Title: Comparison of osmoregulation in two species of the genus Fundulus
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 Material Information
Title: Comparison of osmoregulation in two species of the genus Fundulus
Physical Description: ix, 114 leaves. : illus. ; 28 cm.
Language: English
Creator: Burnside, Dale Frederick, 1943-
Publication Date: 1969
Copyright Date: 1969
 Subjects
Subject: Osmoregulation   ( lcsh )
Atheriniformes   ( lcsh )
Zoology thesis Ph. D
Dissertations, Academic -- Zoology -- UF
Genre: bibliography   ( marcgt )
non-fiction   ( marcgt )
 Notes
Thesis: Thesis -- University of Florida.
Bibliography: Bibliography: leaves 109-113.
General Note: Manuscript copy.
General Note: Vita.
 Record Information
Bibliographic ID: UF00098406
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: alephbibnum - 000574232
oclc - 13842057
notis - ADA1595

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COMPARISON OF OSMOREGULATION IN

TWO SPECIES OF THE GENUS Fundulus













By
DALE FREDERICK BURNSIDE


A DISSERTATION PRESENTED TO THE GRADUATE COUNCIL OF
THE UNIVERSITY OF FLORIDA
IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE
DEGREE OF DOCTOR OF PHILOSOPHY














UNIVERSITY OF FLORIDA
1969
























For

MY PARENTS
who gave mo life

and for

LAURA
who shares my life












ACKNOWLEDGEMffNTS


I would like to express my sincere appreciation to Dr. Frank G.

Nordlie. Without his academic and personal counseling, I could not

have completed this research.

Dr. Robert M. DeWitt has liberally provided equipment and has

dealt with the many problems of organizing my committee.

I would like to acknowledge Dr. Brian K. McNab and Dr. Daniel

Belkin for giving generously of their time in serving on my committee

and evaluating many of my ideas.

My thanks go to all my fellow graduate students for providing the

academic atmosphere in which I have carried out my research. I would

especially like to thank Mr. Charles G. Yarbrough and Mr. Carl M. Colson

for many stimulating conversations. I would like to thank Mr. Colson,

Mr. Richard S. Fox III, and Mr. Paul Maslin for helping with the

collection of animals.

I would like to thank my darling wife, Laura, for her patience and

understanding and for helping type the final manuscript.

Finally thanks are due to Mr. and Mrs. Gerrald Gantt for providing

a typewriter for final processing of the manuscript.












TABLE OF CONTENTS

Page

ACKNOWLEDGENENTS. . . . . . . . . . . . iii

LIST OF TABLES . . . . . . . . . . v

LIST OF FIGURES . . . . . . . . . . vii

INTRODUCTION. . . . . . . . . 1

MATERIALS AND METHODS . . . . . . . . . . 4

Collection, Maintenance, and Acclimation of Fish . 4
Plasma Samples . . . . . . . . . . . 5
,e-termination of Blood Plasma Concentration. * * 5
Activity . . . . a * * * *. * 7
Intact Animal Oxygen Consumption ,, . . . . 8
Oxygen Consumption of Excised Gills, . . .* . 13
Gill Surface Area. . . . . . . . 15
Permeability . .. .. . . . . . 16
Statistical Methods . ............... 17

RESULTS . . . . . . . . . . . . . 18

Activity . . . . . . . . . . ...* 18
Concentration of Blood Plasma. .. . ... . . 18
Intact Animal Oxygen Consumption . . . . . 25
Oxygen Consumption of Excised Gills. ... . . . 51
Comparison of Variations in Oxygen Consumption of
Intact Animals and Excised Gills ... . . . 76
Apparent Permeability . . . . . ....... 78
Anatomical Relationships . . . . 83

DISCUSSION. . . . . . . . . . . . . 95

LITERATURE CITED. . . . . . . . . . . . 109

BIOGRAPHICAL SKETCH . . . . . . . . . . . 114












LIST OF TABLES


Table Page

1 THE DOUBLE LOGARIT~IIC RELATIONSHIP BETWEEN OXYGEN
CONSUITTION AND WEIGHT OF INTACTr F. similis AT
FOUR SALINITIES. . . ... . . . . * 34

2 STATISTICAL COI PARISONS OF SLOPES AND INTERCEPTS OF
THE DOUBLE LOGARIThICPRLATIOI1SHIP BET-EEN OXYGEN
CONSUEP2TIOhN AND WEIGHT OF INTACT F. similis AT FOUR
SALINITIES . . ... .. * . . . . 35

3 THE DOUBLELOGARITHI -C RELATIONSHIP BETWEEN OXYGEN
CONSUI'PTION AN'D ViTTGHT OF INTACT F. chrsotus AT
FOUR SALIIIITIES. .. . . . . . . . *

4 STATISTICAL COi PARISONS OF SLOPES AND INTERCEPTS OF
THE DOUBLE LOGARITHIJC RELATIONSHIP BETWEEN OXYGEN
CONSUNiPTION AND WEIGH. OF INTACT F. chrysotus AT FOUR
S LINITIES . . . . . . . . 46

5 INTERSPECIFIC STATISTICAL COIALRISOIS OF SLOPES AND
INTERCEPTS OF THE DOUBLE LOGA.ITHITC RELATIONSHIP
BETWEEN OXYGEN CONISUITION AND WEIGHT OF INTACT
ANIMALS AT FOUR SXLINITIES . . . . . . . 47

6 OXYGEN CONSUl-PTION OF INTACT F. similis AS A FUNCTIION
OF BODY SIZE ANDA SALINITY. . . .. . . . . 48

7 OXYGEN CONSUTI'.ION OF I-TACT F. chrysots AS A FUNCTION
OF BODY SIZE AND SALINITY. . . . . . . . 49

8 THE DOUBLE LOGARITHPIC RELATIONSHIP BETWEEN OXYGEN
CONSUlJPTION ANID DRY -WIGHT OF EXCISED GILLS OF
F. sinilis AT FOUR SALINITIES. . . . . . 60

9 STATISTICAL COj-ARISONS OF SLOPES AND II!TERCEPTS OF
THE DOUBLE LOGLRIThi-C RELATION:S!IP E TiEI' OXYGEN
CONSUJPTION AND DRY EIGHT OF EXCISED GILLS OF
F. similis AT FOUR SALTIITIES. . . . . . . 61

10 THE DOUBLE LOGARITIi-C RTI.ATICNSHIP BETWEEN OXYGEN
CONSUMPTION AND DRY 'WEIGHT OF EXCISED GILLS OF
F. c sotus AT FOUR SAJINITIES. .. . . ... . 70





Table Page

11 STATISTICAL COMPARISONS OF SLOPES AND INTERCEPTS OF
THE DOUBLE LOGARITHMIC RELATIONSHIP BETWEEN OXYGEN
CONSUMPTION AND D Y WEIGHT OF EXCISED GILLS OF
F. chrSsoJus AT FOUR SALINITIES.. ...... ... 72

12 INTERSPECIFIC STATISTICAL COMPARISONS OF SLOPES AND
INTERCEPTS OF THE DOUBLE LOGARITHMIC RELATIONSHIP
BETWEEN OXYGEN CONSUMPTION AND DRY WEIGHT OF EXCISED
GILLS AT FOUR SALINITIES . . . . . . . . 73

13 OXYGEN CONSUMPTION OF EXCISED GILLS OF F. sim.lis
AS A FUNCTION OF BODY SIZE AND SALINITY. . . . . 74

14 OXYGEN CONSUMPTION OF EXCISED GILLS OF F. chrysotus
AS A FUNCTION OF BODY SIZE AND SALINITY. . . . . 75

15 COMPARISON OF PERCENTAGE CHANGE IN OXYGEN CONSUMPTION
OF INTACT ANIMALS AND EXCISED GILLS AS A FUNCTION
OF SALINITY. . . . . . . . . . . . 77

16 APPARENT PERMEABILITY OF F. similis AS A FUNCTION OF
BODY SIZE AND SALINITY . . . . ..... ... 79

17 APPARENT PERMEABILITY OF F. chrysotus AS A FUNCTION OF
BODY SIZE AND SALINITY .. . . . . . . . 81

18 ANATOMICAL RELIONSHIPS . . . . . . . 88

19 INTERSPECIFIC CO PARISON OF SLOPES AND INTERCEPTS OF
ANATOMICAL RELATIONSHIPS . . . . . . . . 89

20 ENERGY EXPENDITURE FOR OSMOREGULATION (POTTS, 195). . 102

21 ENERGY EXPENDITURE OF F. similis FOR OSEOREGULATION. . 103

22 ENERGY EXPENDITURE OF F. chrysotus FOR OSOREGPULATION. 104











LIST OF FIGURES

Figure Page

1 System for measuring the oxygen consumption of
intact animals. (a) cooling coil; (b) filtration
flask; (c) airstone; (d) circulation pump; (e)
oxygen probe; (f) rubber stopper; (g) 100-ml sample
bottle; (h) magnet; (i) air hose; (j) magnetic
stirrer. . . * * * . * . . 10

2 Activity record of F. sirmlis after placement in res-
piration chambers. Data are the totals from records
of five fish. The period of minimal activity is
indicated, . . . * . * . 20

3 Activity record of F. chrysotus after placement in
respiration chambers. Data are the totals from records
of five fish. Tho period of animal activity is
indicated, , @ e ,* , @ e 22

4 Concentration of the blood plasma of both species as
a function of the concentration of the medium. Two
standard deviations are indicated around each mean.. . 24

5 Oxygen consumption of intact F. similis in fresh
water as a function of body weight. Slope = 0.6896 +
0.0755. Intercept = -1.0655 + 0.0436. Correlation
coefficient = 0.91.. . . . . . . . . . 27

6 Oxygen consumption of intact F. similis in 1/3 sea
water as a function of body weight. Slope = 0.4916 +
0,0772. Intercept = -0.9508 + 0.0274. Correlation
coefficient = 0.81.. . . . . . . . . . 29

7 Oxygen consumption of intact F. similis in 2/3 sea
water as a function of body weight. Slope = 0.5432 +
0.0927. Intercept = -0.9281 + 0,0294. Correlation
coefficient = 0.77.. , . . .. . a 31

8 Oxygen consumption of intact F. similis in full sea
water as a function of body weight. Slope = 0.5801 +
0.0625. Intercept = -0.8239 + 0.0469. Correlation
coefficient = 0.89.. . . . . . . . . . 33


vii







9 Oxygen consumption of intact F. chrysotus in fresh
water as a function of body weight. Slope = 0.3578 -
0.0348. Intercept = -0.9208 + 0.0118. Correlation
coefficient = 0.91. . . .. . . .. 37

10 Oxygen consumption of intact F. chrysotus in 1/3 sea
water as a function of body weight. Slope = 0.3430 +
0.0518. Intercept = -0.9066 + 0.0148. Correlation
coefficient = 0.81. . ..... . . . . . 39

11 Oxygen consumption of intact F. chrysotus in 2/3 sea
water as a function of body weight. Slope = 0.4119 +
0.0511. Intercept =-0.9066 +; 0.0215. Correlation
coefficient = 0.86. . . . . . . . . 41

12 Oxygen consumption of intact F. chrysotus in full sea
water as a function of body weight. Slope = 0.5077 +
0.0424. Intercept = -1.0177 + 0.0126. Correlation
coefficient = 0.93. . . . . . .. . . 43

13 Oxygen consumption of excised gills of F. similis
in fresh water as a function of dry weight of gills.
Slope = 0.3151 0.1311. Intercept = 0.7121 + 0.1325.
Correlation coefficient = 0.47. . . . . . 53

S14 Oxygen consumption of excised gills of F. similis
in 1/3 sea water as a function of dry weight of gills.
Slope = 0.5817 + 0.0673. Intercept = 0.4751 + 0.0685.
Correlation coefficient = 0.87. . . . 55

15 Oxygen consumption of excised gills of F. similis
in 2/3 sea water as a function of dry weight of gills.
Slope = 0.4144 + 0.0764. Intercept = 0.7526 + 0.0888.
Correlation coefficient = 0.77. . . . . . . 57

16 Oxygen consumption of excised gills of F. similis
in full sea water as a function of dry weight of gills,
Slope = 0.2453 + 0.0592. Intercept = 0.9531 + 0.0685.
Correlation coefficient = 0.66. . . . . . 59

17 Oxygen consumption of excised gills of F. chrysotus
in fresh water as a function of dry weight of gills.
Slope = 0.3436 + 0.1216. Intercept = 0.6022 + 0.0801.
Correlation coefficient = 0.60. . . . . . . 63

18 Oxygen consumption of excised gills of F. chrysotus
in 1/3 sea water as a function of dry weight of gills.
Slope = 0.1383 + 0.3402. Intercept= 0.7599 + 0.1991.
Correlation coefficient = 0.11. . . . .. . . 65


viii


Figure


Page








19 Oxygon consumption of excised gills of F, chrysotus
in 2/3 sea water as a function of dry weight of gills.
Slope = 0.5173 + 0.1845. Intercept = 0.5533 + 0.0979.
Correlation coefficient = 0.59. . . . . 67

20 Oxygen consumption of excised gills of F. chrysotus
in full sea water as a function of dry weight of gills.
Slope = 0.4584 + 0.1315. Intercept = 0,6302 + 0.0937,
Correlation coefficient = 0.67. , . . . . 69

21 Surface area of gills of F. similis as a function of
body weight. Slope = 0,8457 + 0.0307. Intercept
2.4330 0.0204. Correlation coefficient = 0.99, 85

22 Surface area of gills of F. chrysotus as a function of
body weight. Slope = 1.1753 0,0294. Intercept
2.3623 + 0.0133. Correlation coefficient = 0.99. . 87

23 Dry weight of gills of F. similis as a function of
body weight. Slope = 0.9874 j 0.0545. Intercept =
0.8940 + 0,0201. Correlation coefficient = 0.89. . 92

24 Dry weight of gills of F. chrysotus as a function of
body weight. Slope = 0.9671 t 0,1022. Intercept =
0.7021 + 0.0284. Correlation coefficient = 0.65. . 94


Figure


Page










INTRODUCTION


The integrity of any organism is maintained by the functioning of

homeostatic systems. Various parameters are physiologically controlled

at levels differing from environmental levels. Because an organism is

an open system, energy is required not only to establish these internal

levels but also to maintain them. The energetic cost of regulating ary

parameter is a function of:

1. the difference between the maintained level and

the environmental level

2. the degree to which the animal is open to the

parameter

3. the efficiencies of the controlling mechanisms.

The osmoregulatory system is one by which body fluid concentration

can be maintained at a relatively constant level irrespective of varia-

tions in the concentration of the external median. General aspects of

osmoregulation have been reviewed by Krogh (1939), Hober et. al. (1945),

Baldwin (1948), and Potts and Parry (1964). Osmoregulation in teleost

fishes has been reviewed by Smith (1932), Black (1951, 1957), Gordon

(1964), and Parry (1966).

To study the problem of teleost osmoregulation, I have chosen

two congenoric Cyprinodont species which inhabit areas of different sa-

linity conditions. The ouryhaline Fndultus similis (Baird and Girard)

lives in salt marshes where salinity conditions are highly variable,







The comparatively stenohaline Fundulus chrysotus (Holbrook) has been

found occasionally in waters of up to 24.7 ppt (Kilby, 1955), but

large populations are restricted to fresh water.

The energetic cost of osmoregulation in fishes is the sum of the

transport work performed by the kidneys, gills, and intestinal tract.

The gills account for the majority of the energetic expenditure

(Margaria, 1931; Potts, 1954). In order to estimate the energetic

cost of osmoregulation, I have measured the oxygen consumption of intact

animals and excised gills as a function of the concentration difference

between the animal and the environment.

The degree to which the osmoregulatory system of an animal is open

to the environment is termed permeability. Because the energetic effi-

ciencies of the controlling mechanisms are not considered in this study,

they have been combined with permeability to yield a compound factor

which I call apparent permeability. If the efficiencies are high or

the permeability is low, the apparent permeability is low. If the

efficiencies are .low or the permeability is high, then the apparent

permeabiltiy is high. Apparent permeability varies directly with permc-

ability and inversely with the efficiencies of the controlling

mechanisms.

Parry (1958) has suggested that body size is an important factor

affecting the ability to osmoregulate. This factor has been almost

completely ignored in osmoregulatory studios. In the present study,

body size is considered by making plots of log oxygen consumption-log

body weight at different salinities. This type of analysis has been

carried out by Hickman (1959) for fishes and by Gilchrist (1956), Rao

(1958), and Lumbye (1958) for various invertebrates. For tissue res-







piration studies, the dry gill tissue weight is correlated with oxygen

consumption and with body weight in order to consider the influence of

body size.


Purposes

The purposes of this study areas

1. to determine the precision of osmoregulation of two

closely related species of fish which are from habi-

tats of different salinity conditions

2. to consider the energetic cost of osmoregulation for

these two species by measuring the oxygen consumption

of intact animals and excised gills as a function of

salinity

3. to calculate apparent permeability as a function of

body size and salinity

4, to compare the magnitude and direction of changes in

the oxygen consumption of intact animals and excised

gills as a function of body size and salinity.












MATERIALS AND METHODS


Collection, Maintenance, and Acclimation of Fish


Seines were used to collect F. similis from tidal creeks near Cedar

Key, Florida. The salinity at times of collection varied from 8 to

34 ppt. Specimens of F. chrysotus were collected in the Gainesville,

Florida,area from Lake McCloud and River Styx with dipnots, seines, and

small dredges.

Fish were maintained in 10-gallon aquaria. Water was aerated and

filtered. Temperature was regulated between 20 and 250 C. During the

studies with intact animals, a 12-hour light and 12-hour dark cycle was

maintained. For the blood plasma studies and tissue metabolism experi-

ments, light was not controlled. Fish were fed commercial shrimp

pellets.
4
F. si-milis was acclimated to the different salinities in a step-

wise fashion. Usually the salinity at collection was near that of full

sea water. Therefore, the fish were first kept in full sea water from

Marinelanc of Florida (35.5 ppt). Two days later, 3/4 of the fish

were transferred to a dilution of 2/3 sea water to 1/3 fresh spring

water. In another two days, 2/3 of these fish were transferred from

2/3 sea water into a 1/3 sea water mixture. Because F. similis could

not survive 100 spring water, 1/2 of the fish in 1/3 sea water were

transferred into a mixture equaling ca. 1 ppt salinity. After all

transfers had been completed, a period of two weeks was allowed for







full acclimation. Parry (1966) has suggested that only a few hours

are required for fish to become acclimated to salinity.

F. chrysotus was also acclimated in a stepwise fashion. The

process began in fresh water and proceeded into 1/3, 2/3, and full sea

water. A period of two days was allowed between transfers and a period

of two weeks was allowed for full acclimation.



Plasma Samples


Plasma samples were obtained using heparinized capillary tubes.

Fish were removed from the aquaria and excess moisture in the gill re-

gion was absorbed with tissue paper. A hoparinized capillary tube

(1.4-1.6 mm in diameter) was placed adjacent to the isthmus and gently

twisted and pushed into the heart. When the capillary tube met re-

duced resistance and the heart had been pierced, the tube was slightly

retracted. WJhole blood entered the tube.

Samples were centrifuged at 10,000 rev/min for 15 minutes to separ-

ate the plasma from the blood cells. For large samples (all F. similis

and some F. chrysotus), the plasma was separated from the cells by

breaking the tube at the plasma-cell interface. Then the plasma was

transferred to a new capillary tube. Such a separation was too diffi-

cult for small samples and the plasma was frozen in a vertical posi-

tion with the column of cells under it.



Determination of Blood Plasma Concentration


The determination of blood plasma concentration was modeled after

Gross (1954), Hoar (1960), and Pavlovskii (1964). This method is per-







forced by comparing the time of melting of frozen solutions of unknown

osmolar concentration with solutions of known osmolar concentration

when allowed to warm very slowly and linearly in a cold brine (satu-

rated NaC1) bath. Because only 0.001-0.01 rl of sample is needed to

fill a small portion of a capillary tube, this method is desirable for

small quantities.

A 1-gallon battery jar was suspended in a 2.5-gallon bucket which

had a 12.5 cm diameter hole in the bottom. StyrofoaQ. insulation was

placed between the bucket and battery jar. The bucket was elevated by

three legs. Reflected light was passed through the battery jar con-

taining cooled brine. One polaroid plate, 0.030 inches thick and 30%

light transmission, was placed immediately below the battery jar and

another was placed above the battery jar in the "crossed" position.

This allowed 1% light transmission and caused crystals to appear to

glow. Gentle circulation of the brine was provided by an air-driven

stirrer. For suspending samples in the battery jar, a capillary tube

holder was made from a plastic soap dish.

Standard solutions were mixed to known concentrations as follows

according to Harned and Owen (1958, page 492)

Molarity Osmotic Coefficient Osmolarity

0.1 NaCl 1.86 .186

0.2 NaCl 1.84 .368

0.3 NaC1 1.83 .549

0.5 Iacl 1.81 .905

An experimental run consisted of these four standards plus two to

four unknowns. All volumes were approximately equal. The standards

and unlncwns were placed in the holder and frozen with dry ice. The







brine was cooled to between -8 and -100 C with dry ice. The capillary

tube holder was suspended in the cooled bath and the polaroid plates

were positioned. The bath was allowed to warm at the rate of ca. 1 C

every half hour during the critical temperature range from -2 to 00 C.

The thawing of ice crystals was observed and time was recorded as the

last crystal melted.

Then the concentrations of the unknowns could be determined from

a graph of osmolar concentration and time of thing of standards.

The accuracy of this method is + 5-10 mosm/liter.





The system for measuring the effects of experimental handling-ono

the activity of fish consisted of a 6.3 volt light source with an infra-

red filter and a Hunter photorelay connected to an Esterline Anxgu

event recorder.

The experimental fish was placed in a flask (either 250 or 500 ml,

depending upon the size of the fish) between the infrared light source

and the photorelay. Whenever the fish broke the beam, the event was

recorded. During an experiment, water was kept aerated with an air-

stone and temperature was maintained constant at 200 C. All experi-

ments were continued for 24 hours in the dark.

Five individuals of each species were used. The number of events

on the recorder paper was- grouped into 1-hour intervals. Data from

each of five fish of the sane species were added together for each

hour interval. The total number of times that the beam was broken per

hour was graphed against time. The period of minimal activity was

thus determined.







Intact Animial OQ:yen Consunption


Apparatus

The system for measuring the oxygen consumption of intact animals

was modified from that of Fry and Hart (1947).

The respiration chamber consisted of a one-arm suction filtration

flask fitted with a one-hole rubber stopper through which a glass tube

was passed. The end of the glass tube inside the flask was positioned

as far away from the arm as possible (see Figure 1). Two sizes of

flask were used (250 and 500 ml),depending upon the size of the fish.

A 19-liter aquarium served as both the temperature bath and the

resevoir for the circulating medium. One end of the aquarium was re-

moved and replaced by plexiglass through which a hole was drilled. A

glass tube, 5 cm long, was passed through this hole around which aquar-

ium cement was applied to prevent leakage. Both ends of the glass

tube were connected to tygon tubing. The longer tube on the outside

of the aquarium passed through a circulation pump and back into the

aquarium. The shorter tube on the inside was connected to the glass

tube of the respiration flask. When the circulation pump was turned on,

water left the flask via the glass tubing and was replaced by water

entering through the arm of the flask. See Figure 1 for flow direc-

tions.

Water circulated through the cooling coil from a constant tempera-

ture bath held at 200 C. An air pump vented through airstones main-

tained a high oxygen concentration in the aquarium water.

To provide insulation and darkness, the aquarium was placed in a

large Styrofoaz cooler which was painted flat black on the inside.








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Determination of Dissolved Oxygen

The Precision galvanic cell oxygen analyzer reads linearly in

milliamperes of electricity flowing between cathode and anode, It meas-

ures oxygen concentration to within 0.1 ppm at constant temperature.

The analyzer's flow requirement of 1.0 ft/sec was met with an electro-

magnetic stirrer (see Figure 1).

Approximately 200 ml of water at the samo temperature and salinity

as the water in the 19-liter aquarium was vigorously aerated overnight.

A Winkler determination for dissolved oxygen indicated that this water

was consistently 99-100l saturated. This water was used as a standard

of reference for the oxygen analyzer readings.

Readings in milliamperes were taken for 100 ml of the standard

water and for 100 ml of an unknown sample, The following calculation

yielded dissolved oxygen in the unknown samples


milliamperes-sample X amount dissolved at saturation
milliacperes-standard for given temperature and salinity


Saturation values were read from a nomogram which was supplied with

the oxygen analyzer.


Experimental Procedure and Analysis

The experimental animals were fasted for 24 hours prior to their

use. Each fish was placed in a full respirometer flask which was

stoppered and connected to the tygon tubing inside the aquarium. The

circulation pump was turned on and adjusted to give a flow rate of

100-150 ml/min. This rate had no visible effects cn the fish and was

fast enough so that the water in the flask would stay as vell aerated

as the water in the aquarium. The fish was allowed to remain in this

condition until the period of minimal activity had been reached.






At the beginning of an experiment, a 100 ml sample of water was

removed through the outside tygon tubing and oxygen concentration was

determined. The system was then stopped from flowing for 30 minutes

and a second sample was then taken from inside the flask. The dead

space (tygon and glass tubing) was cleared and a 100 ml water sample

was collected through the outside tube. The oxygen concentration was

determined and subtracted from the first determination. The system was

then allowed to flow for 30 minutes and the entire procedure was re-

peated from the beginning. Five such determinations were made during

a 4.5-hour period. Fish were superficially dried with paper towels and

weighed after experiments on a TorsioQ balance to a precision of 0,01 g.

Experiments with blue dye demonstrated that no mixing of water

entering and leaving the flask occurred while drawing samples.

The average of five readings was considered to be an estimate of

routine oxygen consumption as defined by Beamish and Mookherjii (1964).

The log of oxygen consumption (ml/hr) was correlated with the log of

body weight (g) for each species at four different salinities.

Log oxygen consumption = A log body weight + log B

where

A = increase in log oxygen consumption relative to

increase in log body weight

B = amount of oxygen consumed by a 1.0 g fish.

Special Considerations

Fry (1957) has suggested that the following variables may affect

the oxygen consumption of fish:

1. experimental temperature and thermal history

2. oxygen and carbon dioxide tensions







3. seasonal influences

4, diurnal variations

5. activity

6. nutritional state

7. sex

8. body weight

9. environmental salinity.

In this study, an attempt was made to consider each of those fac-

tors and to allow only body size and environmental salinity to vary.

Diurnal variations, sex, and seasonal influences are not discussed else-

where in this paper. Experiments were performed between 0800 and 1200

(Eastern Standard Time) and between 2000 and 2400 (Eastern Standard

Time). No difference was apparent in the results between these two

groups. Sex of individuals was determined, but was found not to sig-

nificantly affect oxygen consumption. The duration of these studies

did not permit the observation of possible seasonal influences on

oxygen consumption.



Oxygen Consumption of Excised Gills


Eperimental Setup

The oxygen consumption of excised gills was determined using a

Gilson differential respirometer. This is a closed system in which

the reaction vessels are separated from a compensation chamber by a

manometer (Umbreit et al., 1956). Fifteen ml capacity flasks were

used. The center well of the flask contained a piece" of folded filter

paper which had been saturated with 10% KOH. The main compartment of

the flask contained the gills and 2.0 ml of water of the desired

salinity.






Experimental Procedure

The gill apparatus was excised as a unit. A single cut was made at

the caudal end of the isthmus. The isthmus was pulled cephalad exposing

the gill apparatus. After the gill apparatus had been teased loose

using a scapel and dissecting needles, it was separated from the fish

with a forceps and divided into right and left halves. This division

allowed the tissue to lie flat in the manometer vessels and be completely

covered by the medium.

.Oxygen consumption was corrected to standard conditions. Meas-

urements were made at 200 C and barometric pressure was recorded at the

beginning and end of each experiment.

A period of thirty minutes was allowed for equilibration and then

readings were taken at 15-minute intervals for a period of 2.0 hours.

At the end of each experiment, the gills were removed from the manom-

eter vessels and the surplus liquid was absorbed from the tissue with

paper towels. The gills were placed in small unstoppered vials and

dried for one week at 1100 C in a drying oven. At which time, the dry

tissue weight was determined with a Mettle )analytical balance to a

precision of 0.1 mg.


Analysis

For each species in each medium, a plot of log dry gill weight (mg)

against log oxygen consumption (,ul/hr) was made.

Log oxygen consumption = A log dry gill weight + log B.

The wet body weight of each fish was also recorded and log dry

gill weight (mrg) was plotted as a function of wet body weight (g).

Log dry gill weight = A log wet body weight + log B.







Special Considerations

Krebs (1950) has discussed two important aspects of tissue

metabolism
1. the choice of medium

2. treatment of tissues.

In the present study, the medium was an artificially prepared sea

water (Svedrup et al., 1942) and dilutions were made with distilled

water to produce salinities which would be consistent with those of

the blood plasma studies. The gills were excised as an intact unit

and tissue damage was minimal.


Gill Surface Area


The method of determining the surface area of the gills was

according to Gray (1954) and Hughes (1966). Intact fish were weighed

and their gills removed. The number of filaments on each gill arch

was counted for the gill apparatus on one side of the fish. The average

length of these filaments was determined by measuring every fifth fila-

ment on each gill arch. The average number of lamellae per mm was

determined by making five counts on the filaments of each gill arch.

The average area of the lamellae was determined by sampling ten lamellae

per fish. These were drawn by means of a ZeichentubuiP and a planimeter

was used to determine surface area. The lamellae are triangular in

cross section. Because these drawings are two dimensional, their areas

were multiplied by a factor of three to get total surface area. It is

believed that this is an underestimate of the true surface area be-

cause the triangular cross sections have curved sides. The formula for

gill surface area is:







SA = 8 X F L X 1 X sa

where:

SA = total gill surface area in mm2

8 = total nunbor of gill arches

-F = average number of filaments per gill arch

L = average length of filaments in mm

1 = average number of laiellae per mm

sa = average surface area of lamellao in mm2.

For each species, log surface area (nm2) was correlated with log

wet body eight (g).

Log surface area = A log wet weight + log B.

Fish and gills were stored in 10% formalin during these measure-

ments and correction factors for shrinkage and weight change Twre

applied according to Parker (1963).



Permeability


Gill permeabilities were inferred from other data by using the

equation of Potts (1954):

W = KA (B-M) R T In B/M

whe re

W = work in cal/hr

K = permeability in moles/mm2-molar differonco-hr

A = surface area in mm2

B = blood concentration in moles/liter

M = medium concentration in moles/liter

R = 1.987 cal/C-mole

T = absolute temperature in oK.







This equation may be rewritten
W
K = A(B-II) R T In B/,

Work was derived from gill oxygen consumption readings by multi-

plying data by 4.8 cal/ml 02. It was decided better not to subtract

a small amount of energy used for maintenance of the gill cells, because

Harvey et al. (1967) have show that this portion of the metabolism is

also instrumental in ion transport. Surface area of the gills was known

from the relationships in Table 18. Molar concentrations of the medium

and blood were calculated from osmolar concentration data.

Assumptions upon which these calculations are based are considered

in the discussion.



Statistical Methods


Regression lines were calculated according to the least squares

method for curvilinear regression (Miller and Freund, 1965).

Regression coefficients are given with one standard error of the

estimate and are compared by the method given in Bailey (1959).

Intercepts are given with one standard deviation according to

Snedecor (1965). Both the variation around the sample mean and the

variation of the regression coefficient are taken into account.

Comparison of intercepts involved the use of "pooled" variances from

the regression and the test statistic for intercepts according to Miller

and Freund (1965).

Sample standard deviations from regressions were calculated for

each regression line according to Snedecor (1965).

Means of the blood plasma samples were compared using a standard

"t" test statistic for hypotheses concerning two means after Miller and

Freund (1965).












RESULTS


Activity


Both speciesare most active immediately after being placed in

the respiration flasks. Activity then decreases to a minimal level

and eventually increases until the end of the experiment (see Figures

2 and 3). Between 11 and 15 hours after the beginning of the experi-

ments, F. similis exhibits minimal activity which is approximately 40%

of the original level of activity. The period of minimal activity for

F. chrysotus occurs between 7 and 18 hours after being placed in res-

piration flasks, and is loss than 1% of the original activity. Meas-

urements of routine oxygen consumption were made during these periods

of minimal activity.



Concentration of Blood Plasma


As the concentration of the medium is raised from 25 mosm/litor

to 1,000 mosm/liter, the concentration of the blood plasma of

F. similis increases from 339. 15.9 mosm/litor to 397 + 7.1 mosm/liter

(Figure 4). At CM] = 370 mosm/liter, the mean fish plasma concen-

tration equals that of the medium. All means are significantly dif-

ferent from each other except those of fish in 1/3 and 2/3 sea water.

The concentration of the blood plasma of F. chrysotus increases

from 246 + 22.8 mosm/liter to 459 26.3 mosm/liter as the medium.



























Figure 2.


Activity record of F. similis after placement in respiration
chambers. Data are the totals from records of five fish.
The period of minimal activity is indicated.
















































Tim~ hours


400-


3501-


3001-


2501-


2001-


1501
(]Q!


I- I


I I


2U





























Figure 3. Activity record of F. chysotus after placement in respira-
tion chambers. Data are the totals from records of five
fish. The period of minimal activity is indicated.

















251



201


Timo-hours

















C
0


0

cs






0






aS
0
o














4-1;
0








0








okd






00







o
P0






OQ
0
^1
C
^! T

























































Ci
AZ!,vulouis-ems-uld P0018


ar



I,,
0
U)

'0r
Qa







is increased from 15 nosm/liter to 660 mosm/liter. At [M] = 980

miosm/liter, the blood plasma concentration drops to 297 25.3 mosm/

liter (Figure 4). The point of osmoneutrality is at [M- = 405 mosm/

liter = B] All means are significantly different from one another

(P<.001).


Intact Ainmal Oyen Consmaption


There are significant variations in the oxygen consumption of

intact F. sidilis in response to variations in salinity. The results

are presented graphically in Figures 5 8 and are summarized in

Table 1. The slopes of the log oxygen consumption-log body weight

regressions are lowest in 1/3 sea water, 0.4916, and increase as

IB M increases. In fresh water, the slope is 0.6896, an increase

of 40%. In 2/3 sea water, the slope increases 10% to 0.5432 and in

sea water it is 0.5801, an increase of 30%. There are no significant

difference among these slopes (T.able 2).

The intercepts of these regression lines increase from -1.0655

in fresh water (0.086 ml/hr) to -0,8239 in full sea water (0.150

nl/hr), All intercepts are statistically different from each other,

except those in 1/3 and 2/3 sea water (Table 2).

The slopes of the log oxygen consunption-log body weight re-

gressions for F. chrysotus follow the same pattern as for F. similis.

See Figures 9 12 and Table 3. The slopes are minimal in 1/3 sea
water, 0.3430, and increase as (B MI increases. In fresh water,

the slope- is 0.3578, a 10% increase over the slope in 1/3 sea water.

In 2/3 sea water, it increases 12% to 0.4119 and it is 0.5077, a 15%

increase, in full soa water. Statistically, the slope in sea water

























Figure 5. Oxygen consumption of intact F. similis in fresh water as
a function of body weight. Slope = 0.6896 + 0.0755.
Intercept = -1.0655 + 0.0436. Correlation coefficient =
0.91.

































fresh water


0.02


Body WVight -g


























Figure 6.


Oxygen consumption of intact F. similis in 1/3 sea water as
a function of body weight. Slope = 0.4916 + 0.0772.
Intercept = -0.9508 + 0.0274. Correlation coefficient =
0.81.

























* S


113 sea water


Body Woight-g


N020

i


0.021


0.01

























Figure 7. Oxygon consumption of intact F. similis in 2/3 sea water as
a function of body weight. Slope = 0.5432 + 0.0927,
Intercept = -0.9281 + 0.0294. Correlation coefficient =
0.77.





















0.40
*
0.30



**
bo
S0.20-




0.10- e




0.0

0
0.05 -
02/3 a2 water


x
0

0.02 -




0.O 1 I._,,.. ,. .|....i.11 *
0.1 0W2 0.5 1 2 3 4


Mody Weight-g


























Figure 8. Oxygen consumption of intact F. similis in full sea water as
a function of body weight. Slope = 0.5801 + 0.0625.
Intercept = -0.8239 + 0.0469. Correlation coefficient
0.89.


























c .0





0.02
0 01 I s I I










0.1 0.2 0.5 1 2 5 10

Body Weight-g











0 o 0 0
0 0 0 0
n Hl l






O O O O
* *



S o o0 0



O O O O
-o c- O
o0 CO C \ L0



U) +1 +1 +1 +1
0 o
'.rO O O 0

F 0 0 O 0 0
0H r- 0 0 0
N I I I I



o r


0 o



H O O O O


V, C9- c C0
U) *+1 +1 +1 +1








E-4 0 0

4)9 4 0
SH N n







EOi

9 r;


O

0
O0Rr


0
0\ N l U N












TABLE 2

STATISTICAL COI-PARISONS OF SLOPES AND INTERCEPTS OF
THE DOUBLE LCGARITIMIC RELATIONSHIP BETWEEN OXYGEN CONSUI.iPTION AND
WEIGHT OF INTACT F. similis AT FOUR SALINITIES




Comparison Slopes Intercepts



f-1/3 P>.05 *P<.05


f-2/3 P>.05 *P<.01


f-3/3 P>.05 *P<,O01

1/3-2/3 P>.05 P>.05


1/3-3/3 P>.05 *P<.01

2/3-3/3 P>.05 *P<.05
.V


*Considered to be statistically significant.




























Figure 9,


Oxygen consumption of intact F. chrysotus in fresh water as
a function of body weight. Slope = 0.3578 + 0.0348.
Intercept = -0.9208 + 0.0118. Correlation coefficient =
0.91.



































fresh water


2 34


Body Weight-g


0.01 L
0.1


0.5


























Figure 10. Oxygen consumption of intact F. chrysotus in 1/3 sea water
as a function of body weight. Slope = 0.3430 + 0.0518.
Intercept = -0.9066 + 0.0148. Ccrrelation coefficient =
0.81.










































0.01 I I I I I I
0.1 0.2 0.5 1 2 3 4


Body Weight -g




























Figure 11. Oxygen consumption of intact F. chrysotus in 2/3 sea water
as a function of body weight. Slope = 0.4119 + 0.0511.
Intercept = -0.9066 + 0.0215. Correlation coefficient =
0.86.











































Body Weaght-g


























Figure 12.


Oxygen consumption of intact F. chrysotus in full sea water
as a function of body weight. Slope = 0.5077 + 0.0424.
Intercept = -1.0177 + 0.0126. Correlation coefficient =
0.93.




























O

0-40


EH



oB











S-


+1


o














P4
0


















4,
0
k4H



















U
0


4 g
Ot
CO
0


o
o









O




O
0
rI


0
o



I







o0
o

CO-



o
+1












0
O 4
























N
+1

CO

































N


,N N N/


cH (1N <^
H- CN (r%


O









0
C-'


0'


0
+1
N
N

.0
H







O
0
+1
N
0


go
CrI \0







differs from both the slope in fresh water (P<.01) and the slope in

1/3 sea water (P<.05). The hypothesis that any other differences in

slopes exist is rejected (P>.05). See Table 4.

The intercepts of the regression lines for F. chrysotus do not

show as much variation as those for F. similis. They are -0.9208

(0.120 ml/hr) in fresh water and -0.9066 (0.124 ml/hr) in 1/3 and 2/3

sea water. However, the intercept decreases in full sea water to

-1.0177 (0.096 rml/hr), a result which is significantly different from

all other intercepts (P<.001). See Table 4,

An interspecific comparison of slopes at each salinity shows

that they differ in fresh water (P<.001). In all other salinities, a

difference in slopes does not exist (P>.05). The intercepts differ in

fresh and full sea water (P<.001), but do not differ in 1/3 and 2/3

sea water (P>.05). See Table 5.

Tables 6 and 7 show oxygen consumption of F. similis and'

F. chrysotus of five sizes at four different salinities. These values

are calculated from the relationships given inTables 1 and 3.

Changes are calculated relative to values in 1/3 sea water which is

nearest to being osmotically equivalent to the blood plasma.

Oxygen consumption is lowest in fresh water for similis

weighing from 0.5 to 4.0 g. A 4.0 g fish has approximately the same

oxygen consumption in fresh and 1/3 sea water. A 10.0 g fish shows

a 21,1% increase in oxygen consumption in fresh water. In 2/3 sea

water, fish of all sizes show increases in oxygen consumption (1.6%

to 18.6%). In full sea water, increases range from 25.0% for a 0.5 g

fish to 63.8% for a 10.0 g fish. All fish in a hyperosmotic medium

show increases in oxygen consumption relative to 1/3 sea water. Small












TABLE 4


STATISTICAL COMPARISONS OF SLOPES AND INTERCEPTS OF
TIE DOUBLE LOGARITIHflC RELATIONSHIP BETTVEEN OXYGEN CDNSUIPTION AND
WEIGHT OF INTACT F. chrysotus AT FOUR SALINITIES




Comparison Slopes Intercepts


f-1/3 P>.05 P>.05

f-2/3 P>.05 P>.05


f-3/3 *P<.oi *P<,ool

1/3-2/3 P>.05 P>.05


1/3-3/3 *P<.05 *P<.O01

2/3-3/3 P>.05 *P<.001


*Considerod to be statistically significant.












TABLE 5

INTERSPECIFIC STATISTICAL COMPARISONS OF SLOPES AND INTERCEPTS OF
THE DOUBLE LOGARTIINIC RELATIONSHIP BETWEEN OXYGEN CONSUMIFTION AND
WEIGICT OF INTACT ANhIMALS AT FOUR SALINITIES




Medium Slopes Intercepts


f *P<,001 *P<.001


1/3 P>.05 P>.05


2/3 P>.05 P>.05


3/3 P>.05 *P<.001


*Considered to be statistically significant.












* *
C? u- \ "

N n+

C; (3


0

I o




















+0
ccA












I 0
"

















+ o


0

0













0


0 0 0 0









* *0


0 H CM l
Cn M \ N C\ H







0 0 0 0 0
* 0 0 0 0
0 r- CM 0
H


* * *


+ + + H




0 0 0 0 0
1 1
0000


cs





I o

CO)


-t


*da
-H
H

* 0





CQ H







0 *
0O




4rx


0
0





*H




0 (
o
-.0












'I 0
C


0:
0,

a
2










to
\0 13
I o



o







0
to






II
1 0
I| -
0
( 1











II








+0
* -


to-
" ^









'-A


I I I I


0

O 2








0
H 0
I 1





S0
CO


o0


CO NM -- ,I. n C-
03 r r r
0 0 0 0 0






I I I



O H H
CO 0 0C 0
o& o \ 1 0
0 r-! r-i r'l N -I




0 0 0 0 0








O 0 0 04 0


o 4o o o
S
O 0 H H N


I I I I I

c0 Cm \0 Co c-
0 Co 0\ r-
0 0 0 0 0





e7. -C - 'A
00 0 CO0



.& H 0 N in
I I + +


o 0 0 0 0


Id
0
0

0


0a
o H

*rl





S*r
go







pc

OQ
g







fish (loss than 4.0 g) in a hypoosmotic medium show decreases in

oxygen consumption,while large fish (more than 4.0 g) in the same

salinity show increases in oxygen consumption.

The oxygen consumption of F. chrysotus is relatively stable from

fresh to 2/3 sea water and then drops in full sea water. In fresh

water, all fish show slight decreases in oxygen consumption ranging

from 4.2% for a 0.5 g fish to 2.Z2 for a 2.0 g fish. In 2/3 sea water,

fish below 1.0 g show a slight decrease in oxygen consumption,and fish

above 1,0 g show a slight increase in oxygen consumption. In full sea

water, all fish show decreased oxygen consumption with the greatest

depressions occurring in the smallest fish.

The slopes obtained for the double logaritmic relationship be-

tween oxygen consumption and body weight are lower for both species

in all sclinities than those reported for other fishes. Job (1955)

found slopes of 0.802 0.856 for Salvelinus fontinalis. Hickman (1959)

found a slope of 0.86 to represent the effect of weight on the oxygen

consumption of the starry flounder, Platichthys stellatus. He also

found slope values of 0.84 for lemon sole, ?arohys vetulus, and 0,90

for sanddab, Citharichthys stigLmaus. O"Hara (1968) found slopes of

0.717 and 0.710 for Loomis machrochirus and Lepomis fgbbosus re-

spectively. Beamish aeid Mookherjii (1964) found an average slope of

0.85 for Carassius auratus. Only the value inferred from data of

Wells (1935) for Fndulus parvipinnis approaches the values in this

study. His data imply a slope of 0.50. All other studies concerning

the dependence of oxygen consumption on weight in fishes have been

conducted using larger fish than those in this study and only data of

Wells are for fish of a comparable size range.






Oxygen Consurption of Excised Gills

The slopes of the regression lines of log oxygen consumption-log

dry gill weight for F. sinilis are maximal (0.5817) in 1/3 sea water

and decrease as JB MI increases (Figures 13 16 and Table 8).

In fresh water, the slope decreases 46s to 0.3151. In 2/3 sea water,

it is 0.4144, a 29% decrease. In full sea water, the slope is minimal

at 0.2453, a 57Z decrease. Statistically, the slope in full sea

water differs from both the slope in 1/3 sea water (P<.001) and the

slope in 2/3 sea water (P<.05). See Table 9.

The intercepts of these regression lines for F. similis are

minimal in 1/3 sea water (0.4751) and increase as IB Mi increases.

In fresh water, the intercept increases 50% to 0.7121. Intercepts

are 0.7526 and 0.9531 in 2/3 and 3/3 sea water respectively. These

correspond to increases of 57p and 101% over the intercept in 1/3

sea water. The intercept in 1/3 sea water differs from the intercept

in 2/3 sea water (P<.02) and from the intercept in full sea water

(P<,001). See Table 9.

The gills of F. chrysotus were more difficult to handle because

they were smaller (1 20 mg dry weight) than gills of F. similis

(1 157 nig dry weight). Only 55% (66 of 120) of the excised gills
survived the operation and consumed oxygen when placed in the res-

pironmeters. Therefore, the following results are of questionable

value.

The slopes of the regression of log oxygen consumption-log dry

gill weight for F. chrysotus vary neither directly nor indirectly

with IB MI (Figures 17 20 and Table 10). The slope is minimal in



























Figure 13. Oxygen consumption of excised gills of F. similis in fresh
water as a function of dry weight of gills. Slope =
0.3151 + 0.1311. Intercept = 0.7121 + 0.1325. Correlation
coefficient 0.47,
























20



0.0

c
. 10 -
E

c

uo 5r
~f re~
x
0

2-





12 5 10


Dry Gill vJfrit~ht mng


ish water


a



























Figure 14. Oxygen consumption of excised gills of F. sim7lis in 1/3
sea water as a function of dry weight of gills. Slope =
0.5817 + 0.0673. Intercept = 0.4751 + 0.0685. Correlation
coefficient = 0.87.
































1/3 sea water


200


Dry Gill Weight -mg


2001


a00



I20


8 10


1 5


* &


4o


1 2


5 10


























Figure 15.


Oxygen consumption of excised gills of F. similis in 2/3
sea water as a function of dry weight of gills. Slope =
0.4144 + 0.0764. Intercept = 0.7526 + 0.0888. Correlation
coefficient = 0.77.






























0
g


a


E


C
c


0


Dry Gill Wcight-mg



























Figure 16. Oxygen consumption of excised gills of F. similis in full
sea water as a function of dry weight of gills. Slope
0.2453 + 0.0592. Intercept = 0.9531 + 0.0685. Correlation
coefficient = 0.66.




















2001-


50



20 sva


10

sea water
5-


2


I I


5 10


I I I


I I I-


Dry Cill Wight-mg


1 2


50 uu 0a0v








I10


0 0
oo


R H
H




CO
W CO







co




0 3
H


0



O



0


C1









+4
Q






c-










+1


4
0












C)
*H


+1






0






U
o<
C-


0^ ClV (n


0 C-
n \O
C'6 \o


C\('4N t


c\




O-


+1
*
0









+1





o-l


*
0
+1



0


0o







Co
'o

0







0
o

+1



c0






O0
0
o


+1



0












TABLE 9
STATISTICAL COIMPARSONS OF SLOPES AND INTERCEPTS OF
THE DOUBLE LOGARITHI-C RELATIONSHIP BETWEEN OXYGEN CONSU1IPTION AND
DRY WEIGHT OF EXCISED GILLS OF F. similis AT FOUR SALINITIES




Comparison Slopes Intercepts


f-1/3 P>.05 P>.05

f-2/3 P>.05 P>.05

f-3/3 P>.05 P>.05

1/3-2/3 P>.05 *P<.02

1/3-3/3 *P<.ooI *P<.ooi

2/3-3/3 *P<.05 P>.05

*Considered to be statistically significant.



























Figure 17. Oxygen consumption of excised gills of F. chlysotus in
fresh water as a function of dry weight of gills.
Slope = 0.3436 + 0,1216. Intercept = 0.6022 + 0.0801.
Correlation coefficient = 0.60.
































9*

fresh wat


frGsh E wat


Dry Gill Weight-mg


201-


10


a'







0
0
"^
a:
P


12 5 2_~;__e~~__ dV_


5

























Figure 18. Oxygen consumption of excised gills of F. chysotus in 1/3
sea water as a function of dry weight of gills.
Slope = 0.1383 + 0.3402. Intercept = 0.7599 + 0.1991.
Correlation coefficient = 0.11.
























20-




o
0.



0
E



a 5-
0


c
0 %

1/3 sea water

o 2




1 10 2I
1 2 5 10 20


Dry Gill Weight-mg

























Figure 19. Oxygen consumption of excised gills of F. chrysotus in 2/3
sea water as a function of dry weight of gills.
Slope = 0.5173 + 0.1845. Intercept = 0.5533 + 0.0979.
Correlation coefficient = 0.59.
































2/3 sea wator


Dry Gill Weight mg



























Figure 20.


Oxygen consumption of excised gills of F. crSysotus in
full sea water as a function of dry weight of gills.
Slope = 0.458- + 0.1315. Intercept = 0.6302 + 0.0937.
Correlation coefficient = 0.67.











































Dry Gill Weight -mg


























H



4~-3


0
0






oa


F-I




0H"I
o




gr


+)















0
+1

o

o

H







W

+1















vl
0






0O
o





O o
S E


fk% R ((:
H C'J CC)









C11 10 0
H- R^ \0 C
C'r^ '^0


0 1o C- C
-l r- r- r-


0%G
U-N
r-
O




O
C0



O
Co








o
O


+1
CNI


\0








H
0
+1



0


o 0

+1 +1



o o
* *

0 0







1/3 sea water, 0.1383. In fresh water, it increases 148% to 0.3436.

In 2/3 sea water, the slope is 0.5173, a 2714 increase. However, in

full sea water the slope drops to 0.4584, a 231% increase over the

slope in 1/3 sea water. There are no statistical differences among

these slopes (Table 11).

The intercept of this relationship for F. chrysotus is minimal in

2/3 sea water, 0.5533. It increases 9% to 0.6022 in fresh water. In

1/3 sea water the slope is nis.dmal at 0.7599, an increase of 385 over

the slope in 2/3 sea water. The intercept for the relationship in

full soa water is 0.6302, 5% greater than the intercept for the re-

lationship in 2/3 sea water. Statistically, none of these intercepts

differs from each other (Table 11).

Coparing slopes and intercepts at each salinity for the two

species shows that the intercepts differ in sea water (P<.01). See

Table 12.

Tables 13 and 14 show the amount of oxygen consumed by excised

gills of various sizes of fish in four different salinities. Gill

weight is calculated for each size fish based on the relationships in

Table 18. Oxygen consumption is derived from the relationships in

Tables 8 and 10. For a small F. stmilis (0.5 g), oxygen consumption

of the gills increases as JB MI increases. A 1.0 g fish also shows

increases relative to 1/3 sea water when it is in a hyperosmotic

medium, but there is no rise in gill oxygen consumption when it is in

a hypoosmotic medium. Fish above 1.0 g show decreases *' in

gill oxygen consumption in fresh water. Fish up to ca. 4.0 g show

increases in gill oxygen consumption during hypoosmotic regulation.

Above this weight, decreases in gill oxygen consumption occur and












TABLE 11

STATISTICAL COMPARISONS OF SLOPES AND INTERCEPTS OF
THE DOUBLE LOGARITHMIC RELATIONSHIP BETWEEN OXYGEN CONSUMPTION AND
DRY WEIGHT OF EXCISED GILLS OF F. chrysotus AT FOUR SALINITIES




Comparison Slopes Intercepts


f-1/3 P>.05 P>.05

f-2/3 P>.05 P>.05


f-3/3 P>.05 P>.05

1/3-2/3 P>.05 P>.05


1/3-3/3 P>.05 P>.05

2/3-3/3 P>.05 P>.05


*Considered to bo statistically significant.




73







TABLE 12

INTERSPECIFIC STATISTICAL COMPARISONS OF SLOPES AIND INTERCEPTS OF
THE DOUBLE LOGARITHMIC RELATIONSHIP BETWEEN OXYGEN CONSUMPTION AND
DRY WEIGHT OF EXCISED GILLS AT FOUR SALINITIES


I edium Slopos Intercepts


f ?>.05 P>.05


1/3 P>.05 P>.05


2/3 P>.05 P>.05


3/3 P>.05 *P<,.O


*Considered to be statistically significant.















N 3






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become greater as (B MI increases. The results for F. clrysotus

show increases in gill oxygen consumption relative to 1/3 sea water

for all fish in full sea water and for fish larger than 0.5 g in 2/3

sea water. A 0.5 g fish shows a decreased gill oxygen consumption in

2/3 sea water. Fish weighing 1.0 g or less have depressed oxygen

consumption in fresh water. Fish weighing 1.5 g or 2.0 g show in-

creases in gill oxygen consumption in fresh water.

Oxygen consumption of excised gills increases less than propor-

tionately as dry gill weight increases. Slopes range from 0.2453 to

0.5817 for F. si2ailis and from 0.1383 to 0.5173 for F. chrysotus.

Studies of Weymouth et al. (1944), Bertalanffy and Perozynski (1951,

1953) and Vernberg (1954, 1956) state that, at least in some tissues,

weight specific metabolic rate decreases as body size increases.

More specifically, studios with gill tissue by Dehnel and McCanghran

(1964) and Holmes and Stott (1960) have indicated slopes of 0.33 and

0.86 respectively. These are considerably greater than the slopes

found in this study.



Comarison of Variations in Oxygen Consumption of
Intact Animals and Excised Gills


Changes in oxygen consumption of intact animals do not parallel

changes in oxygen consumption of excised gills as a function of

salinity (Table 15). In fresh water, intact F. similis shows reduc-

tions in oxygen consumption of animals weighing loss than 4.0 g.

Animals below 1.0 g show increased gill oxygen consumptions, but

animals above 1.0 g show decreased gill oxygen consumption. In 2/3

sea water, increases in the oxygen consumption of intact animals are







TABLE 15

COIPTARISON OF PERCENTAGE CHANGE IN OXYGEN CONSUiSPTION OF
INTACT ANI,-ALS AND EXCISED GILLS AS A FUNCTION OF SALINITY


Fundulus similis Fundulus chrysotus

Wt Intact Excisgd Wt Intact Excised
(g) animalgill (g) animal gill


Lresh water

-33.9%

-23.2%

-12.6%

+0.9%

+21.1%

2/3 sea water
+1.6%

+5.4-

+9.2%

+13.1%

+18.6%

.3.2 sea water
+25.0%

+33.9%

+41.7%

+50.6%

+63.8-


+19.5%

0.0%

-17.0%

-31.0%

-46.0%



+50.0%

+34.0%

+19.0%

+6.6%

-8.4%



+89.0%

+51.0%

+19.0%

-5.2e

-30.1%


0.50

0.75

1.00

1.50

2.00



0.50

0.75

1.00

1.50

2.00



0.50

0.75

1.00

1.50

2,00


fresh water

-4.2%

-3.6%

-3.3%

-2.8%

-2.2%

2/3 sea water

-4.0

-1.9%

0.0%

+2.8%

+5.0%

3/3 sea water

-30.6%

-26.0%

-22,6%

-17.5%

-12.7%


0.50

1.00

2.00

4.00

10.00



0.50

1.00

2,00

4.00

10.00



0.50

1.00

2.00

4.00

10.00


-15.7-
-8.4%

-3.1%

+5.0%

+11.2%



-11.2%

+3.3%

+14.6%

+32.9%

+47.9%



+0.5%

+14.0%

+24.5%

+41.1%

+54.1%


*Data are given as a percentage change in oxygen
consumption relative to oxygen consumption in 1/3 sea water.







present in all sizes of fish and the percentage increase becomes

larger as body size increases. Oxygen consumption of excised gills

also shows increases, but the percentage decreases with increasing

body size, until at 10.0 g there is an 8.4% reduction in gill oxygen

consumption. In 3/3 sea water, all F. similis show increased oxygen

consumption by intact animals and the percentage change relative to

1/3 sea water increases with increasing body size. Fish weighing

2.0 g or less show increased gill oxygen consumption, but 4.0 and

10.0 g fish show decreased gill oxygen consumption.

All sizes of F. chrysotus show decreased oxygen consumption of

intact animals in fresh water. Animals weighing 1.0 g or less also

show decreased gill oxygen ccnr.omption. However, animals weighing

either 1.5 or 2.0 g show increases in gill oxygen consumption. In

2/3 sea water, oxygen consumption of intact animals decreases for

animals of less than 1.0 g and increases for animals weighing more

than 1.0 g. Oxygen consumption of gills of animals of all sizes

except 0.50 g shows increases. In 3/3 sea water, oxygen consumption

of intact animals of all sizes is reduced,while gill oxygen consumption

of all animals is elevated.

There is no apparent correlation between changes in oxygen con-

sumption of intact animals and changes in oxygen consumption of

excised gills for either of the species.



Apparent Pormo ability


Apparent permeabilities which have been calculated with the

equation of Potts (1954) are given in Tables 16 and 17. In all












TAhLE 16

APPARENT PERPJEABILITY OF F. sinjiis AS A
FUNCTION OF BODY SIZE AND SALINITY



B-M = +314-. B-M = +25'

Wt Surface area Permeability change Permeability
(g) (mm2) (,/m2M-.r) % (M/mm-M-hr)

0.50 150 4.11 X 10-6 -99.0% 3.84 X 10-4

1.00 251 2.82 X 10"6 -99.15 3.17 X 10-4

2.00 490 1.93 x 10o6 -99.3% 2.61 X 10-

4.00 887 1.32 X 10-6 -99.4% 2.51 X 10-4

10.00 1,941 0.80 X 10-6 -99.5% 1.66 X 10"4

#Difference between the medium concentration and
blood plasma concentration in milliosmoles/liter.

Moles per square millimeter per molar concentration
difference per hour.












TABLE 16 (Extended)


B-M = -31f7

Perrmeability
(1/mmn -M-hr)


5.06 x 10"6


3.72 X 10-6


2.73 X 10-6


2.00 X 10-6


1.33 X 10-6


change



-98.7%


-98.8%


-99.o0%


-99.1%


-99.25


B-M = -6o30

Perme ability*
(M/mm2-IM-hr)


2.23 X 10-0


1.46 X 10-6


0.95 X 10-6


0.62 X 10-6


0.35 X 10-6


change
-


change
%




-99.5%


-99.6%


-99.7%

-99.8%


_ __I__ __ __ __ __


~CII~_ _














TABLE 17

APPARP NT PERPEABILITY OF F. chr2otus AS A
FUNCTION OF BODY SIZE AND SALINITY


B-M = +231# B-M = +66#

Wt Surface area Permeability* change Permeability*
(g) (rm2) (1,1/p2-M-hr) % (./2ra.M-br

0.50 102 4.36 X 10-6 94.5 7.36 x 10-5


0.75 164 3.11 x 10-6 94.0% 5.14 X 10-5

1.00 230 2.45 X 10-6 93.6% 3.82 X 10-5


1.50 371 1,73 x 10-6 93.1% 2.51 X 10~5

2.00 520 1.35 X 10-6 92.7% 1.85 X 10-5


TDifference between the rIndium
blood plasma concentration in


concentration and
milliosmoles/liter.


*Moles per square millimeter per molar concentration
difference per hour.















TABLE 17 (Extended)


B-M = -201#

Permebility*
(/M/ma -K-hr)


1.16 x 10-5


8.79 X 10-6


7.25 x 10-6


5.51 X 10-6


4.53 x 10-6


change



35.3%


82.9%


81.0%





75.5%


B-M = -683

Permeability*



1.17 X 10-6


8.73 X 10-7


7,05 x 10-7


5.26 X 10-7


4.25 X 10-7


change
-


change



98.5%


98.3%


98.2%


98.0%


97.7%


-- -- ------- ---


--------


---


- -- -- -- -- -- ----







salinities for both species, the smallest fish have the greatest

permeabilities, Percentage decreases relative to permeabilities in

1/3 sea water are also given in Tables 16 and 17. The percentage

decrease from 1/3 sea water increases with body size in all salinities

for F. similis. For F. chrysotus, this factor decreases with body

size in all salinities.


Anatomical Relationships


The log gill surface area-log body weight relationship for

F, similis indicates that gill surface area increases less than

proportionately as body weight increases (Figure 21 and Table 18).

The slope equals 0.8457 and the intercept is 2,4330. A similar

analysis for F. chrysotus shows that gill surface area increases

more than proportionately as body weight increases. The slope equals

1.1753 and the intercept is 2.3623 (Figure 22 and Table 18). The

slopes differ statistically from each other, as do the intercepts

with P<,01 in both cases (Table 24).

The most notable studies concerning gill surface area of fishes

are by Gray (1954), Steen and Kruysse (1964), and Hughes (1966).

Gray (1954) found surface areas ranging from 188 mm2/g for Lophopsetta

maculatus to 1773 mm2/g for Brevoortia tyrannus, Steen and Kruysse

(1964) found that a 550 g eel had 530 cm2 of gill surface area or

1,075 v 2/g. Hughes (1966) found surface areas ranging from 168 mm2/g
for a 24 g Callio ymlus lyra to 845 nm2/g for an 85 g Clupea harengas.

The surface area of F. similis ranges from i54.3 mm2/g for a 9.25 g

fish to 313.4 nm2/g for a 1.46 g fish. The surface area of F, chry-

sotus ranges from 148.6 mm2/g for a 0.36 g fish to 279.7 rm2/g for a

3.88 g fish.



























Figure 21. Surface
weight.
0.0204.


area of gills of F. similis as a function of body
Slope = 0.8457 + 0.0307. Intercept = 2.4330 +
Correlation coefficient = 0.99.












































Body vWoiht-0



























Figure 22. Surface area of gills of F. chrysotus as a function of
body weight. Slope = 1.1753 + 0.0294. Intercept
2.3623 + 0.0133. Correlation coefficient = 0.99.















































Cody Weibht -









o o" c
O0 0\ Co
o 0 0
0 C)^ CL- CO








O O O\ o
+ *








0 0 0



C N H N
o o o o
+1 0i CS Ct `0
o o 0 0
4 +1 +1 +1 +1

p0 C' \0 C 0 .

S*
&. H Co0 O 0

HH

0

.0 0 0 O

1 +1 +1 +1 +1


no o C
00 H 0







S0 0 0 N 0
HN N 01 '10
) rl C\ CD 0















c4 C

0 C -)



S * *
r 0r 0 -H1



ao 0 M
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89








TABLE 19

INTERSPECIFIC COMPARISON OF SLOPES AND
INTERCEPTS OF ANATOMICAL RELATIONSHIPS


Relationship Slopes Intercepts



Surface area-Body wt # *P<.01 *P<.01


Dry gill uv-Body t# \ P>.05 *P<.001


SDouble logarithmic relationship.


* Considered to be statistically significant.






Only Price (1931) has previously studied the gill surface area

of a fish as a function of body weight and has found a slope of 0.78

for Micropterns dolomiou. This is lower than the slopes of 0.846 for

F. similis and 1.175 for F. chrysotus.

Dry gill weight increases proportionately with body weight in

both F. similis and F. chrysotus (Figures 23 and 24). The slopes are

0.9874 for F. similis and 0.9671 for F. clhrsotus. These are not

significantly different from each other (P>.05). The intercepts of

this relationship are 0.8940 for F. similis and 0.7021 for F. chrysotus.

The intercepts are significantly different from each other (P<.001).

These anatomical relationships and statistical comparisons are

reviewed in Tables 18 and 19.




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