRH can be attributed to 100% luteolysis that occurred in these cows rega rdless of breed. Cows that ovulated to GnRH would have also had follicle turnover (Macmilla n and Thatcher, 1991) resulting in development

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116 of an actively growing follicle at PGF2 . It is interesting to note that at PGF2 , cows that ovulated to GnRH had similar sizes of ovarian follicles which would eventually ovulate with similar estrogen concentrations compared to cows that did not ovulate to GnRH. Comparison of HeatWatch estrous detection to visual es trous detection yielded similar estrous responses during the 5 d period after PGF2 . However, visual estrous detection failed to detect estrus in 12.5% (5/40) of cows that HeatWatch detected in estrus. Similarly, Rae et al. (1999) also observed 12% more Angus and Brahman Angus heifers in estrus using the HeatWatch compared to visual observation. Gear y et al. (2000) reported similar estrous detection rates between HeatWatch and visual observation in Bos taurus cows. However, in a study by Stevenson et al. (1996), visu al estrous detection failed to detect estrus in 37% of the heifers that were detected in estrus by HeatWatch. It should be noted th at in the current study, four Brangus cows responded to the synchroniza tion protocol and ovulated during the 5 d after PGF2 , but were not detected in estrus by either HeatWatch or visual estrous detection. These cows were considered to have undergone a “s ilent estrus,” which is common in cattle of Bos indicus breeding (Dawuda et al., 1989; Lamothe-Zavale ta et al., 1991). Ther efore, the decreased efficiency of detecting estrus by visual observatio n and an increased incidence of “silent estrus” are primary causes of the decreased es trous response obser ved in cattle of Bos indicus breeding synchronized with the Select Synch protocol compared to Bos taurus cattle. Characteristics associated with behavioral estrus were different between Angus and Brangus cows. Although Angus and Brangus cows had similar intervals to estrus, the duration of estrus was greater for Angus compared to Br angus. In contrast, Rae et al. (1999) reported a tendency for Brahman Angus heifers to have a greater interval to estrus compared to Angus heifers synchronized with an es tradiol/progestin treatment at insertion of a 9 d norgestomet

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117 implant. Additionally, Rae et al. (1999) observed greater durations of estrus for Brahman Angus (11.9 h) compared to Angus (8.5 h) heifers, which is in contrast with the current study. Total mounts received during estrus were greate r for Angus (49.0) compared to Brangus (21.5) in the current study. On the contrary, Rae et al . (1999) observed a greate r number of mounts for Brahman Angus (37) compared to Angus (19) heifers. One possible reason for the contradicting results between the current study and the Rae et al. (1999) st udy is that the heifers in the Rae et al. (1999) study were managed as a single group compared to breeds being managed as separate groups in the present study. Breed has a signifi cant effect on estrous characteristics, as well as social behavior, particularly when ca ttle of different breeds are comingled (Landaeta-Hernandez et al., 2004), and this may have attributed to the difference in breed response between the two studies. Conception rates were similar for Angus and Brangus cows, which are similar (Geary et al. (2000) but slightly less (Stevens on et al., 2000) than reports in Bos taurus cows synchronized with the Select Synch protocol without a CIDR . Conception rates for Brangus cows were 33.9% greater than a study by Lemaster et al. (2001) in Bos indicus Bos taurus cows synchronized with the Select Synch protocol without a CIDR. Whether a cows ovulated or failed to ovulate, had no effect on conception rates, regardless of breed. Therefore, it appears that if a cow exhibits estrus, regardless of bree d, conceptions rates are similar. Breed had no effect on synchronized pregnanc y rate. Synchronized pregnancy rates were similar for Angus cows compared to Geary et al. (2000) in Bos taurus cows synchronized with the Select Synch protocol wit hout a CIDR, whereas, synchronized pregnancy rates for Brangus cows were 33.8% greater compared to Bos indicus Bos taurus cows synchronized with the Select Synch protocol without a CIDR (Lemaster et al., 2001). Whether the addition of the

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118 CIDR to the Select Synch protocol in the presen t study and the study reported in Chapter 3 is the primary reason for the considerable increase in synchronized pregnanc y rates in postpartum cattle of Bos indicus breeding will need to be evaluated. The ability of the CIDR to induce estrous cyclicity in some anestrous cows (L ucy et al., 2001; Larson et al., 2006) has been reported to contribute to more cows exhibiting estrus and being inseminated. The cycling Angus cows had a 19.6% greater synchronized pregnanc y rate compared to noncycling Angus cows, which agrees with Stevenson et al. (2000) who reported a 27.4% greater synchronized pregnancy rates in cycling compared to noncycling Bos taurus cows. Lemaster et al. (2001) also reported greater synchronized pregnancy rates in cycling compared to noncycling Bos indicus Bos taurus cows . This differs to what was observed in the current study, where noncycling Brangus cows had a 28.3% greater synchronized pregnancy rate compared to cycling cows. The reason for the increased synchronized pregnancy rates in the noncycling Brangus cows was due to the fact that a high percenta ge of the cows ovulated to GnRH, had CL regression, exhibited estrus, and conceived. In summary, cycling status influenced the number of cows ovulating to GnRH on d 0. For this reason, ovulation rates in Angus cows were lower than previous reports in Bos taurus cows. However, ovulation rates in the current st udy were greater than th ose previously reported in Bos indicus Bos taurus cows. Size of follicle ovulated to GnRH was similar between breeds and similar to reports in both Bos taurus and Bos indicus Bos taurus cows. Luteal regression in Angus cows was greater than Brangus cows. Luteal regression was greater for cows that ovulated to GnRH, likely due to increased progesterone conc entrations at PGF2 . Cows that ovulated to GnRH had a greater estrous respons e allowing for more cows to be inseminated, resulting in a greater synchronize d pregnancy rate for cows compared to cows that failed to

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119 ovulate to GnRH. HeatWatch estrous detection yielded a sim ilar estrous response to visual observation across both breeds. However, the Angus cows had a longer estrus with more mounts than did Brangus cows. Size of the dominant fo llicle that eventually ovulated along with its growth rate from PGF2 to AI was similar across breeds. However, Brangus cows had decreased estrogen concentrations at PGF2 compared to Angus cows. Implications This study demonstrates the importance of GnRH treatment at the start of a synchronization protocol, as cows that ovulated to GnRH had greater luteal regression, estrous response, and synchronized pregnancy rates. Th e number of cycling cows at the start of a synchronization protocol influenced ovulation rate to Gn RH and tended to increase estrous response, regardless of breed. Brangus cows ha d shorter durations of estrus, fewer total mounts received during estrus, and a gr eater incidence of “s ilent estrus” compared to Angus cows suggesting that a timed-AI is important in cattle of Bos indicus Bos taurus breeding in order to maximize the opportunity for cows to get pregnant during a Select Synch protocol with a CIDR.

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120 0 5 10 15 20 25 30 35 4860728496102114Cows in estrus, % Angus Brangus Figure 5-1: Effect of breed (Angus vs . Brangus) on the interval from PGF2 to the onset of HeatWatch estrus. Estrous response is reported as th e percentage of cows in estrus during each time interval within a breed divided by the total in estrus within a breed. Cows received GnRH concurrent with a CIDR followed by PGF2 7 d later. (Breed P > 0.05) Interval from PGF2 to onset of estrus, h

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121 0 5 10 15 20 25 30 35 40 45 4860728496102114Cows in estrus, % Ovulated to GnRH No ovulation to GnRH Figure 5-2: Effect of ovulation status to GnRH on interval from PGF2 to onset of estrus. Estrous response is reported as the percentage of cows in estrus during each time interval within ovulation status divided by the to tal in estrus within ovulation status. Cows received GnRH concurrent with a CIDR followed by PGF2 7 d later in Angus and Brangus cows. Estrous response by ovulation status to GnRH tended to be different P = 0.07 as determined by survival analysis. Interval from PGF2 to onset of estrus, h

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122 Table 5-1 Effect of breed and estrous cy cling status on ovulati on rates to GnRH and ovulatory follicle size (LS mean SE) in Angus and Brangus cows synchronized with a 7 d CIDR treatment with prostaglandin F2 (PGF2 ) administered at CIDR removal.a Variable n Follicles ovulating to GnRH, %b Ovulatory follicle size, mm, (range)c Angus 31 45.2 12.5 0.6 (10 to 16) Cycling 23 56.5 12.7 0.7 (10 to 16) Not Cycling 8 12.5 10 2.4 (all 10 mm) Brangus 22 45.5 12.5 0.7 (10 to 15) Cycling 12 50.0 12.6 1.0 (10 to 15) Not Cycling 10 40.0 12.3 1.3 (12 to 13) P values Breed P > 0.05 P > 0.05 Cycling Status P < 0.05 P > 0.05 Breed Cycling Status P > 0.05 P > 0.05 a All cows received GnRH at initiation of the 7 d CIDR treatment, with PGF2 administered at the time of CIDR removal. Estrus was detected for 5 d, and cows that exhibited estrus were AI approximately 8 to 12 h later. b Percentage of cows that ovulated to GnRH on d 0 divided by the total treated. c Size of the largest follicle on d 0 that ovulated by 48 h later..

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123 Table 5-2 Effect of breed and ovulation status at GnRH (Ovulated – OV vs. No ovulation NoOV) on follicle sizes (LS mean SE) and estrogen concentrations (LSmean SE) for follicles ovulating after PGF2 in Angus and Brangus cows synchr onized with a 7 d CIDR treatment with prostaglandin F2 (PGF2 ) administered at CIDR removal.a Variable Follicle size at PGF2 , mm (range)b Estrogen concentration, at PGF2 , pg/mLc Follicle size at AI, mm (range)d Estrogen concentration at AI, pg/mLe Angus 12.5 0.5 (6 to 18) 3.5 0.2 16.4 0.4 (13 to 20) 3.1 0.2 OV 12.5 0.7 (9 to 18) 3.0 0.3 16.3 0.5 (14 to 20) 3.0 0.4 No-OV 12.6 0.7 (6 to 18) 3.9 0.3 16.4 0.5 (12 to 20) 3.3 0.4 Brangus 12.6 0.6 (10 to 18) 2.4 0.3 16.1 0.5 (12 to 20) 3.1 0.3 OV 11.9 0.8 (10 to 13) 2.3 0.4 16.2 0.6 (13 to 19) 2.8 0.4 No-OV 13.4 0.9 (11 to 16) 2.5 0.4 16.0 0.8 (13 to 19) 3.5 0.5 P values Breed P > 0.05 P < 0.05 P > 0.05 P > 0.05 OV status P > 0.05 P > 0.05 P > 0.05 P > 0.05 Breed OV status P > 0.05 P > 0.05 P > 0.05 P > 0.05 a All cows received GnRH at initiation of the 7 d CIDR treatment, with PGF2 administered at the time of CIDR removal. Estrus was detected for 5 d, and cows that exhibited estrus were AI approximately 8 to 12 h later. b Size of follicle on d 7 that will be the eventual ovulatory follicle. c Estrogen concentration on d 7. d Size of ovulatory follicl e at the time of AI. e Estrogen concentration at the time of AI. f Follicle growth rate from d 7 to the day of AI (mm/d).

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124 Table 5-3 Effect of breed and ovulation status at GnRH on progesterone concentrations (LS means SE) at CIDR removal, CL regression rate, a nd incidence of silent estrus of Angus and Brangus cows synchronized with a 7 d CIDR treatment with prostaglandin F2 (PGF2 ) administered at CIDR removal.a Treatments Progesterone d 7, ng/mL CL regression, % (n)b Silent estrus Angus 5.0 0.6 100.0 (27) 1 Ovulated 5.1 0.8 100.0 (14) 1 No ovulation 4.9 0.8 100.0 (13) 0 Brangus 5.5 0.7 88.2 (22) 4 Ovulated 6.0 1.0 100.0 (10) 0 No ovulation 4.6 1.1 71.4 (7) 4 P values Breed P > 0.05 P < 0.05 P = 0.06 Ovulation status P > 0.05 P = 0.07 P > 0.05 Breed Ovulation status P > 0.05 P < 0.05 P < 0.05 a All cows received GnRH at initiation of the 7 d CIDR treatment, with PGF2 administered at the time of CIDR removal. Estrus was detected for 5 d, and cows that exhibited estrus were AI approximately 8 to 12 h later. b Percentage of cows that regressed their CL di vided by the total number of cows that had a CL on d 7. c Number of cows that ovulated after PGF2 without displaying estrus.

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125 Table 5-4 Estrous characteristics as determined by HeatWatch of Angus and Brangus cows synchronized with a 7 d CIDR tr eatment with prostaglandin F2 (PGF2 ) administered at CIDR removal. With the ex ception of estrous response estrous ch aracteristics are pr esented as LS means SE.a Breed n Estrous response, %b Interval from PGF2 to onset of estrus, hr, minc Duration of estrus, hr, mind Total mounts during estruse Angus 31 80.7 63 h 46 m 19 h 50 m 11 h 4 m 4 h 51 m 49.0 4.9 Brangus 22 68.2 65 h 28 m 18 h 40 m 7 h 30 m 3 h 57 m 21.5 6.4 P values P > 0.05 P > 0.05 P < 0.05 P < 0.05 a All cows received GnRH at initiation of the 7 d CIDR treatment, with PGF2 administered at the time of CIDR removal. Estrus was detected for 5 d, and cows that exhibited estrus were AI approximately 8 to 12 h later. b Percentage of cows disp laying estrus 5 d after PGF2 of the total treated. c Time from PGF2 administration to the first mount of estrus, as determined by HeatWatch. d Time from the first mount of estrus to the last mount of es trus, as determined by HeatWatch. e Total mounting events which occurred dur ing estrus, as determined by HeatWatch.

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126 Table 5-5 Effect of breed and estrous cycli ng status on estrous, conception and pregnancy rates of Angus and Brangus cows synchr onized with a 7 d CIDR treatment with prostaglandin F2 (PGF2 ) administered at CIDR removal.a Treatments n Estrous response, %b Conception rate, %c Synchronized pregnancy rate, %d Angus 31 80.7 80.0 64.5 Cycling 23 91.3 76.2 69.6 Not cycling 8 50.0 100.0 50.0 Brangus 22 68.2 80.0 54.6 Cycling 12 66.7 62.5 41.7 Not cycling 10 70.0 100.0 70.0 P values Breed P > 0.05 P > 0.05 P > 0.05 Cycling status P = 0.10 P < 0.05 P > 0.05 Breed Cycling status P = 0.06 P = 0.10 P = 0.09 a All cows received GnRH at initiation of the 7 d CIDR treatment, with PGF2 administered at the time of CIDR removal. Estrus was detected for 5 d, and cows that exhibited estrus were AI approximately 8 to 12 h later. b Percentage of cows disp laying estrus 5 d after PGF2 of the total treated. c Percentage of cows pregnant to AI of th e total that exhibited estrus and were AI. d Percentage of cows pregnant during the synchronized breeding of the total treated.

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127 Table 5-6 Effect of breed and ovulation status at GnRH on estrous, conception and pregnancy rates of Angus and Brangus cows synchronized w ith a 7 d CIDR treatment with prostaglandin F2 (PGF2 ) administered at CIDR removal.a Variable n Estrous response, %b Conception rate, %c Synchronized pregnancy rate, %d Angus 31 80.7 80.0 64.5 Ovulated 14 92.9 76.9 71.4 No ovulation 17 70.6 83.3 58.8 Brangus 22 68.2 80.0 54.6 Ovulated 10 90.0 88.9 80.0 No ovulation 12 50.0 66.7 33.3 P Values Breed P > 0.05 P > 0.05 P > 0.05 Ovulation status P < 0.05 P > 0.05 P < 0.05 Breed Ovulation status P > 0.05 P > 0.05 P > 0.05 a All cows received GnRH at initiation of the 7 d CIDR treatment, with PGF2 administered at the time of CIDR removal. Estrus was detected for 5 d, and cows that exhibited estrus were AI approximately 8 to 12 h later. b Percentage of cows disp laying estrus 5 d after PGF2 of the total treated. c Percentage of cows pregnant to AI of th e total that exhibited estrus and were AI. d Percentage of cows pregnant during the synchronized breeding of the total treated.

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128 CHAPTER 6 CONCLUSIONS AND IMPLICATIONS A common estrous synchr onization system used in beef cat tle is administration of GnRH followed 7 d later with a PGF2 treatment, also known as the “S elect Synch” protocol. The overall effectiveness of the protocol can be im proved with the addition of a CIDR between the GnRH and PGF2 treatments. Cows can either be insemi nated after a detected estrus for 5 d or estrous detection can last for 3 d with a timed-AI plus GnRH at 80 h after PGF2 for all cows that did not exhibit estrus. The majority of research conducted with the Select Synch protocols with a CIDR have been carried out in Bos taurus cattle. Therefore, the objec tives of these experiments were to evaluate effectiveness of the Select Synch with a CIDR and (o r) Select Synch with a CIDR combined with timed-AI in Bos taurus, Bos indicus, and Bos indicus Bos taurus cows and heifers. The comparison of a new vs. a once-used CIDR and PGF2 treatments of cloprostenol sodium and dinoprost tromethamine were also investigated. Furthermore, follicular development, ovulation rates, horm one concentrations, and estrous characteristics were evaluated to try to better characterize the actions of the Select Synch protocol between cattle of Bos indicus breeding (Brangus) and Bos taurus breeding (Angus). In Chapter 3, two experiments were conducted in postpartum lactating Bos indicus Bos taurus cows. In Experiment 1, cows were synchroni zed with the Select Synch + CIDR protocol, consisting of a GnRH treatment and CI DR insertion on d 0, followed by a PGF2 treatment and CIDR removal on d 7. Estrous wa s detected 3 times daily for 5 d and cows were inseminated 8 to 14 h after exhibiting estrus using se men from a single AI sire. In a 2 2 factorial design, cows were administered either a ne w CIDR or once-used CIDR and PGF2 treatments of cloprostenol sodium or dinoprost tromethamine. Estrous response, conc eption, and synchronized pregnancy rates were similar for the main effect s of a new CIDR compared to once-used CIDR,

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129 cloprostenol sodium compared to dinoprost tromethamine, and for th e simple treatment effects. There was a significant effect of days postpartum (DPP) on estr ous response, conception, and synchronized pregnancy rates. Cows with long postpartum intervals ( 70 d) had greater responses compared to cows with medium postp artum intervals (50 to 69 d), which had greater responses compared to s hort postpartum intervals ( 50 d). There was also a significant effect of interval from PGF2 to onset of estrus. Cows that exhib ited estrus at and after 80 h after PGF2 had decreased conception rates compared to cows that exhibited estrus earlier. In Chapter 3, Experiment 2, Bos indicus Bos taurus cows were synchronized with the Select Synch + timed-AI with a CIDR protocol . Cows were synchr onized using the same treatments (new CIDR vs. once-used CI DR and cloprostenol sodium vs. dinoprost tromethamine) in a 2 2 factorial design. After PGF2 administration, estrus was detected 3 times daily for 3 d and cows were inseminated 8 to 14 h after exhibiting estrus. Cows not displaying estrus by 73 h after PGF2 were timed-AI between 76 and 80 h. Estrous response was similar for new CIDR vs. once-used CIDR and cloprostenol sodium vs. dinoprost tromethamine treated cows. Conception rates tended to be greater (14.1%) fo r the new CIDR compared to once-used CIDR; whereas, cloprostenol sodium and dinoprost tromethamine treatments had similar conception rates. Timed-AI pregnancy rates were similar for new CIDR vs. once-used CIDR and cloprostenol sodium vs. dinoprost trom ethamine. Synchronized pregnancy rates were similar for new CIDR compared to once-used CIDR and cloprostenol sodium vs. dinoprost tromethamine. Simple treatment effects did no t influence estrous response, conception, timedAI, or synchronized pregnancy rates. Synchronized pregnancy rates were 13.7 % greater for Experiment 2 compared to Experiment 1. The increase in synchronized preg nancy rate was due in part to the increased

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130 conception rates and addition of a timed-AI in E xperiment 2. One reason conception rates were probably greater was because estrous dete ction and AI ended at 73 h after PGF2 in Experiment 2, thereby removing cows that would have exhi bited estrus between 73 and 120 h which had the potential for decreased conception rates as observe d in Experiment 1. Furthermore, addition of a timed-AI in Experiment 2 also allowed for the af orementioned cows to be inseminated earlier as well as insemination of cows that had not exhibi ted estrus. Accordingly, the incorporation of a timed-AI in combination with 3 d of estrous detection in the Select Synch synchronization protocols used in Bos indicus Bos taurus cows appears to be imperative. Cloprostenol sodium and dinoprost tromethami ne appeared to be equally effective luteolysins in both experiments. However, there tended to be an increase in conception rate for a new CIDR compared to a once-used CIDR in Ex periment 2. Since cow numbers were small in Experiment 2, additional research will have to be conducted to determine if the decrease in conception rate is significant. If a once-used CIDR does yield decreased conception rates and possibly lead to decreased sync hronized pregnancy rates, pr oducers must balance the cost savings of using a new CIDR each time a cows is bred at a cost of approximately $10.00 per cow or a once-used CIDR which allows for tw o synchronizations for $10.00. Days postpartum clearly played a si gnificant role in the response to the synchronizat ion protocol a nd synchronized pregnancy rates in both experiment s. Consequently, producers must pay close attention to when cows calve in relation to when synchronization protocols are initi ated. The decreased conception rates observed in Experiment 1 for cows that exhibited estrus > 72 h after PGF2 suggest that ovarian follicular development at PGF2 probably has a significant e ffect on fertility of the oocyte ovulated. This supports th e observation that the Select Synch protocol with a CIDR should not be used in cattle of Bos indicus breeding. Therefore, it a ppears that the combination

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131 of estrous detection and AI combined with a timed-AI at 80 h is a good option for synchronizing cattle of Bos indicus breeding. Follicle development in cattle of Bos indicus breeding appears to uniquely altered by the combination of GnRH and the CIDR synchroniza tion treatments, and additional research is needed to evaluate their effects. Data from E xperiment 1 suggest that an additional 24 to 36 h difference in follicle dominance appears to have detrimental effects on oocyte fertility. This implies that oocytes in cattle of Bos indicus breeding are very sensitive and any increase in duration of dominance beyond what is normal could have a negative effect on oocyte quality. To further evaluate this, additional experiment s will need to be conducted in cattle of Bos indicus breeding where cows are synchroni zed and oocytes are aspirated at designated intervals after PGF2 to evaluate the oocyte integrity, growth rates, and survivability in culture to provide some insight to the reduced fertility of these follicles. In Chapter 4, two studies evaluated the Select Synch + timed-AI with a CIDR protocol in cows (Experiment 1) and heifers (Experiment 2) of Bos taurus (Angus) , Bos indicus (Brahman) , and Bos indicus (Brahman) Bos taurus (Angus) breeding over 2 years. Animals were synchronized and inseminated using the same protocol as Chapter 3, Experiment 2. Prostaglandin F2 treatments of either cloprostenol s odium or dinoprost tromethamine were administered at CIDR removal. In Experiment 1, estrous response and concep tion rate were similar for cloprostenol sodium and dinoprost tromethamine treatments. However, cloprostenol sodium treated cows had 14.3% greater timed-AI pregnancy rates and 11% greater synchronized pregnancy rates compared to dinoprost tromethamine treated cows. Breed effected estrous response, with Angus and Brangus cows exhibiting a greater estrous response compared to the other Brahman Angus

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132 breed combinations and Brahman cows. However, breed did not influence conception, timed-AI, or synchronized pregnancy rates. Cow age e ffected estrous response and conception rates. Older cows (age > 5; range 5 to 15 years) ha d a greater estrous res ponse compared to younger cows (3 years old), whereas, younger cows had a greater conception rate compared to older cows. Cow age did not influence timed-AI or sy nchronized pregnancy rates. Days postpartum influenced estrous response, conception, and sync hronized pregnancy rates. Cows with longer postpartum intervals ( 75 d) had greater responses compared to medium postpartum interval (56 to 74 d) cows, which had greater responses co mpared to short postpartum interval cows ( 55 d). These experiments along with experiments discu ssed in Chapter 3 indicate the importance of having cows of Bos indicus breeding with longer postpartum in tervals when a synchronization protocol is initiated. This howev er, has a caveat. In order to ma intain a yearly calving interval, cows have an approximate window of 80 d after calving in which to get pregnant. If producers wait too long after calving to institute a synchroni zation protocol, they are at risk of having cows with extended calving intervals. In Experiment 2, heifers had similar estr ous responses, concep tion, and synchronized pregnancy rates for cloprostenol and dinoprost. Dinoprost tromet hamine treated heifers had a 22.4% greater timed-AI pregnancy rate compared to cloprostenol sodium tr eated heifers. Breed effected estrous response, but there was no trend for heifers of increasing Bos indicus breeding having a decreased estrous response. Additional replications will ha ve to be conducted with the breed combinations used in Experiments 1 and 2 of Chapter 4 to determine wh at effect breed has on response to the Select Synch + timed AI with a CIDR. This will also include the comparison of cloprostenol sodium

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133 vs. dinoprost tromethamine. Once these experime nts are conducted, it should be possible to determine if a different approach is needed in synchronizing cattle of Bos indicus breeding. In Chapter 5, the Select Synch with a CIDR protocol was evaluated in two-year old postpartum lactating Bos taurus (Angus) and Bos indicus Bos taurus (Brangus) cows. Cycling status was determined prior to the start of synchronization. Animals were synchronized and inseminated using the Select S ynch protocol, with all cows re ceiving a new CIDR and dinoprost tromethamine. With the start of synchroniza tion designated as d 0, cows were examined by ultrasonography and blood samples were taken on da ys 0, 2, 7, at the time of AI, and 8 days after observed estrus. Ultrasonography exams were used to evaluate follicle sizes, ovulation rates following GnRH and PGF2 , follicle growth rates, and lute al volumes. Blood samples were evaluated for progesterone and estrogen concentr ations, as well as luteal regression. Estrous characteristics were monitored using the HeatWatch estrous detection system, as well as visual observation 3 times daily in the pastures. Estr ous response, conception, synchronized pregnancy, and breeding season pregnancy rates were also evaluated. At the start of the experiment, a simila r number of Angus and Brangus cows were cycling. Breed did not influence ovulation rate to GnRH, but a gr eater number of cycling cows ovulated to GnRH compared to noncycling cows. Size of the follicle ovulated was similar for Angus and Brangus, as well as cycling and nonc ycling cows. On d 7, size of the eventual ovulatory follicle was similar for Angus and Bra ngus, as well as for cows that ovulated and failed to ovulate to GnRH. Estr ogen concentrations on d 7 were greater for Angus compared to Brangus cows, but were not influenced by cycling status or ovulation status to GnRH. Number of cows that had a functional CL on d 7 was similar for Angus and Brangus, but a greater number of cycling cows had a CL compared to noncycling cows. Luteal regression was greater

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134 for Angus compared to Brangus cows, and for cows that ovulated to GnRH compared to cows that did not ovulate to GnRH. Interval from PGF2 to onset of estrus was similar for Angus and Brangus cows. However, Angus cows exhibited a greater estr us duration and had a gr eater number of mounts during estrus compared to Brangus cows. F our Brangus and one Angus cow underwent “silent estrus.” Estrous distribution was similar for A ngus and Brangus, however, cows that ovulated to GnRH tended to have a more synchronous estrus than cows that failed to ovulate to GnRH. Estrous response was similar between Angus and Brangus, however cows that ovulated to GnRH had a greater estrus response compared to cows that failed to ovulate, regardless of breed. HeatWatch estrous detection was similar to visual estrous detection. Size of the eventual ovulatory follicle on d 7 and at the time of AI, as we ll as follicle growth rate from d 7 to the time of AI, were similar for Angus and Brangus, as well as for cows that ovulated and failed to ovulate. Ovulati on rate after PGF2 was similar for Angus and Bra ngus, but a greater number of cows that ovulated to GnRH ovulated following PGF2 compared to cows that failed to ovulate to GnRH. Conception rates were similar for Angus and Brangus, as well as for cows that ovulated and failed to ovulate to GnRH. Synchronized pregnancy rate was similar for Angus and Brangus, and cycling and noncycling cows. Co ws that ovulated to GnRH had greater synchronized pregnancy rates compared to cows that failed to ovulate to GnRH. Thirty-day pregnancy rates were similar fo r Angus and Brangus, but breeding season pregnancy rates tended to be greater for Brangus compared to Angus cows. Days postpartum influenced breeding season pregnancy rates, with short ( 60 d) postpartum cows having decreased pregnancy rates compared to medium (61 to 99 d) and long ( 100 d) postpartum cows.

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135 This study emphasizes the importance of a GnRH treatment at the start of a synchronization protocol, as cows that ovulated to GnRH had greater luteal regression, estrous response, and synchronized pregnancy rates. Ho wever, the question that still remains is how effective GnRH is in initiat ing ovulation in cattle of Bos indicus breeding. The current study suggests that this percentage is still less in cattle of Bos indicus breeding compared to reports in the literature for Bos taurus cattle. Additional studies need to be conducted to determine this. The importance of having a large number of cycli ng cows at the start of synchronization was also demonstrated. Cycling cows had greater ovulation rates to GnRH and tended to have a greater estrous response, regard less of breed. However, Brangus co ws had a shorter estrous duration, fewer mounts received during estrus, and more silent heats compared to Angus, which collectively indicate that estrus can be more difficult to detect in cows of Bos indicus breeding and limit the effectiveness of a Select Synch program. Collectively, the experiments presented empha size the importance of incorporation of a timed-AI to synchronization protocols designed for cattle of Bos indicus breeding. Decreased and (or) delayed estrous responses in these cows could lead to ovulation of potentially aged and less fertile follicles and decrease d synchronized pregnancy rates. By inseminating all cows that have not displayed estrus by 72 h after PGF2 at a timed-AI, cows that would have had a delayed estrus are induced to ovulate earlier and cows undergoing a silent estrus are inseminated and given an opportunity to become pregnant. When preparing to implement a synchronization protocol, it is important for producers to know where cows are in relation to calving, as postpartum interval has a significant influence on the responses to a s ynchronization protocol.

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157 BIOGRAPHICAL SKETCH Regina D. Esterman was born September 3, 1984 in Salem, Virginia. Regina is the oldest of two children of Jim and Lavonne Esterman, of West Chesterfield, New Hampshire. During her childhood, Regina lived in Virginia, Tenness ee, Louisiana, and Alabama, before finally settling in Ohio. She attended Northwest High School in Canal Fulton, Ohio, and was an active member of Summit County 4-H, United States P ony Club, and the local horse show circuits. In addition to these activities, Regina participated in the post-secondary option her junior and senior years of high school, attending The Ohio State University Agricult ural Technical Institute fulltime. While attending Ohio State, Regina wa s awarded 2002 Outstanding Student of the Year. In 2002, Regina received her A.S. from Ohio State concurrently with gradation from high school. Regina then enrolled at Colorado State Universit y. During her studies at Colorado State, Regina was a member of Kappa Alpha Theta sorority, Colorado State Polo Team, and the Collegiate Horseman’s Association. Upon gr aduation in 2004, Regina began a Master of Science degree at the University of Florida, under the mentorship of Dr. Joel Yelich. During her time at Florida, Regina was active in the Animal Science Gr aduate Student Association, serving as Vice President. Regina’s future plan s include pursuing her Ph.D. at Un iversity of Florida, continuing with Dr. Joel Yelich as a USDA fellow.