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Group Title: Bulletin University of Florida. Agricultural Experiment Station
Title: Effect of stilbestrol on udder development, pelvic changes, lactation and reproduction
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 Material Information
Title: Effect of stilbestrol on udder development, pelvic changes, lactation and reproduction
Series Title: Bulletin 440 ; University of Florida. Agricultural Experiment Station
Physical Description: 35 p. : ill. ; 23 cm.
Language: English
Creator: Marshall, Sidney P ( Sidney Paul ), 1916-
Becker, R. B.
Arnold, P. T. Dix
Sanders, D. A.
Publisher: University of Florida Agricultural Experiment Station
Place of Publication: Gainesville, Fla.
Publication Date: January, 1948
Copyright Date: 1948
 Subjects
Subject: Diethylstilbestrol   ( lcsh )
Cows -- Reproduction   ( lcsh )
Milk yield   ( lcsh )
Genre: government publication (state, provincial, terriorial, dependent)   ( marcgt )
bibliography   ( marcgt )
non-fiction   ( marcgt )
 Notes
Statement of Responsibility: Sidney P. Marshall ... et al..
Bibliography: Includes bibliographical references (p. 33-35).
General Note: Cover title.
 Record Information
Bibliographic ID: UF00026846
Volume ID: VID00001
Source Institution: University of Florida
Holding Location: University of Florida
Rights Management: All rights reserved by the source institution and holding location.
Resource Identifier: ltuf - AEN6168
oclc - 18254150
alephbibnum - 000925517

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Bulletin 440 January, 1948

UNIVERSITY OF FLORIDA
AGRICULTURAL EXPERIMENT STATION
HAROLD MOWRY, Director
GAINESVILLE, FLORIDA


EFFECT OF STILBESTROL ON UDDER
DEVELOPMENT, PELVIC CHANGES,
LACTATION AND REPRODUCTION

SIDNEY P. MARSHALL, R. B. BECKER, P. T. DIX ARNOLD
and D. A. SANDERS






















Fig. 1.-Method of measuring "udder fullness" from the left pinbone to
the "front of udder."


TECHNICAL BULLETIN


Single copies free to Florida residents upon request to
AGRICULTURAL EXPERIMENT STATION
GAINESVILLE, FLORIDA









BOARD OF CONTROL ECONOMICS. AGRICULTURAL
J. Thos. Gurney, Chairman, Orlando C. V. Noble, Ph.D., Agri. Economists
N. B. Jordan, Quincy Zach Savage, M.S.A., Associate"
Thos. W. Bryant, Lakeland A. H. Spurlock, M.S.A., Associate
J. Henson Markham. Jacksonville D. E. Alleger, M.S., Associate
Hollis Rinehart, Miami D. L. Brooke, M.S.A., Associate
W. F. Powers, Secretary, Tallahassee R. E. L. Greene, Ph.D., Agri. Economist
H. W. Little, M.S., Assistant
EXECUTIVE STAFF
EEOrlando, Florida (Cooperative USDA)
J. Hillis Miller, Ph.D., President of the
University' G. Norman Rose, B.S., Asso. Agr. Economist
H. Harold Hume, D.Sc., Provost for Agr.3 J. C. Townsend, Jr., B.S.A., Agr. Statistician'
Harold Mowry, M.S.A., Director J. B. Owens, B.S.A., Agr. Statisticians
L. O. Gratz, Ph.D., Aast. Dir., Research J. F. Steffens, Jr., B.S.A., Agr. Statistician2
W. M. Fifleld, M.S., Asst. Dir., Admin.
J. Francis Cooper, M.S.A., Editora ECONOMICS, HOME
Clyde Beale, A.B.J., Associate Editors Ouida D. Abbott, Ph.D., Home Econ.'
Jefferson Thomas, Assistant Editors R. B. French, Ph.D., Biochemist
Ida Keeling Cresap, Librarian
Ruby Newhall, Administrative Manager5' ENTOMOLOGY
Geo. F. Baughman, M.A., Business Manager A. N. Tissot, Ph.D., Entomologist'
Claranelle Alderman. Accountants H. E. Bratley, M.S.A., Assistant

MAIN STATION, GAINESVILLE HORTICULTURE
AGRICULTURAL ENGINEERING G. H. Blackmon, M.S.A., Horticulturit'
Frazier Rogers, M.S.A., Agr. Engineers F. S. Jamison, Ph.D., Truck Hort.
J. M. Johnson, B.S.A.E., Asso. Agr. Engineer' H. M. Reed, B.S., Chem., Veg. Proc.
J. M. Myers, B.S., Asso. Agr. Engineer Byron E. Janes, Ph.D., Asso. lort.
R. E. Choate, B.S.A.E., Asst. Agr. Engineer' R. A. Dennison, Ph.D., Asso. Hort.
A. M. Pettis, B.S.A.E., Asst. Agr. Engineer R.K. Shwalter, M.S., Asso. Hrt
Albert P. Lorz, Ph.D., Asso. Hort.
AGRONOMY R. H. Sharpe, M.S., Asso. Hort.
R. J. Wilmot, M.S.A., Asst. Hort.
W. E. Stokes, M.S., Agronomist' R. D. Dickey, M.S.A., Asst. Hort.
Fred H. Hull, Ph.D., Agronomist Victor F. Nettles, M:S.A., Asst. Hort.6
G. E. Ritchey, M.S., Agronomists F. S. Lagasse, Ph.D., Asso. Hort.'
G. B. Killinger, Ph.D., Agronomist L. H. Halsey, B.S.A., Asst. Hort.
H. C. Harris, Ph.D., Agronomist
R. W. Bledsoe, Ph.D., Agronomist PLANT PATHOLOGY
"M. E. Paddick, Ph.D., Agronomist ,.
Paddick, Ph.D.enbrer, Ph.., A ssociate W. B. Tisdale, Ph.D., Plant Pathologist''
C. Litenberger, Ph.D. Associate Phares Decker, Ph.D., Asso. Plant Path.
W. A. Carver, Ph.D., Associate
Wr A. Cark, B.S., Assistat Erdman West, M.S., Mycologist and Botanist
Lillian E. Arnold, M.S., Asst. Botanist
ANIMAL INDUSTRY SOILS

A. L. Shealy. D.V.M., An. Industrialist' 2
R. B. Becker, Ph.D., Dairy Husbandman F S th Ph.D. Mrobiologist
E. L. Fouts, Ph.D., Dairy Technologist' Gaylord M. Volk, Ph.D., Chemist
A. Sdrs, PDV.M., Veterinarian J. R. Henderson, M.S.A., Soil Technologists
D. Sanders, D.V.M., Veterinarian J R. Neller, Ph.D., Soils Chemist
. Swanson, D.V.M., Parasitologist Nathan Gammon, Jr., Ph.D., Soils Chemist
C. E. Bell, Ph.D., Associate Chemist
N. R. Mehrhof, M.Agr., Poultry Husb. C .., Asso. o
G K. Davis, Ph.D., Animal Nutritionist A. Carrgan, B.S., Asso. Biochemist
G ock, Ph.D.., An. sAn. d a H. W. Winsor, B.S.A., Assistant Chemist
R. S. Glasscock, Ph.D., An. Husbandmans Thornton, Asso. Microbiologist
Geo. D. Thornton, M.S., Asso. Microbiologist
P T. Dix Arnold, M.S.A., Asst. Dairy Husb." _I, .
P T. A d, .. At. Dr B R. E. Caldwell, M.S.A., Asst. Chemists
C. L. Comar, Ph.D., Asso. Biochemist J. B. Cromartie, B.S.A., Soil Surveyor
L. E. Mull, M.S., Asst. in Dairy Tech.58 B. Cromarie BS., l Surveyor
Katherine Boney, n.S., Asst. Chem.
Katherine Boney, B.S., Asst.'Chem. Rl W. Leighty, B.S., Asso. Soil Surveyor
J. C. Driggers, B.S.A., Asst. Poultry Husb.
Glenn Van Ness, D.V.M., Asso. Poultry
Pathologist 'Head of Department.
S. John Folks, B.S.A., Asst. An. Husb.3 2 In cooperation with U. S
W. A. Krienke. M.S.. Asso. in Dairy Mfs. sCooperative, other divisions, U. of F.
S. P. Marshall, Ph.D., Asso. Dairy Hush." In Military Service.
C. F. Simpson, D.V.M., Asso. Veterinarian 6 On leave.





/









BRANCH STATIONS .I. C. Hoffman, M.S., Asso. Hort.
C. B. Savage, M.S.A., Asst. Hort.
NORTH FLORIDA STATION, QUINCY D. L. Stoddard, Ph.D., Asso. Plant Path.
J. D. Warner, M.S., Vice-Director in Charge W. A. Desnoyers, B.S., Asst. Hydrologist
R. R. Kincaid, Ph.D., Plant Pathologist
W. I. Chapman. M.S., Asso. Agron. SUB-TROPICAL STA.. HOMESTEAD
R. C. 3ond, M.S.A., Asso. Agronomist Geo. D. Ruehle, Ph.D., Vice-Director in
L. G. Thompson, Ph.D., Soils Chemist Charge
Frank S. Baker, Jr., B.S., Asst. An. Husb. D. 0. Wolfenbarger, Ph.D., Entomologist
Kelvin Iorward, M.S., Entomologist Francis B. Lincoln, Ph.D., Horticulturist
Robt. A. Conover, Ph.D., Asso. Plant Path.
Mobile Unit, Monticello R. W. Harkness, Ph.D., Asst. Chemist
R. W. Wallace, B.S., Associate Agronomist Milton Cobin, B.S., Asso. Hort.

Mobile Unit, Marianna W. CENT. FLA. STA., BROOKSVILLE
R. W. Lipscomb. M.S., Associate Agronomist C. D. Gordon, Ph.D., Geneticist in Charge'

Mobile Unit, Wewahitchka RANGE CATTLE STATION. ONA
J. B. White. B.S.A., Associate Agronomist W. G. Kirk, Ph.D., Vice-Director in Charge
E. M. Hodges, Ph.D., Associate Agronomist
Mobile Unit. DeFuniak Springs D. W. Jones, B.S., Asst. Soil Tech.
R. L. Smith, M.S., Associate Agronomist H. J. Fulford, B.S.A., Asst. An. Hush.

CITRUS STATION, LAKE ALRED CENTRAL FLORIDA STATION, SANFORD
R. W. Ruprecht, Ph.D., Vice-Director in
A. F. Camp. Ph.D., Vice-Director in Charge Charge
W. L. Thompson. B.S.. Entomologist A. Alfred Foster, Ph.D., Asso. P1. Path.
J. T. Griffiths. Ph.D., Asso. Entomologist J. W. Wilson, Sc.D., Entomologist
R. F. Suit. Ph.D., Plant Pathologist Ben F. Whitner, Jr., B.S.A., Asst. Hort.
E. P. Ducharme, M.S., Plant Pathologist"
R. K. Voorhees, M.S., Asso. Horticulturist WEST FLORIDA STATION, MILTON
C. R. Stearns, Jr., B.S.A., Asso. Chemist H. W. Lundy, B.S.A., Asso. Agronomist
James K. Colehour, M.S., Asst. Chemist
T. W. Young, Ph.D., Asso. Horticulturist
J. W. Sites, M.S.A., Horticulturist FIELD STATIONS
H. O. Sterling, B.S., Asst. Horticulturist Leesburg
J. A. Granger, B.S.A., Asst. Horticulturist G. K. Parris, Ph.D., Plant Path. in Charge
H. J. Reitz, M.S.. Asso. Horticulturist
Francine Fisher, M.S., Asst. PI. Path. Plant City
I. W. Wander, Ph.D., Soil Chemist A. N. Brooks, Ph.D., Plant Pathologist
A. E. Willson, B.S.A., Asso. Biochemist
R. W. Jones, Asst. Plant Path. Hastings
J. W. Kesterson, M.S., Asso. Chemist A. H. Eddins, Ph.D., Plant Path. in Charge
C. W. Houston, Ph.D., Asso. Chemist E. N. McCubbin, Ph.D., Horticulturist
R. N. Hendrickson, B.S., Asst. Chemist
E. H. Bitcover, M.A., Soils Chemist Monticello
L. C. Knorr, Ph.D., Asso. Histologist S. O. Hill. B.S., Asst. Entomologist'
Joe P. Barnett, B.S.A., Asst. Hort. A. M. Phillips. B.S., Asso. Entomologist'
J. C. Bowers, B.S., Asst. Chemist
D. S. Prosser, Jr., B.S., Asst. Hort. Bradenton
R. W. Olsen, B.S., Biochemist, Con. Res. J. R. Beckenhach, Ph.D., Horticulturist in
F. W. Wenzel, Jr., Ph.D., Supervisory Chem. Charge
R. W. Jones, M.S.A., Asst. Plant Path. E. G. Kelsheimer, Ph.D., Entomologist

EVERGLADES STA., BELLE GLADE David G. Kelbert, Asso. Horticulturist
E. L. Spencer, Ph.D., Soils Chemist
R. V. Allison, Ph.D., Vice-Director in Charge Robert O. Magie, Ph.D., Gladioli Hort.
F. D. Stevens, B.S., Sugarcane Agron. J. M. Walter, Ph.D., Plant Path.
Thomas Bregger, Ph.D., Sugarcane Donald S. Burgis, M.S.A., Asst. Hort.
Physiologist
B. S. Clayton, B.S.C.E., Drainage Eng.2 Lakeland
J. W. Randolph, M.S., Agricultural Engineer Warren O. Johnson, B.S., Meteorologist'
W. T. Forsee, Jr., Ph.D., Chemist
R. W. Kidder, M.S., Asso. An. Husb. 1 Head of Department.
T. C. Erwin, Assistant Chemist 2 In cooperation with U. S.
Roy A. Bair, Ph.D., Agronomist Cooperative, other divisions, U. of F.
C. C. Seale, Asso. Agronomist In Military Service.
N. C. Hayslip, B.S.A., Asso. Entomologist On leave.













TABLE OF CONTENTS

Page

INTRODUCTION ..............- ......... ......... ........ .... ... ........................... 5


LITERATURE REVIEW .................. .. .................... ........................................ 5

Udder Development and Lactation .............. ............................... 5

Effect on Milk Composition .............. ...... ........ ..... .............. 8

Effect on Sexuality -......-.........- .......-................................ 10

PLAN OF INVESTIGATION ............... ...... ............................... ............. 12


RESULTS ....- ......... ...............................................-..................- 13

Injection Rates ...... -----....... ........--.................--- ............. ..... 13

Changes in Conformation .........-.......-----.......-................ ..... 13

M ilk Y fields .................... ...................- ..........-........... ...... 18

Composition of M ilk ....................... ........... ....................................... 20

Previous Breeding and Reproductive History ........................................ 23

Sexual Development ..-....................... --...- ..-...... .............. 25

Reproductive Organs ........................--.. ..- .................-... 25

Later Observations ................................. ..... .................... ................. 25

DISCUSSION AND SIGNIFICANCE ..................................... .. .................... 30

SUMMARY AND CONCLUSIONS ........................................ ..... ...------32

ACKNOWLEDGMENTS ................. .............................-..-----. 32

LITERATURE CITED .............................. ...................................................... 33









EFFECT OF STILBESTROL ON UDDER DEVELOPMENT,
PELVIC CHANGES, LACTATION AND REPRODUCTION
SIDNEY P. MARSHALL, R. B. BECKER, P. T. DIX ARNOLD
and D. A. SANDERS

INTRODUCTION
In 1938 a synthetic estrogen (7)1 made and tested in Eng-
land opened new avenues of investigation dealing with lactation
and reproduction in mammals. This substance, diethylstilbestrol
(subsequently called stilbestrol) has been tested with rats,
guinea pigs, goats, cattle and fowls, and is being used thera-
peutically with human beings. In virgin animals it causes
proliferation of mammary tissues, false heat, ovarian quies-
cence, relaxation of the broad ligaments and distortion of the
rump and tailhead in cattle. There is need for more complete
observation of the effects of stilbestrol upon milk production
and reproduction. Heifers and dry cows were assigned to
investigate the commercial application of stilbestrol treatment.

LITERATURE REVIEW
UDDER DEVELOPMENT AND LACTATION
Although many observations have been reported concerning
use of stilbestrol with laboratory animals, fewer reports have
dealt with milk goats and dairy cattle. Lewis and Turner
(22, 23) injected stilbestrol subcutaneously, rubbed an alcohol
solution on the udder skin'and implanted pellets subcutaneously
in kids and yearling goats, noting that it caused development
of mammary tissue and started milk secretion in virgins but
appeared to depress lactation with 2 goats in milk. Lewis (21)
believed in 1942 that continued treatment with stilbestrol pro-
longed lactation in both virgin and older goats.
Daily doses of 0.25 milligram of stilbestrol stimulated lacta-
tion in Toggenburg goats (24), while doses of 1 to 4 milligrams
depressed milk production.
Folley, Watson and Bottomley (13) brought a virgin female
goat into lactation by rubbing 1 gram of 1 percent stilbestrol
dipropionate on the udder thrice weekly for 30 days. Milk yield
increased to 1.5 liters daily and then decreased slowly. Omitting
stilbestrol treatment had no effect on the course of the lactation
1 Italic figures in parentheses refer to Literature Cited in the back of
this bulletin.







6 Florida Agricultural Experiment Station

curve for the next 20 days, after which there was a sharp
decline not immediately checked by applying double the dose.
Milk yield increased when the original treatment was resumed.
The milk was normal in nitrogen partition and percentage of
lactose, with fat and non-fatty solids above 6 and 10 percent,
respectively. Two other virgin goats responded similarly and
a virgin heifer gave a small amount of colostrum. These were
the first tests with this estrogen that produced lactation in
virgin goats in the absence of prolactin administration. Folley,
Watson and Bottomley (15) used ointments containing 1.0, 2.5
and 5.0 percent of stilbestrol, rubbed on the udders of virgin
goats, which caused their development and prolonged lactation
without use of anterior pituitary extract. They (16) obtained
as much as 80 milliliters of milk daily from 2 nulliparous Dairy
Shorthorn heifers 181/ and 20 months old, following inunction
of the udder 3 times weekly with 5 grams of 1 percent stilbestrol
dipropionate ointment. The yield dropped to 20 milliliters but
later reached a peak of 170 milliliters soon after treatment
ceased. It declined subsequently as in a normal lactation. Casein
and globulin contents were low but the fat content approached
normal.
In the first practical treatment of cattle, Walker and Stanley
(31) obtained as high as 14 and 16 pounds of milk daily from
2 heifers injected with stilbestrol dipropionate alone or in con-
juction with testosterone propionate. One of the heifers yielded
2,880 pounds of milk subsequently in 8 months. A Jersey heifer
was stimulated into lactation after 5 months' treatment with
both materials, and in 50 days increased from 196 to 3,738
milliliters of milk containing 5 percent of butterfat.
Reece (28) injected 5 milligrams each of stilbestrol dipro-
pionate and testosterone dipropionate twice weekly for 6 weeks
into a 30-months-old Holstein heifer, then used the first estrogen
only for 5 weeks additional. The udder developed; milking
began when the injections were discontinued and eventually
this animal yielded 4,517 pounds of milk from the second to
eighth months of lactation. A calf nursed part of the milk
during the first month of lactation. Stilbestrol dipropionate
alone was injected for 13 weeks into a 33-months-old Jersey.
Milking began after the injections ceased and from 0.4 to 1.2
pounds of milk were obtained daily for 12 days. A calf was
allowed to nurse 9 days. A peak of 33.7 pounds of milk was
obtained on the 95th day of lactation.







Effect of Stilbestrol on Udder Development 7

Petersen and associates (27) removed thyroids from 2 cows,
causing clinical myxedema, and administered 30 milligrams of
stilbestrol on alternate days for 31 and 21 days, without pro-
ducing udder development. After recovery on raw thyroid feed-
ing, stilbestrol administration resulted in milk production in
13 and 18 days in both cows. Peak yields of 10.4 and 14.4
pounds of milk were obtained, as against 8 pounds for a Holstein
heifer treated similarly. They regarded failure of response to
stilbestrol during myxedema, and response following thyroid
therapy, as not due to a synergistic action of thyroxine and
stilbestrol.
When synthetic estrogens were administered by mouth, using
50 to 200 milligram doses daily in the drinking water or as
crystals in the feed, for 23 weeks, Folley and Malpress (10)
obtained peak yields ranging between 2.5 and 19 pounds of milk
daily from Ayrshire heifers. With each group lactation in-
creased after stilbestrol was withheld from the feed. Daily doses
of 50 milligrams were ineffective. "At the best, the peak yields
were unsatisfactory" for commercial production, in the judg-
ment of the workers. In other work (9) stilbestrol tablets were
implanted subcutaneously in open heifers and dry cows on farms.
Milking began once daily on the tenth day following implanta-
tion, increasing to twice daily when yields of 5 pounds were
attained. Responses in maximum daily yields varied from 1 to
30 pounds of milk. A Shorthorn yielded 7,395 pounds of milk
in a 365-day lactation. They concluded: "It seems probable
that no advantage accrues from leaving the oestrogen tablets
in situ once the lactation curve has reached its highest level;
on the contrary, removal of .the tablets at this point frequently
caused a sudden rise in milk yield indicating a transition of the
oestrogenic action from a stimulative to an inhibitory phase."
Spriggs (30) implanted stilbestrol in 50-milligram tablets in
varying amounts subcutaneously in 5 heifers and 1 dry cow
on local farms in southern England. Milking began 12 days
after implantation. Rapid development of the udders and widely
variable milk yields resulted. He observed that cows were
generally less responsive than heifers; peak of lactation was
attained more slowly than following normal calving; no changes
occurred in response upon removal of the tablets and there was
no increase in milk yield following withdrawal of the implants.
Milk production resulting from subcutaneous implantation of
stilbestrol tablets was as follows:








8 Florida Agricultural Experiment Station

Stil- Days
Animal bestrol Im- Absorbed Total Lactation
Tablets planted per Day Milk _
Heifers: rains milligrams pounds days
Heifers:
Guernsey ....... 4.0 94 5.6 2,893 273 dry
Jersey .....-........ 4.0 80 6.4 7,058 435*
Shorthorn ....... 4.0 77 8.8 none -
Shorthorn ........ 4.0 90 10.3 5,174 365*
Shorthorn ........ 2.5 90 4.9 6,649 365*
Cow:
Brit-Friesain.... 2.5 115 3.0 6,484 301

Still in milk.

Farmers regarded the treatment as not economically practical
with a sufficient proportion of animals.
Petersen (26) cited earlier work to the effect that: "In most
cases with sterile heifers as large udders and levels of milk
flow can be attained artificially as would be expected following
a normal calving. In older sterile dry cows only occasionally
as large udder development and satisfactory levels of milk pro-
duction obtained from the diethylstilbestrol treatment."

EFFECT ON MILK COMPOSITION
Shortly after announcement of the new synthetic estrogen
diethylstilbestrol by Dodd and associates (7), its effect was
tested by Folley and Watson (12) by implanting 1 gram sub-
cutaneously in a 1,100-pound Shorthorn cow advancing in lacta-
tion. The daily milk yield was practically unaffected. An in-
crease in percentage of lactose accompanied a significant rise
in the non-fatty solids. The content of phosphatase rose and fell
sharply.
Folley and associates (14) drenched 1 cow with stilbestrol in
water, injected others subcutaneously with the dipropionate
ester, and implanted powdered stilbestrol subcutaneously. They
observed a temporary increase in solids-not-fat and to a lesser
extent in butterfat percentages. Milk yields decreased when
large doses of stilbestrol were administered and some pregnant
cows aborted.
The milk of goats stimulated to lactation by stilbestrol in-
unction was analyzed by Folley, Watson and Bottomley (15).
They observed from analyses of milk obtained during the first
14 weeks of inunction that the casein was low, globulin high, fat
and solids-not-fat high initially and then lower. They com-
mented: "These changes are qualitatively similar to those which





Effect of Stilbestrol on Udder Development 9

occur normally during the transition from colostrum to milk
in the goat", except that the changes were more gradual.
Spielman, Ludwick and Petersen (29) injected 10 to 100 milli-
grams of stilbestrol (in cottonseed oil) intramuscularly in 4
cows over a 30-day period. Butterfat and lactose increased with-
out affecting the total yield of milk. No change was apparent
in casein and total proteins.
A group of investigations sponsored by the Agricultural Re-
search Council, in England, dealt with lactation and milk com-
position. A single injection of stilbestrol per animal was em-
ployed by Parkes and Glover (25) with heifers. The slow-acting
esters appeared more effective in stimulating udder growth and
lactation, possibly due to their action over a long period. The
initial milk was colostral in character. One heifer attained a
peak of 12.5 pounds of milk daily in the 13th week, another
8.5 pounds daily at 20 weeks, and 5 others below 5.5 pounds.
Of 10 heifers in another trial, only 2 yielded over 2 pounds of
milk daily at any time, and that with 2 doses of the slowest-
acting ester. Of 7 heifers in the third trial, 2 yielded as much
as 12.8 and 8.7 pounds daily at their peak following injection
of the stilbestrol dilaurate. One injection appeared inadequate
for maximum lactation in heifers.
Folley and Malpress (11) treated 16 heifers orally and 7 by
subcutaneous implantation of pellets containing synthetic oestro-
gens. The first milk was colostral, high in solids-not-fat and
low in butterfat percentage. The change to normal composition
appeared more rapid with the higher yielding individuals (5
pounds of milk daily). Low casein and high "soluble protein"
(presumably mostly globulin) initially were typical of colostrum
in the early stages of artificially induced lactation. Lactose and
chloride contents likewise changed slowly to the values in normal
milk. Phosphatase activity decreased rapidly to the level in
normal milk. Samples of the milk were supplied to Lawson and
associates (20) who were able to extract and assay small amounts
of the estrogen from it. None of their results suggested suffi-
cient estrogen present in milk to be harmful to man, in their
belief.
Hammond and Day (18) implanted stilbestrol, or hexestrol,
tablets in 140 open dry cows and heifers on farms in England.
A re-analysis of their published records indicated that the re-
sponses were quite variable, a few individuals not coming into
milk, while two-thirds of the treated heifers attained average





10 Florida Agricultural Experiment Station

yields during the 15th week after implantation of 10 to 29.8
pounds of milk daily. About 60 percent of the individuals
yielded more milk in the 15th week than in the 10th, 7th or 4th
week, suggesting slower attainment of maximum yields than
usually is expected. Neither maximum nor total milk yields
appear to equal no-mal production.
Treatment of cows in advanced lactation with stilbestrol tablet
implants led to prompt drying off. Complete records were un-
obtainable on some animals under farm conditions but fairly
complete statements concerning production were given for 91
heifers and 5 cows. Six heifers treated 16 to 33 weeks did not
come into milk. Thirty-two heifers produced between 4,000 and
9,808 pounds of milk in 41 to 80 weeks (average of 57 weeks),
while yields of the remainder were at lower levels.
The above review indicates wide differences in the responses
of individual animals, heifers generally coming into lactation
when treated with this estrogen. Cows responded irregularly
and at less satisfactory levels than did heifers.

EFFECT ON SEXUALITY
Folley and co-workers (14) described the nymphomania:
"Two of the three non-pregnant Ayrshire cows, which received
subcutaneous injections of 1 g. diethylstilbestrol in aqueous
suspension exhibited, some time after the injections, typical
symptoms of nymphomania, namely, raised tail-head, sunken
pelvic bones, and a persistent or frequently recurring tendency
to ride, or be ridden by, other cows. Palpation of the ovaries of
these two cows per rectum revealed no evidence of the presence
of ovarian cysts or indeed of any abnormality whatever."
Lewis (21) believed that stilbestrol, unless given in low dos-
age, tends to cause the ovaries to become inactive while under
treatment. He warned that stilbestrol may cause abortion in
pregnant cows.
A goat fed 1.25 milligrams daily of stilbestrol for the last 73
days of pregnancy dropped a normal kid (23). The ovaries of
1 goat under treatment for 428 days were degenerated until
normal ovarian tissue was absent, while another under treat-
ment for 427 days had several small follicles on the ovaries.
Another goat had normal ovaries, the treatment having been
discontinued 72 days. The ovaries of 3 were quiescent, though
otherwise normal, and the ovaries of a sixth goat had a corpus
luteum.








Effect of Stilbestrol on Udder Development 11

Petersen and Boyd (1) noticed that the ovaries of sterile
heifers became quiescent during stilbestrol administration. Ani-
mals showed apparent estrus, but since ovulation was inhibited
they indicated that such animals should not be bred on apparent
estrus.
Folley and Malpress (10) examined the ovaries of heifers over
a 12-month period. Within 2 weeks to 2 months of treatment
with stilbestrol the ovaries became inactive. About 3 months
after treatment ceased the ovaries were functional. Out of 30
treated, 25 and possibly 27 heifers conceived when bred, many
at first service. Four heifers showed periodic prolapse of the
uterus, 2 having to be slaughtered. Elevation of the tailhead
occurred in all but 3 animals treated, the condition persisting
even after the next calving.
Six esters of stilbestrol were injected subcutaneously into
heifers by Parkes and Glover (25). The doses were 0.05 to 1.0
gram equivalent of stilbestrol in saline solution. One-half inch
longer growth of teats accompanied the larger injections. The
ovaries became quiescent soon and remained so until 712 to 13
weeks after treatment ceased. Three conceived at pasture later.
Quiescence of the ovaries and subsequent functional activity
were observed with 6 heifers injected similarly.
Hammond and Day (18) treated 140 open cows and heifers
subcutaneously or by implantation with either stilbestrol or
hexestrol and observed that follicles ceased developing during
that period. Ovulation was resumed from 5 to 56 days after
the implants were removed.
The uteri of stilbestrol-treated animals frequently lost their
tone, this tending to return when ovarian activity was resumed
after treatments had ceased. The pelvic ligaments relaxed,
followed by elevation of the tailhead, as occurs in some cows
with cystic ovaries. This condition persisted for some time
after treatment ceased. Frequent mounting caused 5 fractures
of pelvic bones among 48 heifers in 1 experiment. Part of the
heifers conceived after stilbestrol administration was discon-
tinued.
With 14 heifers and cows, following subcutaneous implants
of stilbestrol tablets, Folley and Malpress (9) observed nympho-
maniac behavior usually within 14 days of treatment. Asso-
ciated with it were relaxation of the sacro-sciatic and sacro-iliac
ligaments, and the attendant raising of the tailhead and lower-
ing of the coxal tubers (pinbones). Three of these 6 animals








12 Florida Agricultural Experiment Station

suffered fractures of the pelvis. The ovaries became inactive
until about 2 months after treatment ceased. One out of 11
animals conceived subsequently.
Cowie (6) described the pelvic changes observed above in the
words: "There is a general relaxation of the pelvic ligaments
and the pelvis becomes tilted forward on the head of the femur,
thus depressing the coxal and sacral tubers and elevating the
ischial tubers. In addition, the sacro-iliac ligaments are relaxed
and the sacrum is tilted forward, the lumbo-sacral articulation
being bent ventrally and the sacro-coccygeal articulation raised.
The sacro-sciatic ligaments are relaxed, producing the character-
istic hollow appearance of the sacral region. Trauma from per-
sistent coital mimicry was responsible for the fractures with
the changes in pelvic morphology as a probable predisposing
factor."
Spriggs (30) noted excessive early sexual activity in 5 heifers
and 1 cow implanted with stilbestrol tablets. The pelvic liga-
ments relaxed, with elevation of the tailhead and lowering of
the coxal tubers. Ovaries became quiescent and small and uteri
lost tone until some months after withdrawal of the tablets.
The animals were held in individual pens so there was no chance
of injury from coital mimicry.

PLAN OF INVESTIGATION
To study effects of stilbestrol on lactation and reproduction,
4 groups of cattle were used, including a total of 14 open heifers
and 5 open dry cows. Stilbestrol was dispersed in cottonseed
oil (wesson oil) in a concentration of 10 milligrams per milliliter.
This was injected subcutaneously in the neck of each experi-
mental animal, using 1 milliliter daily for 4 to 6 injections, then
2 milliliters in each of 3. injections per week until lactation began.
Upon lactation the injections were discontinued with 6 of the
heifers. They were continued with 8 other heifers and 1 cow
for 32 to 89 days after milking began, being gauged to dis-
continue near the expected peak of milk production. The supply
of stilbestrol became exhausted before 1 heifer that responded
slowly could attain peak production.
Certain body measurements were taken at regular intervals
to note changes in udder size and height of tailhead. Photo-
graphs were taken to show changes in the development of the
mammary system, rump and tailhead. Some rectal examina-
tions were made of the reproductive organs. When lactation








Effect of Stilbestrol on Udder Development 13

began the animals were milked twice daily by machine; milk
records were kept and milk samples analyzed at regular intervals.
Ultimately all experimental animals were slaughtered for
examination of the reproductive organs. The lumbar and sacral
bones from a normal cow with those from an experimental
heifer are compared in Fig. 8.

RESULTS
INJECTION RATES
The quantity of stilbestrol-in-oil injected and number of in-
jections given each animal are stated in Table 1. From 13 to
21 milliliters of stilbestrol-in-oil were used with 12 of the heifers
to develop the udder and initiate milk secretion. Two heifers-
Nos. 795 and 823-responded very slowly, requiring about 9
weeks and 53 to 55 milliliters of stilbestrol-in-oil for the udders
to develop and for milk secretion to be indicated by fullness of
the teats. One cow-No. 679-failed to respond with udder
development and reacted adversely when the injection rate was
increased to 4 milliliters thrice weekly. She was eliminated
from the test after the injection of 110 milliliters of stilbestrol-
in-oil over 71 days, because of rapid decline in physical condition.
The majority of heifers developed udder tissue rapidly, as
shown in Fig. 2. The udders filled and teats were distended,
so that milking was begun usually on the 14th to 17th day.
The animals were milked twice daily by machine.
Four of the 5 cows made little evident udder development
but milking finally was begun on the 59th day. Mastitis occurred
among this group, as indicated later, which caused 3 cows to
be discontinued.
CHANGES IN CONFORMATION
The depth between the rump and cleft of the udder and be-
tween the rump and tip of the right rear teat were measured
regularly with body calipers. To indicate something of udder
development (fullness), measurements were taken with a steel
tape from the right pinbone to the demarcation at the front of
the udder, as shown in Fig. 1 (front cover). Since the "front
of the udder" in heifers is an arbitrary point because of fat
deposits, 3 or more measurements were taken each time and
averaged.
With the second and subsequent groups of animals, height
of tailhead above the pinbones was determined with a caliper.








TABLE 1.-AMOUNTS AND NUMBER OF SUBCUTANEOUS INJECTIONS OF STILBESTROL-IN-OIL* BEFORE AND DURING LACTATION
WITH OPEN HEIFERS AND DRY COWS. .

Interval from Total Volume
Animal Age at First Injection Initial Injection Amount and Number of Injections Injections
to Lactation Before Lactation I During Lactation
Yrs. Mos. Days Days 1 ml. 2 ml. 4 ml. 2 ml. ml.
SInjections Discontinued with Lactation
Heifers
765 4 1 23 17 5 6 .... 0 17
774 3 11 18 17 5 6 .... 0 17
779 4 1 19 22 5 7 .... 0 19
789 4 8 0 14 5 4 .... 0 13
795 4 6 24 62t 5 24 .... 0 53
818 3 9 22 25 5 8 ... 0 21
Cows
435 12 1 21 59** 6 20 .... 0 46
537 8 8 14 59** 6 20 0 46
552 8 5 12 59** 6 20 0 46
679 5 9 18 ... $ 6 20 5 0 66
Injections Continued Into Lactation Period
Heifers
695 5 2 13 17 5 6 .... 36 89
760 4 3 7 17 5 6 .. 36 89
764 4 5 11 17 4 5 ... 36 86
803 4 5 9 17 5 5 ... 39 93
808 4 5 24 15 5 4 ... 29 71
813 3 10 4 17 5 5 .. 39 93
823 4 4 11 64t 5 25 .... 14 83
824 4 3 23 15 5 4 .... 20 53
Cow
692 5 4 20 59** 6 20 ... 37 110

10 milligrams of stilbestrol per milliliter of cottonseed ("wesson") oil.
"** Possibly milking could have been begun earlier.
SIndividuals responded slowly.
.t Animal was losing flesh abnormally, and was dropped from test after 71 days; did not come into milk.








Effect of Stilbestrol on Udder Development 15




















































FP -




Fig. 2.-Rear and side views of heifer No. 774 before stilbestrol-in-oil
injections (left center and upper) and 17 days later (right center and
lower), showing rapid development of the udder.







16 Florida Agricultural Experiment Station

All measurements of heifers were averaged in 2 groups.
Group A included data obtained from 6 Jersey heifers with
which injections of stilbestrol were discontinued when milking
began. Group B numbered 8 Jersey heifers with which injec-
tions were continued until the 32nd to 89th day in lactation.
Two less animals in each group are represented in the tailhead
measurements.
Some individuals in Group B (injections continued into lacta-
tion) made greater udder development than Group A, even prior



































Fig. 3.-Before (above) and 5 days after a 61-day period of injections
with stilbestrol-in-oil. The tailhead of No. 824 raised rapidly and the
animal walked with a stiff gait, the stiffness disappearing slowly after
injections ceased.








Effect of Stilbestrol on Udder Development 17

to initiation of lactation. The averages of individual measure-
ments included both bodily growth and development of udder
and teats. Rapid initial development took place in -the first
3 weeks, followed by smaller increases during the next 2 months.
A typical example is shown in Figure 2. The tailheads of ex-
perimental animals raised progressively in each instance while
injections continued and afterward returned slowly toward the
original position. Heifers in Group B, with the continuing in-
jections, showed more than double the change in height of tail-
head that occurred with Group A.
As the tailhead of heifer No. 824 in Group B raised she walked
with a stiff gait. (See Figure 3.) The stiffness disappeared
slowly after injections ceased. Heifer No. 695 suffered a double
partial dislocation of the ilio-sacral ligaments and a broken


















1_












Fig. 4.-At 91 days in lactation, No. 695 suffered a double partial dislocation
of the ilio-sacral ligaments and a broken tailhead.









18 Florida Agricultural Experiment Station

tailhead, which terminated her record at 91 days in lactation.
(See Figure 4.) The average changes in measurements are
summarized in Table 2.

TABLE 2.-CHANGES IN DIMENSIONS AND IN HEIGHT OF TAILHEAD OF JER-
SEY HEIFERS FOLLOWING SUBCUTANEOUS INJECTIONS WITH STILBESTROL-
IN-OIL.

Approxi-
mate Right Pinbone Tailhead to
Time Rump to Cleft Rump to Tip of to Front of Top of
from of Udder Rear Teat Udder Pinbones
Initial
Injection A B A I B I A [ B [ A B
Days Cm. Cm. Cm. Cm. Cm. Cm. Cm. Cm.
Initial 0 0 0 0 0 0 0 0
7 0.1 5.8 0.5 2.1 0.6 5.6 0.1 0.6
14 3.4* 7.4 4.6 6.2 3.8 10.0 1.4 1.9
21 5.8** 7.6 6.6 7.3 4.6 9.3 1.1 1.6
28 5.9** 8.0 6.9 7.6 4.7 11.3 1.3 2.0
35 6.4 9.0 7.5 9.3 4.5 11.5 1.4 2.4
42 7.0 9.5 7.9 9.9 3.3 11.6 1.4 2.5
56 7.4 9.6 8.5 10.0 5.0 11.8 1.0 2.8
68 8.6* 11.4* 9.9 12.0 6.8 12.6 0.9 3.7
78 8.9 11.4 10.6 12.2 7.5 13.4 0.7 4.2
89 8.8 11.0* 10.8 11.3 6.5 12.4 0.2 3.9
99 9.4 11.8* 11.2 12.6 7.8 10.9 0.9 4.6
103 ... **
106 *... ***
121 .. ... ... .... 0.5 4.0
151 .... ... .... 0.5 3.6

A-Group is the average for 6 heifers; 4 in last column A.
B-Group is the average for 8 heifers; 6 in last column B.
The asterisks indicate the number of animals with which injections were discontinued
at initiation of lactation in Group A, or after the planned interval during lactation in
Group B.

MILK YIELDS
The udders of non-pregnant heifers developed rapidly in 12
of the experimental animals, 2 other heifers developing more
slowly. Secretion began soon and the teats filled with milk.
This occurred in 14 to 25 days and, with 2 animals, in 62 and
64 days. The animals were milked by machine, beginning on
the 14th to 64th day, depending on the reaction of individual
animals. Dry cows gave less evidence of udder development
and milking was begun on the 59th day. Later, it was the
opinion that milking could have begun sooner with the cows.
Some of the heifers, except Nos. 795 and 823, possibly could
have been milked sooner.









17 INITIAL LACTATIONS, AVERAGE
-o 7 LACTATIONS, INJECTIONS CONTINUED
INJECTIONS DISCONTINUED WITH 1 ANIMAL x
6 LACTATION S, INJECTION DISCONTINUED ---
x'-.. x .
















2AY5 IN MILK
S 0





0 So 1W zoo 3

Fig. 5.-Average milk yields in first lactations after injection of stilbestrol-in-oil; with injections continued into a
the lactation period; with injections discontinued; and lactations following normal calvings without administration
of stilbestroi.








20 Florida Agricultural Experiment Station

Milk yields were assembled by 10-day periods and average
lactation curves were computed. The peak of production (high-
est 10-day total) occurred at 30 to 100 days after lactation
began. The animals began milk production at a lower daily
level and appeared to attain the peak yield somewhat more
slowly than following normal calving, as shown by the average
lactation curves in Figure 5. This peak was higher and pro-
duction more persistent with the heifers in which injections
continued into the lactation period. No significant changes in
milk yield were noted after 170 days in lactation, the last
injection of any heifer occurring on the 106th day of lactation.
For purposes of comparison, an average lactation curve was
computed for 17 Jerseys in the Station herd in the first lac-
tation (2).
Lactation curves were assembled for each individual but that
for No. 695 was omitted from the computed average curves,
as explained later. The milk yields in 300 days ranged between
2,838 and 4,151 pounds for the heifers, while from 1 cow 4,080
pounds were obtained, as stated in Table 3. Both average lacta-
tion curves followed a similar trend after the animals were in
milk about 5 to 6 months. Since continuation of lactation is
influenced materially by the mechanical stimulation of milking
(5, 19), continuation of milk secretion long after injections of
the estrogen ceased may be explained as maintained by mechani-
cal stimulation.
COMPOSITION OF MILK
Individual milk samples were obtained from the experimental
animals at regular intervals, the irregularity with which the
several animals came into milk affecting the intervals from
initiation of lactation to the sampling dates. These milk samples
were analyzed for total solids, butterfat (by the Babcock
method), total protein and lactose. The analyses were assembled
by time intervals into 2 groups and averages were computed.
These averages are presented in Table 4 for the Jersey heifers
in which injections of stilbestrol were discontinued at lactation
and those in which injections were continued longer.
The first milk was typical of colostrum in both appearance
and composition. Total protein was high, butterfat variable
and lactose slightly low on the day milking was begun. These
changed to normal quite rapidly.
Composition of the colostrum and changes to normal milk
were comparable with those reported for the milk of 11 dairy






TABLE 3.-MILK YIELDS OF JERSEY HEIFERS AND COWS INJECTED SUBCUTANEOUSLY WITH STILBESTROL-IN-OIL.

Time in
Injection Period Lactation to Peak Milk Production Daily Yield
Peak 10-Days' 10-Days' ____ at Time of
Animal Before During Milk Yield In 300 Entire Lactation Slaughter
Lactation Lactation Production Days Lactation Length
S Days Days Days Pounds I Pounds Pounds | Days Pounds
Injections Discontinued with Lactation
Heifers
765 17 0 100 129.9 3,366.6 5,571.0 644 dry -4
774 17 0 60 180.8 3,602.4 3,607.2 309 dry
779 22 0 30 174.4 3,686.1 5,366.6 597 dry
789 14 0 50 131.8 2,770.7' 3,440.4 439 4
795 62* 0 30 177.8 4,145.3 5,030.5 392 11
818 25 0 80 138.0 ............ 1,835.7 211 dry
Cows o
435 59** 0 .... ........ ......... 315.7t 52t dry
537 59** 0 ... ........ ........ 339.6t 52t dry
552 59** 0 ...... ,683.8t 155t dry
697 71 0 ........ (did not come into milk) ......
Injections Continued Into Lactation Period
Heifers
696 17 86 50 242.8 ............ 1,722.9 91$ dry
760 17 86 60 178.3 3,417.3 3,430.3 309 dry
764 17 89 40 214.6 3,704.4 5,730.4 554 dry
803 17 89 50 170.0 2,867.3 3,107.4 400t dry
808 15 67 80 189.5 3,812.5 4,897.3 439 7
813 17 89 6 50 161.0 2,838.5 4,261.0 633 dry
823 64* 32 40 155.0 3,361.6 4,029.1 390 5
824 15 46 70 183.1 4,151.9 5,571.3 439 9
Cow
692 59** 99 50 225.5 4,080.2 5,121.8 393 10
Individuals responded slowly.
** Possibly milking could have begun earlier.
SMastitis occurred during lactation; part of lactation of heifer 803 not affected.
1 Double partial dislocation of ilio-sacral ligaments terminated record.
SSupply of stilbestrol exhausted.









22 Florida Agricultural Experiment Station

TABLE 4.-AVERAGE COMPOSITION OF MILK FROM HEIFERS WITH WHICH
STILBESTROL-IN-OIL INJECTIONS WERE DISCONTINUED AT LACTATION, OR
CONTINUED INTO THE LACTATION PERIOD.

Injections Injections
Discontinued at Lactation* Continued Into Lactation*
Days Days
in Total But- Total Lac- in Total But- Total Lac-
Lacta- Solids ter- Pro- those Lacta- Solids ter- Pro- those
tion fat tein tion fat tein
Per- Per- Per- Per- Per- Per- Per- Per-
cent cent cent cent cent cent cent cent

1 16.71 5.13 7.18 3.26 1 18.48 8.00** 6.29
10 15.76 5.30 5.76 4.65 11 14.88 5.01 4.41 4.58
18 15.21 5.47 4.18 5.01 20 15.68 6.08 3.93 5.08
30 14.43 4.91 3.76 5.13 32 15.16 5.71 3.77 4.89
46 14.50 5.13 3.70 4.85 47 15.09 5.65 3.64 4.85
59 14.49 5.18 3.71 5.07 61 15.27 5.69 3.73 4.97
73 14.76 5.33 3.92 4.76 76 15.32 5.88 4.17 4.77
88 14.37 5.00 4.10 4.54 91 14.82 5.34 4.32 4.52
100 14.37 5.03 3.81 5.02 104 14.29 4.95 3.71 4.97
115 14.79 5.20 3.74 4.89 118 14.07 5.01 3.76 4.73
129 15.05 5.95 3.78 4.94 133 14.31 5.23 3.89 4.82
142 14.63 5.36 3.70 4.99 145 14.10 4.93 3.79 4.64
158 14.85 5.48 3.84 4.66 162 13.66 4.54 3.42 4.83
170 15.26 5.81 3.97 4.91 173 14.47 5.18 3.75 4.78
184 14.93 5.53 4.19 4.94 188 14.40 5.23 3.98 4.82
199 15.45 6.10 ...... 4.77 205 14.63 5.42 ...... 4.14
212 14.41 4.99 4.01 4.57 215 14.30 5.18 3.80 4.85
225 15.94 6.43 4.48 4.50 231 14.98 5.98 3.98 4.52
237 15.24 6.08 4.05 4.71 240 15.04 5.72 4.37 4.51
251 15.81 6.46 ...... 5.14 254 14.91 5.72 ...... 4.99
270 14.77 5.83 4.06 4.23 270 15.08 5.96 4.22 3.73
294 14.78 5.50 4.06 4.50 298 13.98 5.48 4.39 4.44
316 14.54 5.25 3.82 4.80 318 14.30 5.27 3.93 4.49
335 14.43 5.38 3.59 4.33 338 14.73 5.67 .....-
350 14.51 5.15 4.64 4.64 354 14.38 5.42 4.24 4.29
364 14.75 5.55 ..... ..... 366 14.29 5.27 ..
388 14.74 5.30 4.74 4.51
400 14.86 5.50 ....
413 15.37 5.90 3.85 4.91
427 15.12 5.70 ..
443 14.39 5.10 4.48 4.18
455 15.05 5.50

Since the heifers started into lactation on different days, and milk samples were
obtained on the same dates regardless of time in lactation, the values presented represent
averages for the 2 groups. Analyses with the group in which injections were discontinued
at lactation represent 2 animals on the 335th day, and only heifer No. 765 on the 388th day
and after. Six animals were among those with injections discontinued at lactation, and 8.
with which the injections were continued into the lactation period.
** A single butterfat test of 13.5 percent affects this average.

cows by Eckles and Shaw (8), those cited by associates of Rogers
(4) and by Grimmer (17).
The colostrum from 1 animal contained 13.5 percent of butter-
fat, increasing the average of her group to 8.0 percent. How-
ever, the group of 8 heifers with injections continued from 32







Effect of Stilbestrol on Udder Development 23

to 89 days into lactation yielded milk with a higher average fat
percentage generally than did the group in which injections
were discontinued at lactation. No other marked differences
were noted in composition of milk between the 2 groups.

PREVIOUS BREEDING AND REPRODUCTIVE HISTORY
Only open animals, as shown by examination, were selected
for use in the stilbestrol investigation. Most of the heifers were
doubtful breeders, having failed to conceive from 2 to 7 services,
and all of them were from 46 to 62 months of age. Several
animals had cystic ovaries or for some reason were not observed
in heat to be bred, while breeding was withheld with 2 normal
heifers in order that they might be utilized in these trials. The
breeding histories of these animals are given in Table 5.

TABLE 5.-PREVIOUS BREEDING AND REPRODUCTIVE HISTORY OF ANIMALS
IN STILBESTROL INVESTIGATION.

Interval
Previous Services During Interval from Last
Animal Calvings I Prior to Injection Injection to
SInjections Period Breeding Date
SDays Days
Heifers
765 none 3 17 90, 363
774 none 2 17 not bred
779 none 3 22 126, 530
789 none 2 14 not bred
795 none 5 62 not bred
818 none 4 22 not bred
Cows
455 8 4* 57 not bred
537 4 2* 57 not bred
552 5 2* 57 not bred
679 2 2* 71** discontinued
Heifers
695 none 5 103 not bred
760 none 4 103 6
764 none 6 106 263
803 none 2 106 pregnant when slaughtered
808 none 2 82 not bred
813 none 7 106 100
823 none 6 96 not bred
824 none 5 61 not bred
Cow
692 1 6* 158 14

Services since last calving.
** Udder still flabby: discontinued from trial.












Fig. 6.-Frequency of sexual excitation in Jersey heifers and cows during and following the period of subcutaneous
injection of st:lbestrol-in-oil.
Injections D:scontinued with Lactation
Heifers *
765 DL X X
774 X DL
779 X DL
789 DL X X
795 X X X X X X XX DL
818 DL
Cows
436 X. X X X D'L
537 XX X X DL
552 XX X XXX X DL X X
679 X X XXXX X X X D
Injections Continued Into Lactation Period
Heifers
695 L XX XXX X X X XX XX DX
760 XL X XX X X X XX XX X X X X X X D
764 XX L X X X XX XX D
803 L X X X D
808 H H L XXX X X D
813 LX X X DX X
823 X X XXX X L D
824 L X X XX X X D X
Cows
692 X X X X X X L X XXX X XX XX X XX D to
Days from First Injection .
0 10 20 30 40 50 60 70 80 90 100 110 120 130 0
X-Sexual excitation.
L-Lactation began.
D-Injections discontinued.








Effect of Stilbestrol on Udder Development 25

SEXUAL DEVELOPMENT
One of the first evidences of reaction to stilbestrol injections
was sexual excitation among both the heifers and cows, the
occurrence and frequency of which are shown in Figure 6.
In true estrus, cows and heifers in the Station dairy herd seldom
manifest much excitement. Animals injected with stilbestrol
varied in reaction. Sexual excitation was intense, other animals
mounting the individual repeatedly in coital mimicry. The period
of excitation persisted even as long as 4 days in an instance,
for 3 days in 3 instances and 2 days in 12 instances. Within
the lots there was almost continual sexual manifestation during
the early period of injections. Rectal examinations of the re-
productive organs of selected animals indicated non-function
of the ovaries in this period. It was planned to breed the ani-
mals 90 days after initiation of lactation to simulate a 12-month
calving interval and thereby obtain lactation curves which would
be comparable with those from the herd generally. At the end
of this 90-day interval most of the animals were quiescent. Six
heifers and 1 cow were served a total of 9 times, resulting in
only 1 conception. (See Table 5.) It must be borne in mind
that most of the 14 heifers were regarded as slow or question-
able breeders when selected for this investigation. On this
account, the poor history of conceptions cannot be attributed
to the stilbestrol. The condition of the reproductive organs at
slaughter will be discussed later.

REPRODUCTIVE ORGANS
All animals were examined prior to selection to determine the
condition of the reproductive organs. Only non-pregnant animals
were chosen. All but 2 of the heifers were considered to be
problematical as breeders, based on breeding history. The 2
heifers had been slow to conceive. They had shown estrus
shortly prior to their selection but breeding had been withheld
purposely for this investigation. Only open heifers were desired,
so as to allow a full lactation period of at least 10 months'
duration. The condition of the reproductive organs as examined
initially is shown in Table 6.

LATER OBSERVATIONS
With the raised tailheads caused by the stilbestrol-in-oil in-
jections, and persistent coital mimicry by animals weighing 700
pounds and above, there was risk of injury. In fact, heifer No.









26 Florida Agricultural Experiment Station

TABLE 6.-CONDITION OF REPRODUCTIVE ORGANS OF COWS AND HEIFERS PRECED-
ING, DURING, OR WHEN SLAUGHTERED AFTER TERMINATION OF THE STILBESTROL
INVESTIGATIONS.

Date of
Animal Exami- Right Ovary Left Ovary Remarks
nation

Cows
435 3-20-1943 Surface smooth; surface irregular initial examination
C. L.**
7- 8-1943 small; fibrous C. L.
7-30-1943 -- in estrus
8- 3-1943 uterine wall thick- -slaughtered
ened; slightly
congested

537 3-20-1943 C. L. smooth initial examination
2- 1-1944 cystic
3- 4-1944 -- slaughtered

552 3-16-1943 small small
3-20-1943 fibrous; thick smooth
2- 1-1944 pedunculated
neck; deeply
imbedded C. L.
3- 7-1944 -slaughtered

679 3-20-1943 cystic small cysts
1- 6-1944 -- -slaughtered

692 3-20-1943 crater formation surface irregular fat around ovaries
2- 7-1944- cystic
6-13-1944 barren; slaughtered

Heifers
695 12-16-1942 C. L. smooth; normal
size
1-22-1943-- open; preliminary
2- 8-1943 unchanged unchanged (before milking)
viscous mucous
vaginal dis-
charge
3-20-1943 slightly fibrous unchanged
7- 6-1943 in estrus
8- 3-1943 uterine wall slaughtered
thickened

760 12-16-1942 C. L. smooth; small
1-22-1943 -- open; preliminary
2- 8-1943 smooth; normal appears function- in estrus
ing
3-20-1943 small; nodular smooth
1-21-1944 open; in estrus
2- 1-1944 small small
3- 7-1944 slaughtered; uterus
quite small
764 3-20-1943 C. L. removed preliminary
4-17-1943 -- open
3-23-1944 -- in estrus
4-26-1944 -- in estrus; bred
11-22-1944 cystic C. L. uterus small;
slaughtered









Effect of Stilbestrol on Udder Development 27

TABLE 6.-CONDITION OF REPRODUCTIVE ORGANS OF COWS AND HEIFERS PRECED-
ING, DURING, OR WHEN SLAUGHTERED AFTER TERMINATION OF THE STILBESTROL
INVESTIGATION.-Continued.

Date of
Animal Exami- Right Ovary Left Ovary Remarks
nation_

765 12-16-1942 normal normal (was in estrus on
(normal) 11-22-1944)
1-22-1943 -- -open; preliminary
2- 8-1943 normal smooth; about good uterine tone
half-size
3-20-1943 C. L. normal
2- 7-1944 -in heat; bred
6-26-1944 cyst large
11-22-1944 cyst, C. L.; center normal slaughtered
degenerating

774 12-16-1942 smooth; small; smooth; small
fibrous
1-22-1943 -- open; preliminary
2- 8-1943 follicle smooth; normal good uterine tone
3-20-1943 small smooth; small good uterine tone
2- 1-1944 C. L.
3- 7-1944 -- slaughtered

779 3-20-1943 large C. L. removed -
10-26-1943 in estrus; bred
2-27-1945 C. L. deeply im- normal slaughtered
bedded; cystic
degeneration
begun
789 10- 4-1943 normal C. L. preliminary
2-27-1945 adhesions -- slaughtered

795 10- 4-1943 C. L. preliminary
2-27-1945 cystic C. L. deeply im- slaughtered
bedded
803 3-20-1943 C. L. normal horns of uterus
same size
4-17-1943 open
6-13-1944 found 5.8 lb. fetus; slaughtered
breeding date
not known

808 10- 4-1943 large cyst forma- pedunculated preliminary
tion
2-27-1945 cystic; peduncu- cystic slaughtered
lated; C. L. de-
generating;
adhesions
813 3-20-1943 -- -open; preliminary
4-19-1943 enlarged normal horns small
6-26-1943 cystic small
8-23-1943 accidentally bred
2- 1-1944 -- open
_2-27-1945 fibrous adhesions small C. L. slaughtered







28 Florida Agricultural Experiment Station

TABLE 6.-CONDITION OF REPRODUCTIVE ORGANS OF COWS AND HEIFERS PRECED-
ING, DURING, OR WHEN SLAUGHTERED AFTER TERMINATION OF THE STILBESTROL
INVESTIGATION.-Continued.
Date of
Animal Exami- Right Ovary Left Ovary Remarks
I nation_

818 3-20-1943 enlarged normal horns same size
6-13-1944 -- slaughtered
823 8-27-1943 bred on this date,
prior to selec-
tion; did not
conceive
2-27-1945 cystic degenera- cystic degenera- slaughtered
ation of C. L. tion
824 10- 4-1943 C. L. -- preliminary
(normal) 11-18-1943 -- open
12- 2-1943 C. L. injections started
8-27-1945 cyst deely im- cystic degenera- slaughtered
bedded tion

Injections of No. 695, 760, 765 and 774 began on Jan. 24. 1943; of 764. 779, 803, 813
and 818 on April 24, 1943; of 789, 795, 808, 823 and 824 on December 2, 1943. Injections
of cows No. 435, 537, 552, 679 and 692 began on March 20, 1943. Other pertinent facts
concerning reproductive history, and injections are listed in other tables.
** C. L. indicates a corpus luteum present.

695 suffered a double partial dislocation of the ilio-sacral carti-
lages sufficient to cripple her. After injury, she walked with
difficulty and had to be slaughtered. The federal veterinary













-I--





t;I;er, 242 IaU:% alrter illnje.tl:,r,. b-gan.







Effect of Stilbestrol on Udder Development 29

inspector diagnosed the nature of the injury upon her carcass
as a double partial dislocation.
The sacrum of heifer No. 824 likewise was distorted seriously.
She began to walk with a stiff or stilted gait. Injections during
lactation were discontinued immediately and she improved
gradually.
When heifer No. 760 underwent similar extreme sacral changes
she was photographed in mid-lactation (Figure 7); slaughtered
under inspection (when dry) and a rear quarter of her carcass
was retained for examination, along with the corresponding

Fig. 8.-The lumbo-sacral cartilage is wider at the ventral border
in No. 760 (see arrow), and the rump tilted about 4.6 higher than in
normal cow No. 516 (above). (Photo by Dr. W. B. Tisdale.)















?- -j .. -~.- 4 *--;-

-. o.- : -
N'~r







30 Florida Agricultural Experiment Station

quarter of a normal cow, No. 516, that had borne 8 calves. No.
760 had injections over a 97-day period and was slaughtered
312 days after the injections were discontinued. These parts
were photographed (Figure 8) and measurements made of the
angle of the lumbar to the sacral vertebrae. Both carcasses
had been hung by the hocks and chilled. This tended to reduce
the differences in alignment of rump to loin. Even with this
modification of alignment, the sacrum of No. 760 showed 4.6
degrees higher elevation of the tailhead than that of the normal
cow.
Externally, the live animals showed some relaxation of the
broad ligaments between the sacrum and the ischium, some-
what as occurs prior to calving. The weight of the barrel there-
fore had an advantage in leverage which drew the loin downward
and tended to tilt the pelvis correspondingly. When this hap-
pened a corresponding compensation occurred with the ilio-sacral
cartilage to the extent shown (Figure 7).
The measurements of height of tailhead, taken with body
calipers, showed that when the injections were continued longer,
greater changes occurred in these relations. After injections
were discontinued these parts tended to return slowly toward
normal. The measurements were discontinued when the ani-
mals were turned to pasture at the close of the lactation studies.
Subsequently, the animals were slaughtered and reproductive
organs of most animals were examined. It was found that
heifer No. 803 was pregnant, though no service date had been
recorded. It was learned later also that 2 or 3 of the heifers
had been in heat on pasture but instructions for breeding had
not been complied with. The scarcity of labor during wartime
interfered with completion of some of the plans.
Weights and conditions of the ovaries are noted in Table 7.

DISCUSSION AND SIGNIFICANCE
Use of the estrogen stilbestrol with virgin heifers to cause
lactation may be justified in case valued purebred animals fail
to conceive within the expected time. The changes subsequent
to its injection were thought by several investigators to have
some effect upon the ovaries, possibly beyond the period of
quiescence that accompanies its use. Subsequently, some ques-
tionable breeding animals have conceived and become normal
members of the herds.









TABLE 7.-POSTMORTEM EXAMINATIONS OF OVARIES OF HEIFERS SLAUGHTERED AFTER STILBESTROL-IN-OIL INJECTIONS
WERE DISCONTINUED.

Amount of Interval Since Right Ovary* __Left Ovary*
Heifer Stilbestrol-in- Last Injection Total Corpus Net Total Corpus Net
No. Oil Injected Weight Luteum Weight Weight Luteum Weight
M1. Days Gms. | Gms. Gms. Gms. G Gms. Gms.
Injections Discontinued with Lactation

695 89 89 Uterine wall thickened and congested
760 89 307 uterus quite small
764 86 472 5.40 ...... .5.40 9.87 4.42 .....
803 93 280 5.8 lb. female fetus
808 71 372 14.69 5.43 ...... 8.31 cystic 7.26
813 93 569 8.80 1.36 6.58 4.86 0.39 4.27
823 83 358 15.00 5.38 8.67 8.13 cystic 7.25
824 53 393 11.29 0.94 5.29 7.47 cystic 5.00

Injections Continued Into Lactation Period

765 17 394 12.18 4.33 5.65 ...... 5.65
774 17 392
779 19 561 12.87 5.56 6.40 5.45 ...... 5.45
789 13 439 8.71 0.23 8.26 16.29 6.14 8.92
795 53 391 9.84 cystic 7.89 9.31 2.31 6.92
818 21 396
Fluid released from cysts and/or follicles accounts for loss between total weight, less corpus luleum. and net weight.

I-







32 Florida Agricultural Experiment Station

Rapid development of secretary tissue in the udder and initia-
tion of secretion occurred in the majority of virgin heifers.
However, cows responded far less uniformly, and even adversely
in a few instances, to administration of the estrogen.
It is regarded that the low level of milk production resulting
from this estrogen-produced lactation is insufficient to justify
its use in commercial dairy herds when considered from an
economic viewpoint.
Administration of high levels of stilbestrol to already lac-
tating goats and cows was considered by other workers to depress
lactation.
SUMMARY AND CONCLUSIONS
The synthetic estrogen stilbestrol was injected subcutaneously
as an oil-suspension into 5 open cows and 14 open heifers.
The mammary tissues of most of the heifers developed rapidly
and began secretion. The first-drawn milk was true colostrum,
changing rapidly to the composition of normal milk. Plotted
average lactation curves showed that peak of production was
attained more slowly than normal, and at lower levels. Where
the injections were continued into lactation slightly higher
levels of milk production were attained. Following discontinu-
ance of injections lactation of the 2 groups of heifers came to
the same trend and continued so, apparently maintained by the
mechanical stimulation of milking.
Sexual excitation unaccompanied by true ovarian function
was an early effect of stilbestrol injections. This accompanied
a period of ovarian quiescence, after which apparently ovulation
occurred. Since the 14 heifers used in this investigation were
considered mainly to be slow breeders with questionable repro-
ductive histories, and since 2 or 3 showed estrus later and were
not bred, no conclusions should be drawn from this work as to
the effect on reproduction. One of the slow breeders was preg-
nant when slaughtered.

ACKNOWLEDGMENTS
Stilbestrol to initiate this investigation was supplied by Dr. W. E.
Petersen of the University of Minnesota. Later, material was donated by
Merck and Company. The late Clarence Robinson and employees connected
with the dairy herd were responsible for the daily care of animals and the
production records. Facilities of the Dairy Products and Nutrition Labora-
tories were made available for milk analyses. Certain photographs were
taken by Dr. W. B. Tisdale and J. F. Cooper. Swift and Company slaught-








Effect of Stilbestrol on Udder Development 33

ered all experimental animals under federal inspection and made parts
of 2 carcasses available for further study. Dr. W. E. Petersen read and
criticized the manuscript. For all of these contributions and courtesies
the authors express their appreciation and thanks.

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34 Florida Agricultural Experiment Station

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Effect of Stilbestrol on Udder Development 35

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