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The interrelationship of growth, sexual development, feed intake and plasma lipids in the black-tailed… Stewart, Sheila Frances 1963

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THE INTERRELATIONSHIP  OF GROWTH, SEXUAL DEVELOPMENT,  FEED INTAKE AND PLASMA LIPIDS IN THE BLACK-TAILED DEER (Odocoileus hemionus columbianus), AND THE WISTAR RAT  by  SHEILA FRANCES STEWART B . A . , The University of B r i t i s h Columbia, 1948  A THESIS SUBMITTED IN PARTIAL FULFILLMENT OF THE REQUIREMENTS FOR THE DEGREE OF MASTER OF SCIENCE in the Department of ZOOLOGY'  We accept this thesis as conforming to the required standard  THE UNIVERSITY OF BRITISH. COLUMBIA October, 1963  In presenting  t h i s t h e s i s i n p a r t i a l f u l f i l m e n t of  the requirements f o r an advanced degree at the U n i v e r s i t y  of  B r i t i s h Columbia, I agree that the L i b r a r y s h a l l make i t f r e e l y a v a i l a b l e f o r reference  and  study.  I f u r t h e r agree that  per-  mission f o r extensive copying of t h i s t h e s i s f o r s c h o l a r l y purposes may  be granted by  h i s representatives,,  the Head of my  written  Department  of  permission.  The U n i v e r s i t y of B r i t i s h Columbia,. Vancouver 8, Canada. Date  by  I t i s understood that copying,'or p u b l i -  c a t i o n of t h i s t h e s i s f o r f i n a n c i a l gain without my  Department or  s h a l l not  be  allowed  iii  ABSTRACT Based upon the premise that growth (weight change) i s of a seasonally c y c l i c a l nature i n the deer (Odocoileus), and that this change i s primarily in the fat compartments of the body, a study of the plasma l i p i d s is undertaken to ascertain i f they r e f l e c t the seasonal changes.  The feed intake  is measured throughout the experimental period. Preliminary studies are carried out on the Wistar rat to develop a technique and f a c i l i t a t e a quick prognosis of the assumption.  In this  study, i t i s observed that because of the different sex effects plasma cholesterol levels must be determined. The prepubertal growth i n the male and female rat is characterized by a phasic type of growth upon which is imposed another pattern of weight accretion also of a phasic, but more frequent nature. The larger pattern of growth is referred to as the Main phasic period of growth, and the more frequent one as the Interim phasic period of growth. The weight, feed and l i p i d relationships of the female Main phases are similar to those of the male Interim phases, and both are believed to r e s u l t from the energy balance required for weight accretion. The same r e l a t i o n s h i p s , i n the female Interim phases are similar to those of the male Main phases, and to these are attributed the occurrence of sexual development. Similar observations on the sexually mature animals i n d i c a t e , i n the female, that the Interim phasic type of growth is synchronous with the estrus c y c l e .  Whereas, i n the male no definite conclusions are made.  In the deer study s i x , year o l d , animals of the species Odocoileus hemionus columbianus (Vancouver Island genotype)  are observed for a  iv period of twelve months.  In both sexes weight increases  from approximately  March to October when a decrease i n feed intake and weight gain occurs. In the e a r l i e r period of weight gain (March - August) both sexes increase their feed intake, but the plasma l i p i d s remain the same, that is a high plasma ' f a t ' and a low plasma cholesterol. In the l a t t e r part of the weight gain period, August to October, both sexes decrease their feed intake accompanied by a r i s e i n plasma ' f a t ' , but no change occurs i n the level of plasma cholesterol. At the point of maximum gain, i n October, a difference between the sexes occurs. 'fats'  fall.  "fats' r i s e .  The males decrease their feed intake further and the plasma Whereas the females increase their feed intake and the plasma Both sexes show a marked increase in the plasma cholesterol  level. After October, the rut period ensues and is characterized by a loss of weight and a low feed intake i n both sexes.  The l a t t e r regardless of an  adequate supply. In the males, the period of weight loss extends to December, and an expected r i s e i n the plasma ' f a t '  is seen i n only one of the three animals,  the others being missed because bleeding i s not carried out between October and December.  In the l a t t e r month the plasma cholesterol  is  s i g n i f i c a n t l y lower than i n October. In the females,  the period of weight loss extends to March, and is  never as extensive as i n the males.  The plasma ' f a t '  levels r i s e i n Dec-  ember and f a l l i n March when the nadir of the growth curve is reached.  In  this period the plasma cholesterol levels are varied and appear to depend upon the estrus phase of the animal. By March, the feed intake is increased i n both sexes, but i t is not  V  u n t i l afterwards that weight accretion begins.  The plasma l i p i d levels  are low i n a l l the animals. It i s concluded that weight and plasma l i p i d changes are  closely  a l l i e d to the yearly recrudence and regression of gonadal a c t i v i t y . The plasma ' f a t '  levels r i s e to augment the c a l o r i c requiremehtscof  the animal i f the feed intake is not adequate, both when weight is being gained and l o s t .  Whereas the plasma cholesterol appears to r e f l e c t  state of sexual a c t i v i t y . otherwise i t is  low.  That is the l e v e l is raised i n the ' r u t  1  the period,  xx i  ACKNOWLEDGMENT The writer wishes to express her sincere thanks and appreciation to Dr. I . McTaggert Cowan, Professor and Head of the Department of Zoology, for making this work possible and for his assistance, and i n s p i r a t i o n through her study of Zoology. To Dr. A. J . Wood, Professor of Animal Husbandry, Department of A g r i c u l t u r e , for his continual encouragement throughout the Thanks are also due him for providing the f a c i l i t i e s  experiment.  i n which to work  and the small animals necessary for the preliminary t e s t s . To Dr. H. C. Nordan, Research Associate i n the Department of Zoology, for  suggesting the subject of this t h e s i s , and who has i n addition provided  constant advice and guidance, without which the thesis could not have been completed. To the men, i n the Central Animal Depot, for their assistance i n supplying animals, feed and carrying the heavy loads. To those students, both of the Department of Agriculture and Zoology, for help i n weighing and bleeding the deer.  I should not have succeeded  without their help. Many thanks, too, are due to Mrs. A l i c e Schonfeld, secretary to the Central Animal Depot, for her u n f a i l i n g help whenever called upon.  vii  TABLE OF CONTENTS y.xx  ACKNOWLEDGMENT INTRODUCTION  1  PART A.  7  THE RAT G e n e r a l Comments on F a t D i g e s t i o n A b s o r p t i o n and Utilization  8  MATERIALS I II III  14 Animals  15  Diet  15  Housing  . .  IV  Weighing Procedure  V  B l e e d i n g Procedure  15  V a g i n a l Smears  16  VI  . . . . . . . . . . . . . . . . . .  15  METHODS  15  17 D e t e r m i n a t i o n o f T o t a l Plasma L i p i d s  . . . . . . . . .  D e t e r m i n a t i o n o f T o t a l Plasma C h o l e s t e r o l . . . . . . . . . . TREATMENT OF RESULTS  17 20 23  THE MALE PREPUBERTAL RAT  .  28  RESULTS AND DISCUSSION  . .  28  Body Weight  29  Body Weight and Plasma 'Fat'  29  Body Weight, Plasma ' F a t ' and Feed Intake  . . . . . . .  39  Body Weight and T o t a l Plasma C h o l e s t e r o l and T h e i r Relationship  to the Other Parameters  Terminal  ............  P o i n t o f I n t e r i m Phase  42 42  Onset P o i n t o f I n t e r i m Phase . . . . . . . . . . .  42  P o i n t s o f J u n c t i o n o f the Main Phase  42  Plasma 'Fat' to Plasma C h o l e s t e r o l R a t i o  (F/C)  P o i n t s o f J u n c t i o n o f the Main Phase Terminal  P o i n t o f the I n t e r i m Phase  Onset P o i n t o f the I n t e r i m Phase  ....... . . . .  43  ......  43  . . . . . .  44  . .  44  viii Intermediate Points Within the Interim Phase . .  44  Feed Intake and the Total Plasma Cholesterol Levels  46  THE FEMALE PREPUBERTAL RAT  55  I  56  II  BODY WEIGHT A.  A r i t h a r i t h Representation of the Growth Curve . . . .  56  B.  A r i t h l o g Representation of the Growth Curve . . . .  64  RELATIONSHIP OF BODY WEIGHT AND PLASMA 'FAT A.  B.  Interim Phasic Period of Growth. . . . . . . . . . . . .  III  B.  66  2.  Terminal Points of Junction  66  Main Phasic Period of Growth  67  Points of Junction  67  Points of Junction i n A r i t h l o g Growth Curve at 51 Days of Age  67  . . . . . . .  67  Interim Phasic Period of Growth . . . . . . . . . . . . . . .  68  1.  Onset Point of Junction  2.  Terminal Points of Junction  . . . . . . . . . . . . .  68 69  Main Phasic Period of Growth 1.  C.  66  Onset Point of Junction  RELATIONSHIP OF BODY WEIGHT AND FEED INTAKE A.  65  1.  1. C.  . .. .  1  .  Points of Junction  70 70  Points of Junction i n A r i t h l o g Growth Curve at 51 Days of Age . . . . . . . . . . . . . . . . . . . .  71  IV  RELATIONSHIP OF BODY WEIGHT AND PLASMA CHOLESTEROL . . . .  72  V  RELATIONSHIP OF BODY WEIGHT AND PLASMA CHOLESTEROL TO OTHER PARAMETERS WITHIN THE SEPARATE PHASIC PERIODS OF GROWTH . .  72  A.  B.  Interim Phasic Period of Growth  72  1.  Terminal Points of Junction  72  2.  Onset Points of Junction . . . . . . . . . . . . . .  73  Main Phasic Period of Growth 1.  C.  . .  74  Points of Junction  74  Intermediate Points i n the Interim Phasic Period of Growth 1.  One Day After Onset Points  .  74 74  ix  VI  2.  Two Days After Onset Points  . . . . . . . . . .  3.  One Day Prior to Terminal Point  . . . . . . . .  75  4.  Two Days Prior to Terminal Point . . . . . . . .  75  THE EFFECTS OF HEAT ON THE DIFFERENT PARAMETERS  VII VIII  75  . . . . .  75  THE EFFECTS OF INANITION ON THE DIFFERENT PARAMETERS  . .  76  PLASMA 'FAT' TO PLASMA CHOLESTEROL RATIO; F/C RATIO  . .  77  A.  Main Phasic Period of Growth 1.  B.  . . . . . . . . . . . .  78  Points of Junction  78  Interim Phasic Period of Growth  . .  78  THE SEXUALLY MATURE MALE RAT  89  I  90  II  III  BODY WEIGHT . . . RELATIONSHIP OF BODY WEIGHT AND FEED INTAKE A.  Weight Gain i n Groups 1 and 2 . . . . . . . . . . . . .  B.  Weight Loss in Groups 3 and 4  97 97 1  Weight Gain i n Group 1 1.  High Feed Intake  2.  Low Feed Intake  97 98  . . .  98 98  B.  Weight Gain i n Group 2 . ..  C.  Weight Loss i n Group 3  99  D.  Weight Loss i n Group 4  100  ..  98  RELATIONSHIP OF BODY WEIGHT, FEED INTAKE, PLASMA 'FAT' AND TOTAL PLASMA CHOLESTEROL A.  V  .  RELATIONSHIP OF BODY WEIGHT, FEED INTAKE AND PLASMA 'FAT A.  IV  97  . .  Weight Gain i n Group 1  100 100  1.  High Feed Intake  100  2.  Low Feed Intake  101  B.  Weight Gain i n Group 2  101  C.  Weight Loss i n Group 3  101  D.  Weight Loss in Group 4  102  1.  Low Plasma Cholesterol  2.  High Plasma Cholesterol  TOTAL PLASMA CHOLESTEROL AND FEED INTAKE  102 . . . . . . . . . . . . .  103 103  X  VI  THE CHRONOLOGICAL OCCURRENCE OF EACH GROUP  103  THE SEXUALLY MATURE FEMALE RAT .  108  FLUCTUATIONS OF WEIGHT, PLASMA LIPIDS AND FEED INTAKE RELATIVE TO THE SEPARATE PHASES OF THE ESTRUS CYCLE  118  I  118  EARLY METESTRUS 1 PHASE OF THE ESTRUS CYCLE A.  Body Weight Changes  B.  Body Weight and Feed Intake Relationships  C.  Body Weight, Feed Intake and Plasma Fat Interrelationships . . . . . .  123  Total Plasma Cholesterol i n Relationship to Other Parameters . . . . . . . . . . . . . . . . . . . . .  127  D. II  . . . .  120  A.  Body Weight Changes  B.  Body Weight and Feed Intake Relationships  C.  Body Weight, Feed Intake and Plasma 'Fat' Interrelationships  124  Total Plasma Cholesterol i n Relationship to Other Parameters . . . . .  127  119 . . . .  121  METESTRUS 2 PHASE OF THE ESTRUS CYCLE A.  Body Weight Changes  B.  Body Weight and Feed Intake Relationships  C.  Body Weight, Feed Intake and Plasma 'Fat' Interrelationships  125  Total Plasma Cholesterol i n Relationship to Other Parameters  128  D.  IV  118  METESTRUS 1 PHASE OF THE ESTRUS CYCLE  D. III  . . . . . . . . .  . . . . .  119  . . . .  121  DIESTRUS PHASE OF THE ESTRUS CYCLE A.  B.  C.  Body Weight Changes  119  1.  F i r s t Day of Diestrus  119  2.  Second Day of Diestrus  120  Body Weight and Feed Intake Relationships  . . . .  121  1.  F i r s t Day of Diestrus  121  2.  Second Day of Diestrus  122  Body Weight, Feed Intake and Plasma 'Fat' Interrelationships . . . . . . . . . . . . 1.  F i r s t Day of Diestrus  2.  Second Day of Diestrus  125 125  . . . . . . . . . . .  126  xi D.  Total Plasma Cholesterol i n Relationship to Other Parameters 1.  128  F i r s t Day of Diestrus  128  2. . Second Day of Diestrus V  VII  129  PROESTRUS PHASE OF THE ESTRUS CYCLE A.  Body Weight Changes  B.  Body Weight and Feed Intake Relationships  C.  Body Weight, Feed Intake and Plasma 'Fat' Interrelationships  127  Total Plasma Cholesterol i n Relationship to Other Parameters . . . .. . . . . . . . . .  129  D.  VI  .  120 . . . .  122  ESTRUS PHASE OF THE ESTRUS CYCLE B.  Body Weight and Feed Intake Relationships  . . . .  D.  Total Plasma Cholesterol i n Relationship to Other Parameters  130  PLASMA 'FAT' TO PLASMA CHOLESTEROL RATIO; F/C RATIO . . .  PART B.  THE DEER  122  130  '.  136  INTRODUCTION  137  MATERIALS  138  I II III IV  Animals  138  Diet  138  Housing  138  Weighing Procedure  138  METHODS  .  I II  I  Determination of Total Plasma Lipids  139 .  139  Determination of Total Plasma Cholesterol . . . . .  139  THE MALE DEER  .  INTRODUCTION  141 142  Antler Growth  142  Rutting Period  142  RESULTS  . . . . . . . .  Body Weight  146 146  xii Feed Intake  146  Plasma 'Fat'  147  Total Plasma Cholesterol DISCUSSION  .  .  148 149  Seasonal Fluctuations of Weight, Feed Intake, and Plasma Lipids Relative to Sexual A c t i v i t y  . . . . . .  149  A.  Spring  149  B.  Summer.  150  C.  Autumn  150  D.  Winter  151  E.  Spring  152  SUMMARY  153  II  156  THE FEMALE DEER  INTRODUCTION  157  RESULTS  161  Body Weight  161  Feed Intake  161  Plasma 'Fat'  . . . . . . . .  162  Total Plasma Cholesterol  162  DISCUSSION . . . . . . . . . . . . . . . . . . . . . . . . . . . . . .  . .  163  Seasonal Fluctuations of Weight, Feed Intake, and Plasma Lipids Relative to Sexual A c t i v i t y . .. . .. . . .  163  A. . Spring B.  . .  Summer  163  C. . Autumn  164  D.  Winter  165  E.  Spring  166  SUMMARY  APPENDIX  BIBLIOGRAPHY  163  •. .  . .-  168  171  184  xiv  LIST OF FIGURES Figure PART A . i 1-5  THE RAT  Growth Curve for a Representative Male of Odocoileus hemionus columbianus A r i t h a r i t h Representation Prepubertal Rats. Feed Intake GMS/Animal; Plasma Cholesterol  of Growth Curve for Male  GMS/100GMS of Body Weight.  (Total) MGM%  Plasma ' F a t '  6-7 8-12  MGM7o  F/C Ratio  30  A r i t h l o g Representation of Growth Curve for Male Prepubertal Rats . . . . . . . . . . . . . . . . . . .  35  A r i t h a r i t h Representation Prepubertal Rats. Feed Intake, GMS/Animal; Plasma Cholesterol  of Growth Curve for Female GMS/100GMS of Body Weight.  (Total) MGM%  Plasma ' F a t '  13-14  3  MGM7<,  F/C Ratio .  57  A r i t h l o g Representation of Growth Curve for Female Prepubertal Rats . . . . . . . . . . . . .  62  SEXUALLY MATURE MALE RATS 15-17  18  19-20  Body Weight, Feed Intake and Plasma Lipids for Eight Consecutive Days  91  Sexually Mature Male Rats: Occurrence of the 'Groups'  The Chronological . . . . . . . . . . . . . . .  94  Sexually Mature Male Rats:  Control Animals . . . . . .  95  SEXUALLY MATURE FEMALE RATS 21-25  26-27  Body Weight, Feed Intake and Plasma Lipids for Ten Consecutive Days  110  Sexually Mature Female Rats:  115  Control Animals . . . .  XV Figure 28  1  Sexually Mature Female Rats: Weight Accretion, Feed Intake, Plasma Lipoids During a Complete Estrus Cycle Prepubertal Male Rat, Triphasic Relationship of Feed Intake and Plasma Lipids . . . . . . .  PART B. 1-3 4-6  117  .  Appendix  .  143  THE DEER  Male Deer: Weight Accretion, Plasma L i p i d s , Feed Intake for Eleven Months . . . . .. . . . . . . . . . . . . Female Deer: Weight Accretion, Plasma L i p i d s , Feed Intake for Eleven Months  . 158  xvii  LIST OF TABLES Table PART A . I II-VI  THE RAT  Composition of Diet Levels of Weight Accretion, Plasma Lipids and Feed Intake, Prepubertal Male Rats: Rat No. 1  47  Rat No. 2 . . . .  48  Rat No. S •  VII  VIII  IX  X-XIV  22  .  49  Rat No. 5  50  Rat No. 6  51  Weight Changes: Plasma Lipids and Feed Intake Levels of Male (Prepubertal) Rats. At Junction of Two Main Phasic Periods of Growth . . . . . . . . . . . . . . . . Weight Changes: Plasma Lipids and Feed Intake Levels of Male (Prepubertal) Rats. A.  At the Terminal Points of the Interim Phase of Growth  B.  At the Onset Points of the Interim Phase of Growth  53  Weight Changes: Plasma Lipids and Feed Intake Levels of Male (Prepubertal) Rats. Intermediate Points i n Interim Phase of Growth  54  Levels of Weight Accretion, Plasma Lipids and Feed Intake, Prepubertal Female Rats: Rat No. 1  .  Rat No. 2  . .  80  .  81  Rat No. 3 Rat No. 6  82 ....  Rat No. 7 XV  52  Weight Changes :^ Plasma Lipids and Feed Intake Levels of the Female (Prepubertal) Rats. At Junction of two Main Phasic Periods of Growth . . . . . . . . . . . . .  83 84  85  xviii Table XVI  XVII  Weight Changes: Plasma Lipids and Feed Intake Levels , of the Female (Prepubertal) Rats.  XX  XXI  XXII  XXIII  XXIV  At the Terminal Points of the Interim Phase of Growth  B.  At the Onset Points of the Interim Phase of Growth . . . .  86  Weight Changes: Plasma Lipids and Feed Intake Levels of the Female (Prepubertal) Rats. Intermediate Points i n the Interim Phase of Growth.  XVIII  XIX  A.  A.  One Day Post Onset Point  B.  One Day Prior to Terminal Point  C.  Two Days Prior to Terminal Point . . . . . .  D.  Animals Subjected to Heat ( 8 2 ° F ) for Three Successive Days  E.  Animals Subjected to Inanition for Twentyfour Hours  A.  Sexually Mature Male Rats. Showing Levels of Body Weight, Plasma Lipids and Feed Intake on Eight Consecutive Days.  B.  Changes Occurring i n Body Weight, Lipids and Feed Intake Levels for Eight Consecutive Days . . . .  87  88  105  Sexually Mature Male Rats. Showing Weight Gained or Lost and the Accompanying Plasma L i p i d and Feed Intake Changes According to "Group" Arrangement . . . . . .  106  Sexually Mature Male Rats: Control Animals. Showing Actual and Compared Values, for Weight and Feed Intake During the Eight Consecutive Days the Test Animals were Observed. . . . . . . . . . . . . . . . . . . . . . .  107  Sexually Mature Female Rats. Showing Body Weight Plasma L i p i d s , Feed Intake, F/C Ratio and Phases of Estrus Cycle for Ten Consecutive Days  132  Sexually Mature Female Rats. Test Animals. Actual Values and Changes Occurring i n Plasma L i p i d s , Body Weight and Feed Intake Arranged According to the Phase of the Estrus Cycle . . . . . . .  133  Sexually Mature Female Rats. Control Animals. Showing Body Weight and Feed Intake Levels for Ten Consecutive Days . . . . . . .  134  xix Table XXV  Sexually Mature Female Rats. Control Animals. Showing Weight and Feed Intake Levels Arranged According to the Phase of the Estrus Cycle  PART B.  135  THE DEER  I  Composition of Diet  140  II  Male Deer. A. Weight Accretion, Plasma L i p i d s , and Feed Intake at Two Monthly Intervals for One Year. B. Changes Occurring i n the Parameters at Two Monthly Intervals  154  Male Deer. (mgm7o)  155  III IV  V  Plasma Lipids as Milligrams Percent  Female.„Deer.A. Weight Accretion, Plasma L i p i d s , and Feed Intake at Two Monthly Intervals for One Year. B. Changes Occurring i n the Parameters at Two Monthly Intervals ... . . . . . . . . . . . . .  169  Female Deer. (mgm7c)  170  Plasma Lipids as Milligrams Percent  2. INTRODUCTION Growth of a 'Seasonally C y c l i c a l nature  1  occurs i n adults and young of  both the male and female deer (Odocoileus hemionus) (115).  That which occurs  in the prepubertal animal (0. hemionus columbianus) was described by Cowen et a l (21) as a phasic type of growth.  In this period three separate phases were  observed, each successive one showing a diminished rate of weight accretion. This same phasic pattern of growth was described by Brody (13) i n the albino r a t ; Wood and Groves (116)  i n the preweanling p i g , and Krebs (62)  in  reindeer fawns. The t h i r d phase of the prepubertal growth curve terminates at the same time as the male deer shows signs of antler bud formation (21) at approximately 100 days.  Cowen et a l interpreted the occurrence of antler buds as the i n i t i a l signs  of puberty.  However, Meschake et a l (80) noted i n the reindeer (Rangifer  tarandus) that antler growth occurs long before sexual maturity. . Similarly Wislocki (112)  investigating the white-tailed deer (Odocoileus virginianus  borealis) presumed that the p i t u i t a r y gland stimulates antler growth a number of months before gonadal a c t i v i t y , thereafter,  the antlers become dependant upon the  gonads (112) . Wood et a l (114)  demonstrated that during post pubertal l i f e the adult male  deer (0. hemionus) continues to grow steadily with decreasing r a t e .  The most  obvious feature of weight change, however, is an annual cycle of gain and loss that is r e p e t i t i v e through at least six years probably throughout l i f e (Fig i ) . The precise timing of the phasic response varies s l i g h t l y between different individuals of the same race and more between different r a c i a l  stocks.  Meschake and Wislocki demonstrated i n reindeer and V i r g i n i a deer respect i v e l y that soon after the velvet is rubbed from the antlers the male becomes aggressive i n i t s behavior.  Meschake demonstrated also that at this time there  is a recrudence of t e s t i c u l a r development and an increased urinary Zimmerman  BODY  WEIGHT  IN  POUNDS  4. chromagen reaction; 30 - 45 mg/liter from the spring l e v e l of 12 - 15 mg/liter. After the postpubertal animal reaches the point of maximum weight accretion, at or about the time that f u l l sexual a c t i v i t y i s regained, winter loss of weight occurs (25).  a-yearly autumnal and  This loss i s accompanied by a 'compulsary'  reduction of feed intake which continues u n t i l spring. Inspection of carcasses of deer from the UBC  colony  (male and female)  k i l l e d at the period of maximum weight gain reveal marked depots of subcutaneous, perirenal and mesenteric f a t .  Animals k i l l e d i n the early spring show a marked  decrease i n the fat compartments. It i s presumed, therefore, that the c y c l i c a l changes i n weight of the postpubertal deer (Odocoileus  hemionus) i s due p r i n c i p a l l y to attendant changes i n  the deposition and mobilization of body f a t .  It i s further presumed that the  fat i s mobilized to maintain the c a l o r i c requirements of the animal when there i s a diminished feed intake (30, 41, 49) during the autumn - winter period. Because of the changes that occur i n the fat compartments, i t seems reasonable to assume that the plasma l i p i d levels r e f l e c t these changes. present  The  study i s therefore planned to explore the nature and timing of events  in the annual weight cycle of the deer as they r e f l e c t t o t a l plasma ' f a t ' and t o t a l plasma c h o l e s t e r o l . A number of workers (18, 37, 39, 86) reported some sex differences i n the plasma cholesterol levels associated with the sexual cycle and believe i t to be related to gonadal hormone action.  This leads to the decision to explore  both the cholesterol and non-cholesterol  f r a c t i o n i n both sexes.  The deer i s a p a r t i c u l a r l y suitable animal i n which to study these phenomena because of the very pronounced r u t t i n g period.  Also because of the  well documented d e t a i l s of change i n weight and i n food intake that are available as a result of the work done at the University of B r i t i s h Columbia laboratory.  5. Males of the genus Odocoileus at t h i s l a t i t u d e s t r i p the velvet from t h e i r antlers during the period from mid August to late September.  The actual time  d i f f e r s s i g n i f i c a n t l y between c e r t a i n r a c i a l stocks and to a degree i n d i v i d u a l l y . Antlers continue to increase i n weight for perhaps a month a f t e r the velvet i s removed, but they are e s s e n t i a l l y mature at that time.  Rutting behavior begins  shortly a f t e r and i s accompanied by enlargement of the muscle tissues of the neck, decrease i n food intake, aggressive behavior, in the presence of the doe.  and overt sexual  behavior  The rut i n the male p e r s i s t s i n our colony from  September or October through u n t i l the loss of the antlers during the period between February and A p r i l . The sexual cycle of the female, at this l a t i t u d e , involves a prolonged anestrus extending usually from January or February, or l a t e r , u n t i l October or November.  There then follows two,  three or more estrus periods, each one  apparently b r i e f i n duration and separated by a period of 26 - 28 days.  The  precise d e t a i l s of the timing of the events i n the female cycle has not been determined. The female shows few overt signs of sexual a c t i v i t y and vaginal smears are not attempted. To f a c i l i t a t e the interpretation of events as encountered i n Odocoileus i t i s decided f i r s t to explore the changes i n plasma ' f a t ' and plasma cholesterol levels i n the laboratory r a t .  This animal o f f e r s many advantages i n that i t i s  e a s i l y handled, the stages i n the reproductive cycle are well known and can be e a s i l y i d e n t i f i e d , and estrus periods are frequent.  Of p a r t i c u l a r importance  also i s the fact that the rat (Wistar) becomes sexually mature within four to six weeks after b i r t h (12, 13, 53, 65).  Hence the period of prepubertal growth  i s greatly accelerated i n comparison to that of Odocoileus which i s approximately 120 days (21).  6. The polyestrus nature of the female rat occasions an estrus period at four to five day i n t e r v a l s .  This gives ample opportunities to determine i f  plasma l i p i d changes occur at estrus, and/or at the other phases.  Whereas,  the doe, a seasonally polyestrus animal, ovulates one to five times over a period of weeks and the rest of the year i s sexually quiescent  (16).  The hormonal control of the sex cycle i s so well documented (33, that interpretation is f a c i l i t a t e d .  73)  The rat has also been used extensively  for the study of the effects of gonadal hormones on the plasma cholesterol levels (18,  37, 38', 39, 71), and growth hormone effects on plasma non-esterified  fatty acids (Nefa)(10, To summarize; 1.  31, 36, 47, 48, 56, 57, 68, 89, 90,  110).  the use of the rat i n this study is primarily threefold:  The rat shows similar patterns of growth and weight changes, as seen in the deer, but the 'events' are greatly telescoped in time.  2.  The a v a i l a b i l i t y of the animal, and the ease of handling f a c i l i t a t e the development of the methods.  3.  Evidence of the relationship of weight change and plasma l i p i d levels can be ascertained long before i t can i n deer.  Because the rat as well as the deer is used, this study is i n two parts: Part A pertains to the results obtained on the r a t ; on the deer.  Part B to those obtained  PART A  THE RAT  8. PART A  THE  RAT  GENERAL COMMENTS ON FAT DIGESTION, ABSORPTION AND  UTILIZATION  The plasma l i p i d s have their o r i g i n from two sources, one exogenous, i s i n the food, and  the other endogenous, i e i n the f a t depots of the body.  Usually only the endogenous fats are used for energy and  the factors that  influence their mobilization from the f a t depots are many and Digestion and Absorption of Fat from the Intestine after  varied.  Ingestion.  The fats ingested by the omnivore are usually t r i g l y c e r i d e s , l e c i t h i n s , phospholipids,  cholesterol, cholesterol esters and free f a t t y acids.  The f a t s ,  other than cholesterol and i t s esters are rendered capable of absorption by hydrolysing  action of the digestive enzyme l i p a s e , the presence of b i l e  the subsequent formation of polarised p a r t i c l e s .  the  and  The absorbed fats are  usually t r i g l y c e r i d e s , p a r t i a l l y hydrolysed mono-and diglycerides (40,  95),  and free f a t t y acids s p l i t from the glycerol moiety of the above mentioned mono- and diglycerides.  The  form of sodium taurocholate  cholesterol f r a c t i o n requires b i l e s a l t s in the and a free fatty acid, usually an oleate, both of  which must be present at the same time and  in the proper molar concentrations  for the optimal transfer of cholesterol through the i n t e s t i n a l mucosa as an ester. (105)  I t has been shown by experiment that the cholesterol ester i s  more readily absorbed by the mucosa than the free cholesterol (99).  However,  free cholesterol does appear to pass through the i n t e s t i n a l mucosa. The  f a t p a r t i c l e s absorbed are believed to pass through the minute canals  in the cuticular surface of the v i l l i of the small i n t e s t i n e , i e between the m i c r o v i l l i , and (141).  The  then into the subadjacent cytoplasm of the columnar epitheliums  f a t p a r t i c l e s coalesce  into larger droplets, which move through the  c e l l toward the Golgi apparatus; then toward the nucleus and f i n a l l y out of the  9. c e l l into the space between adjacent c e l l s and thence into the lamina propria, the lacteals and subsequently the lymphatic c a p i l l a r i e s .  The fats are s t i l l i n  the same form as when absorbed but are combined with proteins to produce what i s c a l l e d chyle, the p a r t i c l e s of which are c a l l e d chylomicrons,  (27).  The chylomicrons enter the Vascular System by way of the thoracic duct in the region of the innominale vein and make t h e i r way to the l i v e r v i a the hepatic vein.  This pathway is the most commonly used by the t r i g l y c e r i d e s and phospholi-  pids, and the only pathway used by c h o l e s t e r o l .  The former can also reach the  l i v e r by way of the hepatic p o r t a l vein, which originates in the c a p i l l a r y network i n each v i l l u s .  The phospholipids, especially l e c i t h i n , are thought to a i d  in maintaining the f i n e l y emulsified state of the chyle. appear i n the chyle after absorption of f a t ,  These phospholipids w i l l  regardless of t h e i r having been i n -  gested, and so i t is believed that they are synthesised in the mucosa of the small intestine  (27).  The Endogenous Source of Plasma L i p i d s .  Fat Synthesis in the L i v e r .  The chylomicrons pass into the l i v e r c e l l s either intact or hydrolysed, i t is believed i n the former condition, as no hydrolysis of neutral fat has been demonstrated e x t r a c e l l u l a r l y under normal conditions (64).  Inside the l i v e r  c e l l l i p o l y s i s occurs, and the free fatty acid moiety and some of the exogenous glycerol is used i n the synthesis of phospholipids (103).  .The remainder of the  glycerol combines with -.'.endogenous fatty acids (from fat depots) to form neutral fats.  The mitochondria of the l i v e r c e l l are believed associated with this pro-  cess (64).  The t r i g l y c e r i d e s formed are stored i n the l i v e r c e l l s as neutral  fats or as lipoproteins u n t i l such time as they t r a v e l v i a the c i r c u l a t o r y system to other tissues to be stored i n fat globules (64,  103).  Some w i l l be  u t i l i z e d immediately or stored in adipose t i s s u e . It is interesting to note that after ingestion of fat the vascular levels are raised, f i r s t by the exogenous f r a c t i o n , secondly by endogenous non-esteri-  10. f i e d fatty acids, mobilized,  and making t h e i r way to the l i v e r to combine with  the exogenous glycerol moiety, and t h i r d l y by the endogenous neutral fats, newly synthesised  by the l i v e r .  In the rat i t i s some six hours post  ingestion  before the l i p i d serum levels return to normal (27). Exogenous cholesterol w i l l , a f t e r leaving the thoracic duct,, enter the c i r c u l a t i o n , and be stored i n the l i v e r "pool", which i n turn supplies and maintains the blood l e v e l , i t s main channel of excretion i s from the l i v e r as b i l e acids.  Endogenous cholesterol i s synthesised  by the l i v e r , and helps to maintain  the l e v e l of the cholesterol "pool" of the l i v e r (9) when the exogenous supply i s low. Fat ingestion has been demonstrated by many investigators (27) to cause an increased plasma cholesterol l e v e l , although others have not been able to confirm t h i s finding possibly due to the time that elapsed between ingestion and measurement,, or to the nature of the fat ingested.  Cholesterol i s found i n most  tissues of the body, but so f a r i t s synthesis can only be attributed to the l i v e r , and  i n some lesions of individuals affected by atherosclerosis. Cholesterol i s found i n fat depots, but only i n small quantities, as the  fat of these tissues i s p r i n c i p a l l y of the neutral type. commonly cerebrosides,  Organ fats are more  cholesterol and phospholipids (27).  Fat Storage. The fats entering the fat depots do so by means of active transport (97). As no hydrolysis has been demonstrated outside the c e l l i t i s believed that the neutral fats and other l i p i d s enter the adipose tissue intact (103, 64). However, when endogenous f a t t y acids are required l i p o l y s i s occurs within the fat c e l l (92, 93, 96) and the glycerol moiety i s freed for intermediary carbohydrate metabolism, and the free f a t t y acid (lipoprotein) goes by way of the c i r c u l a t i o n to the l i v e r where i t i s used i n t r i g l y c e r i d e synthesis . Neutral fats have been known to be synthesised  (102).  i n the adipose tissue,  11. subsequent to the deposition of glycogen (109). The composition of the neutral fats deposited in adipose tissue c h a r a c t e r i s t i c of the species, as well as the s i t e of deposition,  is  ie unsaturated  fats are found p e r i p h e r a l l y , and the more saturated are deposited deeper i n the body. The actual mechanisms involved in fat deposition and mobilization are not c l e a r l y understood.  Several have been suggested  (77).  Causes for Deposition of Body F a t . Although the mechanisms of fat deposition are not known i t is common knowledge that fat depot formation is due to a positive energy balance (77, This positive energy balance can be brought about by overeating, diet,  insufficient  Najjar (82)  exercise or a combination of any of these.  82).  an unbalanced  According to  overeating-can be stimulated by psychological, metabolic or physio-  l o g i c a l malfunction either of a genetic or of an environmental o r i g i n . Factors that effect the Mobilization and Deposition of Endogenous Fats and thus change the plasma levels are many and v a r i e d .  One of the prime movers of fat  from the depots is the imposition of a state of fasting (28,  31).  It is known  that the energy derived from t h i s fat is capable of maintaining the metabolic needs of the animal (46, ion of carbohydrates  50), whereas a low plasma fat ensues after the ingest-  (74).  It is also well known that the growth hormone (STH) w i l l mobilize fat as non-esterified fatty acids (Nefa)(10, 31, 56, 68, 90,  110) for the purpose of  providing endogenous energy for the protein anabolic a c t i v i t y attributed to i t (47, 48, 57, 89, 90),  and i n doing so w i l l raise the p l a s m a . l i p i d levels  as much as four or five times higher than normal (90).  (Nefa)  It has also been noted  in some instances when growth hormone injections are prolonged that the plasma cholesterol levels are increased (44,  78,  110).  Control of fat deposition and mobilization has been demonstrated under the  12. influence of adrenocortical and medullary hormones, i n s u l i n , and thyroid hormone. The adrenocortical hormones, l i k e the growth hormone w i l l i n h i b i t fat synthesis and at the same time promote lipogenesis  (66).  The adrenalectomized  animal increases fat synthesis but is unable to mobilize the deposited f a t .  (43)  Clements and Schaeffer (19) were able to demonstrate that epinephrine had some effect on fat mobilization, and more recently Gordon and Cherkes (46) showed by i n v i t r o experiments that epinephrine is responsible for the release of Nefa from adipose t i s s u e .  Insulin, on the other hand, decreases the outflow  of Nefa from adipose tissue (11),  and in the diabetic r a t , increases the uptake  of fat by the tissue (104). The thyroid hormones, thyroxine and t r i i o d o t y r o s i n e , were reported by Aldersberg (1),  Deuel (27) and Rosenman (94)  to lower t o t a l serum l i p i d  levels,  but with new methods of identifying fats i t has recently (91) been shown that there is an increase i n Nefa when thyroid a c t i v i t y is increased. For many years i t has been known that the plasma cholesterol levels are effected by the degree of thyroid a c t i v i t y .  In the hyperthyroid state there  is  increased destruction and excretion of cholesterol by the l i v e r which lowers the plasma levels (84),  and the opposite condition prevails in the hypothyroid  state. It is evident that any hormonal a c t i v i t y that causes increased metabolism and thus raises the c a l o r i c requirements of the animal w i l l increase the  lipid  f r a c t i o n of the blood, primarily by increasing the non-esterified f a t t y acids '(•Nefa),which are r e a d i l y oxidized by the tissues. The non-esterified fatty acid response i n both male and female i s quite similar, hence the plasma levels d i f f e r very l i t t l e between sexes, but may vary from species to species, because of the type of fat ingested, animal and possibly the genotype.  the age of the  13. Plasma c h o l e s t e r o l  l e v e l s , on the o t h e r hand, w i l l  to the sex of the animal (37, 9 9 ) .  show v a r i a t i o n s  according  "Femaleness" f o r i n s t a n c e i s a s s o c i a t e d  with  h y p e r c h o l e s t e r o l e m i a (99, 38, 72), i f e s t r a d i o l benzoate i s a d m i n i s t e r e d t o a male r a t the plasma c h o l e s t e r o l  l e v e l i s elevated  (18),  while i f testosterone  i s a d m i n i s t e r e d t o a female r a t i t lowers the plasma c h o l e s t e r o l 38,  55).  Okey e t a l (86) r e p o r t e d a c y c l i c v a r i a t i o n i n c h o l e s t e r o l  the human female and o t h e r s (87) r e p o r t e d the same i n the r a t . estrogen secretions before ovulation, pid  l e v e l (2, 18,  levels.  immediately  w i t h both a r i s e i n the c h o l e s t e r o l and p h o s p h o l i -  The drop i n e s t r o g e n l e v e l s a t o v u l a t i o n  plasma c h o l e s t e r o l  The maximum  o f the l a t t e r occur d u r i n g the f o l l i c u l a r phase,  and c o i n c i d e  levels i n  i s accompanied by a low  l e v e l (87).  B e s i d e s the hormone c o n t r o l o f f a t d e p o s i t i o n t i s s u e t h e r e i s t h e a f f e c t of nervous c o n t r o l  (45).  and m o b i l i z a t i o n E x c e s s i v e nerve  i n adipose stimulation  causes a l o s s o f f a t from adipose t i s s u e , whereas i f the t i s s u e i s denervated fat  fails  t o be m o b i l i z e d a t the same r a t e .  Diets for  a r e i n s t r u m e n t a l i n changing the plasma l i p i d  i n s t a n c e i t i s w e l l known t h a t  of f a t t y a c i d s fat  ingested  l e v e l s (4, 20, 88, 99),  a carbohydrate d i e t w i l l  and thus lower the plasma l i p i d  l e v e l s (74).  spare t h e o x i d a t i o n A l s o the type of  has been shown by many t o a f f e c t the plasma c h o l e s t e r o l  levels.  (5, 18) Abnormal c o n d i t i o n s sectomy (16),  such as c a s t r a t i o n  thyroidectomy  (75),  oophorectomy (18),  (32) and adrenalectomy cause r a i s e d  lipid  hypophylevels i n  r a t s , men and dogs e s p e c i a l l y the plasma c h o l e s t e r o l f r a c t i o n . In t h i s study o f the W i s t a r r a t weight changes, f e e d smears (•?), and plasma l i p i d first  intake  l e v e l s a r e observed i n both sexes.  t h r e e measurements w i l l be u s e f u l  compared.  vaginal  Each of the  i n e s t a b l i s h i n g events, i e of growth,  puberty, and the female sex c y c l e , and t o these t h e plasma l i p i d be  levels,  levels w i l l  MATERIALS AND METHODS  15, MATERIALS AND METHODS  I  Animals: Male and female weanling rats (Wistar) from the University of B r i t i s h Columbia colony are used.  Both sexes are chosen from  two l i t t e r s , (10  females and 8 males) born one day apart. II  Diet:  1  UBC Rat Ration #10 - 63. This diet i s the stock diet f o r the whole colony. . See table I f o r composition. Ill  Animal.Housing: The rats are housed i n d i v i d u a l l y i n wire mesh cages arranged i n t i e r s . There are four t i e r s , each containing f i v e rats of one sex. The top t i e r i s 60 inches from the f l o o r , and the bottom t i e r 25 inches from the f l o o r . Each week the rats are moved down one t i e r so that by the end of two months they spend two weeks at each l e v e l . Water and feed are provided ad libitum except f o r the s i x hours of fast from 11:30 a.m. to 5:30 p.m. Controls, also l i t t e r mates are housed beside the test animals i n the same t i e r s , and are fasted f o r the same s i x hours.  IV  Weighing Procedure: A l l rats are weighed between 9 - 10 a.m. f o r the s e l f - a c c e l e r a t i n g or prepubertal growth period, and at 12:30 p.m. f o r the cycling or postp u b e r a l period.  Feed intake records are made at the same time.  animals and feed are weighed on a Mettler balance. V  Bleeding Procedure:  Both  16. Prepubertal Growth Period On almost every alternate day at 2:30 p.m. the rat to be tested i s taken from i t s cage and placed i n a holding device made of p l a s t i c (Econo-cage, #88, #90, #91,. Econo-cage D i v i s i o n Maryland P l a s t i c s , Inc. New York).  On most occasions the animal enters the 'holder' readily,  but i f a struggle occurs the animal i s removed from the test on that day. About 0.4 - 0.6 mis of blood i s taken from the t a i l after being swabbed with isopropanol. When this i s f i n i s h e d the animal i s returned immediately to i t s own cage and fed. Postpubertal Growth Period The same procedure i s used for these animals as described above, except that they are bled daily at 5:30 p.m. These animals are the former Control group for the prepubertal growth period. Blood C o l l e c t i o n The blood i s c o l l e c t e d i n a 'Red Tip' heparinized c a p i l l a r y tube (1.3 - 1.5 mm x 75 mm size, Adam Clay Inc., New York 10,. N.Y.)  One end  is stoppered with plactocene and centrifuged i n a microcapillary c e n t r i fuge (International Equipment Co., Boston, Mass. Model M.B.).  After 10  minutes of centrifuging the haematocrit i s read on a microcapillary reader (International Equipment Co., Boston, Mass., Model CR), after which the tube i s broken i n order to separate the plasma and packed  cells.  Vaginal Smears of Postpubertal Female Rats At  12:30 p.m. every day for 13 consecutive days a vaginal smear i s  taken from the ten rats, both Test and Control.  This i s started when the  animals reach the l e v e l i n g - o f f stage i n their weight gain, at 81 and 82 days of age depending on the l i t t e r .  17. Procedure for Vaginal Smears The animals are held on the lap of the technician with only s l i g h t pressure from the hand.  The t a i l i s held high and a glass eyedropper  containing 0.5 - 1.0 ml of physiological saline i s inserted into the vagina.  The saline is washed in and out of the vagina twice and the  dropper extracted.  Immediately the animal i s released and the vaginal  wash i s studied for the c e l l types present. The method of Long and Evans (73)  i s used to describe the  different  phases of the c y c l e . Methods of Determination for Plasma ' F a t ' and Cholesterol A Micromethod for the Determination of Total Lipids i n Serum and Plasma by Bengt Swahn (106) The method i s based on the colouring of plasma on f i l t e r paper with Sudan Black i n 50 - 55% ethanol. . The dye dissolves p h y s i c a l l y i n the l i p i d s and thereby colours them. The t o t a l amount of l i p i d s i n plasma may be determined as  follows:  A small amount of plasma (20 ) i s deposited by means of a pipette onto a f i l t e r paper which is then allowed to dry i n the a i r .  The  f i l t e r paper is placed in a bath containing an ethanol solution of Sudan black.  After the l i p i d s have time to take up the dye the  paper is washed i n dilute ethanol and dried i n the a i r .  The dye is  eluted from the paper with a mixture of 207 acetic acid i n absolute o  ethanol and the eluate i s then compared with suitable standards i n a spectrophotometer.  (Beckman DU)  Reagents: 1.  Saturated Solution of Sudan Black  18. To 0.1' gm of Sudan Black B. add 100 ml of 60% solution of ethanol. Heat to b o i l i n g , s t i r r i n g c o n t i n u a l l y . After cooling, f i l t e r through an Munktell Nr. 00 f i l t e r paper at least twice to separate off any undissolved Sudan Black. F i n a l concentration of alcohol w i l l be 50 - 55%,. Rinsing f l u i d 50% ethanol Eluent 20 volumes per cent Acetic Acid i n absolute  ethanol.  Standard Solution: T r i o l i e n in absolute  ethanol.  Apparatus: a)  Beckman Spectrophotometer,  DU, with 1 cms. Cuvettes  (quartz)  b)  Three shallow glass dishes for colouring and r i n s i n g .  Saranrap is  used for a cover. Technique: Whatman f i l t e r paper N r . l . , into 30 cms lengths.  Electrophoresis s t r i p s , 4 cms wide cut  This s t r i p is divided into 2 cm. squares along one  edge and further divided by 2 cms long incisions extending from the edge toward the centre of the s t r i p . Twenty micrograms (20 ) of plasma and standards are deposited by means of a micropipette in the centre of each square.  Three squares are kept  free of plasma to provide a blank. The plasma and standard are allowed to dry i n the a i r , and then are placed in clean glass dishes which contain 100 mis of saturated sudan black solution i n 50 - 55% ethanol, where i t remains undisturbed for three (3) hours.  The paper i s then removed, drained and placed i n a  similar dish containing 100 mis of 50%, wash solution for 15 minutes.  At  19. f i r s t the dish is t i l t e d once or twice but no more.  This l a t t e r procedure  is repeated u n t i l the paper has been in three successive baths.  After  the t h i r d washing the spot of serum deposited on the paper w i l l appear blue-black on a l i g h t blue background.  The f i l t e r paper i s allowed to  dry by hanging i n the a i r . A cut down the centre longitudinal l i n e of the paper w i l l  separate  the 2 cms squares, which are then placed separately i n test tubes containing 4 mis of eluent and stoppered with corks (not rubber). Elute for 60 minutes.  The extinction value of the blue solution i s  then read against Ethanol-acetic acid i n a Beckman Spectrophotometer i n a 1 cm cuvette at 590 nyu. Calculations:  The extinction value for the blank is subtracted from the  extinction values for the plasma and t r i o l i e n spots.  The amount of t o t a l  l i p i d s i n the serum is given in mg/100 ml by the following formula::*L i p i d content (mg/100 ml) of t r i o l i e n solution extinction value for t r i o l i e n spot Extinction Value :  where  log Io I  =  log Io I  =  KCL  o p t i c a l density  K  =  value of Standard  ie Constant.  C  =  concentration of unknown  L  =  1 cm. length of cuvette  x  extinction value of plasma spot.  20. A Micromethod for the Determination of Total Plasma Cholesterol: (96) Reagents: 1.  FeSo^ - Acetic Acid:Several grains of FeSo^  711^0 i s added to one l i t r e (1 1.) of  reagent grade g l a c i a l acetic acid.  After a few minutes of mixing the  solution i s f i l t e r e d and a clear solution i s obtained.  This reagent  is stable for many months at room temperature. 2.  Concentrated ^ S o ^ " A n a l y t i c a l grade.  3.  Cholesterol Standard: ing  A series of standards are prepared by dissolv-  p u r i f i e d cholesterol i n acetone-ethanol.  The solution i s kept  t i g h t l y stoppered and renewed frequently. Technique: Whatman f i l t e r paper Nr. 1 i s used, and the same design of p a r t i t i o n i s used as i n t o t a l plasma l i p i d s . 20 #of plasma i s applied to 2 cms squares of f i l t e r paper with a micropipette, and i s allowed to dry i n the a i r . The 2' cms. squares of f i l t e r paper are cut free and each i s placed i n a small test tube containing 4 mis. of the saturated ferrous sulphate solution i n g l a c i a l acetic a c i d .  The t o t a l cholesterol i s eluted by  allowing the plasma spotted squares to remain i n the reagent for at least one hour. Three m i l l i l i t r e s (3 ml.) of cholesterol-ferrous sulphate mixture i s transferred to a clean test tube and 1 ml of concentrated sulphuric acid i s added from a syringe and needle with force to f a c i l i t a t e quick mixing.  The tubes are stoppered (not rubber) and mixed again.  pink colour develops " after 10 minutes.  A salmon  21. A reagent blank i s prepared by eluting unspotted f i l t e r paper (2 cms. square) with the colour reagent and R^SO^. The standard cholesterol i s pipetted d i r e c t l y into the ferrous sulphate colour reagent and ^SO^  added.  The test solutions are then transferred to 1 cm cuvettes of four m i l l i l i t r e (4 ml.) capacity and the o p t i c a l density i s read at 490 in a Beckman Spectrophotometer. Calculations: OD OD  x s  C C  x s  X  V V  x s  m^u  TABLE I  COMPOSITION OF UBC RAT RATION # 10 - 63 COMPONENT  PER CENT  Ground barley  15.00  Ground wheat  20.00  Wheat bean  10.00  Wheat germ meal  5.00  Oat groats  10.00  Fish meal (707=)  12.50  Soya meal (50%)  5.00  Linseed o i l meal  2.50  Vitagrass  5.00  Skim milk powder  5.00  Brewers yeast  1.20  Irradiated yeast  0.10  Steamed bone meal  0.50  Iodized s a l t  0.50  Stabilized fat  2.50  Molasses (cane)  5.00  Iron oxide  0.20  Dry vitamin 'A'  (spray)  100,000 I.U.  TREATMENT OF RESULTS  24. TREATMENT OF RESULTS  Plasma L i p i d  Determinations.  The t o t a l plasma cholesterol (TPC)  l e v e l of each individual sample i s  determined as well as the t o t a l plasma l i p i d s (TPL'), and the difference between the two i s used as an estimate of the non-cholesterol or " f a t " f r a c t i o n of the blood, ie.. TPL - TPC =  (TPF)  TPF.  Haematocrit. As the blood volume of .these animals can not be determined without  inter-  fering with the growth or feed intake levels, the blood l i p i d levels were a l l standardized to an haematocrit  value of 50%.  It i s anticipated that this com-  pensates for any d i l u t i o n or dehydration that ensues because of p h y s i o l o g i c a l changes. S p e c i f i c Gravity of Plasma. The s p e c i f i c gravity of the plasma i s assumed to be 1.028 this value the number of grams of l i p i d per 100 gms  (111) and from  of plasma i s calculated.  Feed Intake. The feed intake i s expressed  as grams per animal, and grams per unit of  body weight on the days i n which the animal i s bled. In this way  the r e l a t i o n s h i p of body size to feed intake can be seen  more e a s i l y . Body Weight. Daily body weights are plotted for both male and female rats during both the pre- and postpubertal growth periods. Treatment of Relationships of Plasma L i p i d Values to one another and to Feed Intake during the Prepubertal Growth Period. The three parameters, plasma 'fat', plasma cholesterol and feed intake are  25.  plotted separately one against the other to determine i f any r e l a t i o n s h i p . occurs.  I f there i s any doubt as to the d i r e c t i o n of the trend, c o r r e l a t i o n s  are calculated by the Method of Least Squares, and their p r o b a b i l i t i e s by the Student 'T' Test. It i s seen that i n the prepubertal male r a t the plasma ' f a t ' and feed intake values when treated as above show negative,correlation.  Similarly, when  plasma ' f a t ' and plasma cholesterol are plotted there i s a negative c o r r e l a t i o n . However, when plasma cholesterol (x) and feed intake (y) values are plotted no trend i s evident.  Instead i t i s seen that at one cholesterol l e v e l there are  a number of different feed intake levels spread over a considerable range. However, i t i s assumed that i f the plasma fat (F) shows a r e l a t i o n s h i p both to the feed intake (Fl) and to the plasma cholesterol (C) then there may be some relationship between the plasma cholesterol and the feed intake. ie.  if  \FT A  F/C  Ratio The r e l a t i o n s h i p of plasma ' f a t ' to plasma cholesterol i s further deve-  loped to form a r a t i o (F/C).  This i s done by calculating the r e l a t i v e values  of the two parameters (F & C) by a r b i t r a r i l y setting the lowest value at 100 percent (0/0).  The r e l a t i v e plasma ' f a t l e v e l i s then divided by the 1  r e l a t i v e plasma cholesterol l e v e l (|j) and the product  i s referred to as the  F/C Ratio. When this i s done each point on the feed intake/plasma i s assigned a corresponding  "F/C r a t i o " value.  pattern of d i s t r i b u t i o n can be seen.  cholesterol graph  From this a r e l a t i v e l y good  That i s when the "F/C r a t i o " i s high the  26. many levels of feed intake appear to have no effect upon the plasma cholesterol value, but when the F/C r a t i o i s low, regardless of the feed intake l e v e l , the plasma cholesterol l e v e l i s high. Each "F/C r a t i o " i s tabulated, see appendix, Table v i , with the associated r e l a t i v e values of each of the three parameters (F, C & F l ) .  It can be seen  from this arrangement, that as the "F/C r a t i o " increases the plasma ' f a t increases and the plasma cholesterol decreases, a decreased  1  and there i s a trend toward  feed intake.  At this juncture a mean value i s calculated for each of the associated parameters (F, C & F l ) for a s p e c i f i c "F/C Ratio".  When the mean values are  treated by the Method of Least Squares the following correlations are observed: (male prepubertal rats) F/C Ratio : feed intake  -  negative  F/C Ratio : plasma cholesterol  -  negative  F/C Ratio : plasma fat  -  positive  Plasma Fat : feed intake  -  negative  -  positive  *(Plasma cholesterol : feed intake  * This c o r r e l a t i o n e x i s t s only because of the "F/C Ratio" arrangement (see i n appendix, Table v i ) . Graphic  Representation  In order to show the r e l a t i o n s h i p of the three parameters graphically the same mean values are prepared as percentage values and plotted on triangular co-ordinate paper ( F i g . 1", Appendix).  i  1  Female Prepubertal Rats The three parameters (F, C & F l ) are treated i n the same fashion and F/C r a t i o s are calculated.  The results are similar to the males for "F/C r a t i o s "  (relative) from 2.0 - 3.7 but are the opposite for "F/C r a t i o s " of a lower value (0.62 - 1.9).  27. Graphic Representation of the Prepubertal Growth Period. It has been customary for investigators i n the laboratory at the University of B r i t i s h Columbia (21, 62), and elsewhere (14) to express the three main phasic periods of prepubertal growth on a semilogarithmic g r i d .  Where the age  in days i s regressed against the logarithm of body weight. .See F i g s . and 13 - 14.  However, as this method of expression did not a i d i n the interpre-  tation of a l l our r e s u l t s we use the a r i t h - a r i t h g r i d as w e l l . and Figs.  6-7  See F i g s .  1-5  8-12.  In the a r i t h - a r i t h graph frequent changes i n the rate of growth are shown to advantage i f a number of small curves are drawn.  (See F i g . 2, a - d.)  We c a l l each of these an Interim phase of growth because many occur within the main phasic periods of growth.  The l a t t e r are shown on each a r i t h - a r i t h  graph by means of an arrow. Each interim phase of growth i s drawn so that the line of one that of i t s neighbour.  intersects  From the point of intersection "a" (See F i g . 2) growth  progresses slowly for twenty-four  hours, "b".  A f t e r which the rate increases  for one to two days, "c", and then slows i n the twenty-four  hours p r i o r to the  termination of the ".Interim phase, "d". The point of intersection, "a", sometimes has the f e e d - l i p i d r e l a t i o n ship of the termination of the Interim phase and sometimes that of the onset of a new phase.  This i s to be expected for the change i s gradual rather than  abrupt as the intersection of the l i n e implies.  RESULTS AND DISCUSSION  THE MALE PREPUBERTAL RAT  29. RESULTS AND DISCUSSION: THE MALE PREPUBERTAL RAT  Body Weight. The daily arithmetic accretion of weight from 30 to 90 days of age i n the male prepubertal rat i s shown i n F i g s . 1 - 5 .  The Semilogarithmic  t i o n of these same periods and animals are shown i n F i g s . 6 - 7 ,  representa-  where the age  in days has been regressed against the logarithm of body weight i n grams.  The  l a t t e r graphs show the three phases of growth i n the prepubertal s e l f - a c c e l e r a ting growth period reported by Brody (13) i n albino rats, and by Cowan et a l (21) i n the deer, and Wood et a l (116) i n the preweanling  pig.  Representation  such as t h i s shows the o v e r a l l decreasing rate of growth more f u l l y than does the arithmetic graphs.  The Semilogarithmic graph, however, masks the interim  changes of daily weight accretion and the Interim phases that occur within the three 'main' phases. In the a r i t h - a r i t h representation the lines are drawn (see Representation of Results) i n an a r b i t r a r y fashion to show the Interim phases. i n the l i n e indicate the junction between two Interim phases.  The  'breaks'  This point of  junction usually occurs between a period of fast growth i n the l a t t e r part of one phase, and a period of slow growth at the onset of another phase. It i s believed that this method of segmentation of the prepubertal growth curve w i l l serve to a i d i n the explanation of the apparently fluctuating levels of plasma l i p i d s to be described below. Body Weight and Plasma Fat: The weight changes that occur i n a twenty-four hour period and the accompanying plasma ' f a t ' levels are shown i n Tables II - VI.  The actual body  weights and the plasma ' f a t ' concentrations are represented graphically for  (S RAT  340 r  AGE IN  DAYS.  FIG. 1  SELFACCELERATING GROWTH PERIOD-  1.  d*RAT 2.  AGE IN DAYS.  FIG:2  S E L F - A C C E L E R A T I N G GROWTH PERIOD.  DAYS OF AGE.  FIG. 3  SELF-ACCELERATING GROWTH PERIOD.  AGE IN DAYS.FIG. 4  SELF-ACCELERATING GROWTH PERIOD.  d*6  AGE IN  DAYS FIG.5  SELF-ACCELERATING GROWTH PERIOD.  RAT-  400r  400r 300 200  100 90 80 70^ 30  <53 40  50  60 70 80 AGE IN DAYS FIGURE-6 SELF-ACCELERATING GROWTH PERIOD IN MALE RATS,  90  AGE IN DAYS.Fl GURE't SELF-ACCELERATING GROWTH PERIOD IN MALE RATS.  37. each animal i n F i g s .  1-5.  The plasma l i p i d determinations  are compared with the weight changes  that occur i n the twenty-four hours before the animal i s bled.  This i s done  as i t i s believed that the plasma l i p i d s are a result of the growth made and not the cause of growth to be made.  It i s known that the growth hormone (STH),  in conjunction with other anterior p i t u i t a r y and e x t r a - p i t u i t a r y gland hormones mobilizes fat from the fat depots of the body (10, 31, 47, 57, 89, 90,  110).  This enables the animal to maintain a growth energy increment over and above the energy required for maintenance.  Growth occurs primarily, therefore,  because of the STH effect, and secondarily because of the raised plasma 'fat'; both of which may  be stimulated simultaneously by the  STH.  Unfortunately i t cannot be stated, from our r e s u l t s , that the plasma 'fat* levels (Tables II - IV) show a p o s i t i v e c o r r e l a t i o n to the weight accumulated.  Although there i s a s u f f i c i e n t trend to indicate that when the rate of  gain i s high the plasma ' f a t ' l e v e l i s high, and conversely with a low growth rate.  The lack of c o r r e l a t i o n i s believed caused i n some instances by the  depressing effect of the feed intake on the plasma ' f a t known that the consumption of carbohydrates Also i t may  1  l e v e l (48, 90).  has a fat 'sparing  1  It i s  e f f e c t (30,  74).  be because the energy p o t e n t i a l mobilized by an i n t r i n s i c or external  factor i s not r e a l i z e d i n weight gain.  The factors can be hormonal ( i n t r i n s i c ) ,  or environmental effects, such as temperature or pressure or lack of adequate feed, etcetera. l o s t , eg. Rat No.  It i s possible that under these circumstances 1 at 75 days (Table I I ) ;  at 46 days (Table V);  weight can be  No. 4 at 39 days (Table IV);  No. 5  No. 6 at 39 days (Table V I ) .  Plasma ' f a t ' levels are low when the rate of gain i s low.  Such periods  of growth stasis occurs at the onset of each interim phase ( F i g . 2, ' b ) . 1  This  i s followed i n the middle of the phase ( F i g . 2, *c') by a rapid rate of growth and the maximum plasma ' f a t ' l e v e l s .  Following t h i s , but before the phase  38. terminates, (Fig.  2,  the rate of weight gain and the plasma ' f a t  1  levels drop s l i g h t l y  'd').  The point at which the lines describing the interim phases intersect (Fig  2, 'a')  i s often the terminal point of one phase or the onset point of  another phase.  The amount of weight accumulated i n the previous  twenty-four  hours and the l e v e l of plasma ' f a t makes i t quite clear as to which one i t i s . 1  The cause of a sudden drop i n both the growth rate and the plasma ' f a t ' i s not known.  The close s i m i l a r i t y of time of occurrence within a l i t t e r sug-  gests an heritable f a c t o r . rats at or near f o r t y days.  One  of the main a l t e r a t i o n s i n rate occurs i n male  At this time there i s other evidence that the  animals are entering puberty (13, 29, 53).  The animals under observation show  a coarser hair growth and a marked increase i n the size of the t e s t i c l e s . Hooker (53), by means of h i s t o l o g i c a l studies, found a marked difference in t e s t i c u l a r a c t i v i t y at 40 days amongst the rats i n his colony. suggests i s the time of the onset of puberty.  It i s i n t e r e s t i n g to note that  Leonard et a l (66) found that growth hormone (STH) interstitial  This, he  i s antagonistic to the  c e l l stimulating hormone (ICSH or LH).  If this i s true i n the  reverse then possibly the high levels of ICSH i n the anterior p i t u i t a r y e f f e c t the STH and decrease i t s l e v e l of synthesis and secretion (99) thus causing a f a l l in the growth rate.  Possibly this i s the reason for the f a l l i n plasma ' f a t '  which i s observed at forty days. Follicle  stimulating hormone (FSH) which i s also increasing i n  tion i n the blood at the same time i s not considered  concentra-  antagonistic to STH  The Main as well as the Interim breaks i n the growth curve may  (67).  be indica-  tive of a c y c l i c growth hormone e f f e c t caused by the antagonistic e f f e c t s of ICSH(LH).  Solomon and Greep (102), however, found l i t t l e difference i n the  content or potency of STH  i n the anterior p i t u i t a r y between the age of 10 -  630  39. days ( i n r a t s ) .  This does not, however, necessitate a similar l e v e l of secretion  throughout the period. There i s also the p o s s i b i l i t y that the lower growth rate and plasma ' f a t  1  i s an obligatory reaction brought about by the exhaustion of the body f a t supply (endogenous energy), and regardless of the STH effect growth i s unable to progress. Raben et a l (90) showed that one of the prime functions of STH i s to mobilize the non-esterified fatty acids (Nefa) from the f a t depots of the body.  They also showed that the fat depots can become exhausted when con-  tinuous injections of growth hormone are given.  Greenbaum (48) reported that  rats receiving limited feed when injected with STH stop growing after f i f t y days because of the exhaustion of their endogenous energy supply.  The limited  feed prevents the accretion of further fat depots from which the metabolism of the animals can draw for growth. In order to accumulate a further supply of endogenous energy, eg. as body f a t , i t i s necessary for the animal to be i n a state of p o s i t i v e energy balance.  This can be achieved i n two ways, either by an increase i n energy  intake or a decrease i n output.  From our r e s u l t s , i t appears that the prepu-  b e r t a l male animal accomplishes weight gain as fat i n both these fashions. Body Weight, Plasma ' f a t ' and Feed Intake. The relations of the l e v e l of feed intake to weight accretion f o r those days when the animals are bled are recorded i n Tables I I - VI and are expressed as the number of grams of feed per 100 grams of body weight. (Figs. 1 - 5 )  Graphically  the feed intake i s expressed as the t o t a l amount (gms) consumed  per animal i n the twenty-four hours of weight accretion, and also as the number of grams per 100 grams of body weight. In the Appendix the daily feed intake i n grams per animal i s tabulated, Tables i - v, and the r e l a t i o n s of plasma ' f a t and feed intake can be seen i n 1  40. the triangular coordinate graph, F i g . 1'. It can be seen i n F i g s . 1 - 5  that as the weight of the animal increases  i  the amount of feed consumed increases, but decreases per unit of weight.  The  l e v e l of feed intake, however, does not show a p o s i t i v e c o r r e l a t i o n to the amount of weight gain i n a twenty-four hour period.  This occurs because a high  l e v e l of feed intake causes the growth rate to diminish temporarily; a high energy cost i n a s s i m i l a t i o n (48).  there i s  Conversely when the rate of growth  i s at i t s greatest the feed intake f a l l s , and the energy i s supplied by an endogenous source; the plasma ' f a t . 1  et a l (90).  This corroborates the findings of Raben  There i s , therefore, a negative c o r r e l a t i o n of the rate of weight  gain and the l e v e l of feed intake. Within the Interim phase, the r e l a t i o n s between the three parameters are quite c l e a r l y defined. . At the onset weight accretion and the plasma ' f a t ' are low and the feed intake i s raised.  When acceleration of weight ensues  (Fig. 2, 'c') the plasma ' f a t l e v e l r i s e s and the feed intake l e v e l drops. 1  The p r i n c i p a l action appears to be between the plasma fat and the feed intake. Their levels 'seesaw' back and forth, one,ratio conducive to fast growth, the other to a slower rate of growth. The slow rate of growth and high feed intake i s conducive to an increase in the fat compartments of the body. in their work on the preweanling p i g .  This i s born out by Wood and Groves  (116)  They found that the body fat increases  as a percentage of body weight at the onset of a period of phasic growth and diminishes when the rate of weight gain i s accelerated. What causes the sudden increase i n feed intake or the change i n the other parameters i s not known.  It i s known, however, that repeated injections of  STH w i l l cause the fat depots to become exhausted (90).  Therefore i t may  be  41. possible that a 'hypothalamic fat centre', suggested by Murray (81), causes an increase i n feed intake to ensue when STH can no longer raise the Nefa Value of the blood. hormones.  This action can be likened to the 'feed back* mechanisms of many  The high feed intake or the low'fat', or both, would be the cause  of the decrease weight gain.  Then, when s u f f i c i e n t fat i s accumulated and the  plasma ' f a t ' can assume a c e r t a i n l e v e l (to effect the 'hypothalamic centres') the feed intake f a l l s and a faster growth rate can be resumed. There i s also the p o s s i b i l i t y , . as mentioned e a r l i e r , that the secretions of STH from the hypophysis i s i n h i b i t e d by the increase i n secretion or biosynthesis of some other hormone (ie FSH and/or ICSH).  This decreases the  stimulating effect upon the fat depots and allow the accumulation  of fat i n  this compartment. A high l e v e l of synthesis of some hormone i n the anterior p i t u i t a r y may also lower the biosynthesis of the thyroid stimulating hormone and hence lower the metabolism of the animal. to this suggestion.  However, a r i s e i n feed intake i s not conducive  Nor does i t seem l i k e l y that the weight gain has ceased  due to a state of stress brought about by the high rate of growth.  For i n  stress the adrenocorticoids decrease fat synthesis, raise the blood sugar by preventing u t i l i z a t i o n of carbohydrates  by the tissue and increase ketosis,  whereas the ACTH i s capable of mobilizing the fat depots. It can be seen i n the Tables I I - VI that a loss of weight occurs when there i s a low feed intake and a r e l a t i v e l y high plasma fat l e v e l . .An animal in a state of fasting i s capable of mobilizing s u f f i c i e n t Nefa (28, 41, 46, 48, 50) to s a t i s f y the c a l o r i c requirements for maintenance.  In doing so no growth  ensues, but often a loss from the fat compartments lowers the t o t a l weight of the animal.  This can be seen i n animal #1 on days 73 and 75.  42. Body Weight and Plasma Cholesterol and their r e l a t i o n s h i p with the other parameters. Irrespective of the amount of weight gain achieved the l e v e l of the plasma cholesterol does not appear to be affected d i r e c t l y . . At both the onset and terminal points i n the Interim phases the cholesterol values are high (Table VIII) whereas, at the junction of the Main periods of phasic growth there i s a low cholesterol (Table V I I ) . The terminal point of an Interim phase i s characterized by having accomplished a moderately high rate of growth i n the previous twenty-four hours; a concommitant r i s e i n the plasma ' f a t l e v e l ; 1  a feed intake which i s r a i s e d  from the previous day and which fluctuates with a negative c o r r e l a t i o n to the plasma f a t ;  and a high plasma c h o l e s t e r o l .  The onset point of an Interim phase i s characterized by a lower rate of weight accretion;  a lowered plasma ' f a t ' l e v e l ;  a raised feed intake (from  previous day) and a raised plasma cholesterol l e v e l . The points of junction of the Main phasic periods of growth are d i s t i n c t i v e from the other periods by a generally low rate of growth; high l e v e l of plasma 'fat';  a relatively  a feed intake which i s lowered from the previous  day, and p r i n c i p a l l y d i s t i n c t i v e because of a low plasma c h o l e s t e r o l . This shows very well that the different breaks i n the growth curve are s i g n i f i c a n t and d i s t i n c t i v e . plasma cholesterol l e v e l s .  The differences are emphasised primarily by the The Interim phases appear to have a d i s s i m i l a r  physiological basis for their occurrence phases.  than does the Main or p r i n c i p a l  It appears that the former i s caused by the different energy demands  of the metabolism to s a t i s f y maintenance and growth increments,  whereas, the  l a t t e r i s an inherent physiological function superimposed upon the other, possibly due to an i n i t i a l sex hormone e f f e c t .  This i s not to say that the  43. Interim phases are not inherent as w e l l . and 6 the day of occurrence F/C Ratio:-  In l i t t e r mates 1 and 2, and 4, 5,  i s often s i m i l a r .  In the section 'Presentation of Results  1  methods and reasons for  the c a l c u l a t i o n of an F/C r a t i o are outlined. . In Tables VII - IX, which are a refinement presented  of the other f i v e Tables  (Tables II - VI) i t can be seen that the F/C r a t i o s become concentrated  according to value and what may  be the b i o l o g i c a l a c t i v i t y .  ' The points of Junction of the main growth phase have an F/C r a t i o i n the Actual range of 2.0 - 2.4 or a Relative range of 1.29 are doubtfully placed.  to 1.55,  except two which  These are among the higher l i m i t s of the range of F/C  r a t i o s and denote a low plasma cholesterol, and a high plasma'fat' l e v e l , the l a t t e r always 2 - 2.4 times higher than the plasma c h o l e s t e r o l . Correlations between the F/C r a t i o and the other parameters are calculated, as described e a r l i e r (Presentations).  For the present group of animal under  consideration they are as follows: F/C Ratio :- Feed intake  = negative  F/C Ratio :  Plasma cholesterol = negative  F/C Ratio;;  Plasma ' f a t  1  = positive  With the plasma values of ' f a t ' and cholesterol arranged according to t h e i r r a t i o (see Appendix, Table v i ) to one another,. and subsequently  their  b i o l o g i c a l a c t i v i t y , the following r e l a t i o n s also become evident: Plasma ' f a t  Feed intake  = negative  Plasma cholesterol: Feed intake  = positive  Plasma ' f a t ' : Plasma cholesterol  = negative  1  :  Correlation of the F/C r a t i o and weight accretion cannot be made as the l a t t e r i s unrelated to either the plasma ' f a t ' levels or the cholesterol l e v e l s . That a r e l a t i o n s h i p did exist i s not doubted, but because we have no knowledge  44. of the energy cost of maintenance or the composition  of gain i t i s not evident.  The Terminal Point of the Interim phase of Growth. The F/C r a t i o at the terminal point of an Interim phase of growth are shown i n Table VIII.  The levels are i n the medium range compared to the Main  phasic periods, and the onset point of the Interim phase. 1.3 - 1.9 and the Relative range i s 0.86  - 1.2.  cholesterol i s high r e l a t i v e to the plasma ' f a t ' .  The Actual range i s  This indicates that the plasma As this i s a period of moder-  ately high growth the plasma fat i s s i m i l a r i l y h i g h . An interesting r e l a t i o n s h i p that e x i s t s at the terminal point and nowhere else, i s that an increase i n the weight gain i n twenty-four hours i s p o s i t i v e l y correlated with the l e v e l of feed intake i n that time.  It may  dence because of the small number of animals and occasions. may  only be a c o i n c i -  Alternatively i t  be that maintenance i s dealt with exclusively by the endogenous energy, and  the weight gain by the exogenous energy source, which i n this instance may fat.  This i s a situation often encountered i n the female r a t .  be  Kochakain (61)  relates the c a l o r i c value of the feed intake to weight gain as fat after excess testosterone treatment has ceased. Onset point of Interim phase of Growth. The F/C r a t i o s at the onset point of an Interim phase of growth (Table VIII) occurs i n the lowest range of the three points of junction so far mentioned, as well as a l l other points within the Interim phase. values i s 0.96  - 1.1,  and the Relative range i s 0.68  The Actual range of  - 0.71.  The plasma  cholesterol values are actually and r e l a t i v e l y high i n comparison with the now  lower plasma ' f a t l e v e l s . 1  Because of the negative c o r r e l a t i o n of these  two l i p i d values one l e v e l f a l l s as the other r i s e s . Intermediate  points within the Interim phase of Growth.  The intermediate points that exist between the onset and terminal points  45. of an Interim phase are found to have F/C r a t i o values i n the medium and high range,, but never i n the low range of the onset point which i s unique. It appeared that the F/C r a t i o s occur i n a d e f i n i t e pattern within an Interim phase, this information i s derived from the composite r e s u l t s of a number of animals bled on alternate days.  This derived pattern shows the  maximum F/C range occurs i n the middle of the phase ( F i g . 2, 'c') having risen slowly from the nadir at the onset medium range ( F i g . 2, 'b ) 1  to the zenith.  ( F i g . 2, 'a  1  or 'b') through a  After this high range i s reached  the F/C values f a l l again to the medium range where the Interim phase terminates. If we compare the points of i n t e r s e c t i o n (or junction) of the Interim growth phase as delineated on the a r i t h - a r i t h graph, with the points of i n t e r section of the three main phases on the a r i t h - l o g graph of growth we f i n d that they d i f f e r markedly i n their F/C r a t i o .  This seems to emphasize that the  two  points of apparent change i n growth are not similar and do not describe the same phenomenon. We have ascribed to them a different b i o l o g i c a l a c t i v i t y on t h i s b a s i s . The suggestion i s that the p r i n c i p a l or main growth rate change may the onset of puberty.  be due to  If i t i s an androgen e f f e c t we would expect a low  plasma cholesterol (18, 39) which i s indeed the case.  Yu Min Chen (18) demon-  strated the a b i l i t y of t h i s hormone to lower the plasma cholesterol i n intact females and i n castrated males. Kochakain (60) showed that high levels of androgen (testosterone propionate) lower feed intake.  This a c t i v i t y also appears evident i n our animals at  the point of i n t e r s e c t i o n i n the Main phase on the a r i t h - l o g graph., We have seen i n our own r e s u l t s , and from other reports (28, 41, 46,  48,  50) on the effect of fasting, that a low feed intake stimulates a high plasma 'fat . 1  Also that the protein anabolic a c t i v i t y of androgens (60) raise this  46. level.  Therefore, i t i s not surprising to f i n d this effect at the 'break' or  junction of the main phasic period of growth. Increase i n the rate of growth i s usually the outcome of an increase i n the l e v e l of androgen, and yet we see a d e f i n i t e decrease i n the rate of accretion.  This suggests that sexual maturity i s i n h i b i t i n g fast growth.  Kochakain  (60) found that high levels of androgens lower the weight of the male rat, but t h i s i s as a result of the loss of f a t . As suggested e a r l i e r , an anterior p i t u i t a r y 'cut out' of STH may  be caused by the increasing biosynthesis of  gonadotropens occuring at t h i s time (8).  It i s known that i n an hypophysec-  tomised animal the epiphyseal plates i n the long bones close, but i f i n j e c t i o n s of STH are given the plates stay open and the animal can increase i t s skeletal size (108).  The suggestion here then i s that the rate of growth i s diminished  as a result of i n i t i a l 'cut out ICSH (LH) or FSH. rate then, may  Sohval (100)  1  of STH by the anterior p i t u i t a r y gonadotropen suggested that t h i s i s a p o s s i b i l i t y .  Growth  progress more slowly, e s p e c i a l l y i n the long bones.  It appears then from the lipid-feed-growth relationships that this  may  be an androgen e f f e c t . Feed intake and Plasma Cholesterol l e v e l s . There i s no indication, other than a r i s e i n feed intake, to suggest what causes an increase i n plasma c h o l e s t e r o l . However, i t must be remembered that t h i s c o r r e l a t i o n exists by v i r t u e of the F/C r a t i o which we believe may be an indication of a s p e c i f i c b i o l o g i c a l a c t i v i t y . a c t i v i t y other than the feed intake i s responsible.  This means that some  TABLE II LEVELS OF WEIGHT ACCRETION, PLASMA LIPIDS, AND FEED INTAKE IN MALE RAT. 1.  Weight * Changes GM.  Plasma Fat mgm7o  Feed Intake gm/lOOgms of B.W.+  Plasma Cholesterol mgmT.  F/C Ratio Actual  Relative  Age in Days  - 1.3  212.0  5.2  108.0  2.0  1.29  73  + 1.9  246.0  8.14  110.5  2.2  1^9  40  + 2.0  182.5  11.40  176.0  1.0  0.69  47  + 3.0  164.9  8.84  84.0  2.0  1.29  61  + 7.4  319.0  12.30  144.0  2.2  1.49  45  - 1.3  330.0  8.50  100.0  3.3  2.12  75  + 8.8  234.5  9.04  136.5  1.7  1.08  68  +11.2  261.0  11.70  157.0  1.7  1.08  50  +11.0  317.0  8.40  119.0  2.7  1.74  66  +18.2  299.0  11.00  141.0  2.1  1.39  52  *  Weight gained or l o s t i n twenty-four hours.  +  B.W. = body weight  TABLE III LEVELS OF WEIGHT ACCRETION PLASMA LIPIDS, AND FEED INTAKE IN MALE RAT. 2.  Weight * Changes GM.  Plasma Fat mgm7o  Feed Intake gm/lOOgms of B.W.+  Plasma Cholesterol mgm7o  F/C Ratio Actual  Relative  Age in Days  10.6  107.0  1.29  1.29  40  + 1.2  209.0  + 1.3  190.0  9.05  127.0  1.5  0.97  47  + 1.6  220.0  7.90  100.0  2.2  1.43  61  + 2.3  188.0  4.60  101.0  . 1.9  1.20  73  + 4.7  234.8  8.27  135.5  1.8  1.18  68  + 4.9  194.0  8.45  176.0  1.1  0.71  50  .+ 5.0  235.0  7.07  96.0  2.4  1.55  75  + 5.5  200.5  8.20  148.5  1.4  0.91  52  .+ 8.2  263.5  7.16  114.0  2.3  1.50  66  + 8.9  288.0  10.95  116.0  2.4  1.55  45  -  164.5  -  157.5  1.0  0.68  38  -  233.0  8.15  125.0  1.9  1.2  59.0  Weight gained or l o s t i n twenty-four  +  B.W. = body weight  hours.  TABLE IV LEVELS OF WEIGHT ACCRETION, PLASMA LIPIDS,, AND FEED INTAKE IN MALE RAT. 4. Feed Intake gm/lOOgms of B.W..+  Plasma Cholesterol mgm7o  F/C Ratio Actual  Relative  Age in Days  11.5  129.8  1.4  0.91  46  10.4  89.5  2.0  1.29  39  5.36  103.0  1.6  1.06  72  155.5  8.30  122.5  1.3  0.86  67  + 4.0  217.0  7.02  96.0  2.3  1.50  74  + 4.4  180.0  8.15  163.0  1.1  0.71  49  + 6.5  259.0  8.65  106.0  2.4  1.55  60  + 7.8  206.5  8.20  145.5  1.4  0.91  51  + 7.8  194.5  11.95  129.4  1.5  0.97  44  Weight * Changes GM.  Plasma Fat mgm7o  - 3.3  179.2  - 2.2  179.5  + 1.8  171.0  . + 3.9  *  Weight gained or l o s t i n twenty-four hours  +  B.W. = body weight  TABLE V LEVELS OF WEIGHT ACCRETION, PLASMA LIPIDS, AND FEED INTAKE IN MALE RAT. 5.  " Weight * Changes GM.  Plasma Fat mgm7o  Feed Intake gm/lOOgms of B.W.+  Plasma Cholesterol mgm%  F/C Ratio Actual  Relative  Age" in Days  - 0.4  238.0  8.2  132.0  1.8  1.18  46  + 3.7  258.0  7.52  137.0  1.9  1.20  67  + 3.7  138.0  11.80  103.0  1.3  0.86  39  + 3.9  148.0  8.30  154.0  0.96  0.68  49  + 5.4  224.0  7.92  98.0  2.3  1.50  60  :+ 5.7  196.0  10.45  132.0  1.5  0.97  44  + 7.2  224.0  7.30  112.0  2.0  1.29  65  :+ 9.0  298.0  7.70  136.0  2.2  1.43  51  255.0  7.92  109.0  2.3  1.50  58  153.5  1.1  0.71  35  153.5  1.1  0.71  37  —  163.5 172.5  -  *  Weight gained or l o s t i n twenty-four hours.  +  B.W. = body weight  TABLE VI LEVELS OF WEIGHT ACCRETION, PLASMA LIPIDS, AND FEED INTAKE IN MALE RAT. 6.  Weight * Changes GM.  Plasma Fat mgm7o  Feed Intake gm/lOOgms of B.W.+  Plasma Cholesterol mgm%  F/C Ratio Actual  Relative  Age in Days  - 3.2  201.0  5.87  119.5  1.7  1.08  72  - 1.1  238.0  9.80  120.0  2.0  1.29  39  +12.7  202.0  11.90  154.0  1.3  0.86  44  + 4.6  243.0  10.70  154.0  1.6  1.06  46  + 5.5  297.0  6.90  120.0  2.6  1.71  74  + 6.1  223.0  8.25  150.0  1.5  0.97  67  + 6.6  157.0  7.66  110.0  1.4  0.91  60  + 8.5  185.0  9.72  168.0  1.1  0.71  49  + 8.6;  200.0  7.01  122.0  1.6  1.06  65  +15.1  210.0  9.94  155.0  1.4  0.91  51  -  200.0  8.50  165.0  1.2  0.77  58  *  Weight gained-or l o s t i n twenty-four hours.  +  B.W. = body weight  TABLE VII WEIGHT CHANGES:  PLASMA LIPIDS AND FEED INTAKE OF MALE (PREPUBERTAL) RATS.  AT JUNCTION OF TWO 'MAIN  1  Animal No. 1  2  4  5  . Weight * Changes (gm)  Plasma Fat mgm7o  Feed Intake GMS/100GMS +  Plasma Cholesterol mgm%  F/C Ratio Actual Relative  Age Tested  + 1.9  39 Approx.  246.0  8.14  110.5  (2.2)  1.43  40  + 3.0  62  164.9  8.84  84.0  (2.0)  1.29  61  + 1.2  39 or 40  209.0  10.60  107.0  (2.0)  1.29  40  + 1.6  60  220.0  7.90  100.0  (2.2)  1.43  61  -  38 or 39  179.5  10.40  89.5  (2.0)  1.29  39  + 6.5  60  259.0  8.65  106.0  (2.4)  1.55  60  + 3.7  37 & 38  138.0  11.80  103.0  58  255.0  7.92  109.0  (2.3)  1.50  58  1.1  38  238.0  9.8  120.0  (2.0)  1.29  39  + 6.6  61  157.0  7.66  110.0  2.2  mm  6  Day of Junction  PHASIC PERIODS OF GROWTH  -  *  Weight gained or l o s t i n twenty-four  +  of body weight  hours.  39  60  TABLE V I I I WEIGHT CHANGES:  PLASMA LIPIDS AND FEED INTAKE OF MALE (PREPUBERTAL) RATS.  A. . AT TERMINAL POINTS OF INTERIM PHASE OF GROWTH  Animal No.  Weight * Changes GM.  Plasma 'Fat' mgm7  Feed Intake GMS/100GMS o f B.W. +  0  Plasma Cholesterol  mgm7o  F/C R a t i o Actual Relative  Age in Days  1  +11.2 + 8.8  261.0 234.0  11.70 9.04  157.0 136.5  1.7 1.7  1.08 1.08  50 68  2  + 4.7  234.5  8.27  135.5  1.8  1.18  68  4  + 7.8  206.5  8.20  145.5  1.4  0.91  51  5  + 3.7  258.0  7.52  137.0  1.9  1.20  67  6  + 6.1 +12.7  223.0 202.0  8.25 11.90  150.0 154.0  1.5 1.3  0.97 0.86  67 44  B.  1  AT ONSET POINTS OF INTERIM PHASE OF GROWTH  2.0  182.5  11.40  176.0  1.1  0.71  47  2  .+ 4.9  194.0  8.45  176.0  1.1  0.71  50  4  + 4.4  180.0  8.15  163.0  1.1  0.71  49  5  + 3.9  148.0  8.30  154.0  0.96  0.68  49  6  + 8.5  185.0  9.72  168.0  1.1  0.71  49  * .+  * +  Weight gained o r l o s t B.W. + body weight  i n twenty-four h o u r s .  TABLE IX WEIGHT CHANGES:  PLASMA LIPIDS AND FEED INTAKE OF MALE (PREPUBERTAL) RATS.  THE (PREPUBERTAL) MALE RAT:  INTERMEDIATE POINTS IN INTERIM PHASE OF GROWTH Age in Days  Weight Changes (GM)  Plasma 'Fat* mgm7o  Feed Intake gm/lOOgm of B.W..+  Plasma Cholesterol mgm7o  + 7.4 +18.2 +11.0 - 1.3  319.0 299.0 317.0 212.0  12.30 11.00 8.40 5.20  144.0 141.0 119.0 108.0  2.2 2.1 2.7 2.0  1.49 1.39 1.74 1.29  45 52 66 73  2  + + + + + +  8.9 1.3 5.5 8.2 2.3 5.0  288.0 190.0 200.5 263.5 188.0 235.0  10.95 9.05 8.20 7.16 4.60 7.07  116.0 127.0 148.5 114.0 101.0 96.0  2.4 1.5 1.4 2.3 1.9 2.4  1.55 0.97 0.91 1.50 1.20 1.55  45 47 52 66 73 75  4  + + + +  7.8 3.9 1.8 4.0  194.5 155.5 171.0 217.0  11.95 8.30 5.36 7.02  129.4 122.5 103.0 96.0  1.5 1.3 1.6 2.3  0.97 0.86 1.06 1.50  44 67 72 74  5  - 0.4 + 9.0 .+ 7.2  238.0 298.0 224.0  8.2 7.7 7.3  132.0 136.0 112.0  1.8 2.2 2.0  1.18 1.43 1.29  46 51 65  6  + 4.6 +15.1 + 8.6 - 3.2 + 5.5  243.0 210.0 200.0 201.0 297.0  10.7 9.94 7.01 5.87 6.9  154.0 155.0 122.0 119.5 120.0  1.6 1.4 1.6 1.7 2.6  1.06 0.91 1.06 1.08 1.71  46 51 65 72 74  +  B.W. = body weight  Animal No. 1 T  1  o  n  F/C Ratio Actual Relative  RESULTS AND DISCUSSION  THE FEMALE PREPUBERTAL RAT  56.  RESULTS AND DISCUSSION THE FEMALE PREPUBERTAL RAT  Body Weight The daily l e v e l of weight accretion from t h i r t y to eighty days of age i s shown f o r f i v e female rats on an A r i t h a r i t h grid i n F i g s . 8 - 12, and on an A r i t h l o g g r i d i n F i g s . 13 and 14.  In these l a t t e r graphs the age i n days  i s regressed against the log of the body weight. The l i n e s drawn i n a l l graphs are a r b i t r a r y , as i t i s believed that the different growth trends were quite c l e a r .  Also, by not using s t a t i s t i c a l  methods s l i g h t changes from the normal trend are not o b l i t e r a t e d . The daily weights i n grams f o r each animal and t h e i r age i n days are enumerated i n Tables v i i i - x i , found i n the appendix. A.  A r i t h a r i t h Representation of the Growth Curve. From F i g s . 8 - 12 i t can be seen that weight accretion i n the female rat progresses at a slower rate than i n the male.  However, l i k e  the male there i s a r e l a t i v e l y fast rate of growth to approximately t h i r t y - e i g h t or thirty-nine days of age, after which many variations i n rate occur.  This v a r i a t i o n i s represented by a number of broken lines  dividing the growth curve into small phases of growth which we w i l l , from henceforth, c a l l the Interim phasic growth periods.  (See also the  section on prepubertal growth i n the male r a t s ) . The d i s t i n c t change i n the growth pattern at t h i r t y - e i g h t or thirty-nine days coincides with the time of onset of puberty as described by Hooker (52) i n h i s colony of rats, and by Brody (14) i n the albino rat, and we believe f o r the male l i t t e r mates of these females. The fluctuating growth pattern made by the small Interim phases  A G E IN  D A Y S . FIG.8.  S E L F - A C C E L E R A T I N G G R O W T H PERIOD-  A G E IN DAYS. F I G . 9 . S E L F - A C C E L E R A T I N G G R O W T H PERIOD-  $ 3 RAT P L A S M A FAT  PLASMA  CHOLESTEROL  FEED INTAKE  G MS/ANIMAL  G M S / I O O GMS X  P  B O D Y WEIGHT.  --or 30  40  50  6 A G E IN  0  7  0  DAYS.  80" FIG.iO.  SELF-ACCELERATING G R O W T H PERIOD.  90  9  6. RAT.  P L A S M A FAT  220 190  200  170  180  150  x  130F  rj]i60 >  Q 110 O CD140  <S)  120 FEED 100 • 70 RATI07  .9  19  INTAKF  G MS/ANIMAL  15 80  •  ) 10  CO P  60  ^ '  5. 30  40  5 0 6 0 " 70" A G E I N D A Y S . FIG.11.  SELF-ACCELERATING  GMS/IOOGMS BODY W E I G H T 80"  G R O W T H PERIOD  90  £  5-1 30  . 40  . 50  60  80  70  A G E IN DAYS. FIG.12. SELF-ACCELERATING GROWTH  PERIOD-  . 90  200r  A G E IN DAYS. FIGUREM3. S E L F - A C C E L E R A T I N G GROWTH PERIOD IN F E M A L E  RATS.  '  40  '  »  •  '  50  I  60 AGE  SELF-ACCELERATING  IN  •  70 DAYS  80  90  FIGURED.  GROWTH P E R I O D IN  FEMALE  RATS.  64,  is reminiscent of the growth rate pattern seen during estrus c y c l i n g in the sexually mature female r a t .  From approximately  forty to sixty  days the timing of the cycle i s e r r a t i c , that i s the Interim phasic periods vary i n length.  A f t e r sixty days the timing more c l o s e l y re-  sembles the four to f i v e day c y c l i c periods (12, 65). In Tables XVI - XVIII the weight changes i n the twenty-four hours p r i o r to blood taking are recorded for the different points within the Interim phase.  The pattern of weight accretion during each phase  appears to begin slowly, often with a regression of weight occurring at and/or after the onset, that i s only a f t e r forty days of age. After the loss of weight or slow gain, weight gain increases u n t i l the terminal point of the Interim phase i s reached.  The greatest amount of gain  being achieved two days p r i o r to the terminal point. It can be seen ( i n Table XVIII) that on days 64 and 65 there i s a large drop i n weight. . These appear to be caused by unduly high environmental temperatures that occur on three successive days during the test period.  The only other examples of a weight loss of such magnitude  occurs when feed i s not supplied for a twenty-four hour period. A r i t h l o g Representation  of the Growth Curve  In the female rats, as i n the males, another pattern of growth i s introduced by an a r i t h l o g representation of weight changes, F i g s . .13 and 14. In these graphs the small Interim phasic periods of growth do not show c l e a r l y but a larger more general pattern i s expressed of three intersecting l i n e s .  that consists  The points of junction occurring on the  f o r t i e t h , s i x t i e t h and eightieth days approximately.  The exact age i s  marked by arrows on the a r i t h a r i t h graphs of the individual  animals.  65,  From each of these points of junction a d e f i n i t e decrease i n the rate of gain takes place.  We w i l l , henceforth, c a l l the phases that occur  between these points the Main phasic periods of growth. In one l i t t e r (Rats No. 1, 2 and 3) a d e f i n i t e increase i n the rate of growth occurs at the age of f i f t y - o n e days.  I t s relationship  to the other points of junction i s not f u l l y understood.  However,  twenty-four hours p r i o r to t h i s happening aureomycin was administered to the drinking water because one animal coughed and sneezed more than usual.  It i s believed, because of the changes i n growth, that this one  l i t t e r was suffering from a s u b c l i n i c a l i n f e c t i o n .  The other l i t t e r  (Rats No. 6 and 7) shows no such change i n weight accretion.  It w i l l  be seen i n a later description, however, that the feed intake and the plasma cholesterol levels are effected i n a l l the animals. II Relationship of Body Weight and Plasma  'Fat . 1  The effect of weight accretion on the l e v e l of plasma 'fat* i s shown graphically i n F i g s . 8 - 12, and numerically i n Tables X - XIV.  In the l a t t e r  the weight accumulated i n the twenty-four hours p r i o r to bleeding i s tabulated. There i s no direct c o r r e l a t i o n between the weight accumulated i n twenty-four hours and the plasma ' f a t l e v e l .  That i s , there i s never a s p e c i f i c l e v e l  1  of plasma ' f a t for a s p e c i f i c weight gain. 1  This i s because the l e v e l of  plasma ' f a t ' i s the result of other metabolic demands besides that of growth. However, i t i s usual to f i n d a high plasma ' f a t ' when the growth rate i s high.  At such a time the non-esterified fatty acid f r a c t i o n (Nefa) i s ,  according to Raben et a l (90^ a more e f f i c i e n t source of energy for growth than an exogenous source where digestion and assimilation compete with the growth increment for the available energy. A high l e v e l of plasma ' f a t also occurs when the animals lose weight 1  66. as f a t . Many investigators (28, 41, 46, 48, 50) have shown that i n a n i t i o n i s the p r i n c i p a l factor for this happening.  They have been able to demonstrate  the presence of a raised Nefa f r a c t i o n , and have shown that t h i s energy mobilized.from  the animal's fat stores i s s u f f i c i e n t to support i t s maintenance.  On days forty-one and forty-two, i n l i t t e r s two and one respectively, the animals are without  feed but adequate water.  shows us that the plasma'fats'are of inanition marked by an 'x'). a loss of weight;  This i s unintentional but  raised by inanition (Figs. 8 - 12; days A l l the animals except No.  1 and No. 6 show  a loss which appears to be p r i n c i p a l l y from the fat stores  of the body. A.  Interim phasic Growth Period. 1.  Onset Points of Junction (Table  XVI)  Weight accretion at t h i s point i s low.  The plasma 'fats,'  however, are moderately high, but lower than at any other point in the Interim phase.  These two parameters fluctuate together  in a p o s i t i v e fashion, but whether t h i s i s a direct r e l a t i o n s h i p i s not known, as i t w i l l be seen that the feed intake i s also involved.  A loss of weight i s accompanied by a low plasma 'fat',  while those points that show a gain i n weight generally are accompanied by a higher plasma ' f a t . 1  This might suggest that a loss  of weight i n t h i s instance i s not a loss from the fat compartments of the 2.  animal.  Terminal Points of Junction. Weight accretion p r i o r to t h i s junction i s always p o s i t i v e and r e l a t i v e l y high; e a r l i e r i n the phase.  the greatest accumulation  usually occurring  The plasma ' f a t ' i s concomitantly  high  although these two parameters do not fluctuate together as they  67. do i n the previous point discussed, ie the onset point. B.  Main Phasic Period of Growth. 1.  Points of Junction of the Main Phasic Growth Period.  (Table  XV)  The plasma fat levels at t h i s junction are low which i s different from the males.  At f o r t y days the rate of weight  accretion i s r e l a t i v e l y high, and higher than at sixty days of age, commensurate with which the range of plasma fats i s also higher.  This difference may  be due d i r e c t l y to age,  ie the  a b i l i t y to grow at a greater rate before f o r t y days of age, or possibly to the composition  of the gain.  It i s possible that the low plasma 'fats' occur because weight i s being accumulated i n the fat compartments, and not being mobilized as energy for protein growth. G.  .Points of Junction i n Growth Curve at Fifty-one days of  Age.  These points are believed to be unnatural, t h e i r occurrence i n Rats Nos.  1, 2 and 3, a r i s i n g because of the use of aureomycin i n the  drinking water. The weight accretion that occurs twenty-four hours before this point i s r e l a t i v e l y low, as i s also the plasma ' f a t ' .  Similar levels  are found only at the points of junction of the Main phasic period. We have no explanation for this observation.  However, i t should be  r e c a l l e d that the Main phase points are probably not related to estrus cycles whereas the Interim phase points are believed to be so governed. There i s therefore no reason to expect similar events to characterize the two classes of phases i n the growth behavior. I l l Relationship of Body Weight and Feed Intake. The r e l a t i o n s h i p of weight accretion and feed intake i s represented  graphically i n F i g s . 8 - 12.  The feed intake i s expressed as the amount of  food consumed per animal and per 100 grams of body weight.  Daily body weights  and feed intake are tabulated and can be found i n the appendix Tables v i i xi.  Tables X - XIV show the twenty-four hour accumulation  accompanying feed intake per 100 grams of body weight.  of weight and the  Whilst i n Tables XV -  XVIII the parameters are arranged according to their p o s i t i o n i n the Interim phase or the Main phasic period. The feed intake per animal changes very l i t t l e i n i t s levels, other than to fluctuate, from t h i r t y to seventy-plus  days.  However, the amount of  feed consumed per 100 grams of body weight decreases as the animal grows. A.  Interim Phasic Period of Growth. 1.  Onset Point of Junction. Generally there i s no c o r r e l a t i o n between the rate of weight accretion and the amount of feed consumed i n twenty-four hours.  This i s due p r i m a r i l y to our ignorance of the c a l o r i c  requirements of maintenance.  However, at the onset point of the  Interim phase the rate of growth and the l e v e l of feed intake are low and both fluctuate together i n a p o s i t i v e fashion.  These  two low l e v e l l e d parameters are accompanied by a moderately high plasma ' f a t , but lower than at any other point i n the Interim 1  phase.  Although the weight and feed intake levels are i n extremes  of the plasma 'fat' they a l l appear to fluctuate i n p o s i t i v e accord. The moderately high plasma ' f a t ' probably results from the  low  feed intake. The weight loss that sometimes occurs at the onset point appears to result either from i n s u f f i c i e n t feed intake or from i n s u f f i c i e n t fat stores to support more than maintenance.  A high  69.  plasma ' f a t ' on this occasion suggests that the weight loss i s from the fat compartments, but the actual levels are low enough to suggest that l i t t l e fat i s being mobilized. When there i s a gain i n weight there i s a higher feed i n take and higher plasma ' f a t ' .  In the males at the onset point  the rate of gain and the plasma 'fats* are low, similar to the females, but the l e v e l of feed intake i s increased.  This suggests  that a different endogenous and exogenous energy balance occurs i n the females, possibly as a r e s u l t of the female sex hormones. 2.  Terminal Points of the Interim Phase. As described e a r l i e r , weight accretion at the terminal point i s always positive and moderately high, but never as high as at some intermediate points i n the Interim phase.  This moderate  growth i s accompanied by a low l e v e l of feed intake, which i s often at i t s lowest l e v e l when the weight gain i s at i t s highest. However, there i s no general negative c o r r e l a t i o n . The energy for the growth appears to be supplied by a very high plasma ' f a t ' l e v e l :  higher at the terminal points than at  any other point i n the Interim phase.  The feed intake and plasma  'fat' relationships are therefore the usual low-high relations of these two parameters during good growth.  There i s also, what  appears to be t y p i c a l only to the female rat, a tendency for an increase i n feed intake to cause an increase i n the plasma ' f a t ' . That i s the plasma ' f a t and feed intake fluctuate i n positive 1  accord. however.  There appears to be no e f f e c t upon the rate of growth, Whereas i n the male rat an increase i n the feed intake  causes the plasma ' f a t ' to decrease with a concommitant drop i n  70. weight a c c u m u l a t i o n .  T h i s i m p l i e s t h a t the male r e l i e s more on  endogenous energy ( f a t ) f o r growth, whereas the female appears t o be capable  of u t i l i z i n g an exogenous source without  i n t e r f e r e n c e from the energy e x p e n d i t u r e it  the  a more e f f i c i e n t  endogenous energy source. 'fat  1  However,  the female never  same h i g h r a t e of weight g a i n as the male.  as mentioned e a r l i e r ,  Since,  growth ensues from  an  I t i s a l s o p o s s i b l e t h a t the f e e d -  - weight r e l a t i o n s i n the female are more conducive to f a t  storage  i n the t i s s u e s .  That i s to say the female i s g a i n i n g  weight p r i n c i p a l l y as f a t , and  B.  of a s s i m i l a t i o n .  i s p o s s i b l e t h a t t h i s i s the reason why  achieves  too much  the male p r i n c i p a l l y as p r o t e i n .  Main P h a s i c P e r i o d of Growth. 1,  The  P o i n t s of J u n c t i o n of the Main P h a s i c P e r i o d s of Growth. At f o r t y and  t o the p r e v i o u s  s i x t y days the f e e d i n t a k e i s r a i s e d r e l a t i v e  day.  weight b e i n g r e c o r d e d The  The  highest  l e v e l s per  100  grams of body  f o r the f o r t y day p e r i o d (Table  XV).  r a t e of weight g a i n i n the twenty-four hours p r i o r  the Main p h a s i c p o i n t s of j u n c t i o n covers  a wide range, hence  t h e r e i s no evidence  of a r e l a t i o n s h i p between the r a t e of  weight a c c r e t i o n and  feed intake.  weight i s gained  However, at s i x t y days  than at f o r t y days and  grams of body weight i s c o n c o m i t a n t l y l e v e l s are lower than at any subsequently  a negative  but when a h i g h e r also higher.  less  the f e e d i n t a k e per  lower.  The  r e l a t i o n s h i p of f a t and  100  plasma ' f a t '  o t h e r time i n the growth  feed intake i s recorded  to  curve,  feed i s evident,  the plasma ' f a t ' i s  T h i s i m p l i e s t h a t the l e v e l of plasma ' f a t ' i s  dependent upon the feed intake, ie i t i s p a r t i a l l y of an exogenous  nature. The conditions found at this point i n the female are the  opposite to those i n the male at the same point.  However, they  are s i m i l a r to the onset point i n the male Interim phase.  It  w i l l be remembered that we had considered t h i s l a t t e r point to be r e l a t e d to the energy requirements pertaining to growth and maintenance, and the points of junction of the Main phases to pertain to sexual development.  There i s every p o s s i b i l i t y that  in the female the Interim phases are concerned with sex development and the Main phases with the energy requirements of the growth proper, A possible explanation of the differences i n the size of the phases pertaining to the growth proper i n the two  sexes i s  that the very fast growth i n the male animal requires more energy from the system, subsequently the body fat compartments become exhausted more frequently and growth i s interrupted, during which time energy i s stored (116).  Because the female grows at  a lesser rate these interruptions occur less often, and because the demands upon her fat depots are less she i s generally a f a t t e r animal than the male. Fifty-one Days of  Age.  No c o r r e l a t i o n of weight accretion and feed intake i s seen at this age.  The feed intake i s high per unit of body weight  and the plasma ' f a t ' i s low.  Because this plasma ' f a t ' and feed  intake picture i s so similar to the points of junction of the Main phasic period of growth we can assume the l a t t e r to be more  7-2. c l o s e l y a l l i e d t o the energy exchange of the growth proper  and  not t o sex development. IV  R e l a t i o n s h i p of Body Weight and Plasma C h o l e s t e r o l . As seen i n the males, i r r e s p e c t i v e of the amount of g a i n i n body weight achieved,  the l e v e l of the plasma c h o l e s t e r o l d i d not appear t o be a f f e c t e d  directly. From T a b l e s XVI - XVIII, t h e i r occurrence t e r o l i s lowest point.  V  i n the I n t e r i m phase, i t i s e v i d e n t  a c c o r d i n g to  t h a t the plasma c h o l e s -  two days p r i o r t o the t e r m i n a l p o i n t , and r i s e s at the t e r m i n a l  This higher  then f a l l s ,  where parameters a r e arranged  l e v e l remains on the day of onset  of the next phase and  t o r i s e a g a i n at the t e r m i n a l p o i n t .  Body Weight:  Plasma C h o l e s t e r o l and t h e i r R e l a t i o n s h i p t o o t h e r Parameters  w i t h i n the S e p a r a t e P h a s i c P e r i o d s of Growth. A.  I n t e r i m P h a s i c P e r i o d of Growth. 1.  The T e r m i n a l  P o i n t of an I n t e r i m phase i s a p e r i o d of a moderate  r a t e of growth, and the h i g h e s t weight a c h i e v e d w i t h i n the phase. It  i s c h a r a c t e r i z e d by a low f e e d i n t a k e , a v e r y h i g h plasma  'fat , 1  and a h i g h l e v e l of plasma c h o l e s t e r o l . I t i s of i n t e r e s t t o note t h a t a low f e e d i n t a k e l e v e l i s accompanied by a r a i s e d c h o l e s t e r o l v a l u e . s t a t i c r e l a t i o n s h i p e x i s t s , but t h e r e  That i s , a n e g a t i v e  i s no commensurate  relation-  s h i p i f the f e e d i n t a k e changes.  T h i s i m p l i e s t h a t not f e e d but  p o s s i b l y some b i o l o g i c a l a c t i v i t y  i s r e s p o n s i b l e f o r the r a i s e d  l e v e l of plasma c h o l e s t e r o l . When we c o n s i d e r e d  the f e e d - plasma ' f a t ' r e l a t i o n s h i p  e a r l i e r we assumed because one source was  reasonable  of energy was  low t h a t i t  f o r the o t h e r t o be h i g h but no s i m i l a r assumption  can be made with cholesterol because i t s function i s not f u l l y understood, therefore i t s r e l a t i o n s h i p to the other parameters remains obscure. There i s a p o s s i b i l i t y that the high plasma cholesterol values are as a result of a raised estrogen l e v e l at this time, for i t i s known that estrogens w i l l r a i s e the plasma cholesterol (18, 20, 37, 87, 99), and also that they have an e s s e n t i a l f a t t y acid sparing e f f e c t (18).  There have been reports that cholester-  o l functions to carry unsaturated f a t t y acids (35) so that i t s r i s e at t h i s time may be simply for that function. If the estrogens are the reason for the r i s e i n cholesterol we can assume that the terminal point of the Interim phase i s a period of f o l l i c u l a r a c t i v i t y i n the ovary, as i t i s often during t h i s period that the sexually mature c y c l i n g female gains weight (70).  Also high estrogen levels induce a lower feed  intake (65, 58, 59). . In the male the weight-lipid relationships are similar at t h i s time, but the feed intake i s r a i s e d . The Onset Point of the Interim Phase. This point i s characterized by a low l e v e l of weight accretion;  a feed intake lowered from the previous  day; a  moderately high plasma ' f a t , but lower than the terminal point, 1  and a raised plasma cholesterol value, s i m i l a r to the range at the terminal point. The plasma ' f a t ' and plasma cholesterol values r i s e and i n p o s i t i v e accord with the feed intake.  together  The weight and  feed show a similar r e l a t i o n s h i p and so a l l parameters conform.  74. The point of onset of the Interim phase i n the male i s d i f f e r e n t , i n fact i t resembles the Main phasic points of junction of the female.  Thus supporting the assumption that the  Interim phases i n the male and female are as a result of d i f f e r ent b i o l o g i c a l a c t i v i t i e s . B.  The Main Phasic Period of Growth. 1.  The Main Phasic Points of Junction. The points of junction of the Main phasic period of growth are distinguishable from the points of junction of the Interim phase by the very low plasma ' f a t and the very high feed intake 1  levels.  The levels of both these parameters are higher at forty  days than at sixty days.  This i s also true of the plasma  cholesterols, which are on both days high r e l a t i v e to the plasma 'fat'.  A r a t i o which we have not seen previously. The feed intake and the plasma cholesterol are both high,  and as one changes the other conforms i n a l i k e manner. - At the terminal and onset points i n the interim phase the feed intake and plasma cholesterol show negative  correlation,  whereas, a change i n the l e v e l of feed intake has no effect upon cholesterol at the terminal point i t i s e f f e c t i v e at the onset point, showing a change similar to the Main phase points of junction.  Subsequently i t can be seen that a l l the parameters  at three different points of junction are d i s t i n c t i v e i n their relationships to one another. C.  Intermediate 1.  Points i n the Interim Phasic Periods of Growth.  Table XVII  One day post Onset Point or second day of Phase. The growth rate and the feed intake levels are s t i l l  low  on the second day of the Interim phase, but the plasma ' f a t  1  and plasma cholesterol are higher and lower, respectively, than on the f i r s t day. 2.  Two  This possibly suggests a lower estrogen e f f e c t .  days after the Onset of the Interim Phasic Period of Growth. Unfortunately no good r e s u l t s are available for these days  because of a series of accidents, such as unusually high temperatures and a day without adequate food supply. 3.  One day p r i o r to the Terminal Point.  Table XVII  This point i s d i s t i n c t i v e for i t s low plasma cholesterol levels;  low feed intake;  high plasma 'fats' and a moderately  high rate of weight gain. 4.  Two  days p r i o r to the Terminal Point.  Table XVII  This point occurs i n a f i v e day cycle three days a f t e r the onset of the Interim phase, but i n the longer phases i t s occurrence i s two days before termination.  A high rate of weight accretion  and low plasma cholesterol are c h a r a c t e r i s t i c .  The feed intake  ranges from low to high, and the plasma ' f a t ' fluctuates with i t , however, there appears to be no c o r r e l a t i o n of the feed intake with the plasma c h o l e s t e r o l . VI The E f f e c t s of Heat on the different Parameters. Tables XVIII From July 30th to August 1st, 1962,  the temperature i s exceedingly hot,  as a result the a i r conditioning i s not adequate i n the area i n which the animals are housed, and the temperature r i s e s to 80° and 82° F. The female rats Nos. 1, 2 and 3 are bled on the s i x t y - t h i r d day when the temperature i s 80° F.  The plasma 'fats' are found to be high and the  plasma cholesterols very low.  Unfortunately no twenty-four hour records of  weight gain and feed intake are available.  76, On the sixty-fourth and s i x t y - f i f t h day of age (August 1st, 1962) the temperature i s 82° F. f o r the second day i n succession.  A l l the animals  show a weight loss from the previous day, i n the range of 1.7 to 3.2 grams. Weight also i s lost on the day p r i o r to t h i s . Two animals (Nos. 1 and 2) show a loss of 1.7 and 1.8 grams, respectively, and a high plasma ' f a t ' l e v e l ;  a low feed intake and plasma c h o l e s t e r o l .  Whereas, the two animals (Nos. 7 and 3) which lose 2.5 and 3.2 grams, respectively, show low plasma 'fat' l e v e l s ;  a high feed intake but a low  plasma cholesterol value. The plasma ' f a t s ' show a negative s t a t i c relationship with the feed intake as i s so often observed.  However, the levels of plasma cholesterol  are not r e l a t i v e to the feed intake, but appear to be correlated with weight loss.  Formerly, i t i s seen that low values of plasma cholesterol are  associated with a fast rate of growth.  This implies that fast weight change,  whether p o s i t i v e or negative, tends to be associated with low plasma cholestecl levels. VII The E f f e c t s of Inanition on the Different Parameters.  Table XVIII  For a two day period the animals are inadvertently  given feed s u f f i c i e n t  for only a one day period, but an adequate water supply. A l l the animals consume more feed i n two days than i s usual f o r one day, but the amount f a l l s short of the two day supply.  No twenty-four hour weight  accretion levels are recorded, but only one animal does not lose weight at the end of forty-eight hours.  See F i g s . 8 .- 12 where the point i n question  i s marked with an 'x . 1  The most s t r i k i n g effect of inanition i s the exceedingly low plasma cholesterol l e v e l s .  It i s surprising that the plasma ' f a t ' levels are not  higher under the circumstances.  However, i t i s interesting to note that from  77.  • '  this time u n t i l f o r t y - s i x days of age the feed intake and the plasma ' f a t s show a continuous  rise.  1  Again we see than on some occasions the female  gains high plasma ' f a t ' levels from the feed intake;  a female t r a i t .  VIII F/C Ratio. We have seen that d i s t i n c t i v e conditions occur at the three major points of junction, that i s both i n the Main phasic period of growth and the Interim phases. F/C  This d i v i s i o n i s further exemplified by the use of the  ratio.  PHASIC PERIOD  TIME  RANGE OF F/C RATIOS ACTUAL VALUES RELATIVE VALUES  Main  40 days 60 days  0.85 - 1.1 0.35 - 1.4.  1.47 - 1.94 0.62 - 2.45  Interim  Onset Terminal  1.2 - , 1.4 1.6 '- 1.9  2.05 - 2.45 2.74 - 3.26  Onset + 1 day  1.6 - 1.9 -  2.74 - 3.26  Terminal - 1 day Terminal - 2 days  1.6 - 2.0 1.7 - 2.1  2.74 - 3.49 2.90 - 3.71  In the males i t i s the marked changes in plasma cholesterol which emphasize the differences that occur at each point of junction.  Whereas  in the female i t i s the levels of feed intake and plasma ' f a t ' .  For  example, the low F/C r a t i o s which occur at the Main phasic periods are as a result of the low plasma'fat'rather  than an unduly high cholesterol value.  The F/C r a t i o s reveal that as the ' f a t ' values f a l l the cholesterol values r i s e and v i c e versa, so that neither parameter remains s t a t i c while the other changes. When we attempt to correlate the F/C r a t i o s with the other parameters the results are often generalizations that are not applicable to a l l e x i s t i n g instances.  Therefore, we f i n d that i n the female rat the F/C r a t i o i s useful  only as an indication or 'hint at' the b i o l o g i c a l a c t i v i t y taking place, but  78:, not as a consistent indicator of a l l the i n t e r r e l a t i o n s h i p s of the parameters measured.  This i s believed to be so because of the variable nature of the  female estrus cycles. A.  Main Phasic Period of Growth. 1.  The Main Growth Phase points of Junction. At these points the range of F/C r a t i o s are 0.85 (actual) at 38 and 39 days and 0.35 day period.  - 1.4  -  1.1  (actual) at the 60  This implies both a low plasma 'fat' and a raised  plasma c h o l e s t e r o l . This i s true of the forty day period, but at 60 days the very low plasma 'fats' are the more s i g n i f i c a n t as the cholesterol i s high only i n contrast. Because of this difference and the similar results obtained at f i f t y - o n e days we assume that, unlike the male, these points are concerned with the growth of the animal and not with development.  Nevertheless,  sex  some reservations are held on t h i s  point because i t i s soon after the forty day period that cycling occurs, i f only i r r e g u l a r l y .  It i s conceivable, therefore, that  sex development i s concerned, and the closure of the epiphyseal plates i n the long bones, caused by the high estrogen  levels,  diminishes the rate of growth. The high feed intake on these occasions, however, i s not usual with high estrogen a c t i v i t y . (65) and Kochakain (59) who  That i s according to Leathern  have demonstrated a drop i n feed  intake when estrogens are injected into mice and r a t s . B.  Interim Phasic Periods of Growth. The F/C ratios, found i n the Interim phases are never as low as  79* those seen at the points of junction of the Main phasic periods. the onset of the phase the actual values are low, one day to 1.6 - 1.9  (actual);  to the terminal point, 1.7 - 2.1 point to 1.6 - 1.9  (actual).  1.2 - 1.4;  At  rise in  r i s e s t i l l higher about two days p r i o r (actual), and f a l l again at the terminal  This pattern of F/C r a t i o s r e f l e c t s the  different rates of weight accretion throughout the interim phase, and gives some indication of the l i p i d i n t e r r e l a t i o n s h i p , but not of the feed intake. The smaller Interim phases appear to be superimposed upon the inherent growth pattern, but there i s no doubt that these two are i n t e r e f f e c t i v e .  patterns  For i t i s r. well known that a speyed immature  female w i l l grow to a greater size than her intact  counterpart.  TABLE X LEVELS OF WEIGHT ACCRETION; PLASMA LIPIDS; FEED INTAKE.  Weight * Changes (GM.)  Plasma Fat mgm7o  Feed Intake GMS/lOOgms of B.W. +  Plasma Cholesterol mgm7o  FEMALE RAT. NO. 1. F/C Ratio Actual Relative  Age (Days)  - 1.7  239.0  6.8  127.0  1.8  3.1  65  - 0.5  234.0  6.6  130.0  1.8  3.1  74  + 1.8  236.0  7.9  152.0  1.6  2.74  53  + 2.4  266.0  8.0  148.0  1.8  3.10  67  + 2.6  218.0  10.6  182.0  1.2  2.05  44  + 2.6  162.0  10.0  180.0  0.9  1558  51  + 3.9  55100  8.7  143.0  0.35  0.62  60 (hot)  + 4.9  167.0  12.7  176.0  0.95  1.65  39  + 5.3  281.0  10.8  179.0  1.60  2.74  46  + 5.5  242.0  6.7  146.0  1.70  2.90  72  -  223.0  -  117.0  1.90  3.2  63 (hot)  207.0  -  132.0  1.60  2.74  245.0  13.6 (2 Days)  107.0  2.3  75 42  +  B. W. = Body Weight. Weight l o s t or gained i n twenty-four hours.  TABLE XI LEVELS OF WEIGHT ACCRETION; PLASMA LIPIDS; FEED INTAKE.  Weight * Changes (GM.)  Plasma Fat mgm7o  Feed Intake GMS/lOOgms of B.W. +  Plasma Cholesterol mgm%  FEMALE RAT. NO. 2 F/C Ratio Actual Relative  Age (Days)  - 1.8  289.0  3.9  133.0  2.2  3.75  65  - 1.2  198.0  7.5  140.0  1.4  2.45  67  + 0.2  66.0  9.6  138.0  0.48  0.83  60 (hot)  + 0.4  222.5  7.2  141.0  1.6  2.74  74  + 1.0  164.0  11.5  148.0  1.1  1.94  51  + 1.7  263.8  9.7  146.2  1.8  3.15  46  + 3.5  166.0  14.12  156.0  1.1  1.94  39  + 4.2  225.0  10.0  174.0  1.3  2.26  53  + 5.9  221.0  12.5  119.5  1.9  3.20  44  + 6.1  191.0  6.5  159.0  1.2  2.05  72  89.0  2.5  -  -  289.0  117.0  2.5  —  (Fasted 42(for 2 (Days 63-(hot)  -  207.0  132.0  1.6  2.74  77  223.0  * +  13.6  Weight gained or l o s t i n twenty-four hours. B.W. = Body weight.  TABLE XII LEVELS OF WEIGHT ACCRETION; PLASMA LIPIDS; FEED INTAKE. Weight * Changes (GM.)  Plasma Fat mgm7o  Feed Intake GMS/lOOgms of B.W. +  FEMALE RAT. NO. 3.  F7C  Plasma Cholesterol mgm7o  Ratio  Age (Days)  Actual  Relative  1.4  2.45  74  1-3 .  2.45  67  113.3  0.96  1.65  65  - 0.4  175.0  7.07  127.0  - 0.8  164.0  6.63  129.5  - 3.2  108.7  + 1.3  206.0  9.0  122.0  1.7  2.90  44  + 1.5  149.0  9.46  106.0  1.4  2.45  60 (hot)  +11.8  257.0  13.10  120.3  2.1  3.71  53  + 2.4  257.0  5.86  127.0  2.0  3.49  72  .+ 3.0  225.0  8.70  141.2  .1.6  2.74  46  + 4.0  157.5  12.5  185.5  0.85  1.47  39  + 5.5  138.0  13.15(spill?)  135.0  1.0  1.74  51  - 0  189.0  20.0(in 2 days)  115.0  1.6  2.74  42  -  265.8  83.0  3.2  143.0  1.1  (spill)  -  125.0  *  Weight gained or l o s t i n twenty-four hours.  +  B. W. = Body weight.  :  -  63(hot) 82°F 77  TABLE XIII LEVELS OF WEIGHT ACCRETION; PLASMA LIPIDS; FEED INTAKE. Weight * Changes (GM.)  Plasma Fat mgm7o  Feed Intake GMS/lOOgms of B.W. +  Plasma Choles terol mgm7o  FEMALE RAT. NO. 6.  F/C Ratio Actual Relative  Age (Days)  - 6.7  289.0  -  116.7  2.5  - 3:6  126.0  8.4  130.0  0.99  1.71  64  + 0.9  209.0  8.18  166.4  1.3  2.26  52  + 2.1  172.0  174.0  0.99  1.71  50  . + 2.8  177.0  124.0  1.4  2.45  59 (hot)  + 3.8  238.0  10.2  120.0  2.0  3.49  43  + 4.4  237.0  8.4  127.0  1.9  3.26  73  + 4.9  256.5  10.7  145.5  1.8  3.15  45  + 6.6  215.0  6.9  137.0  1.6  2.74  71  + 7.6  124.0  10.0  173.0  0.72  1.25  38  -  225.0  12.3 (2 Days)  71.0  3.2  117.5  1.7  2.90  62 (hot)  148.5  1.9  3.26  76  196.5  10.0 9.24  -  271.0  *  Weights gained or l o s t i n twenty-four hours.  +  B.W. = Body weight.  66  .  41  TABLE XIV LEVELS OF WEIGHT ACCRETION; PLASMA LIPIDS; FEED INTAKE.  Weight * Changes (GM.)  Plasma Fat mgm7o  Feed Intake GMS/lOOgms of B.W. +  Plasma Cholesterol mgm7o  FEMALE RAT. NO. 7. F/C Ratio Actual Relative  Age (Days)  138.0  1.2  2.05  64  148.5  1.2  2.05  50  8.61  168.0  1.4  2.45  52  230.0  8.43  173.0  1.3  2.26  45  + 0.6  241.0  11.92  128.0  1.9  3.26  43  + 1.8  237.0  9.36  127.0  1.9  3.26  66  + 2.4  206.0  9.5  102.0  2.0  3.49  73  + 3.5  262.0  9.4  141.0  1.9  3.26  59 (hot)  + 6.2  180.0  14.68  197.0  0.88  1.58  38  + 6.8  205.0  6.58  118.0  1.7  2.90  71  _  191.0  89.0  2.2  3.75  41  -  222.5  -  123.0  1.8  3.15  62 (hot)  —  174.0  —  121.0  1.4  2.45  76  - 2.5  159.0  - 1.8  173.5  - 1.5  244.0  + 0.4  9.91 11.0  11.9 (2 days)  *  Weight gained or l o s t i n twenty-four hours.  +  B. W. = Body weight.  TABLE XV WEIGHT CHANGES:  PLASMA LIPIDS AND FEED INTAKE LEVEL OF THE FEMALE (PREPUBERTAL) RATS. AT JUNCTION OF TWO MAIN PHASIC PERIODS OF GROWTH  Feed Intake GMS/100 GMS of Body Weight  Plasma Cholesterol mgm7.  167.0 166.0 157.0 124.0 180.0  12.7 14.1 12.5 10.0 14.7  176.0 156.0 185.5 173.0 197.0  0.98 1.1 0.85 0.72 0.88  124-780  10-14.7  156-197  0.72-1.1  + 3.9 + 0.2 + 1.5  51.0 66.0 149.0  8.7 9.6 9.5  143.0 138.0 106.0  0.35 0.48 1.40  0.2-3.9  51-149  8.7-9.6  + 2.6 + 1.0 + 5.5  162.0 164.0 138.0  10.0 11.5 13.2  1.0-5.5  138-164  + 2.1 - 1.8  172.0 173.0  -1.8-4-2.1  172-173  Weight * Changes GMS + + + + +  4.9 3.5 4>'0 7.6 6.2  3.5-7.6  Range  Range  i  Plasma Fat mgm%  10.0-13.2 10.0 11.0 10-11  106-143  F/C Ratio Actual Relative  0.35-1.4  180.0 148.0 135.0  0.9 1.1 1.0  135-180  0.9-1.1  174.0 148.0  0.99 1.2  148,174  0.99-1.2  Weight gained or l o s t i n twenty-four hours prior to blood l e t t i n g .  1.65 1.94 1.47 1.25 1.58  Age in Days 39 (1) 39 (2) 39 (3) 38 (6) 38 (7)  1.25-1.94 0.62 0.83 2.45  60 (1) 60 (2) 60 (3)  0.62-2.45 1.58 1.94 1.74  51 (1). . ... 51 ( 2 ) 51 (3)  .  L l t t C r r l  1.58-1.94 1.71 2.05 1.71-2.05  50 ( 7L i) t t e r 2 5 0  ( 6 )  L l t t e r  1  TABLE XVI WEIGHT CHANGES:  PLASMA LIPIDS AND FEED INTAKE LEVEL OF THE FEMALE (PREPUBERTAL) RATS. A.  Weight Gained or Lost + + + + + + + + +  1.7 1.8 2.4 3.0 3.5 4.9 5.3 5.5 6.6  1.7-6.6  («)•> Plasma Fat  AT TERMINAL POINTS OF INTERIM PHASE OF GROWTH  Feed Intake GMS/100GMS of Body Weight•  263.0 237.0 . 266.0 225.0 262.0 256.0 281.0 242.0 215.0  9.7 9.3 8.0 8.7 9.4 10.7 10.8 6.7 6.9  215-281  6.7-10.8  (mg%) Plasma Cholesterol  F/C Ratio Actual Relative  146.2 127.0 148.0 141.0 141.0 145.0 179.0 146.0 137.0  1.8 1.9 1.8 1.6 1.9 1.8 1.6 1.7 1.6  127-179  1.6-1.9  3.15 3.26 3.15 2.74 3.26 3.15 2.74 2.90 2.74  Age in Days 46 (2) 66 (7) 67 (1) 46 (3) 59 (7) 45 (6) 46 (1) 72 (1) 71 (6)  2.74-3.26  • B. AT ONSET POINTS OF INTERIM PHASE OF GROWTH 1.5 1.2 0.8 0.4 0.4 0.9 2.6 2.8 4.2  244.0 198.0 164.0 209.0 175.0 230.0 218.0 177.0 225.0  -1.5H-4.2  .164-244  + + + + +  8.6 7.5 6.6 ' 8.2 7.1 8.4 10.6 9.3 10.0 7.1-10.6  168.0 140.0 129.5 166.4 127.0 173.0 182.0 124.0 174.0  1.4 1.4 1.3 1.3 1.4 1.3 1.2 1.4 1.3  2.45 2.45 2.26 2.26 2.45 2.26 2.05 2.45 2.26  127-182  1.2-4.4  2.05-2.45  52 (7) 67 (2) 67 (3) 52 (6) 74 (3) 45 (7) 44 (1) 59 (6) 53 (2)  TABLE XVII WEIGHT CHANGES:  PLASMA LIPIDS AND FEED INTAKE LEVEL OF THE FEMALE (PREPUBERTAL) RATS,  AT THE INTERMEDIATE POINTS IN THE INTERIM PHASE OF THE GROWTH CURVE. A. Weight Gained or Lost + + +  0.5 0.4 .1.8 4.4  i -0.5-4-4.4 1 t  (mg7o) Plasma Fat 234.0 222.5 236.0 237.0 223-237  Feed Intake GMS/100GMS of Body Weight 6.6 7.2 7.9 8.4 6.6-8.4  B. + 0.6 + 2.4 + 5.9  •+ 2.4  241.0 257.0 221.0 207.0 196.0 222.5 206.0  0.6-5.9  196-257  -  ONE DAY POST ONSET (mg7») Plasma Cholesterol  F/C Ratio Actual Relative  130.0 141.0 152.0 127.0  1.8 1.6 1.6 1.9  127-152  1.6-1.9  3.15 2.74 2.74 3.26  Age in Days 74 74 53 73  (1) (2) (1) (6)  43 72 44 77 62 62 73  (7) (3) (2) (1) (6) (7) (7)  44 43 71 53  (3) (6) (7) (3)  2.74-3.26  ONE DAY PRIOR TO TERMINAL POINT  9.5  128.0 127.0 119.5 132.0 117.5 123.5 102.0  1.9 2.0 1.8 1.6 1.7 1.8 2.0  3.26 3.49 3.26 2.74 2.90 3.15 3.49  5.9-12.5  102-132  1.6-2.0  2.74-3.49  11.9 5.9 12.5  -  1 C. + 1.3 + 3.8 + 6.8 +11.8 i 1.3-11.8 I  206.0 238.0 205.0 257.0 205-257  TWO DAYS PRIOR TO THE TERMINAL POINT  9.0 10.2 6.58 13.1 6.6-13.1  122.0 120.0 118.0 120.0  1.7 2.0 1.7 2.1  2.90 3.49 2.90 3.71  118-122  1.7-2.1  2.90-3.71  TABLE XVIII WEIGHT CHANGES:  PLASMA LIPIDS AND FEED INTAKE LEVEL OF THE FEMALE (PREPUBERTAL) RATS.  AT DIFFERENT POINTS IN THE INTERIM PHASE OF THE GROWTH CURVE. D. Weight Gained or Lost -  1.7 1.8 2.5 3.2  ! -1.7to-3.2  ANIMALS SUBJECTED TO HEAT (82°F) FOR THREE SUCCESSIVE.DAYS Feed Intake GMS/100GMS of Body Weight  (mg7o) Plasma Cholesterol  239.0 289.0 159.0 108.7  6.8 3.9 9.9 9.9  127.0 133.0 138.0 113.0  1.8 2.2 1.2 0.96  109-289  3.9-9.9  143-138  0.96-2.2  (mg%) Plasma Fat  E. 245.0 223.0 189.0 225.0 191.0  13.6/2 13.6/2 20 12 12.3/2 11.9/2  | -0.2-f4.8  189-245  11.9/2-20/2  48 Hour weight accretion.  3.15 3.75 2.05 1.65  Age in Days 65 65 64 65  (1) (2) (7) (3)  42 42 42 41 41  (1) (2) (3) (6) (7)  1.65-3.75  ANIMALS* SUBJECTED TO INANITION FOR TWENTY-FOUR HOURS  + 4.8 - 0.2 0.0 + 4.5 + 1.9  1  F/C Ratio Actual Relative  107.0 89.0 115.0 71.0 89.0 71-115  2.3 2.5 1.6 3.2 2.2 1.6-2.5  RESULTS AND DISCUSSION  THE MALE SEXUALLY MATURE RAT  90. SEXUALLY MATURE MALE RATS. The r e l a t i o n s h i p of weight accretion, actual feed intake i n grams per animal and the plasma l i p i d s (mgm %) for three male rats .(Nos..3, 7, 8) f o r eight consecutive days are shown i n F i g s . 15 - 17. Numerical representation i s found i n Table XIX.  In these the plasma l i p i d levels are expressed as  gm/100 gms of plasma and the feed intake as gm/100 gms of body weight. Similar representations for f i v e control animals (Nos. 1, 2, 4 and 5) are found i n F i g s . 19 - 20 and Table XXI.  The l a t t e r animals had already been  used i n the s e l f - a c c e l e r a t i n g growth tests, and are l i t t e r mates to the above test  animals. Fig.  18 and Table XX relegate a l l parameters to s p e c i f i c groups  believed to be indicative of the l e v e l of c i r c u l a t i n g gonadal hormone. The male Test animals are tested at the same time as their female l i t t e r mates, but unlike them their growth has not yet 'plateaued'.  However, there  is no doubt that these males are sexually mature at 84 to 92 days of age. I t i s at this age i n the UBC colony that they are used for breeding. I  Body Weight In the eight day test period, the pattern of weight gain of the Test  animals appears to have plateaued,, F i g s . 16 - 17, but the Control animals (Figs. 19 - 20) continue to show weight gain.  This suggests that the effects  of the daily bleeding procedure interferes with the growth of the animal. Because of t h i s , the growth pattern of the Interim phasic type of growth i s obscured i n the Test animals, but can be seen i n the Controls. I t i s possible that the act of bleeding the animals also causes a daily fluctuation of weight, especially evident i n r a t number 7.  However,  Test animal number 3 shows a weight gain pattern almost i d e n t i c a l to Control animal number 2, and so no d e f i n i t e conclusion can be made.  .4  21  20  Sfl  2-2  1-1  15  0-3  RATIO:  F/C  C5*  300  3 . RAT.  P L A S M A FAT  200 F  cn 2  100 P L A S M A CHOLESTEROL  vp!3 0 cn  \  ID  2  / \j.  110  • / \  'V  /  o  r/  3 20 BODY  W E IGHT  31 5^ Z5 CO 15  310  10  3 0 5 J 1 S ,  N>-' 6  <  '  85  2  3 87  ^o"'  A  5 89  F E E D  6  7 91  6  , N T A K E  D A Y S *f T E S T D A Y S *f A G E  S E X U A L L Y M A T U R E M A L E RAT SHOWING UPID;WEIGHT AND FEED INTAKE CHANGES IN 8 DAYS. FIGURE: 1 5 .  1$  I-JB  2S  2.0  2-5  1$  ir.6  1:1  RATIO-  F/t  (5V  RAT-  300 PLASMA  FAT'  200  if)  2  100  20  W  1  0  i  0  A  PLASMA CHOLESTEROL  \  \ \  2 80  325 BODY  84  86  88  90  WEIGHT  D A Y S °f  S E X U A L L Y M A T U R E M A L E RAT.SHOWING LIPIDiWEIGHT AND FEED INTAKE CHANGES IN 8 DAYS. FIGURE: 16.  AGE  23  V£  17  26  3^  K9  24  1-8  R A T I O F/Z  d a RAT  PLASMA 120  A  GO 110 2  \ / \ /  100  350  /  /  \  \/  b'  90  \  /  V/ of  25 BODY  <3  CHOLESTEROL  3^5 U o L  WEIGHT  FEED INTAKE  340 2 84  3 86  4  5  6  88  7 90  §  DAYS°f T E S T D A Y S *f A G E  S E X U A L L Y M A T U R E MALE RAT.SHOWING LIPID;WEIGHT ANO FEED INTAKE CHANGES IN 8 DAYS. FIGURE: 1 7 .  SEXUALLY MATURE MALE RATS: THE CHRONOLOGICAL OCCURRENCE OF THE'GROUPS'  I  2  3 4 5 6 7 8 DAYS OF TEST. FIG. 18..  FIGURf:l9. S E X U A L L Y MATURE M A L E RATS: CONTROLS.  320 29  5 1 9 20 CO CO  o  310 19  303J10 UTTER  M A T E S OF M A L E RAT  3.  330^ CQ CO  2 o  323i  •-BODY WEIGHT.  320  O - F E E D INTAKE  8 DAYS.  FIGURE: 2 0 .  eft  SEXUALLY MATURE MALE RATS: CONTROLS. 325  320 20 CQ  1/5  2 O  JO  3134 13  3I0JI0  ^ 7  OD U. O  330,  LITTER MATES OF MALE RATS 7 & 8.  o t/5'  325 25 DQ  UJ  CO  2 o  Z  320 20  V/ ^  o  3j5Jl5 UJ  g  i  0 i  1  1 2 84  »  '  B.W. ©-BODY WEIGHT. 0-FEED INTAKE. "  3 4 88 86  90  8 DAYS. AGE  97. II Relationship of Body Weight and Feed Intake A.  Weight Gain.  Group 1 and 2.  Table XX  On a l l occasions of testing, except two  (No. 3, 23/8/62;  No.  7,  21/8/62) a p o s i t i v e gain i n weight i s accompanied by a p o s i t i v e feed intake. In the two exceptions, the feed intake i s lowered i n the twentyfour hours during the period of weight gain.  This r e l a t i o n s h i p i s one  that i s often encountered i n the self-accelerating growth period (SAGP), and so i t s occurrence  i n rats number 3 and 7 suggests that they are  s t i l l at t h i s stage of growth at the beginning of the test period, i e . at days 3 and 1, respectively.  Its occurrence,  from time to time, i s  also evident i n the control rats which continue to show a high rate of weight gain.  There i s a p o s s i b i l i t y that a weight-feed r e l a t i o n s h i p  of t h i s nature i s s p e c i f i c to the composition The p o s i t i v e weight-feed  of the gain being made.  r e l a t i o n s h i p observed on the majority  of occasions i s similar to that seen at the terminal point of the Interim phase i n the self-accelerating growth period, but beyond this parameters) the s i m i l a r i t y B.  Weight Loss.  (two  ceases.  Group 3 and  4.  On nearly a l l occasions of weight loss the feed intake i s lowered concomitantly.  Two  instances occur, however, when weight i s lost re-  gardless of an increase i n the feed intake (No. 3, 27/8/62; 25/8/62).  No.  t h i s suggests an increase i n the metabolism of the animal  accompanied by an inadequate exogenous energy source to maintain weight. III  Relationship of Body Weight, Feed Intake and Plasma 'Fat*.  Weight Gain  8,  98, There are four occasions of weight gain with an increase i n feed i n take which are accompanied by an increase i n plasma ' f a t ' as well (Group 1). On a l l other occasions of weight gain the plasma ' f a t ' i s decreased . A.  (Group 2).  Weight Gain and a Rise i n Plasma 'Fat*.  Group 1. 1.  High Feed Intake The four occasions, mentioned above, are found only i n  animals number 3 (22/8/62; 23/8/62).  24/8/62) and number 8 (21/8/62;  The positive weight-feed-'fat  1  relationship of the  three parameters implies that the exogenous source of energy i s not adequate for the weight being made, and so an endogenous source of energy i s required and mobilized. indicative of the composition  It may  also be  of the gain that i s being made.  That i s to say i f the gain i s protein one would expect the plasma 'fat' to r i s e , but i f i t i s fat being gained one would expect a lowered plasma ' f a t * . 2.  Two  animals (No. 3, 23/8/62;  No. 7, 21/8/62) show an  increase i n weight and have a low feed intake, unlike '1', an increase i n plasma ' f a t ' .  and  A pattern usually seen i n SAGP.  This condition has been described by Raben and Hollenberg  (89)  to occur under the influence of STH, where the animal appears to maintain growth by an endogenous energy source.  In t h i s way i t  avoids the excess energy cost of ingestion and absorption which would result from large quantities of food (48). B.  Weight Gain and a Decreased Plasma 'Fat*.  Group 2. Of the two groups, this i s the most common condition found i n  99. rats of 84 - 92 days of age. I t i s a combination of a weight gain and a low plasma ' f a t accompanied by an increase i n the feed intake. 1  the low plasma ' f a t ' implies that there i s an increased uptake of fat by the depots or possibly a decreased  output,  the former, suggests  a weight gain as f a t , whereas the l a t t e r i s indicative of a more ' s t a t i c ' condition i n the fat compartments and growth progressing from the increased exogenous energy source. C.  Weight Loss and an Increased Plasma 'Fat*.  Group 3. In Table XX i t can be seen that i n the majority of cases when there i s a loss of weight there i s an accompanying drop i n feed intake. The high plasma ' f a t ' associated with t h i s i s i n d i c a t i v e of weight being lost as f a t .  This ' f a t provides energy for the maintenance metabolism 1  (30, 41, 42, 46, 50) of the animal. On the days p r i o r to and a f t e r the above instances there i s always weight gain accompanied by a low plasma ' f a t ' and an increased feed intake (Group 2). - We had suggested that this indicates weight gain i n the fat compartments. There i s a p o s s i b i l i t y that the weight loss as fat i s stimulated by high testosterone levels, and the weight gain as fat i s a recuperative measure after the high levels of testosterone subside.  The  findings of Kochakain (60) corroborate t h i s , for he was able to demonstrate that weight i s lost as fat after injections of excess levels of testosterone propionate.  Also that when the i n j e c t i o n s ceased the  animals gained weight as fat and increased t h e i r feed intake. Very recently, 1963, Laron and Kowadlo (63) showed that testosterone has a fat-mobilizing e f f e c t .  lOJO, r  If our hypothesis  i s correct high testosterone values ( i n Group  3) appear to occur almost on every alternate day ( F i g . 18). such as this may  A pattern  be r e f l e c t i n g the p i t u i t a r y gonadal hormone feedback  mechanism. The two occasions (No. 8, 25/8/62; intake i s r a i s e d instead of lowered may  No. 3, 27/8/62) when the feed  be indicative of the amount of  testosterone present. . Weight Loss and a Decreased Plasma ' F a t . 1  Group 4. A loss of weight and a decreased plasma ' f a t ' i s associated with a low feed intake i n t h i s group.  This condition appears to be  diate between the low and high testosterone groups.  interme-  The low ' f a t  1  suggesting that testosterone levels are not as high as i n Group 3, but the low feed intake indicating that i t i s s u f f i c i e n t l y high to diminish the 'appetite' (60). IV Total Plasma Cholesterol. Plasma cholesterol levels are low compared to the female at the same age.  This corroborates the findings of many other investigators (18, 27,  38, 71, 99). A.  The range of values i s from 83 mgm  7 - 125 mgm 0  37,  %.  .Weight Gain and a Rise i n Plasma 'Fat . 1  Group 1. 1.  The p o s i t i v e weight-feed-'fat'  r e l a t i o n s h i p of t h i s Group  i s associated with a r i s e i n the plasma cholesterol and resembles the conditions at the terminal point i n the Interim phase. values ranging from 107 mgm 100 gm of plasma.  "L to 111 mgm  "k or 0.104  - 0.108  However, the r i s e i s not considered  by our method except i n animal number 8 (23/8/62).  The gm/  significant  10 L. On two occasions, i n one animal (No, 3), the plasma cholesterol levels f a l l even though there i s a raised feed intake. 2.  The increase i n weight and plasma ' f a t accompanied by a 1  low feed intake also shows a r i s e i n plasma cholesterol, a l b e i t ins i g n i f icant. It i s believed that this i s a low testosterone period intermediate between the low testosterone Group 2 and the higher testosterone Group 4. B,  This i s represented i n F i g . 18.  Weight Gain and a Low Plasma 'Fat.  Group 2. The increase i n weight concomitant with low plasma ' f a t ' i s accompanied by both an increase i n feed intake and plasma c h o l e s t e r o l . Its occurrence i s invariably after Group 3 and 4 as seen i n F i g . 18. As suggested e a r l i e r , this i s a period of weight gain i n the f a t compartments of the body, and possibly of low levels -of c i r c u l a t i n g testosterone.  F i l l i o s (39) demonstrated that when sexually mature male  rats were castrated the plasma cholesterol levels r i s e .  This i s also  found to be true i n the human male (51, 52, 54, 75). Further, Kochakain (60) found that body weight and feed intake increase when the excess levels of testosterone are lowered.  These findings appear to corrobor-  ate the assumption that Group 2 i s a period of low testosterone a c t i v i t y . C.  Weight Loss and an Increased Plasma 'Fat*.  Group 3. During t h i s period of weight loss and a high plasma ' f a t ' the feed intake and plasma cholesterol are decreased. The range of the l a t t e r i s 0.090 - 0.096 gm/100 gms of plasma, except on one occasion  102, when the feed intake r i s e s i n animal number 3 i t i s 0.114 gm/100 gms of plasma. We have designated this a period where weight loss i s from the fat compartment, and have suggested that the weight- fat'-feed r e l a 1  tionship i s as a result of high testosterone l e v e l s . cholesterol further strengthens  our assumption.  The low plasma  For i t i s now known  that injections of testosterone propionate of p h y s i o l o g i c a l strength given to female rats or castrated males lowers the plasma cholesterol (3, 19, 32). Therefore i t appears that a l l four parameters under observation are i n t h i s period affected by high testosterone levels (60, 63, 3, 19,.32). It i s interesting to note that the above conditions described are similar to the points of junction of the Main phasic period of growth described i n this thesis i n the section on the prepubertal male, rat. D.  Weight Loss and a Decreased Plasma 'Fatj  Group 4. - As seen i n Table XX this intermediate group exhibits both a r i s e and a decrease i n plasma c h o l e s t e r o l . The l a t t e r i s a s i g n i f i c a n t decrease and the former i s i n s i g n i f i c a n t .  Both levels are accompanied  by a decrease i n the feed intake. 1.  Low Plasma c h o l e s t e r o l .  gm/100 gms of plasma.  The range here i s 0.081 - 0.092  The relationships of weight, feed intake  and plasma cholesterol are similar to those seen i n Group 3 which are attributed to high testosterone l e v e l s .  Only the l e v e l of  plasma ' f a t ' i s different and t h i s i s believed to result from the l e v e l of c i r c u l a t i n g testosterone not having reached the same  103, l e v e l as Group 3.  See F i g . 18, animal number 8, on the fourth  day of t e s t i n g . 2.  High plasma c h o l e s t e r o l .  The range of plasma cholesterol  i s 0.106 - 0.119 gm/100 gms of plasma.  Irregardless of the feed  intake being low from the previous day the plasma cholesterol rises, albiet insignificantly. cantly high.  However, the levels are s i g n i f i -  These high levels of plasma cholesterol accompanied  by low growth, low feed intake and low plasma ' f a t are a l l 1  indicative of a low metabolic rate.  The cause of which may be  r e l a t i v e to the l e v e l of c i r c u l a t i n g testosterone i n this group believed to be moderately low, V  intermediate between Group 1 and 3.  Total Plasma Cholesterol and Feed Intake. It appears that the l e v e l of feed intake and plasma cholesterol  fluctuate i n p o s i t i v e accord i n most instances, but negative c o r r e l a t i o n does occur i n a l l the groups except 3 described above.  Therefore there i s no  conclusive evidence on the e f f e c t s of the l e v e l of feed intake on plasma cholesterol. VI  The Chronological Occurrence of Each Group. In F i g . 18, the chronological order of occurrence  can be seen for each Test animal.  Their occurrence  of the four groups  appears to beaconsistent,  for instance Group 3, a low cholesterol period i s usually flanked by the high cholesterol Group 2. Group 2 and 3 are put at the lowest and highest points on the graph, because t h e i r parameters seem to exemplify the e f f e c t s of low and high levels of testosterone, whereas Group 1 and 4 appear to be indicative of a waning or gaining a c t i v i t y of the gonadal hormone.  This l a t t e r assumption i s made  because of the variable results i n the plasma l i p i d s i n these two groups.  104. There' i s a p o s s i b i l i t y that these points on the graph i n F i g . 18 give an indication of the testosterone l e v e l c i r c u l a t i n g at the time designated. They may,  however, only be indicative of a d a i l y energy exchange that i s  associated with growth and maintenance.  TABLE XIX SEXUALLY MATURE (MALE) RATS. SHOWING LEVELS OF BODY WEIGHT, PLASMA LIPIDS AND FEED INTAKE ON EIGHT CONSECUTIVE DAYS.  Date 20/8/62 21/8/62 22/8/62 23/8/62 24/8/62 25/8/62 26/8/62 27/8/62 28/8/62  Day of Test  1 2 3 4 5 6 7 8  (GMS) 3ody BodyhWeight CT3 tf 8 tf 7 314.8 316.8 320.0 317.5 321.0 318.5 322.0 319.7 321.0  309.9 307.3 310.0 312.2 313.7 311.. 0 317.0 313.4 319.0  341.5 350.4 344.5 34777 346.3 344.3 346.0 345.5 349.0  tf 3  Plasma L i p i d Levels GMS/100GMS Plasma "Fat" Choleste r o l CT 8 0" 7 0* 8 CT 7 6 3  Feed Intake GMS/100GMS Body Weight tf 3 tf 8 Cf 7  Ratio: F/C Cf 3 Cf 7 tf 8  .170 .220 .232 .298 .234 .130 .173 .105  .196 .172 .237 .236 .237 .147 .132 .094  6.6 5.67 6.80 6.10 6.56 6.10 6.63 6.70 6.52  1.4 2.1 2.0 2.8 2.2 1.1 1.5 0.9  .250 .150 .274 .201 .321 .204 .214 .208  .122 .107 .115 .104 .106 .122 .114 .122  .108 .092 .108 .119 .091 .110 .090 .114  .107 .099 .096 .119 .096 .100 .081 .090  5.90 5.87 5.88 5.35 5.80 5.77 6.02 5.67 5.95  6.00 6.85 5.45 6.23 5.37 6.04 6.04 5.45 6.01  CHANGES OCCURING IN BODY WEIGHT LIPIDS AND FEED INTAKE LEVELS FOR EIGHT CONSECUTIVE DAYS. 21/8/62 22/8/62 23/8/62 24/8/62 25/8/62 26/8/62 27/8/62 28/8/62  1 2 3 4 5 6 7 8  -2.6 +2.7 +2.2 +1.5 —2.7 +6.0 -3.6 +6.4  +2.0 +3.2 -2.5 +3.5 -2.5 +3.5 -2.3 +1.3  +8.9 -5.9 +3.2 -1.4 -220 +1.7 -0.5 +3.5  +  +  + .050 -.024 -.100 + .012 + .065 + .124 + .066 -.001 -.073 -.064 + .001 + .101 -.104 -.090 -.108 + .043 -.015 + .010 -.068 -.038 -.006  + -.015 -.008 + .008 -.003 -.011 + .013 + .002 -.013 + .016 + .004 -.008 -.019 + .008 + .009  + -.016 + .016 + .011 -.028 + .019 -.020 + .024  -0.93 +1.13 -0.70 +0.46 -0.46 +0.53 +0.07 =0.18  -0.03 +0.01 -0.53 +0.45 -0.03 +0.25 -0.35 +0.28  -0.85 -1.4 +0.78 -0.86 +0.67 0 -0.59 +0.56  1.8 1.8 2.5 2.0 2.5 1.5 1.6 1.1  2.3 1.6  2.6 1.7 3.5 1.9 2.4 1.8  TABLE XX SEXUALLY MATURE MALE RATS. SHOWING WEIGHT GAINED OR LOST AND THE ACCOMPANYING PLASMA LIPID AND FEED INTAKE CHANGES, ACCORDING TO "GROUP" ARRANGEMENT. Weight Gained or Lost  Feed Intake GMS/100GMS of B.W. *  GROUP 1 + 1.5 + 2.0 + 2.2 + 2.7 + 3.2 + 8.9  + + + +  GROUP 2 + 1.3 + 1.7 + 3.2 + 3.5 + 3.5 + 3.5 + 6.0 + 6.4  B.W.  N.B. -  =  +  24/8/62 21/8/62 23/8/62 22/8/62 23/8/62 21/8/62  3 7 3 3 8 8  0.00 0.01 0.45 0.25 0.56 0.53 0.18  -0.038 -0.108 -0.024 -0.001 -0.090 -0.006 -0.104 -0.068  +0.009 +0.019 -0.008 +0.013 +0.004 +0.024 +0.016 +0.008  28/8/62 26/8/62 22/8/62 24/8/62 26/8/62 28/8/62 26/8/62 28/8/62  7 8 7 7 7 8 3 3  - 0.59 +00567 - 0.03 - 0.53 + 0.07  +0.010 +0.101 +0.001 +0.065 +0.043  -0.020 -0.028 -0.013 -0.003 -0.008  27/8/62 25/8/62 25/8/62 23/8/62 27/8/62  8 8 7 7 3  -  -0.073 -0.015 -0.064 -0.100  +0.011 -0.019 +0.002 -0.016  24/8/62 27/8/62 25/8/62 22/8/62  8 7 3 8  -  GROUP 4 - 1.4 - 2.3 - 2.7 - 5.9  +  -0.011  Animal No.  +0.008 -0.015 +0.016  + + + + +  - 316  +0.066  Date  +0.012 +0.050 +0.124  + 0.28  GROUP 3 - 0.5 - 2.0 - 2.5 - 2.5  *  0.46 0.03 0.70 1.13 0.78 0.85  Plasma GM/100GMS of Plasma Fat Cholesterol  0.86 0.35 0.46 1.40  +  Body weight  A s i g n i f i c a n t change i n l i p i d l e v e l  0.012 gm/lOOgm of plasma.  TABLE XXI SEXUALLY MATURE MALE RATS. CONTROL ANIMALS SHOWING ACTUAL AND COMPARED VALUES FOR WEIGHT, AND FEED INTAKE DURING THE EIGHT CONSECUTIVE DAYS THE TEST ANIMALS WERE OBSERVED. Date Date  21/8/62 22/8/62 23/8/62 24/8/62 25/8/62 26/8/62 27/8/62 28/8/62  Body Weight GMS  Feed Intake GMS/100GMS of B.W.  (f 1  0*2  A  6 5  6 1  0*2  328.9 331.0 326.7 320.0 333.5 335.5 333.8 344.0  326.5 328.5 328.5 330.7 331.8 335.5 337.0 338.0  315.3 320.3 318.8 320.8 326.0 329.7 326.7 329.5  309.0 312.8 312.5 321.8 320.0 320.0 324.0 324.5  6.5 7.5 4.45 6.63 6.3 3.66 7.06 6.50  -0.2 +1.0 -3.05 +2.18 -0.33 -2.64 +3.40 -0.56  •6*4  0^5  6.76 6.74 5.60 7.16 6.26 3.85 6.76 6.24  5.6 6.6 5.8 6.6 6.3 6.7 6.7 6.1  5.1 6.2 5.3 6.5 5.9 5.7 6.0 6.5  +0.62 -0.02 -1.14 +1.56 -0.90 -2.41 +2.91 -0.52  -0.8 +1.0 -0.8 +0.8 -0.3 +0.4 0.0 -0.6  -1.2 +1.1 -0.9 +1.2 -0.6 -0.2 +0.3 +0.5  DAILY COMPARISONS 21/8/62 22/8/62 23/8/62 24/8/62 25/8/62 26/8/62 27/8/62 28/8/62  ++1.9 + 2.1 - 4.3 - 6.7 +13.5 + 2.0 - 1.7 +10.2  + + + + + + .+  5.1 2.0 0.0 2.2 1.1 3.7 1.5 1.0  +  -  + + +  -  +  1.4 5.0 1.5 2.0 5.2 3.7 3.0 2.8  +  -  + + + +  1.2 3.8 0.3 8.3 1.8 0.0 4.0 0.5  RESULTS AND DISCUSSION  THE FEMALE SEXUALLY MATURE RAT  109. SEXUALLY MATURE FEMALE RATS In Figs. 21 - 25 the weight, feed intake and plasma l i p i d s levels for 10 consecutive days (83 - 93) i n each animal are shown g r a p h i c a l l y . These same results are recorded numerically i n Tables XXII - XXIII.  In the l a t t e r  the values are arranged within the separate phases of the estrus cycle. Figs. 26 and 27 show graphically the weight and feed intake values of f i v e Control animals for the same period of time.  Tables XXIV and XXV  cite  the above values i n each Control animal within the divisions of the estrus cycle. In Fig.. 28 the average levels of the four separate parameters are represented graphically for the f i v e Test  animals.  When the s e l f - a c c e l e r a t i n g growth period was under observation types of phasic growth were d i f f e r e n t i a t e d .  two  The Main phasic type of growth,  described by Cowan et a l (21) i n the deer and Brody (13) i n the r a t , and a smaller phasic type of growth;  the Interim phasic growth periods.  I t was  suggested that the l a t t e r were as a r e s u l t of the changes wrought by the estrus c y c l i n g . In order to determine whether this assumption i s true, f i v e mature female rats are bled and vaginal smears are made for ten consecutive days. Testing begins only after the body weights have 'plateaued*, that i s when the s e l f - a c c e l e r a t i n g period of growth terminates. Unfortunately the estrus c y c l i c phases are not well established before we s t a r t testing.  However, from the weight change data i t appears to be  either a four or f i v e day cycle depending on the individual r a t . The true early estrus, or time of mating, i s believed to occur i n the very early morning (33) i n our r a t s , and because smears are not made on animals u n t i l 12:30  p.m.  the t y p i c a l estrus smear i s not encountered.  these Instead  $-4>RAT-  8?  '  86  '  88 *! 9 0 F I G . 21 .  '  92  '  S E X U A L L Y M A T U R E F E M A L E RATSHOWING LIPID.WEIGHT.AND FEED INTAKE CHANGESJN 10 DAYS.  DAYS O F A G E .  . 100  170  A / \  8l50 2  /  PLASMA CHOLESTEROL  \  ,  D  \  130  \  1 20' RATIO-  >8  22  1-8  22  23  1-3  30  1*7  20  15  200 BODY WEIGHT  If) (3 195  15  -O 190  .10J  ° ^ « r v - ~  o—\—2  3  $6  FEED INTAKE  a  4—5 88  90  92  10 DAYS°f T E S T DAYS *f A G E  F I G ^ S E X U A L L M A T U R E F E M A L E RAT, SHOWING LIPID,WEIGHT AND FEED INTAKE CHANGES INIO DAYS.  (J)  8 RAT  P L A S M A FAT.  3O0  g200 2 100 PLASMA  I70r  CHOLESTEROL.  q /  150  / \-  o\  ui CD  \  2 130  I  \  /  \  10 1-8  RATIO:  1-8  2-2  3-1  31  24  17  OS  M  V6  205  BODY W E I G H T .  200  FEED I NTAKE.  00  2  CD  195 to A  7  83  85  87  89  8  9 91  FIG.23. S E X U A L L Y M A T U R E F E M A L E RAT.SHOWING LIPID,WEJGHT AND FEED INTAKE CHANGES IN 10 DAYS-  10 D A Y S ° i T E S T D A Y S "f A G E  9  400  9-RAT.  P L A S M A FAT  300 00  '200  100' I 701 PLASMA ..^HOLESTEROL  A  s  /  I 30  .•a  • I 20  RATIO:  21  25  26  35  23  25  13  H  13  ^*BODYWEIGHT.  21 Oi GO  ^205'  15  J  P\  FEED INTAKE. x  \  \  /  /  \  b  2 0 0 10 0'  o—\—W—3—4 83  85  ~%  6  "87  1 89  8  9  TO DAYS "(TEST  91  FIG. 24. S E X U A L L Y M A T U R E F E M A L E RAT.SHOWING LIPID;WEIGHT AND FEED INTAKE CHANGES IN10 DAYS.  DAYS *f AGE.  300  00200 2  100 PLASMA CHOLESTEROL 190 P  / 170  /  \  \  1  GO  /i  ^150  130  ! !  h—J  J  26  RATIO:  17  23  29  22  12  12  1€  14  K>  BODY W E I G H T  210  FEED INTAKE  ^205-1  A..  ID  200J  Q  I &  3  2^v/ 8 5  A 8  3  6 '  FIG  25.  7  7 8~9"  6 "  5 9l  TQ  S E X U A L L Y M A T U R E F E M A L E RAT SHOWING LIPIDlWEIGHT AND FEED INTAKE CHANGES IN K> DAYS  DAYS^TEST DAYS'? A G E  FIGURE:27. SEXUALLY MATURE FEMALE RATS:  CONTROLS.  215-120 X  >o o 210 15 CQ  205-1 lOJ » AGE IN DAYS X  230-1  vy  x o  2 E 2254  8 \ E  v5  2  220 t— x °  UJ  9  215 15  o co 2I0.J|0.J * 4  DAY OF TEST  6  8  10  FIGURE: 28. MATURE FEMALE RATS-' SHOWING A GENERAL PATTERN OF WEIGHT ACCRETION ; FEED INTAKE ; PLASMA Ll PIDS, DURING A COMPLETE ESTRUS CYCLE. plasma cholesterol.  o.2 1 0  / to > o _  feed intake.  * 8.  < O»2  y  %  "Ov  O  VBody weight. R-proestTus E:esrrus EMrearly metestrus. M.-met estrus  2  D.-diesrrus  -3H E / EM  Ml  M2  DI  D2  ESTRUS CYCLIC PHASES  118. we see the metestrus period i n i t s very early stages. simply as Early Metestrus 1.  This we  designated  Also the l a t t e r part of this phase i s seen, ie  Late Estrus, when the t y p i c a l cheesy, clumped e p i t h e l i a l c e l l picture i s e v i dent.  This i s designated Metestrus  1.  Metestrus 2 i s recognized by the pre-  sence of large numbers (+4) of leucocytes, and the diestrus phase by fewer leucocytes, mucus and polygonal c e l l s .  Proestrus i s only encountered  on a  possible four occasions, for i t i s believed to commence i n the early afternoon twelve hours before ovulation, subsequently  12:30  p.m.  et a l (33) found his colony began proestrus at 1:30  i s too early.  - 2:30  Everette  p.m.  The actual bleeding of the animals i s not carried out u n t i l 5:30 which i s f i v e hours after the smear i s made.  p.m.,  I f the timing of the estrus  cycle suggested by Nalbandov (82) and Everette (3) i s applicable to our rats i t means that the blood picture f a l l s within the phase after the one which i s designated.  Proestrus i s the exception.  Because we do not know the exact  time of ovulation i n our colony we have tabulated the results without acknowledging this possible discrepancy of timing. A l l body weights and feed intake levels are accumulated i n the twentyfour hours before the phase indicated. Fluctuations of Weight, Plasma Lipids and Feed Intake Relative to the Separate Phases of the Estrus Cycle. A.  Body Weight. In Table XXII the actual body weights i n grams for each animal i s recorded.  In Table XXIII the amount of weight gain or loss i s  arranged according to the c y c l i c phase i n which the weight i s taken. I  Early Metestrus  1 Phase.  Table XXIII  The animals accumulate weight during this period except on one occasion (No. 8, 21/8/62) when the loss i s n e g l i g i b l e (0.3  gms).  119 » II  Metestrus 1  (or late Estrus) Phase.  A l l the eight occasions i n which Metestrus 1 i s encountered weight i s l o s t .  In one instance as much as seven grams. Similar  weight loss i s seen i n the control animals. III  Metestrus 2 Phase. Metestrus 2 i s a period of weight gain f o r both the Test and Control animals.  However, there are two occasions i n which  weight i s lost (No. 10, 27/8/62;  No. 9, 23/8/62).  See Table  XXIII. This phase occurs shortly after ovulation.  The ovaries  contain corpora lutea and small f o l l i c l e s ,  and the u t e r i  diminish i n v a s c u l a r i t y and c o n t r a c t i l i t y .  According to  Nalbandov (83) the corpora lutea last only six hours i n the unfm-ated female r a t . fourteen hours long IV  However, the actual phase i s ten to (109).  Diestrus. In t h i s phase of the estrus cycle, of 2 to 2% days duration, there is.both weight gain and weight loss.  However, neither  condition occurs on two successive days so that weight gain i s followed by weight loss and vice versa.  One exception i s seen  in animal number 9 on 28/8/62 and 29/8/62 (Table XXII). 1.  F i r s t Day of Diestrus The f i r s t day of diestrus follows the Metestrus 2 phase and of the f i v e Test animals as many gain weight as lose i t . for  The order i n which this occurs i s not constant  any one animal.  For instance rat number 4 on 24/8/62  gains weight on the f i r s t day of diestrus whereas,, on  120. 29/8/62 i t loses weight.  Rat number 5 exhibits the opposite  pattern. The control animals a l l show weight loss on the f i r s t day.  The difference that occurs between the Test and  Control animal suggests that the bleeding process i s interfering, but as the occurrence of the estrus phases appear regular we assume that they are not affected. 2.  Second Day of Diestrus Weight i s both gained and lost on the second day similar to the f i r s t , but the majority of animals show a weight gain.  The l a t t e r i s also true of the Control group.  In the diestrus phase there i s a functional regression o f the corpora lutea, hence a drop i n the progesterone l e v e l occurs.  The u t e r i are small and only s l i g h t l y  c o n t r a c t i l e (109), and the vaginal mucosa i s t h i n . V  Proestrus. Both the Test and Control animals show weight gain at the proestrus phase of the sex cycle. This phase 'heralds  1  the next heat and i s characterized by  preovulatory swelling of the f o l l i c l e s under the influence of LH. Body Weight and Feed Intake. I  Early Metestrus 1. Although weight i s gained during this phase the feed intake shows no d e f i n i t e pattern as can be seen i n Table XXIII.  How-  ever, the general pattern (Fig 28) shows the feed intake to be decreased per unit of weight from the Estrus and Proestrus  levels.  The variations i n feed intake may be a result of the time a f t e r  124. estrus (ovulation) that the measurement i s made.  That i s , the  feed intake decreases as time progresses. II  Metestrus 1. (Late Estrus) The lowered body weights at this time are accompanied by low feed intakes r e l a t i v e to body weight.  Two  exceptions occur i n  animals number 5 and 8 when the feed intake increases, but a loss in weight occurs regardless.  The Gontrol animals show a similar  relationship. III  Metestrus 2. The body weight increases but the feed intake levels continue to f a l l per unit of body weight.  This suggests a low  metabolic rate i n these animals during Metestrus 2. There are three animals that did not gain weight.  Of  these, animal number 9 (23/8/62) exhibits a very low feed intake for two consecutive days and loses weight.  The other two  animals,  number 5 (28/8/62) and 10 (27/8/62), also ingested very low levels of feed i n the two days, but remain at the same weight. The Control animals are generally similar to the Test animals at this time, but there are more occasions when the feed intake i s increased and no occasions of weight l o s s . IV  Diestrus Phase. 1.  Diestrus 1. The Control animals on the f i r s t day of diestrus show a commensurate drop i n the feed levels and the weight. The Test animals, as mentioned above, show as many occasions of weight loss as gain.  However, the majority  show a r i s e i n feed intake, but i t appears that weight i s  122, lost because the r i s e is i n s u f f i c i e n t to ameliorate the effects of the low levels on the previous day. 2.  Diestrus 2. Both the majority of Test and Control animals show' positive weight gain on this day, but the l a t t e r have a high feed intake,  whereas,  cases a low feed intake.  the Test group have in most  The same d i s s i m i l a r i t y i n the  two groups of animals exists on the f i r s t day of diestrus, but i n none of the other phases.  We do not know why this  occurs unless i t is as a result of no bleeding procedure. The weight gain, accompanied by a low feed intake, when compared to the previous day, is often accomplished because of the low metabolic rate of the animal. Proestrus. The often high weight of accretion in the proestrus phase is accompanied by a r i s e in feed intake per unit of body weight. The Control animals are similar i n a l l  respects.  Estrus. We had stated previously that this phase was not encountered in our studies.  However, rat number 5 on 26/8/62 has a smear  which is t y p i c a l of early metestrus 1, but a very high drop i n body weight  (7.5 gms) makes i t a t y p i c a l .  that i t i s early estrus.  Therefore we assume  A t y p i c a l early metestrus 1 smear  occurs on the following day. The loss of weight is accompanied by an increase i n feed intake.  This l a t t e r level is higher than at any other time  during the 10 day testing period i n animal number 5.  The estrus phase i s associated with a heightened  running  a c t i v i t y on the part of the rat caused by an increase i n the l e v e l of estrogen (83).  It i s reasonable  to assume therefore  that the increase i n a c t i v i t y raises the c a l o r i c requirements of the animal, and also contributes to the loss of weight. Unfortunately there are no other occasions to which t h i s observation can be compared. Body Weight, Feed Intake and Plasma 'Fat'. I  Early Metestrus 1. The majority of instances within t h i s phase show weight gain and a r i s e i n feed intake r e l a t i v e to the body weight. The plasma 'fat', however, does not show a constant r e l a t i o n s h i p . This i s unavoidable,  as t h i s phase occurs mostly on the f i r s t  day of testing and i t cannot be compared with values of the previous day.  In the three instances when we can make a compari-  son the phases on the previous day are not s i m i l a r .  For instance  animals number 4 (26/8/62) and 10 (26/8/62) show a decrease i n the l e v e l of plasma ' f a t ' at.early metestrus 1 when the comparison i s made with the diestrus phase, whereas animal number 5 (27/8/62) shows a r i s e i n plasma 'fat' when the comparison i s made with the proestrus phase.  Also, the feed intake i s decreased  this l a t t e r animal and the weight gain i s high.  in  This suggests  a drop i n the metabolic rate from proestrus and further implies that although the feed intake has dropped i t i s s t i l l adequate to support growth, as one i s no longer feeding for the a c t i v i t y increment stimulated by the estrus condition.  The increase i n  plasma ' f a t ' suggests that weight i s gained i n other than the  124. fat compartments of the body. Metestrus 1. Metestrus 1 i s a period of weight loss, lowered feed intake and low plasma ' f a t . 1  The low feed intake and plasma ' f a t ' suggest a lowered metabolic rate.  The lowered plasma ' f a t ' and the decrease i n  weight, which i s sometimes 3.8% of the t o t a l body weight (No. 4, 22/8/62), implies that the loss i s a result of f l u i d and tissue of the post ovulation state rather than f a t .  Weight  loss can be caused by low feed intake but the plasma ' f a t ' i s expected to r i s e . The concept of a low metabolic  rate corroborates the  findings of Soliman and Reinike (101),, and also Brown-Grant (14).  The l a t t e r , demonstrated by means of radio-active iodine  131 (  I) that the thyroid gland was most active at estrus and  f e l l at metestrus.  Farbman (34) and others (69, 71) also re-  ported lowered thyroid a c t i v i t y following a period of high estrogen secretion or administration. There are three occasions pattern i s not observed.  i n this phase when the above  Instead the plasma.'fat'  i s raised  s i g n i f i c a n t l y along with the loss of weight, and there i s either no change or a marked decrease (No. 9, 22/8/62) i n the feed intake from the previous day. This implies that, either there can be a loss of weight as fat i n this phase i f the feed intake drops, or that the time of testing on these three i s e a r l i e r than that of the other  five.  occasions  125. Metestrus 2. A period when weight i s gained, but the feed intake and plasma 'fat' are lowered f o r the second consecutive day. According to Brown-Grant (14) the l e v e l of metabolism decreases u n t i l diestrus, therefore we can assume that the feed intake at t h i s time i s s u f f i c i e n t to cause the animals to be i n positive energy balance.  The low plasma 'fat* may not  only be indicative of the low metabolism but also of an i n creased uptake of fat by the fat compartment.  Thus weight i s  gained as f a t . One animal, number 9 (23/8/62), shows a weight loss along with the low feed intake, and a r i s e i n plasma ' f a t . 1  This i s  the same pattern as noted i n the Metestrus 1 phase for the same animal, which suggests that i t i s an i n d i v i d u a l condition brought about by some cause other than the phase of the sex cycle. Diestrus Phase. 1.  Diestrus 1. The feed intake at t h i s time i s generally raised regardless of weight being lost or gained, but i s highest in the l a t t e r .  The plasma 'fat' shows a r i s e i n both  instances, but i s highest when weight i s l o s t .  With  reference to the l a t t e r animals, t h i s implies that weight i s lost as fat because of i n s u f f i c i e n t c a l o r i e s being ingested to support maintenance.  Whereas, i n the weight  gain group the animals are i n positive energy balance and the r i s e i n plasma ' f a t suggests that the weight gain i s 1  126. other than f a t . Two animals, numbers 8 and 10 (23/8/62) lose 1.3 and 2.5 grams, respectively, and have a lower feed intake concomitant with the usual r i s e i n plasma for  1  fat'.  Except  the low feed intake these animals are similar to the  weight loss group described above. It appears that i n this phase the metabolism of the animals i s raised, but some have not yet increased their feed intake adequately to support maintenance consequently, lose weight.  and,  Other animals have adjusted  adequately to the increased metabolism.  This suggests  that we tested at two different times i n the phase and the general pattern seen i n F i g . 28 i s more usual f o r the f i r s t day of diestrus. Brown-Grant (14) had suggested that the metabolism of rats was lowest at diestrus, but i t does not appear to be the same i n our animals. 2.  Diestrus 2. Weight gain i n this part of the phase i s accompanied by a f a l l i n feed intake i n most instances and a r i s e i n plasma ' f a t . 1  The picture i s similar to the weight gain  group of the f i r s t day of diestrus. The loss of weight in t h i s part of the phase d i f f e r s from the f i r s t day of diestrus. f a l l s and the plasma  That i s , the feed intake  ' f a t ' i s lowered, except i n animal  number 9 when the loss i n weight i s excessive and the plasma ' f a t ' i s high.  As the loss of weight i s not very  127. great i t appears that the feed intake i s adequate to support maintenance but not a gain i n weight.  The  general pattern suggests a decrease i n metabolism which i s more i n keeping with the r e s u l t s of Brown-Grant (14). V  Proestrus. A period when a l l three parameters are increased.  The  r i s e i n plasma ' f a t ' and feed intake are i n keeping both with the increase i n body weight and the r i s e i n the metabolic rate reported to exist at this phase. D  Plasma Cholesterol i n Relationship to other Parameters. I  Early Metestrus 1. As seen i n Table XXIII a high plasma cholesterol i n t h i s phase i s not accompanied by either a constant change or feed intake.  l e v e l of weight  I t , therefore, appears that i t s l e v e l i s  not dependent upon these a c t i v i t i e s , but possibly upon those factors that control the phase of the sex cycle, namely the hormones. Estrogens have been reported by many investigators to cause a r i s e i n plasma cholesterol (18, 34, 37, 39, 99). E s t r a d i o l benzoate, when injected, has been shown to cause weight to be gained as fat (59, 70), and when i n excess to cause weight to be lost as f a t , and to cause the feed intake to drop (59).  Therefore,  the r i s e i n cholesterol as well as the inconsistent results on weight change and feed intake may be contributed by the action of estrogen and i t s l e v e l of concentration. II  Metestrus 1. A period of weight loss, decreased  feed intake, and i n the  128. majority of instances a lower plasma fat and cholesterol than on the previous day.  However, the drop i n cholesterol i s only  s i g n i f i c a n t on two occasions (No. 4, 28/8/62;  No. 8, 22/8/62),  the rest sustain the higher value of the previous day.  This i s  i n keeping with the theory of low thyroid a c t i v i t y described by Brown-Grant (14) to occur at Metestrus 1. The lower cholesterol values i n animals number 4 and 8 corroborate the low levels reported by F i l l i o s (39) to occur i n metestrus when the f o l l i c u l a r a c t i v i t y subsides.  However, i t  i s possible that the l a t t e r instances occur later i n the phase than the others.  For instance, number 8 shows a diestrus vaginal  condition on the following day. Metestrus 2 Phase. This i s a period when weight i s gained as f a t and the metabolism of the animal appears to be low.  This l a t t e r i s  exemplified by weight being gained when the feed intake l e v e l i s lower than i n a period of weight loss.  Further, to corroborate  t h i s condition, the plasma cholesterol values are l i t t l e lower than i n the Metestrus 1, or s i g n i f i c a n t l y raised i n two i n stances, i n number 8, 28/8/62 and i n number 9 28/8/62. Diestrus Phase. 1.  Diestrus 1. The weight gain group of animals a l l show a r i s e i n feed intake and plasma ' f a t but an i n s i g n i f i c a n t change 1  in the plasma cholesterol, with the exception of animal number 9 (29/8/62) which shows a marked increase.  Whereas,  'fat' and feed intake appear to herald an increase i n  129. metabolism i t i s not r e f l e c t e d i n the plasma c h o l e s t e r o l . In the weight loss group nearly a l l animals show the r i s e i n feed intake, and a l l show a r i s e i n plasma ' f a t ' t y p i c a l of this phase.  On three of f i v e occasions  there  is a s i g n i f i c a n t f a l l i n the plasma cholesterol l e v e l as reported by F i l l i o s  (39) to be lowest at diestrus.  These  results also suggest a r i s e i n metabolism (91) and a decrease i n f o l l i c u l a r a c t i v i t y (37). 2.  Diestrus 2. The weight gain group of animals are similar i n most respects to the weight gain group on the f i r s t day of diestrus.  With one exception, the feed intake f a l l s i n  three of the s i x examples c i t e d . The weight loss group on the second day of diestrus decrease their feed intake and plasma ' f a t , whereas, the 1  plasma cholesterol increases on three of the four occasions cited. As stated e a r l i e r , t h i s period of the phase appears to show a decrease i n metabolism and no doubt precedes i n time the weight gain group of the second day of diestrus. This l a t t e r assumption i s made because nearly a l l the Control animals gain weight on the second day of diestrus as seen i n Table XXV. V  Proestrus. The picture presented i n t h i s phase i s similar to the weight gain groups on both the f i r s t and second days of diestrus. Except that the weight gain i s superior.  130. The plasma cholesterol levels are s t i l l r e l a t i v e l y low as the bleeding procedure i s carried out at the very early stages of this phase.  According to F i l l i o s (39) the levels r i s e  toward the onset of estrus, that i s as the preovulatory swelling of the f o l l i c l e procedes. VI  Estrus. As mentioned e a r l i e r the weight loss i s exceedingly high, reaching 7.5  grams.  This i s accompanied by a commensurate drop  i n plasma ' f a t ' from .291 gms  to .210 gms/100 gms  of plasma,  which implies that weight i s being lost as other than f a t . The plasma cholesterol, on the other hand, r i s e s from .126 .158  gms/100 gms  gm -  of plasma, which i s indicative of the high  c i r c u l a t i n g l e v e l of estrogen associated with estrus. VII  F/C Ratio. The F/C r a t i o s do not appear to have a s p e c i f i c l e v e l for each of the separate phases of the estrus cycle.  However, a  general pattern, calculated from the mean of the range for each phase, shows i t to r i s e from a low l e v e l at estrus to a peak at diestrus. reached.  The F/C r a t i o then f a l l s u n t i l proestrus i s  If t h i s i s compared with the Interim phase i n the  prepubertal female rat the equivalent pattern i s seen.  The  lowest F/C r a t i o i s at the onset of the phase and from here r i s e s u n t i l a peak i s reached two days p r i o r to the terminal point.  It then f a l l s u n t i l the terminal point i s reached.  Thus i t appears that the Interim phasic period of growth i n the prepubertal rat has a general F/C r a t i o pattern somewhat similar to that expressed  i n the estrus cycle of the mature female r a t .  131. If a further comparison of a l l l i p i d , weight change and feed intake levels i s made between the two patterns of growth we see that the terminal point i s equivalent to proestrus, and the onset point to early estrus.  Also late estrus coincides with  the day after the onset point, and diestrus with the two days p r i o r to the terminal point.  During active growth, i n t h i s  l a t t e r period, the feed intake i s much higher than i n the sexually mature animal. different composition  Possibly t h i s can be a t t r i b u t e d to the  of growth i n the prepubertal animal.  The Interim phasic type of growth began at approximately forty days i n our colony, therefore we can assume that the estrus cycles began also at t h i s time.  By means of the 'Copula-  tory response' test, i n other than our test animals we found that ovulation accompanies the f i r s t estrus cycles.  This  corroborates the findings of Blandau and Money (12) who established an average age of 49.4 - 5.46 days f o r the occurrence of the f i r s t heat.  TABLE XXII SEXUALLY MATURE FEMALE RATS. . SHOWING BODY WEIGHT,. PLASMA LIPIDS, FEED INTAKE, F/C RATIO AND PHASES OF ESTRUS CYCLE FOR TEN'CONSECUTIVE DAYS Plasma L i p i d Levels GM/100GMS Plasma Cholesterol "Fat"  (GMS) Body Weight  Date  Feed Intake GMS/100GMS of Body Weight  4*  5 *  10*  4*  5 *  10 *  7.40  1.9  1,8  2.6  Early Met.l  Early Met.l  Met.l  6.76  5.16  1.8  2.2  1.7  Met.l  Met.l  Met.l  6.72  6.34  4.43  1.7  1.8  2.3  Met. 2  Met.2  Met.2  .139  7.77  6.75  6.62  2.3  2.2  2.9  Diest. Diest. Diest.  .126  .140  6.45  5.92  6.40  2.1  2.3  2.2  Diest. Diest. Diest.  .158  .158  .170  7.42  7.43  6.65  1.3  1.3  1.2  Pro.  Early Met.l  Early Met.l?  .169  .141  .133  .147  7.42  6.76  5.92  1.8  2.0  1.2  Met.l  Met.l  Met. 2  .221  .231  .133  .131  .147  6.56  5.14  6.86  1.6  1.7  1.6  Met. 2  Met.2  Diest.  .149  .268  .228  .146  .133  .173  7.43  6.60  6.33  1.0  2.0  1.4  Diest, Diest. Diest.  .216  .195  .180  .141  .135  .178  7.04  5.30  6.57  1.5  1.5  1.0  Diest. Diest. Pro.  4-*  5 *  10*  6.00  6.94  6.76  .134  7.23  6.49  .135  .139  5.81  .126  .133  .129  .400  .118  .120  .291  .305  .134  .208  .210  .204  205.7  .246  .260  196.6  209.5  .211  189.9  197.0  206.8  193.4  196.5  208.1  10*  Ratio F/C  4 *  5 *  10*  4*  5*  4*  5 *  20/8/62  186.8  197.6  207.3  21/8/62  191.0  199.0  209.8  .240  .240  .352  .129  .137  22/8/62  184.0  198.0  205.5  .230  .291  .232  .126  23/8/62  187.6  198.7  203.0  .208  .240  .300  24/8/62  193.0  197.0  207.0  .271  .270  25/8/62  189.0  197.5  206.2  .278  26/8/62  190.0  190.0  206.2  27/8/62  189.2  196.6  28/8/62  193.3  29/8/62 30/8/62  10 *  Met. 1  =  Metestrus 1 or late estrus:  Met. 2  =  Metestrus 2  : .+ 3 or + 4 Leucocytes.  Diestrus  : Mucus, polygonal e p i t h e l i a l c e l l s , leucocytes (+1*)  Proestrus  : Polygonal e p i t h e l i a l c e l l s , nucleated squamous epith.  Diest. Pro.  .=  Phases of Estrus Cycle (Vaginal Smears)  + 4 clumped cornified epithelium.  TABLE XXII Cont'd.  Date  (GMS) Body Weight 9*  Plasma L i p i d Levels GM/100GMS Plasma Cholesterol "Fat" 8*  9?  8*  9*  Feed Intake GMS/100GMS of Body Weight 8^  F/C Ratio  9*  Phases of Estrus Cycle (Vaginal Smears)  9*  8 -?  9*  20/8/62  203.3  200.  21/8/62  203.0  208.0  .233  .250  .131  .122  5.42  7.89  1.8  2.1  Early Met.l  Pro  22/8/62  201.3  206.5  .206  .295  .115  .121  5.52  6.67  1.8  2.5  Met.l  Met.l  23/8/62  200.0  204.0  .276  .315  .128  .119  4.60  4.26  2.2  2.6  Diest.  Met.2  24/8/62  202.0  207.0  .349  .433  .114  .123  7.04  6.42  3.1  3.5  Diest.  Early Diest.  25/8/62  199.0  205.1  .338  .312  .109  .136  6.03  6.30  3.1  2.3  Diest.  Diest.  26/8/62  202.2  210.5  .294  .315  .122  .128  7.16  7.12  2.4  2.5  Pro.  Pro.  27/8/62  201.0  203.5  .206  .163  .124  .122  6.41  6.70  1.7  1.3  Met.l  Met.l  28/8/62  202.2  2.Q6.5  .143  .149  .158  .136  5.68  5.51  0.9  1.1  29/8/62  200.3  206.6  .150  .202  ,135  .155  6.35  6.73  1.1  1.3  Diest.  Diest.  30/8/62  203.3  208.0  .217  .272  .138  .152  6.65  5.92  1.6  1.8  Diest.  Diest.  i  6.35  Met. 2  TABLE XXIII SEXUALLY MATURE FEMALE RATS.  TEST ANIMALS.  . ACTUAL VALUES AND CHANGES OCCURRING IN PLASMA LIPIDS, BODY WEIGHT AND FEED INTAKE ARRANGED ACCORDING TO THE PHASE OF THE ESTRUS CYCLE. ESTRUS CYCLE PHASE: Date  Animal No.  A  Plasma Lipids gm/lOOgm Plasma Fat gm/lOOgm Cholesterol.9  + -  -.101  .129 .158 .137 .133 .131 .135 .170  .230 .246 .291 .206 .206 .295 .163 .232  -.010 + .038 + .051 -.027 -.088 + .048 -.152 -.120  .208 .211 .240 .221 .149 .315 .202 .169  -.022 -.035 -.051 -.039 -.063 + .020 -.014 -.035  21/8/62 26/8/62 21/8/62 27/8/62 21/8/62 21/8/62 26/8/62  4 4 5 5 8 10 10  .240 .208 .240 .260 .233 .352 .204  + -  Metestrus 1 (Late Estrus)  22/8/62 27/8/62 22/8/62 22/8/62 27/8/62 22/8/62 27/8/62 22/8/62  4 4 5 8 8 9 9 10  Metestrus 2 (Metestrus)  23/8/62 28/8/62 23/8/62 28/8/62 28/8/62 23/8/62 28/8/62 27/8/62  4 4 5 5 8 9 9 10  Metestrus 1 (Early Estrus?)  -.070  -  -  + .050  + +  Feed Intake/ 100 GMS Body Weight  Weight Gained or Lost  F/C Ratio  + .030  7.23 7.42 6.49 6.76 5.42 7.40 6.65  +1.23 +0.97 -0.45 -0.67 -0.93 +0.64 +0.25  +4.2 +1.0 +1.4 +6.6 -0.3 +2.5 +3.8  1.9 1.3 1.8 2.0 1.8 2.6 1.2  .126 .141 .135 .115 .124 .128 .122 .139  -.003 -.017 -.002 -.016 + .002 + .006 -.006 + .004  5.81 7.42 6.76 5.52 6.41 6.67 6.70 5.16  -1.42 0 +0.27 +0.10 -0.75 -1.22 -0.42 -2.24  -7.0 -0.8 -1.0 -1.7 -1.2 -1.5 -7.0 -4.3  1.8 1.8 2.2 1.8 1.7 2.5 1.2 1.7  .126 .133 .133 .131 .158 .119 .135 .147  0 -.008 -.002 -.002 + .034 -.002 + .014 -.023  6.72 6.56 6.34 5.14 5.51 4.26 6.73 5.92  +0.91 -0.86 -0.42 -1.62 -0.73 -2.41 -1.19 -0.73  +3.6 +4.1 +0.7 0 +1.2 -2.5 +3.0 -0.5  1.7 1.6 1.8 1.7 0.9? 2.6? 1.1 1.2  + .024  + -  -.025  + -  TABLE XXIII Cont'd. ESTRUS CYCLE PHASE Date  Animal No.  Plasma Lipids gm/100gm Plasma Fat gm/lOOgm Cholesterol. 9  Feed Intake/ 100 GMS Body Weight  Weight Gained or Lost  F/C Ratio  Diestrus 1  24/8/62 29/8/62 24/8/62 29/8/62 23/8/62 29/8/62 24/8/62 29/8/62 23/8/62 28/8/62  4 4 5 5 8 8 9 9 10 10  .271 .149 .270 .268 .276 .150 .433 .202 .300 .231  + .063. -.062 + .030 +.047 + .070 + .007 + .118 +.053 +0068 + .062  .118 .146 .120 .133 .128 .135 .123 .155 .129 .147  -.008 + .013 -.013 + .002 + .013 -.023 + .004 + .019 -.010 0  7.77 7.43 6.75 6.60 4.60 6.35 6.42 6.73 4.43 6.86  +1.05 +0.87 +0.41 +1.46 -0.92 +0.67 +2.16 +1.22 -0.73 +0.94  +5.4 -3.4 -1.7 +0.4 -113 -1.9 +3.0 +0.1 -2.5 +3.8  2.3 1.0 2.2 2.0 2.2 1.1 3.5 1.3 2.3 1.6  Diestrus 2  25/8/62 30/8/62 25/8/62 30/8/62 24/8/62 30/8/62 25/8/62 30/8/62 24/8/62 29/8/62  4 4 5 5 8 8 9 9 10 10  .278 .216 .291 .195 .349 .217 .312 .272 .400 .228  + .007 + .067 + .021 -.073 + .073 + .067 -.121 + .070 +.106 -.097  .134 .141 .126 .135 .114 .138 .136 .152 .139 .173  -.016 -.005 + .006 + .002 -.014 + .003 +.013 -.003 +.010 + .026  6.45 7.04 5.92 5.30 7.04 6.65 . 6.30 5.92 6.62 6.33  -1.32 -0.39 -0.83 -1.30 +2.44 +0.30 -0.12 -0.81 +2.19 -0.53  -6.0 +6.5 +0.5 -0.5 +2.0 +3.0 -1.9 +1.4 +4.0 -2.7  2.1 1.5 2.3 1.5 3.1 1.6 2.3 1.8 2.9 1.4  Diestrus 3 - Eg. Occurring on 3rd day  25/8/62 25/8/62  8 10  .338 .305  -.011 -.101  .109 .140  -.005 +.001  6.30 6.40  -1.01 -0.22  -3.0 -0.8  3.1 2.2  Proestrus  26/8/62 26/8/62 21/8/62 26/8/62 30/8/62  5 8 9 9 10  .210 .294 .250 .315 .217  -.081 -.044  .158 .122 .122 .128 .178  + .022 + .013  7.43 7.16 7.89 7.12 6.65  +1.51 +1.13  -  + .003 + .067  -  -.008 +.005  +  +0.82 +0.24  -7.5? +3.2 +8.0 +5.4 " +1.3  1.3 Estrui 2.4 2.1 2.5 1.0  TABLE XXIV SEXUALLY MATURE FEMALE RATS. CONTROL ANIMALS. SHOWING BODY WEIGHT.AND FEED INTAKE LEVELS FOR TEN CONSECUTIVE DAYS.  Date  S  1 20/8/62 21/8/62 22/8/62 23/8/62 24/8/62 25/8/62 26/8/62 27/8/62 28/8/62 29/8/62 30/8/62 Q„ + 2 20/8/62 21/8/62 22/8/62 23/8/62 24/8/62 25/8/62 26/8/62 27/8/62 28/8/62 29/8/62 30/8/62  20/8/62 21/8/62 22/8/62 23/8/62 24/8/62  (GMS) Body Weight  Weight Gained or Lost  Feed Intake /Animal  Feed Intake gm/lOOgms of Body Weight Actual Change  Phase of Estrus Cycle  203. 199. 206.5 204.5 205.5 205.5 208.5 207.0 209.0 208.0 211.0  -4.0 +7.5 -2.0 +1.0 +0.0 +3.0 -1.5 +2.0 -1.0 +3.0  13.0 12.3 16.5 14.5 13.5 14.7 15.3 14.3 13.4 12.7 15.3  6.40 6.18 7.99 7.09 6.57 7.15 7.33 6.90 6.40 6.10 7.25  -0.22 +1.81 -0Q90 -0.42 -+0.;58 +0.16 -0.43 -0.50 -0.30 +1.15  Diestrus Diestrus Proestrus Metestrus Metestrus Diestrus Diestrus Metestrus Metestrus Diestrus Diestrus  200.5 200.0 202.0 205.5 207.0 203.3 200.3 202.5 207.0 206.5 208.0  -0.5 +2.0 +3.5 +2.5 -3.7 +3.0 +2.2 +4.5 -0.5 +1.5  12.0 13.5 13.0 14.0 14.3 13.7 14.2 13.7 14.7 14.2 15.7  5.98 6.75 6.44 6.81 6.91 6.74 7.07 6.76 7.10 6.86 7.55  +0.77 -0.31 +0.37 +0.10 -0.17 +0.33 -0.31 +0.34 -0.14 +0.69  Metestrus Metestrus Diestrus Proestrus Metestrus Metestrus Diestrus Proestrus Metestrus Metestrus  191.0 196.0 197.5 195.5 194.3  +5.0 +1.5 -2.0 -1.2  14.8 13.5 17.3 12.7 15.4  7.75 6.88 8.75 6.50 7.92  -0.87 +1.87 =2.25 +1.42  Diestrus ? ?  1  1. 2. 1. 2.  1. 2. or Estrus 1. 2.  1. 2.  TABLE XXIV Cont'd.  Date $ 3 Cont'd. 25/8/62 26/8/62 27/8/62 28/8/62 29/8/62 30/8/62 .* 6 20/8/62 21/8/62 22/8/62 23/8/62 24/8/62 25/8/62 26/8/62 27/8/62 28/8/62 29/8/62 30/8/62 o + 7 20/8/62 21/8/62 22/8/62 23/8/62 24/8/62 25/8/62 26/8/62 27/8/62 28/8/62 29/8/62 30/8/62  (GMS) Body Weight  Weight Gained or Lost  186.5 191.3 186.7 194.5 189.5 191.5  -7.8 +4.8 -4.6 +7.8 -5.0 +2.0  210.3 222.0 220.5 219.0 225.0 219.7 227.0 227.5 231.5 225.0 230.5  +12.0 + 1.5 -1155 + 6.0 - 5.3 + 7.3 + 0.5 + 4.0 - 6.5 + 5.5  209.0 207.0 211.0 211.5 204.5 210.0 212.5 212.5 212.0 212.0 225.0  + + + +  2.0 4.0 0.5 7.0 5.5 2.5 0 - 0.5 + 0.0 +13.0  Feed Intake /Animal  Feed Intake gm/lOOgms of Body Weight Actual Change  9.9 10.1 12.6 1448 11.2 11.0  5.30 5.27 6.75 7.60 5.91 5.75  15.0 18.8 15.5 16.5 13.0 17.4 18.7 17.2 12.7 17.0  7.10 8.46 7.01 7*01 7.33 5.93 7.66 8.22 7.44 5.65 7.36  15.0  7.18  16.5 12.7  7.81 6.00  16.0 16.8 16.3 14.2 15.3 16.3  7.62 7.90 7.66 6.70 7.22 7.25  -  -  -  -  Phases of Estrus Cycle  -2.62 -0.03 +0.48 +0.85 -1.69 -0.16  Metestrus Metestrus Diestrus Proestrus Metestrus Metestrus  +1.36 -1.45  Diestrus Proestrus Metestrus 1. Metestrus 2. Diestrus Diestrus Proestrus Early Metestrus 1. Metestrus 1. Metestrus 2.  -  -  -1.40 +1.73 +0.56 -0.78 -1.79 +1.71  1. 2.  1. 2.  -  -1.81 -  +0.32 -0„24 -0.96 +0.52 +0.03  Early Metestrus 1. Metestrus 1. Metestrus 2. Diestrus ? Diestrus ? Metestrus 1. 1 Metestrus 2. Diestrus Diestrus Proes trus  TABLE XXV SEXUALLY MATURE FEMALE RATS.  CONTROL ANIMALS.  SHOWING WEIGHT AND FEED LEVELS ARRANGED ACCORDING TO THE PHASE OF THE ESTRUS CYCLE. Date  Animal . No.  Early Metestrus  28/8/62 21/8/62  •' 6 7  Metestrus 1  23/8/62 27/8/62 21/8/62 25/8/62 29/8/62 25/8/62 29/8/62 23/8/62 29/8/62 22/8/62 26/8/62  1 1 2 2 2 3 3 6 6 7 7  24/8/62 28/8/62 22/8/62 26/8/62 30/8/62 26/8/62 30/8/62 24/8/62 30/8/62 23/8/62 27/8/62  1 1 2 2 2 3 3 6 . 6 . 7 7  29/8/62 25/8/62 24/8/62 28/8/62  1 6 7 7  Diestrus 2  21/8/62 30/8/62 23/8/62 27/8/62 27/8/62 21/8/62 26/8/62 29/8/62  Proestrus  22/8/62 24/8/62 28/8/62 28/8/62 22/8/62 27/8/62 30/8/62  Metestrus 2  Diestrus 1  Weight Gained or Lost (GMS)  Feed Intake + or -  + 4.0 - 2.0  -0.78  + +  -0.90 -0.43 +0.77 -0.17 -0.14 -2.62 -1.69  2.0 1.5 0.5 3.7 0.5 7.8 5.0 1.5 6.5 4.0 2.5  + 1.0 + 2.0 + 2.0 + 3.0 .+ 1.5 + 4.8 + 2.0 • + 6.0 + 5.5 + 0.5 + 0.0  -  -  -1.79 +0.32 -0.42 -0.50 -0.31 +0.33 +0.69 -0.03 -0.16  -  +1.71 -1.81 -0.24  1.0 5.3 7.0 0.5  -0.30 -1.40  1 1 2 2 3 6 6 7  - 4.0 .+ 3.0 + 3.5 + 2.2 - 4.6 +12.0 + 7.3 + 0.0  -0.22 +1.15 +0.37 -0.33 +0.48 +1.36 +1.73 +0.52  1 2 2 3 6 6 7  + 7.5 + 2.5 + 4.5 + 7.8 + 1.5 + 0.5 +13.0  +1.81 +0.10 +0.34 +0.85 -1.45 +0.56 +0.03  -  -  -0.96  PART B  THE DEER  137. PART B  THE DEER  Introduction. ' The annual cycle of growth of the b l a c k - t a i l e d deer Odocoileus hemionus columbianus (Vancouver Island Genotype) have been described by Wood et a l (117).  Carcass inspection on a number of occasions during t h i s  cycle of growth makes i t obvious that the changes are p r i n c i p a l l y i n the fat  compartments of the body.  Observations are therefore made of the plasma  l i p i d levels to see i f they r e f l e c t the deposition and mobilization of f a t . The l a t t e r purported to be non-esterified f a t t y acids (Nefa)(42). After s i x years of observation of the deer i n the UBC  colony i t i s  evident that the weight accretion pattern changes r e l a t i v e to the state of sexual a c t i v i t y .  The f u l l extent of the rut season i n these animals  has  not been ascertained h i s t o l o g i c a l l y or physiologically, but from observation i t appears to extend from September to March.  The height of the a c t i v i t y  occurs i n the l a t t e r weeks of October and the f i r s t weeks of November. Wislocki (112) and others (113) have reported on the reproductive physiology of the white-tailed deer (Odocoileus virginianus borealis) and Meschake et a l on the reindeer (Rangifer tarandus scandinavicus).  Although,  by comparison of different species we cannot expect i d e n t i c a l r e s u l t s , especially from different latitudes, there i s a s u f f i c i e n t degree of s i m i l a r i t y i n their sexual behavior to warrant, at this early stage of our studies, some comparison. The ruminant gains most of i t s exogenous supply of energy from v o l a t i l e short-chained f a t t y acids (7).  These are a Metabolic by-product of the  rumen microflora, which u t i l i z e s both the carbohydrate  and protein components  138. of the diet i n t h e i r production. diet of the UBC  However, there i s some doubt that the  colony (Table I) i s conducive to rumination.'  For the purpose of the following discussion i t i s assumed that the digestive process in.our animals i s of a modified ruminant type, and not necessarily that of a t y p i c a l ruminant i n the wild state. Materials. I  Animals:  Six deer of the species Odocoileus hemionus columbianus  (Vancouver Island genotype),  3 males and 3 females are used.  The  animals are a l l 11 months of age at the start of the observations, and a l l are raised i n the UBC II  Diet:  UBC  Deer Feed #36  colony. - 57 (See Table 1).  This diet i s a p e l l e t e d  feed on which a l l deer i n the colony are maintained. Feed and water are provided ad libitum and no fast i s imposed before t e s t i n g . III  Housing: The animals are housed i n d i v i d u a l l y i n pens i n a well constructed covered unit  IV  Weighing  (115).  Procedure:  Before the animals are bled they are immobilized by means of succinyl choline ('Anectine , 1  mg/cc).  Burroughs Welcome (Canada) Ltd., 20  The dosage required f o r complete immobilization i s 0.06  mgm  per kilogram of body weight (85). The  'Anectine  1  i s injected intramuscularly into the brachioce-  phalic muscle midway along the side of the neck (27). The time to immobilization i s approximately  2 - 10 minutes (85).  The animal remains down on the average of 19 minutes, during which time  al39.  the hind limb i s shaved and blood i s taken from the recurrent metat a r s a l vein. Approximately 5 - 1 0  mis of venous blood i s extracted i n  heparinized 'vacutainer' tubes on each occasion. The plasma i s separated out by centrifugation within one to three hours, and then frozen u n t i l used. Methods:  Determination  of Plasma Total L i p i d s and Total Cholesterol.  The plasma i s used to determine t o t a l plasma l i p i d s , applying the micromethod of Bengt Swahn (106). The t o t a l plasma cholesterol f r a c t i o n i s determined by the Ultramicro method of estimation of Searcy, Bergquist and Jung (96). Both values are reported as grams per 100 grams of plasma and as milligrams percent  (mgm % ) .  TABLE 1 COMPOSITION OF UBC DEER FEED #36  COMPONENTS  - 57  PER CENT  Ground Yellow Corn  30.00  Ground Wheat  12.50  Wheat Bran  13.75  Molasses  7.50  Beet Pulp  10.00  Grass Meal  10.00  Soyabran Meal  8.75  Fish Meal  5.50  Bone Meal  1.00  Iodized Salt  1.00  THE MALE DEER  142. THE MALE DEER  Introduction Antler Growth: The antlers of the coastal b l a c k t a i l deer begin t h e i r growth i n late March and early A p r i l , and continue to grow u n t i l August when the velvet is rubbed o f f and they v i r t u a l l y become dead tissue.  The antlers p e r s i s t  in this condition u n t i l well after the rut period has reached i t s peak i n November, being dropped i n late February and early March.  The actual times  of occurrence can be seen on the individual graphs (Figs.  1-3).  Rutting Period: The extent of the rutting period has not been ascertained i n the blackt a i l e d deer but the male animals show aggressive behavior soon after the velvet i s rubbed from the antlers i n August or early September.  In the  wild these animals have been known to cover a female i n September and subsequently young are born as early as A p r i l . occur as late as March.  Also mating i s reported to  However, the usual and most successful time of  mating i s the l a t t e r weeks of October and the f i r s t weeks of November. Hence, the majority of fawns are dropped i n the last week of May and throughout  June. In the UBC colony attempts have been made to mate the animals at many  periods from September u n t i l March, but a l l except one of the successful matings occurred i n the f i r s t weeks of November. No extensive h i s t o l o g i c a l studies of the t e s t i c l e s of the coastal deer have been made.  One report (22) states that free spermatozoa were  present i n the epididymis of an animal shot i n the f i e l d on September 15th.  G 9 - 1 4 2 lUTur?. L I T ! .  CAHADIAN C H A R T S  1 YEAR BY DAYS X 2SO DtVNS. CALENDER YEAR  FIG.I.  2  <  3  2 O  o o  if)  m X  Q O CQ  WEIGHT  MAY  JUNE  5 10 15 20 25  5 10 15 20 25  ACCRETION: PLASMA LI P I D S JULY  5 10 15 20 25  AUGUST  5 10 15 20 25  SEPTEMBER  5 10 15 20 25  :  AND5UPPI  OAKViLXE,  OUT.  M A D E IN C A N A D A  FEED INTAKE FOR OCTOBER  5 10 15 20 25  NOVEMBER  5 10 15 20 25  ELEVEN DECEMBER  5 10 15 20 25  MONTHS. JANUARY  5 10 15 20 25  FEBRUARY  5 10 15 20 25  MARCH  5 10 15 20 25  APRIL  5 10 15 20 25  MAY  5 10 15 20 25  JUNE  5 10 15 20 25  JULY  5 10 15 20 25  G9-142  SUPPLIES.  CANADIAN C H A R T S A N D S U P P L I E S , L T D .  t YEAR BY DAYS X 2SO DIVNS. CALENDER YEAR  OAKVILLE,  ONT.  M A D E IN CANAOA  FIG. 2. WEIGHT ACCRETION: PLASMA LIPIDS: FEED INTAKE FOR ELEVEN MONTHS MAY 5  5  10 15 2 0 2 5  lt 1 5  2b 25  MAY'W  5 10 15 2 0 2 5  5 10 15 2 0 2 5  JUNE'bl  JULY'fel  5 10 15 2 0 25 AUGUST'fel  5 10 15 2 0 2 5 S E P T E M B E R 'fcl  5  10 15 2 0 25 O C T O B E R "61  5 10 15 2 0 2 5 NOVEMBER'61  5 10 15 2 0  25  D E C E M B E R %i  5 10 15 2 0 25 JANUARY  '6*  5  10 15 20 2 5 FEBRUARY  5 10 15 2 0 2 5 MARCH'fel  5 10 15 2 0 25 APRIu'fcZ  5 10 15 2 0 2 5 MAY "62  5 10 15 2 0 2 5 JUNE  5  1 0 15 2 0 J U LY  25  G 9 - 1 4 2 <JP»UFS.  CANADIAN C H A R T S A N D S U P P L ' E ? . L T D .  ITS.  0*KVILLE,  1 YEAR BY DAYS X 2S0 DIVNS. CALENDER YEAR  ONT.  MADE IN C A N A D A  FIG.3 WEIGHT ACCRETION *• PLASMA LIPIDS: FEED INTAKE FOR ELEVEN MONTHSM A Y  5 10 15 20 25  JUNE  JULY  AUGUST  SEPTEMBER  OCTOBER  NOVEMBER  DECEMBER  5 10 15 20 25  5 10 15 20 25  5 10 15 20 25  5 10 15 20 25  5 10 15 20 25  5 10 15 20 25  5 10 15 20 25  5 10 15 20 25 J U L Y '•»«  5 10 15 20 25 AUGUST'tl  5 10 15 20 25 S E P T E M BER 'bl  5 10 15 20 25 OCTOBER 'bl  1U 13 £U  £3  NOVEMBER*'!  • 1U 13 £U £3 DECEMBER't4»  JANUARY  5  3  10 15 20 25  UJ ID  £\J  J A N U A R Y 'bl  FEBRUARY  MARCH  APRIL  MAY  JUNE  5 10 15 20 25  5 10 15 20 25  5 10 15 20 25  5 10 15 20 25  5 10 15 20 25  3  1U 13 £U ^3 FEBRUARY'W  5 10 15 20 25  ¥46* Results: Figs. 1 - 3 present graphic representation of weight accretion, feed intake and plasma l i p i d s ("fats" and cholesterol) at approximately monthly i n t e r v a l s .  two  Numerical representation of the same data as shown i n  Table I I . Body Weight: From May, weight.  1961, u n t i l October of the same year a l l the males gain  The amount of gain being i n the range of 44 - 53 pounds.  From October to December a l l the male deer lose weight.  As much as  38.67, - 64.27. of that gained i n the spring, summer and early autumn is l o s t in the late autumn and beginning of the winter. In the winter period, December to March, animal number 8 continues  to  lose weight to the extent of 69 percent of the autumnal gain. From March u n t i l May,  1962,  a l l animals gain from 35 - 82 percent of  the weight l o s t i n autumn and winter. Feed Intake: The l e v e l of feed intake for each animal i s expressed daily intake for a seven day period.  as the mean  This includes the day of testing  weighing, and three days before and a f t e r .  and  Because this scheme i s used,  days during the winter months when food i s refused do not appear. From June u n t i l August, 1961,  the animals increase their feed intake  but not enough to a l t e r s i g n i f i c a n t l y the feed consumed per unit of body weight. From August to October the feed intake drops i n i t s actual and r e l a t i v e values to the body weight. By December a l l the animals drop their l e v e l of intake by  approximately  M7.. nine ounces to one pound four ounces per day per animal.  In one animal,  number 6, the loss of weight from October to December i s so great (34 lbs.) that the l e v e l of feed intake per unit of body weight appears to increase over the October l e v e l .  Later this animal i s the f i r s t to show signs of  a gain i n weight. By March a l l animals have increased their feed intake a c t u a l l y and r e l a t i v e to body weight. In May, when weight gain i s well under way the l e v e l of feed intake per animal drops a few ounces per day. is not known.  The explanation of t h i s decrease  Animal number 4 lags behind the other two deer i n weight  gain and continues to increase i t s l e v e l of feed intake through the summer. Plasma 'Fat': The plasma ' f a t ' i s the non-cholesterol portion of the plasma l i p i d s . This i s calculated by subtracting the t o t a l plasma cholesterol f r a c t i o n from the t o t a l plasma l i p i d value. The plasma ' f a t ' f r a c t i o n remains constant and r e l a t i v e l y high, 176 to 192 mgm 7,, from June u n t i l October i n animals number 4 and 8. is the period of high weight gain.  This  However, i n August, number 6 shows an  increase i n plasma 'fat', from 176 to 200 mgm %.  This animal i s i n the  process of rubbing the velvet from i t s antlers at the time of testing whereas the other two animals had finished or not yet begun. In October animal number 6 shows a drop i n plasma ' f a t ' from 200 mgm 7. to  120 mgm %.  As mentioned above, the other two animals show no change.  In December, after a period of rapid weight loss, a l l animals have a r e l a t i v e l y low plasma 'fat' l e v e l i n the range of 128 to 160 mgm %.  These  Mi8, levels f a l l again i n March to a range of 112 - 152 mgm into  7, and remain low  May.  Total Plasma Cholesterol From June u n t i l August the plasma cholesterol l e v e l i s lowest (80 mgm  7o - 96 mgm  7»), i n one instance (No, 8) very low (48 mgm  7,).  However,  on October 20th, 1961, a l l three animals show a s i g n i f i c a n t increase i n this l i p i d f r a c t i o n (96 mgm  7, - 144 mgm  "/») .  By December animals number 4 and 8 regain a low l e v e l of plasma cholesterol, ie 80 mgm  7., and exhibit a drop i n the feed intake l e v e l ,  whereas number 6 shows an i n s i g n i f i c a n t drop i n cholesterol from 144 mgm 7> to the  128 mgm  %.  The high l e v e l of cholesterol i s retained possibly because  animal increases the l e v e l of feed intake r e l a t i v e to the body weight,  though i t i s not raised i n i t s actual l e v e l from October.  A picture such  as this results from the very high loss of weight which this animal experiences without a commensurate drop i n feed intake. It i s pertinent to examine the plasma cholesterol levels i n March i n r e l a t i o n to the shedding of a n t l e r s .  In t h i s context, number 4, which had  shed i t s antlers on February 22nd, shows an i n s i g n i f i c a n t r i s e i n cholesterol above the December l e v e l .  Number 8 shed on March 7th and when cholesterol  levels are determined on March 10th they are elevated.  Deer number 6  experiences a prolonged delay between losing the f i r s t antler on February 20th and the second on March 10. lower than those i n December.  In this animal the March values are  On the basis of this data there i s no clear  picture of the relationship between plasma cholesterol and antler shedding.  Discussion: Seasonal Fluctuations of Weight, Feed Intake, and Plasma L i p i d s Relative to Sexual A c t i v i t y . The seasonally c y c l i c a l changes of body weight, plasma l i p i d s and feed intake i n the b l a c k - t a i l e d deer for a period of twelve months appears to p a r a l l e l the recrudence and regression of sexual a c t i v i t y . A.  Spring. From March, when the r u t t i n g season i s over and the antlers are shed, weight loss ceases or i s markedly decreased. number 5 (10/3/62), begins to gain weight. the period when the nadir of growth occurs.  Only one animal,  In most animals this i s Accompanying the cessa-  tion of weight loss there i s an increased feed intake and a low plasma ' f a t ' . . The l a t t e r would suggest either a decrease i n the output of non-esterified f a t t y acids (Nefa) by the fat depots, or i n the.case of weight gain, an increase i n the uptake of Nefa.  There i s every  l i k e l i h o o d that the f a t compartments of the body are exhausted by this time of the year, and so. a decreased output would be i n order. Increased plasma cholesterol levels i n March accompany an increase i n feed intake, and the dropping of the antlers. .Either of these may be casually related to the cholesterol l e v e l s .  There may  also be j u s t i f i c a t i o n for assuming a state of sexual regression (39, 51, 52, 54, 75, 76, 106). In late March and early A p r i l the antler buds appear. growth presumably i s stimulated by the anterior p i t u i t a r y  Their  (FSH)  (112), u n t i l gonadal recrudence i n the late summer, ie.. August. that time their growth comes under the control of testosterone.  At  150. .Summer. During this period of gonadal quiescence a large part of the spring and summer accumulation of weight occurs.  In mid-August the  velvet i s shed from the antlers and i n the next two months growth rate i s double that of the previous two months.  The shedding of  velvet i s , according to Wislocki (112) and others (113), attendant upon recrudence of gonadal a c t i v i t y . levels decrease.  Goncomitantly the feed intake  This l a t t e r occurrence corroborates the e f f e c t s of  testosterone that Kochakain (61) reported i n the laboratory r a t . In animal number 8 there i s a marked decrease i n plasma cholesterol, and substantially low levels i n the other two animals.  According to  a number of investigators (2, 18, 38, 55) with the laboratory rat and with man (75, 106) testosterone has the effect of lowering plasma cholesterol.  The plasma ' f a t values remain high i n August. 1  These  high levels seem to r e f l e c t the high growth rate, and possibly the lower l e v e l of exogenous energy ingested, or both. Autumn. By October the animals reach the high point i n t h e i r annual weights (117). plasma ' f a t * .  The feed intake i s lower than i n August, s i m i l a r l y the The l a t t e r may be indicative of an increase uptake of  fat by the f a t depots. October 20th, the day of testing, i s close to the time of active rut, and we assume that there i s a high l e v e l of c i r c u l a t i n g testosterone as has been demonstrated by Meschake (80) i n the reindeer and Wislocki et a l (113) i n the white-tailed deer.  However, there occurs  at t h i s time i n a l l three animals a s i g n i f i c a n t r i s e i n the plasma  151. cholesterol l e v e l .  This i s a reversal of the supposed e f f e c t of high  testosterone levels upon the plasma c h o l e s t e r o l . Therefore we  may  assume that the c i r c u l a t i n g levels of testosterone are not high on October the 20th, at the moment.  The reasons for this are beyond our comprehension  A f u l l e r understanding  of these animals,  of the reproductive physiology  or more knowledge of weight gained as fat and i t s  e f f e c t s on plasma cholesterol may  help to give an explanation.  We believe, however, that the high cholesterol may some mechanism involved i n sexual recrudence, very high testosterone l e v e l .  result from  possibly the effect of a  This effect may  cause a momentary f a l l  in testosterone and a commensurate r i s e i n c h o l e s t e r o l . However, a low l e v e l of testosterone i n June does not cause a r i s e i n cholesterol when there i s gonadal quiescence.  This suggests that some other  aspect of the hormone balance besides testosterone i s involved. D.  Winter. Two months later, i n December, a l l three animals lose a considerable proportion of the weight gained i n the summer-autumn growth . period.  The feed intake for each animal drops almost a pound per  day i n two animals (No. 6 and No. 8) and nine ounces i n another (No. 4).  It has been observed by us and others (26) that during this  two month period the animals refuse to eat altogether.  Sometimes for  a period of a week or more even though adequate feed i s supplied. There are no plasma ' f a t ' determinations  for this period because  the animals are i n rut and too d i f f i c u l t to handle.  However, they are  expected to be high, since an endogenous source of energy i s necessary to augment the lack of an exogenous source.  Further evidence for this  152. assumption i s presented by the fat depleted state of the carcass of animals after a period of rut and self-imposed  fast.  In December, we f i n d the plasma ' f a t ' levels are low, a decrease i n the output of fat from the depots, possibly because the body fat depots are exhausted.  suggesting decreased  However, i n December  animal number 6 shows a high plasma " f a t r e l a t i v e to a very 1  October l e v e l .  This may  low  be r e f l e c t i n g a weight loss as f a t .  The plasma cholesterol levels are lower than i n October, but only s i g n i f i c a n t l y i n number 8.  This animal shows the highest drop  in feed intake as well for this time, which suggests that t e s t i c u l a r a c t i v i t y i s s t i l l i n progress.  This corroborates the results of  Amoroso and Marshall (6) and others (17) who  state that sexual regres-  sion occurs at or about the time that the antlers are dropped which, in this case, i s not u n t i l February and March. E.  Spring. . In March weight accretion begins again, the feed intake levels increase and the plasma ' f a t levels f a l l . 1  feed and  ' f a t ' i s a f a m i l i a r one  The l a t t e r pattern of  (74), for i f the diet i s s u f f i c i e n t  to s a t i s f y the c a l o r i c needs of the animal the mobilization of plasma 'fats', to any degree, i s not necessary. by an increase i n weight may as f a t .  A low plasma ' f a t ' accompanied  also be indicative of weight being  gained  That i s , there i s an increased uptake of fat by the fat  compartments of the body. Recently i t has been discovered that testosterone raises the plasma ' f a t ' levels (63).  From this we might assume that the  low  levels of plasma ' f a t ' are as a result of low testosterone levels,  and  153. subsequently that i n March the animals are i n a state of gonadal quiescence. Summary: i  The male deer 0. hemionus columbianus shows a seasonally c y c l i c a l growth pattern.  Concomitant with the weight changes are c h a r a c t e r i s t i c  changes i n feed intake, plasma ' f a t ' and plasma c h o l e s t e r o l . The summer period, May to August, i s characterized by weight accretion; antler growth and increasing levels of feed intake per animal per day; a r e l a t i v e l y high-plasma ' f a t ' and a low plasma cholesterol, 48 - 96 mg %. August to October, begins with the shedding of velvet from the antlers and an increase i n the rate of weight gain.  By October the feed  intake i s usually less; the plasma ' f a t ' lower i n range, 120 - 176 mgm 7,, and the plasma cholesterols r i s e to 96 - 144 mgm  7> from 48 - 80 mgm  % in  August. In the autumn period, October to December, rut occurs; the animals lose weight, and the actual value of feed intake drops. are i n a low range of 128 - 160 mgm  The plasma 'fats*  7,, also, the plasma cholesterols are  s l i g h t l y lower than i n October, i e . 80 to 128 mgm 7,. From December to March, weight accretion and the plasma 'fats' are roughly unchanged, but the feed intake i s increased.  The antlers are shed  and two animals show a s l i g h t r i s e i n the plasma cholesterol at the time. May i s characterized by weight gain; antler growth; low plasma fats (128 - 136 mgm  7), and plasma cholesterol levels of 96 mgm 7o and l i t t l e or  no change i n the feed intake from the previous two months. A l l parameters, therefore, appear to show a c h a r a c t e r i s t i c change r e l a t i v e to the c y c l i c growth pattern of the deer, and the growth changes appear to p a r a l l e l the regression and recrudence of gonadal a c t i v i t y .  TABLE I I MALE DEER. A.  Date  WEIGHT ACCRETION, PLASMA LIPIDS AND FEED INTAKE AT TWO MONTHLY INTERVALS FOR ONE YEAR. Feed Intake GMS/100GMS of Body Weight  Plasma L i p i d s , GM/100GMS of Plasma Cholesterol "Fat"  Body Weight (lbs.)  R4  R6  R8  R4  R6  R8  2.83  3.09  2.68  3.6  3.4  3.3  .094  2.94  3.18  2.63  2.0  2.15  2.12  .078  .048  3.02  3.10  2.27  3.11  4.1  2.6  .094  .140  .125  2.56  2.39  2.42  2.15  3.9  3.13  1125  .078  .125  .078  2.14  2.60  1.95  2.6  2.10  2.9  .133  .109  .094  .094  .109  2.40  3.10  2.61  3.1  3.6  3.5  1133  .133  .094  .094  .094  3.08  2.41  2.18  3.9  3.4  3.2  R6  R8  R6  R4  R6  $8  R4  24/5/61  93  101  117  .179  16/6/61  102  106  121  il87  .172  .187  .062  .078  16/8/61  114  127  135  .180  .195  .172  .062  20/10/61  137  154  165  .172  .117  .172  28/12/61  120  120  135  .156  .148  10/3/62  120  125  132  .148  25/5/62  128  . 148  158  .125. B.  Feed Intake lb s/Anima1/Day l b s . oz •  R4  R8  .078  CHANCJES OCClIRRING l l^ THE PARAMETER AT TWO  16/6/61  + 9  + 5  + 4  + .008  -  +  16/8/61  +12  +21  +14  -.007  + .023  -.015  20/10/61  +23  +27  +30  -.008  -.078  0  28/12/61  -17  -34  -30  -.016  +.031  10/3/62  0  + 5  - 3  -.008  25/5/62  + 8  +23  +26  -.023  iVIONTHLY  INTERVAL s.  +0.11  +0.09  -0.05  -1.6  -0.5  -0.7  -.046  +0.08  -0.08  -0.36  +1.11  +1.1  -0.6  + .032 + .062  + .077  -0.46  -0.61  +0.15  kl.04  -0.7  +1.7  -.047  -.016  -.015  -.047  -0.42  +0.51  -0.47  -0.9  -0.15  -1.4  -.015  -.016  +.016  -.031  + .031  +0.26  +0.40  +0.66  +0.10  +0.12  +0.12  0  + .024  0  0  -.015  +0.28  -0.59  -0.44  +0.8  -0.2  -0.3  -.016 0  0  TABLE I I I MALE DEER. PLASMA LIPIDS LEVELS MILLIGRAMS PERCENT (mgm7o)  Plasma " F a t " Date  Plasma  MG7,  R4  Cholesterol  MG7,  R6  R8  R4  R6  R8 .-  24/5/61  184  •-  -  80  -  16/6/61  192  176  192  64  80  96  16/8/61  184  200  176  64  80  48  20/10/61  176  120  176  96  144  128  28/12/61  160  152  128  80  128  80  10/3/62  152  136  112  96  96  112  25/5/62  128  136  136  96  96  96  THE FEMALE DEER  157. THE FEMALE DEER  Introduction. The three female deer, Odocoileus hemionus columbianus, used i n this experiment are the same age as the male deer, that i s , eleven months o l d at the onset of observations. They are weighed, bled, fed and housed i n the same area and at the same time as the males, but i n t h e i r separate cubicles. The female deer does not have antlers or other overt signs of pending sexual maturity, and i t i s therefore more d i f f i c u l t to detect the events in her sexual cycle.  The period of fawn drop, however, permits the assump-  tion that f e r t i l e matings are possible i n some females at intervals between October and May.  The highest occurrence of successful mating at t h i s  latitude (49°) i s i n the middle of November (24). It i s known that ovulation can and does occur four or more times i n the breeding season at intervals of 26 to 28 days i f the doe i s not bred. Cheatum and Morton (16) reported the same c y c l i c period for the white  tail  deer. Each estrus period lasts f o r only a few hours and i t i s only at these times that the female w i l l accept the male.  Therefore, diestrus periods occur  in the mating season as well, at which time there i s r e l a t i v e gonadal quiescence.  This i s unlike the male which i s i n a continuous state of rut,  except possibly f o r short recuperative periods (17).  The anestrus, or  complete gonad quiescent phase, occurs after March and continues u n t i l September or October. Although i t i s d i f f i c u l t to t e l l ,  i t i s believed that the i n i t i a l  G9-142 ••UPFUES,  CANAD'AfV  L T T .  FI6.4. WEIGHT ACCRETIONMAY  5 10 15 20 25  j  U  N  E  5 10 15 20 25  PLASMA  AND OAKVILLE,  t YEAR PY DAYS X 250 D1VNS. CALENDER YEAR  MADE  LI PI DS  :  F E E D INTAKE  FOR ELEVEN MONTHS.  JULY  AUGUST  SEPTEMBER  OCTOBER  NOVEMBER  DECEMBER  5 10 15 20 25  5 10 15 20 25  5 10 15 20 25  5 10 15 20 25  5 10 15 20 25  5 10 15 20 25  S U P P L Y or.r.  I r« C A N * D A  G9-142 UPPLIES, Lrn.  CANADIAN  1 YEAR BY DAYS X 250 DIVNS. CALENDER YEAR  FIG.5  WEIGHT ACCRETION: 5 10 15 20 25  SEPTEMBER  OCTOBER  NOVEMBER  DECEMBER  5 10 15 20 25  5 10 15 20 25  JANUARY  5 10 15 20 25  5 10 15 20 25  5 10 15 20 25  ;  iiiiliiiilmi ...  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TT jr'::: n n I H I Hii fTTTTTiii: l i l j i l l l i i : ; ; ! ! nil n n nm n n nn n n IHi IHi mn in. HH •r ! m .1! H. "i ::= ::n •H •' | •' 'QT>' 5 0 IH!IliiillllillllSilllli Hi: mn HI iiii imIHInil Hi; iHi Hi! ;n;; rrS^Iiiliillii Mni-irUmmnmM .'iifei miOiii m i i n n iiii iiii n n mi HIIHH i m mi mi nn im —,— „ . to ::::):::: iiH Hii|ni:T:HiiiiHrHrijiiii HH H;;|;H; iili iiiltmilllil lirl; ;i;; n n mi1mi . ; mi i m nn iiii n n mi iiii IHI n n Is IHIHi iHi HH Hill iili i i i i • 4 4 m ! n ; . l l ; : : : ::::!::::: ; n i -::::r.:t:;:-. Hiil::i: 5 ::::!:r-r: ' t S - - [ i l i j i E g i m j :t;iH:':LHHl~i:n:t-_^: -A rrif :r_*:::-.:::.-::!::::i;:::::"5: nm Hi! n n inn S i :::: ::::)-: iii: Iiil s i p iHi ;n; nn lilliiiii nm nlini immi iHi HH;H; HH nn Hi !!i;{!!!iliii! -HH i : '1 H HHHiHiiiiiiHiiHiilH;; ::S"~::S:i" :::::::: iinblimiimlnminni ::::|:r.: .130 ••Hliii: iii': HHIHllmilHIl ;;nIIH! 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Iiiii nm inn ;;;;]HH(iiHp:: iniinniiniinnjnninm r— 9 0 HH|iHiiiHi HH.rVi;::HHH|;H= m i m*i; rr:.:- .::i.::j' : : : : ! : . ::r.\::::l:::L IHi|mi iiiii n n m m m - " 1:— .:r I"- j 0 - L; iiii iii: iiii iiii n n iHii m i nm mn iHi Hill mi n n m i n n Hii HH IHI nm I i i " : . : ; n ; ; nniimimiiiHi ::^ln:lilljlill Si iii"; Iiiillill llii m i n n n n . L !' ! "H iH'H; •i-i L ...4 ^-!- -L ! i to 8 0 Kit w iHH im n HHiiHHiHiiHH-SiiiiH nmiiiiiiHnimiinminin! m n \-~\ ;iiiii;;i mi nn m i nn IH! ll;;jini|liii nn iiii mi nil m . LU _• --H~-- -"-t T7 - i-l'IH: iiii ;Hi;iH=;HiiiHiijnii;ini :~;~:l::n|:~;:~:i:—^: IliljiliT lllil ; V i ; — i . . . . ~:iii iiii iiiilpii n i l Hiii Hii inn nnn n IHI HHHH iHi Hi HHHH mi Q f= iinlim !;;l.i- !:; _J ::::,:::: • : • 70 Hii mil iHi n Hiii IHIHH •» O lliifliii JIIIII iiil iii; iiHinnyniiimiHiiinm imimmmmniiminn n n n n mn m i H I n m miimiim H H I I H ; liiiiiii m n m n m i m I^ -o> X i:ii n n H;; i;;;; Hi; m i nn n n HH—HHjIH—ii. 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WEIGHT A C C R E T I O N : MAY 5 10 15 20 25  1015 20 25  MAY 'Ul  JUNE 5 10 15 20 25  5 10 15 20 25  JUNE 'bl  PLASMA  LIPI D S :  MADE IN C A N A D A  F E E D INTAKE  FOR E L E V E N  MONTHS JUNE  JULY  5 10 15 20 25  5 10 15 20 25  5 10 15 20 25  JULY "fol  5 10 15 20 25  5 10 15 20 2b  5 10 15 20 25  AUGUST 'fcl SEPTEMBER "fcl OCTOBER 'bl  10 15 20 25  5 10 15 20 25  NOVEMBER '£» DECEMBER'GI  5 10 15 20 25  JANUARY 'W  5 10 15 20 25  5 10 15 20 25  FEBRUARY 'C2  5 10 15 20 25  MARCH "tZ.  APRIL-M.  10 15 20 25  MAY bZ  10 15 20 25  JUNE  JULY 5 10 15 20 25  10 15 20 25  JULY  161. recrudence of the sex gland i n the females occurs at the same time as the males (17). Like the male, the accumulation of body weight occurs during the summer and autumn and the loss of weight occurs during the mating season. et a l (16) have suggested that body weight gain and increased a c t i v i t y are  11  interrelated expressions  of an altered endocrine  Cheatum  sexual balance."  Results. Body Weight: The d e t a i l s of the annual cycle of changes i n body weight, feed intake and plasma l i p i d s ("fats" and cholesterol) are given on F i g s . 4 - 6  and  Tables IV - V. Feed Intake: The feed intake i s divided into four different periods.  In May  and  June a resurgence of growth i s accompanied by an increase i n feed intake actually and r e l a t i v e to body weight. In summer, June u n t i l August, the feed intake levels f a l l , and yet weight continues to be gained.  This suggests that the animals have a lower  c a l o r i c requirement than previously, or because they are young (12 - 14 months), the growth i s of a s e l f - a c c e l e r a t i n g nature. of growth energy i s secured from an endogenous source,  In t h i s l a t t e r type ie body f a t , and at  this time the plasma 'fats' are r e l a t i v e l y high. During the autumn period, August to October, the feed intake i s equivalent to that of the spring period, with the exception of animal number 1 which shows a low feed intake. Through the winter, from October to March, a l l the animals lower t h e i r l e v e l of feed intake.  This i s a period when the animals are i n a state of  162. negative energy balance and lose weight steadily.  At their lowest weight  in March the feed intake r e l a t i v e to body weight i s actually higher than i n December, thus r e f l e c t i n g a considerably greater energy demand at this time. Plasma  'Fats': In  the spring-summer  period of increasing energy input two of the  experimental animals demonstrate an increasing plasma 'fat'.  The t h i r d  female i s out of phase during the winter and gradually decreases the plasma 'fat'  u n t i l i t reaches the l e v e l of the other two. In  the autumn period, August to October, the plasma fat levels r i s e  s i g n i f i c a n t l y i n number f i v e , or change l i t t l e  i n numbers 1 and 12.  From  October u n t i l December, when weight i s lost and the feed intake i s low, two animals level.  (No. 1 and 12) exhibit a s i g n i f i c a n t r i s e i n the plasma ' f a t ' Whereas, the t h i r d animal, number 5, shows a s i g n i f i c a n t f a l l i n  this component. . Animal number 5 has over the same period as the others lost three and four times as much weight, and so i t may be assumed that the  low plasma 'fat'  i s as a result of an exhausted supply of depot f a t .  In March and May of 1962 the plasma 'fat' the  levels are low relatiV*~/.to  previous periods described.  Total Plasma Cholesterol: The plasma cholesterol levels r i s e abruptly i n October to 80 - 144 mgm %, from the low values of June and August; 6 4 - 8 0 mgm  %.  These l a t t e r  values are similar to the male plasma cholesterol values at the same time. "At no time during our observations of these deer i s the female cholesterol value higher than that of the male. the  This d i f f e r s from results obtained on  laboratory rat by many investigators (37, 38, 72, 99) and on the human. From October to March each animal has i t s own individual pattern of  163. fluctuation, but i n March they a l l f a l l to a common l e v e l similar to that of the males, 96 mgm /o • Discussion. Seasonal Fluctuations of Weight, Feed Intake and Plasma L i p i d s Relative to Sexual A c t i v i t y . The pattern of growth i n the female deer throughout the year p a r a l l e l s that of the male, although the weights achieved are not as great nor i s the loss as dramatic.  The l a t t e r , however, extends over a longer time, as  much.weight being lost from December to March as i n the previous two months. It w i l l be r e c a l l e d that the male lost very l i t t l e weight between ^December and March, i n fact one animal showed a gain. A.  Spring. In the spring, May to June, 1961, after a winter of an inadequate c a l o r i c intake and low plasma 'fat', the l a t t e r plasma component i n creases concomitant with a r i s e i n body weight and. feed intake. The high plasma ' f a t ' accompanying the increased feed intake may be as a result of the absorption from the rumeu of a high concentration of v o l a t i l e short chain f a t t y acids (acetic, propionic and b u t y r i c ) ( 7 ) . Regardless  of the high feed intake and the a c q u i s i t i o n of body weight  the plasma cholesterol i s low, ranging from 64 mgm % to 80 mgm %. B.  Summer. From June to August the l e v e l of feed intake drops, the plasma 'fat'  increases and the rate of weight gain i s high.  As suggested  e a r l i e r , growth i n the young animal i s more e f f i c i e n t when an endogenous source of energy i s u t i l i z e d rather than an exogenous source (47, 90).  In this fashion the animal i s saved the high energy costs  164. of ingesting and absorbing  large amounts of feed.  Similar to the  spring period the plasma cholesterol levels are low (64 - 80 mgm 7») . The spring-summer period therefore, appears to be one i n which there is recuperation and growth, and i t i s believed one i n which there i s sexual quiescence. It i s not known whether these does had already experienced a period of sexual maturity  i n their f i r s t year.  However, i t i s known  (21) that females i n c a p t i v i t y , that are well nourished, are capable of ovulation and reproduction at the age of eight months.  Therefore,  the autumn period to be described may be the f i r s t or the second occasion i n which ovulation has occurred i n these three female deer. C.  Autumn. In October the animals once more increase t h e i r feed intake and continue to gain weight; the plasma • f a t  1  r i s e s i n number 5 or  remains at the same l e v e l as seen i n number 12, whereas animal number 1 shows an i n s i g n i f i c a n t decrease.  For the f i r s t time the  plasma cholesterol i s raised i n a l l three deer. There i s every reason to believe that many does ovulate i n October, and that the r i s e i n plasma cholesterol at t h i s time r e f l e c t s the high estrogen levels attendant upon t h i s recrudence of sexual activity.  Many investigators (18, 20, 39, 87) working with the  laboratory r a t have been able to demonstrate that an increased  level  of. estrogens cause the plasma cholesterol to r i s e . whereas animal numbers 5 and 12 show an increase i n feed intake and plasma 'fat', animal number 1 shows the inverse condition and also a large increase i n plasma cholesterol from 80 mgm °L to 144 mgm %.  165. The low plasma ' f a t ' i n animal number 1 suggests either a decrease i n output or increase i n uptake of f a t by the depots.  The  l a t t e r indicates weight gained as f a t , which'corroborates the findings of Loeb (70), who reported that the administration of estrogen to laboratory rats i s conducive to weight gain i n the fat compartments. A low feed intake i s reported by Kochakain (59) to occur when estrogens are  injected i n excess.  Conceivably then, t h i s animal i s i n the late  f o l l i c u l a r phase of the estrus cycle. others (101) have demonstrated  However, Brown-Grant (14) and  i n the rat that after high levels of  estrogens have taken effect the l e v e l of the thyroid stimulating hormone (TSH) drops suddenly with a subsequent of the animal.  f a l l i n the metabolism  This decreases the feed intake and the plasma ' f a t '  and increases the plasma cholesterol l e v e l s .  If the same r e l a t i o n -  ships are applicable to the deer i t suggests that animal number 1 may be i n the l u t e a l phase.  Whichever i t i s , i t appears that this  animal i s at a different phase of the estrus cycle than the other two. D.  Winter. From October to December, l i k e the males, the females lose weight and begin t h e i r 'compulsary' p a r t i a l f a s t .  The high levels of  plasma ' f a t , i n animals number 1 and 12, indicate that they are 1  losing weight as f a t . It has been suggested that the c i r c u l a t i o n of high levels of f a t (Nefa) contribute d i r e c t l y to the lack of 'appetite' (25), i t may, levels (59).  however, be as a result of the high estrogen  Nevertheless, the high values of plasma ' f a t ' from  October to December are believed to s a t i s f y the c a l o r i c requirements of the animal during t h i s period.  166. The cholesterol levels, i n December, i n these two animals vary. For instance, animal number 12 continues to express a high l e v e l (112 mg % ) , whereas animal number 1 drops from 144 mg % to 96 mg %. This v a r i a t i o n occurs because the levels of estrogen, during the mating season, fluctuates according to the phase of the estrus cycle, and so i t may be that animal number 1 i s i n the diestrus phase. A l l does i n December reduce feed intake to the lowest point i n the annual cycle (Figs. 4 - 6 ) , but consequent  loss of weight i s  greater i n number 5 than i n the other two deer.  In fact the loss i n  weight i n deer number 5 i n two months equals the loss over four months in the other two. significantly.  Also, unlike the other does,. the plasma ' f a t ' drops  Number 5 had on many occasions i n i t s history eaten  very l i t t l e , consequently the plasma.'fat' levels are often high, see August and October Table IV. This had the effect of diminishing i t s f a t storage levels, and so we believe when the winter fast ensues, i t s f a t compartments are more quickly exhausted.  This could account  for the low plasma ' f a t ' levels i n December when the other two animals were mobilizing t h e i r f a t stores f o r t h e i r energy needs. The plasma cholesterol l e v e l i n t h i s animal i s raised only s l i g h t l y above that of the previous month, but i s s i g n i f i c a n t l y higher than the August E.  level.  Spring. In March, a l l animals continue to lose weight, number f i v e much more than the other two.  The plasma 'fats' are decreased, thus, we  believe, indicating a decrease i n output of f a t from the f a t stores of the body, the l a t t e r because the supply i s exhausted.  167. Animals number 5 and 12 show no change i n the actual l e v e l of their feed intake from the low levels of December, whereas the males had by now increased their feed intake levels and ceased to lose weight.  Because weight i s s t i l l being lost i n the females and the  feed intake levels do not f a l l concomitantly  the intake per unit of  body weight increases. Plasma cholesterol levels are low i n animals number 5 and 12, but s i g n i f i c a n t l y high i n animal number 1. Animal number 1, i n March, exhibits the same r e l a t i o n s h i p of feed and l i p i d s that i t had i n October, but i t i s s t i l l losing weight.  In the absence of detailed weights between December  and March we do not know when the upturn It  apparently  occurs.  i s s i g n i f i c a n t , however, that this animal s t i l l exhibits i n  March what we believe to be evidence  of high estrogen output.  We  infer that this animal alone i s i n estrus at the time of testing i n March.  In as much as we do not know how long before or a f t e r estrus  the cholesterol levels are high i t cannot be assumed that the other two does d i d not experience a March estrus. By May the females are i n the anestrus state.  The feed intake  increases, weight i s r i s i n g and the plasma ' f a t ' i s s t i l l low. The suggestion i s that the weight gain i s largely i n the form of f a t , She rapid storage thus reducing the plasma l e v e l s .  Plasma cholesterol  levels i n animals number 5 and 12 remain at the low March l e v e l and are joined there by number 1. The plasma cholesterol levels i n animal number 5 show no dramatic change throughout the whole period of t e s t i n g .  It i s be-  168. lieved that either the changes are missed or that t h i s animal f a i l s to express estrus i n this, i t s second year of l i f e . Summary: In the doe we have shown that during anestrus there i s a gradual gain i n weight while i n the period of recurring estrus there i s weight loss. The feed intake i s high during weight gain i n the early stages of anestrus and just p r i o r to the mating season, but i s lower when growth progression i s high i n the anestrus phase and during the period i n which the animals mate (October - March). The plasma 'fats' appear to r i s e when high growth ensues i n the anestrus periods and feed levels are low, and also when weight i s being lost as fat (December).  Low levels ensue when the source of fat i s  exhausted after the 'female rut' period, and when weight i s gained as fat during the i n i t i a l stages of the sexually quiescent state. The t o t a l plasma cholesterol levels are s i g n i f i c a n t l y high during the 'female rut', and low at times of diestrus and anestrus.  This suggests  that high estrogen levels are responsible for the r i s e . The high plasma cholesterol levels occur often when the feed intake is e s p e c i a l l y low, and are low when the feed intake i s high.  This suggests  that a b i o l o g i c a l factor and not feed intake i n the ruminant i s responsible for the v a r i a t i o n s . Whereas the plasma 'fats' appear to r e f l e c t weight loss and feed intake changes, the plasma cholesterol levels are believed to be indicative of the different phases of the estrus cycle, high i n the f o l l i c u l a r and l u t e a l phase and low i n the diestrus and anoestrus phase.  TABLE IV FEMALE DEER. A.  Date  SHOWING WEIGHT ACCRETION ,iPLASMA LIPID AND FEED INTAKE LEVELS DURING A TWELVE MONTH PERIOD. Feed Intake GMS/100GMS of Body Weight  Plasma Lipids GMS/100GMS Plasma Cholesterol "Fat"  Weight (lbs.) Rl  Rl  R5  R12  3.00  2.60  .062  3.07  .062  .078  .140  .078  .212  .094  .133  .109  .133  .125  R12  R5  &1  R5  R12  2.67  3.2  2.6  2.6  3.07  2.95  3.3  2.13  2.11  2.61  0077  2.53  3.0  0.3  2.11  .109  2.00  2.42  2.74  2.9  2.4  3.5  .094  .109  1.65  1.87  1.66  2.1  2.0  1.15  .156  .094  .094  0.17  2.08  1.76  0.5  2.0  1.15  .094  .094  .094  2.57  3.13  1.92  3.8  3.5  2.4  +0.5  Rl  R5  Rl  R5  R12  24/5/61  102  89  89  16/6/61  105  92  91  .203  .141  .164  . .078  .078  16/8/61  115  105  105  .156  .156  .187  .078  20/10/61  128  119  121  .140  .187  .187  28/12/61  125  107  117  .187  .140  10/3/62  121  96  110  .164  25/5/62  136  110  117  .164 B.  R12  Feed Intake l b s . oz. per day  CHAN GES OCCURRING 1.N THE PARAMETEF.S AT TWO MONTHLY INTERVALS.  16/6/61  .+ 3  + 3  + 2  +  -  -  16/8/61  +10  +13  +14  -.047  +.015  +.023  20/10/61  +13  +14  +16  -.016  + .031  0  28/12/61  - 3  -12  - 4  + .047  -.047  + .025  -.046  + .016  0  10/3/62  - 4  -11  - 7  -.023  -.007  -.103  + .062  0  25/5/62  +15  +14  + 7  0  0  + .016  -.062  0  +  -  + .07  + .47  +0.28  +0.1  +0.7  -.015  +.015  - .46  -2.30  -0.42  -0.3  -2.10  + .062 + .015  +.031  - .61 +1.65  +0.21  -0.7  +2.1  +0.10  - .35  -0.55  -1.08  -0.8  -0.4  -1.6  -.015  -1.48  +0.21  +0.10  -1.12  0  0  0  +2.40  +1.05  +0.16  +3.3  +1.5  +0.5  0  0  TABLE V FEMALE DEER. PLASMA LIPIDS AS MILLIGRAMS PERCENT (mgm%)  Date  Plasma Cholesterol MG7  Plasma "Fat" MG%  0  Rl  R5  R12  Rl  R5  R12  16/6/61  208  144  168  80  80  64  16/8/62  160  160  192  80  64  80  20/10/61  144  192  192  144  80  112  28/12/61  192  144  218  96  96  112  10/3/62  168  136  112  160  96  96  25/5/62  168  136  128  96  96  96  APPENDIX  TABLE i DAILY BODY WEIGHTS,. FEED INTAKE LEVELS AND PLASMA LIPIDS OF PREPUBERTAL 0* RAT. NO. 1. Age in: Days  Weight in GMS.  30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 ^50 51 52 53 54 55  75.3 84.4 89.5  t  Plasma Cholesterol mgm7o  Fat mgm7o  F/C Ratio (Relative)  Feed Intake GMS/Animal  _ _  114.5  -  176  203  .86  129.5 136.0 137.9  105  216  1.39  110.5  246.5  1.43  11.2 11.2  _  147.2 150.0 157.4 163.0 165.0 172.4 174.6 185.8 179.5 197.7 202.5 200.0  -  144.0  319.0  1.43  176.5  182.5  0.68  157.0  261.0  1.08  141.0  299.0  1.39  13.7 22.1 14.9 19.9 18.8 18.8 (19.1) (19.6) (21.7) 21.3 21.8 22.5 18.3  -  Age iri! Days  Weight in GMS.  56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77  198.8  Plasma Cholesterol mgm7o  Fat mgm7o  F/C Ratio (Relative)  -  -  234.5 228.1 231.1  -  253.0 254.8. 258.3 269.3 265.2 274.0  -  277.7 272.8 291.5 290.2 289.8 298.5  -  304.5  Feed Intake GMS/Animal  144.0  251.0  1.08  89.1  164.9  1.29  ^,..., . J  119.0  317.0  1.74  136.5  234.5  1.08  -  -  1.29 108  212  100  330  21.12  -  •21.2 (25.2) 20.4  -  21.4 (27.8) 23.6 17.2 26.3  -  21.2 22.2 15.1 18.3 25.2  •-  -  TABLE i i DAILY BODY WEIGHTS, FEED INTAKE LEVELS AND PLASMA LIPIDS OF PREPUBERTAL cfRAT. NO. 2. Age Days 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54  Weight in GGMS.  Plasma Cholesterol Fat mgm7o  mgm7o  F/C Ratio (Relative)  Feed Intake GMS/Animal  72.3 73899 86.1  --  118.5  195.  195  0.68  129.3 141.6 142.8  157.5  164.5  0.68  -  -  148.6 153.8 162.7 167.0 168.3 174.2 181.0 185.9 186.5 195.0 178.4 198.9  107.0  209.5  1.29  116.0  288.0  1.55  127.0  190.0  0.97  176.0  194.0  0.71  148.5  200.5  0.91  15.1 15.1  -  11.7 22.7 16.0 17.8 17.0 15.2 18.1 17.0 15.7 20.8 16.0  -  15.4  Age inf Days 55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77  Weight in GMS.  Plasma Cholesterol  Fat  F/C Ratio (Relative)  mgrnVo  mgm7o  125.0  233  1.2  100.0  220  1.43  _  211.7  -  -  232.5 238.2 239.8  -  249.6 247.8 255.8 264.0 262.4 267.1  -  272.4 275.0 287.0 289.3 288.6 293.6  -  114.5  263.5  1.5  135.5  234.5  1.18  101.0  188.0  1.20  96.0  235.0  1.55 '  19.0 22.0 19.0 18.1 (29.7) 18.9 19.4 22.1  24.3 17.9 13.3 12.1 20.8  303.0 21.0  TABLE i i i DAILY BODY WEIGHTS, FEED INTAKE LEVELS AND PLASMA LIPIDS OF PREPUBERTAL O^RAT. NO. 4. Age in: Days  Weight in GMS.  30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54  73.9 89.8 -  Plasma Cholesterol Fat mgmT, mgm7o  F/C Ratio (Relative)  Feed Intake GMS/Animal  _  120.5 135.5 141.8 139.6  167.0  179.0  0.71  151.0  189.0  0.86  89.5  179.5  1.29  _  141.3 148.0 155.8 164.5 161.2 170.5 170.0 174.4 180.5 188.3 .195.2 188.0  129.4  194.6  0.97  129.8  179.2  0.91  163.0  180.0  0.71  14555  206.5  0.91  14.5 14.5 - ( 1.1.5 ( 19.3 14.4 18.6 16.8 18.6 16.9 12.3 14.2 21.8 15.4 18.9 27.4 -  Age iif Days  Weight , in GMS.  55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77  210.0 225.0 231.5 238.0  -  249.5 254.0 240.0 265.5 258.5 262.4 271.5 268.0 277.7 279.5 284.0 288.0 292.7 291.5  Plasma Cholesterol mgm7o  Fat mgm7o  F/C Ratio (Relative)  Feed Intake GMS/Animal -  103  223  1.55  106  259.0  1.55  16.3 20.5 20.6  -  96.0  186.0  1.25  122.5  155.5  0.86  103 96.0  171.0  1.06  217.0  1.50  18.7 Spill 25.8 18.0 21.7 19.1 16.9 15.0 23.4 20.2 -  TABLE i v DAILY BODY WEIGHTS, FEED INTAKE LEVELS AND PLASMA LIPIDS OF PREPUBERTAL 0*RAT. NO. 5. Age Itf Days  Weight in GMS.  30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54  82.9 389.7  Plasma Cholesterol Fat mgm7o  mgm7o  F/C Ratio (Relative)  Age Feed Intake GMS/Animal  -  120.7  153.5  163.5  0.71  132.4 131.3 135.0  153.5  172.5  0.71  -  103.0  138.0  0.86  _ _  145.8 148.6 154.3 161.4 161.0 169.3 176.0 179.9 179.5 188.5 190.5 199.7  -  132.0  196.0  0.97  132.0  238.0  1.18  154.0  148.0  0.68  136.0  298.0  1.43  15.9 15.9 ( 10.8( 23.2 14.8 16.1 17.8 18.0 18.9 16.1 18.4 19.6 14.5 17.5 15.9  -  to: Days 55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77 78  Weight in GMS.  Plasma Cholesterol mgm7o  Fat mgmT.  F/C Ratio (Relative)  Feed Intake GMS/Animal  212.0  -  -  -  •-  226.0 232.6 236.0  109  255.0  1.50  98  224.0  1.50  -  247.0 247.8 247.8 255.0 257.5 261.2  -  272.5 267.4 281.0 277.8 282.5 284.5 .  -  -  17.9 19.8 21.0  -  -  112.0  224.0  1.29  137.0  259.0  1.20  17.1 21.1 18.6 17.7 19.6  -  96.0  205.0  1.39  98.0  226.0  1.50  -  -  -  19.2 17.5 13.3 (28.0)Spill 19.8  -  TABLE v DAILY BODY WEIGHTS, FEED INTAKE LEVELS AND PLASMA LIPIDS IN PREPUBERTAL d*RAT. NO. 6. Age Days 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54  Weight in GMS.  Plasma Cholesterol Fat mgm7o mgm?.,  F/C Ratio (Relative)  Feed Intake GMS/Animal  84.3 91.0 _ _  122.6  205.0  179.0  0.57  135.0 141.8 140.7  154  182.0  0.77  -  120  238.0  1.29  _  140.0 144.5 157.2 163.8 168.4 174.4 180.8 189.3 186.7 201.8 204.3 210.0 -  154.0  202.0  0.86  154  243  1.06  168.0  185.0  0.71  155.0  210.0  0.91  13.8 13.8 - ( 10.8( 20.8 15.8 18.7 18.6 18.0 18.9 16.1 18.4 19.6 20.0 17.2 19.3  Age inf Days  Weight in GMS.  55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77 78  225.3 243.3 231.4 248.0 261.2 262.1 261.9 270.5 271.4 277.5 289.5 284.3 290.9 287.7 293.7 299.2 -  Plasma Cholesterol mgm?o  Fat mgm7o  F/C Ratio (Relative)  165.0  200.0  0.77  110.0  157.0  0.91  122.0  200.0  1.06  150.0  223.0  0.97  119.5  201.0  1.08  113.0  297.0  1.71  Feed Intake GMS/Animal 20.6 17.0 19.0 19.7 22.7 19.0 20.0 22.7 21.2 16.9 16.9 21.6 20.6 -  TABLE v i PREPUBERTAL MALE RATS:  PLASMA LIPID AND FEED INTAKE VALUES  ASSOCIATED WITH 'F/C RATIOS' ABSOLUTE VALUES . Plasma gm/lOOgm Plasma Cholesterol "Fat'  F/C 0.96 1.0  .150 .175 .145 .160 .159 .164 .172 .175 .168 .154 .120 .100 .150 .131 .127 .142 .110 .126 .126 .129 .146 .124 .131 .100 .150 .119 .140 .127  n 1.1  n 1.3  n 1.4 n 1,5.  n 1.6  n  'C 'F'  = =  Cholesterol 'Fat'  .144 .178 .152 .165 .175 .180 .189 .194 .187 .204 .152 .134 .197 .172 .175 .200 .153 .176 .190 .191 .218 .185 .196 .167 .237 .195 .228 .207  RELATIVE VALUES Feed Intake gm/lOOgms of Body Weight 8.30 11.40 8.20 9.8 8.15 9.72 8.45 8.45 8.69 9.94 8.30 11.80 11.90 10.5 11.50 8.20 7.66 9.12 11.95 10.45 8.25 9.05 9.92 5.35 10.70 7.00 9.62 8.16  Plasma F/C .68 .66 .68 0.68 .72 .71 .71 .71 .71 .86 .82 .87 .86 .86 .90 .92 .90 .91 .99 .97 .97 .97 0.97 1.09 1.03 1.06 . 1.05 1.06  «, 7.  Cholesterol 172 201 167 184 182 189 198 201 193 177 138 115 172 151 146 163 126 145 143 148 168 143 151 115 172 137 161 146  'Fat' 107 133 113 123 131 134 141 144 137 152 113 100 147 128 131 149 114 131 142 143 163 138 147 125 177 145 „ 170 154  Feed Intake gms/lOOgms of Body Weight 68 93.4 67.2 80.3 66.8 79.5 69.3 69.3 71.2 81.5 68.0 96.7 97.5 85.9 94.2 67.2 63.0 74.8 98.0 86.0 68.0 74.6 81.7 43.9 87.7 57.4 78.7 66.9  % Values for TriGPhase Graph. 'Fl' •c ipi 43 40.2 42.0 41.7 42.9 40.9 43.4 43.6 42.7 37.8 37.3 29.2 35.1 34.9 32.8 38.6 36.2 35.9 31.4 33.7 38.0 35.0 34.5 37.3 34.6 36.6 - 34.9 35.9  22.6 22.2 23.9 22.9 25.6 24.4 25.9 26.2 25.5 27.2 25.7 21.3 25.0 24.8 24.6 29.4 27.4 27.2 25.8 27.0 30.9 28.3 28.0 34.0 30.0 32.5 30.8 31.8 Cont' d.  n  =  mean value  34.4 37.6 34.1 35.4 31.5 34.7 30.6 30.2 31.7 35.0 37.0 49.5 39.9 40.3 42.6 32.0 36.4 37.0 42.8 39.3 31.0 36.7 37.4 28.7 35.4 30.9 34.3 32.3  TABLE v i Cont'd. RELATIVE VALUES  ABSOLUTE VALUES  F/C  1.7 n  1.8  n  1.9  n  n  n  n  1.94  2.0  2.1  2.2  n„ '••&' 'F 'Fl' n 1  Plasma gm/lOOgm P l asma 'Fat' Cholesterol  . .153 .132 .117 .134 .130 .133 .131 .134 .122 .099 .087 .111 .094 .113 .103 .087 .105 .110 .117 .104 .105 .137 .094 .108 .113 .100 .140 .132 .098 .094 .113 = Cholesterol = 'Fat = Feed Intake = mean value 1  .254 .220 .196 .223 .232 .244 .238 .252 .227 .183 .161 .206 .181 .217 .199 .175 .206 .218 .232 .204 .207 .292 .200 .230 .241 .217 .311 .290 .214 .211 .249  Feed Intake gm/lOOgms of BodyyWeight  11.70  8V30  5.90 8.63 8.20 9.00 8.60 11.00 7.50 9.40 8.84 9.19 4.6 10.95 7.78 10.40 5.20 7.30 9.80 10.60 8.66 11.00 4.80 8.20 8.00 7.25 12.22 7.70 7.90 7.00 8.41  Plasma F/C  Cholesterol  'Fat'  1.08 1.08 1.09 1.08 1.16 1.19 1.18 1.22 1.21 1.20 1.20 1.20 1.25 1.25 1.25 1.31 1.27 1.29 1.29 1.27 1.29 1.39 1.38 1.39 1.39 1.41 1.44 1.42 1.42 1.45 1.43  176 152 134 154 149 153 151 154 140 114 100 127 108 130 119 100 121 . 126 134 120 120 157 108 124 129 115 161 152 113 108 130  190 164 146 167 173 182 178 188 169 137 120 153 135 162 148 131 154  163  173 152 155 218 149 172 179 162 232 216 160 157 185  Feed Intake gms/lOOgms of Body Weight  95.9 68.0 48.8 70.9 67.2 73.8 70.5 90.2 61.5 77.0 72.5 75.3 37.7 89.8 63.8 85.2 42.6 59.8 80.3 86.9 71.0 90.2 39.3 67.2 65.6 59.4 100.0 63.0 64.8 57.4 68.9  7o Values for T r i -•Phase Graph. •c 'Fl'  33.3 35.7 37.9 35.6 34.8 33.7 34.3 31.2 34.6 29.7 28.9 31.1 36.4 29.1 32.7 26.3 36.0 33.0 30.5 28.4 30.8 27.3 34.6 30.9 30.9 31.1 28.9 33.0 29.9 30.4 30.7  30.1 32.2 34.6 32.3 33.7 33.6 33.7 32.0 35.0 29.8 28.9 31.4 38.1 30.4 34.3 28.7 38.4 35.6 32.8 30.1 33.1 36.4 40.0 35.5 37.3 36.6 34.9 39.4 35.5 37.1 36.7 Cont'd.  36.6 32.2 27.5 32.1 31.5 32.7 32.1 36.8 30.5 40.5 42.2 37.5 25.5 40.5 33.0 45.0 25.6 31.4 36.6 41.5 36.1 36.2 25.4 33.6 31.7 32.3 36.2 27.6 34.6 32.5 32.6  TABLE v i Cont'd. RELATIVE VALUES  ABSOLUTE VALUES  F/C  Plasma gm/100 gm P l asmaq Cholesterol 'Fat'  Feed Intake gm/lOOgms of Body Weight  Plasma F/C  Cholesterol  'Fat  1  Feed Intake gms/lOOgms of Body Weight  7o Values for T r i -•Phase Graph. 'C 'Fl' F 1  1  2.6  .106 .096 .096 .111 .102 .104 .094 .099 .110  .248 .218 .220 .256 .236 .252 .220 .236 .290  7.90 8.90 36.95 7.16 7.72 8.65 7.07 7.86 6.90  1.52 1.48 1.49 1.49 1.50 1.57 1.52 1.55 1.71  122 110 110 128 118 120 108 114 126  185 163 164 191 176 188 164 176 216  64.8 73.0 57.0 58.7 63.3 70.9 58.2 64.5 56.6  29.9 28.1 30.5 31.4 30.0 28.5 29.8 29.1 30.0  38.1 34.6 37.8 39.4 37.5 37.5 37.8 37.6 43.0  32.0 37.3 31.7 29.1 32.5 34.0 32.4 33.3 27.0  2.7  .116  .309  8.40  1.74  133  231  68.9  28.6  41.5  29.8  3.3  .098  .322  7.65  2.12  113  240  62.7  25.6  45.7  28.7  2.3  n 2.4 n  'C 'F' 'Fl' n  = Cholesterol = 'Fat' = Feed Intake = mean value  TABLE v i i DAILY BODY WEIGHT, FEED INTAKE AND PLASMA LIPIDS OF PREPUBERTAL £ RAT. NO. 1. \ge in Days 30 31 32 33 34 35 36 37 38  39  40 41 42 43 44 45 46 47 48 49 50 51 52 53 54  Weight in GMS.  Plasma Cholesterol Fat mg% mg%  F/C Ratio Actual Relative  Feed Intake GMS/Animal  55.1 66200 66.4  a*  87.1 100.3 105.2 108.2  113 :o  120.6 123.2 127.7 134.0 132.5 135.5 135.4 138.9 141.5 145.2 147.0 150.4  176.0  167.0  0.95  1.65  107.0  245.0  2.30  182.0  218.0  1.20  2.05  179.0  281.0  1.60  2.74  180.0 152.0  162.0 236.0  0.90 1.60  1.58 2.74  13.4 13.4 ( 15.4( 17.5 13.1 13.5 14.5 11.0 11.9 9.7 10.9 14.2 13.5 11.6 13.1  Age in Days 55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77 78  Weight in GMS.  F/C Plasma Ratio Cholesterol Fat mg7o Actual Relative mg?.  -  156.5  -  162.4 166.3 161.5  143.0  51.0  0.35  0.62  117.0  223.0  1.90  3.20  127.0  239.0  1.80  3.10  148.0  266.0  1.80  3.10  -  169.5 169.7 168.0 174.8 177.2 175.0  Feed Intake GMS/Animal  11.9 14.5( 80° 11.3(-82 F - ( - ( 10.6( 11.4 74°F 12.1 13.1  -  -  180.4 184.0 189.5 188.0 187.5 188.5  14.5 11.4 11.8 12.3 -  146.0  242.0  1.70  2.90  130.0  234.0  1.80  3.10  132.0  207.0  1.60  2.74  -  197.3 192.5  -  14.6  TABLE v i i i DAILY BODY WEIGHT,. FEED INTAKE AND PLASMA LIPIDS OF PREPUBERTAL £ RAT. NO. 2.  Age in Days 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54  Weight in GMS.  F/C Plasma Ratio Cholesterol Fat mg% Actual Relative mg%  Feed Intake GMS/Animal  60.0 65.3 70.5  -  m»  _  93.0  -  102.5 106.0 110.2  156.0  166.0  1.1  1.94  89.0  223.0  2.5  119.5  221.5  1.9  3.20  146.2  263.8  1.8  3.15  -  110.4 114.3 120.2 126.3 128.0 129.7 131.0 131.2 133.0 134.0 139.4 143.6 146.8  148.0 174.0  164.0 . 1.1  1.94  225.0  2.26  1.3  15.0 15.0  (  15.0( 17.0 15.1 15.0 12.4 12.8 12.3 10.3 10.8 15.4 13.6 14.4 12.9  Age in Days 55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77 78  Weight in GMS.  Plasma F/C Ratio Cholesterol Fat mg7=> mg% Actual Relative  -  -  154.0  -  -  mm  162.7 162.9 161.0  138.0  66.0  0.48  0.83  -  172.5 170.3 168.5 175.7 174.9 177.6  117.0  289.0  133.0  289.0  2.2  3.75  140.0  198.0  1.4  2.45  159.0  191.0  1.2  2.05  141.5  222.5  1.6  145.5  204.5  1.4  . 2.74  -  195.2 195.8  13.9 15.6 11.3  -  2.5  -  183.5 182.2 188.3 188.4 188.8 189.5  Feed Intake GMS/Animal  2.45  12.3 6.5 12.4 11.1  -  11.9 10.1 13.9 13.6 14.3  14.7  82°F 82°F 82°F 82°F 74°F  TABLE i x DAILY BODY WEIGHT, FEED INTAKE AND PLASMA LIPIDS OF PREPUBERTAL $ RAT. NO. 3.  Age Weight in in Days GMS. 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54  *  Plasma Cholesterol Fat mg% mg/o  F/C Ratio Actual Relative  Feed Intake GMS/Animal  62.7 68.6 72.9  _  90.2 _  97.5 101.5 104.3  -  104.3 112.2 113.5 116.4 119.4 119.9 120.3 120.8 120.0 125.5 127.5 139.3 139.3  Spill  185.5  157.5  0.85  1.47  115.0  189.0  1.64  2.74  122.0  206.0  1.70  2.90  141.2  225.8  1.60  2.74  135.0 120.3  138.0 257.7  1.0 2.1  1.74 3.71  12.7 12.7 - ( 20.9( 11.7 10.2 10.1 10.4 11.3 11.5 6.6 13.5 16.5* 14.7 18.3 13.9  Age in Days 55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77 78  Weight in GMS.  F/C Plasma Cholesterol Fat Ratio mgT, Actual Relative mg7o  -  Feed Intake GMS/Animal  -  147.0  -  12.9  156.0 149.0  2.45  157.5 159.0  106.0  163.9 166.2 163.0 168.0 167.2 166.0  83.2  265.8  3.2  113.3  108.7  0.96  1.65  129.5  164.0  1.3  2.26  -  1.4  -  176.7 177.3  82°F 82°F 82°F 13.3 82°F 16.0* 74°F 11.7 11.1  -  174.6 170.0 172.4 178.4 178.0 178.0  14.9 14.9  127.0  257.0  2.0  3.49  127.0  175.0  1.4  2.45  12-50  143.0  1.1  1.94  11.9 10.1 13.9 12.6 15.2 13.5  DAILY BODY WEIGHT, FEED INTAKE AND PLASMA LIPIDS OF PREPUBERTAL •? RAT. NO. 6  Age Weight in in Days GMS. 30 31 32 33 34 35 36 37 38  .39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54  Plasma  Cholesterol mg?.  Fat mg%  F/C Ratio Actual Relative  Feed Intake GMS/Animal  81.4 87.0  -  112.8  -  123.5 131.1 128.9  173.0  12440  0.72  1.25  -  133.4 136.2 140.0 141.0 145.9 148.5 148.2 151.8 153.5 155.6 160.4 161.3 161.6  Spill  71.0  225.0  3.17  120.0  238.5  2.0  145.5  174.0 166.4  256.5  172.0 209.0  1.8  0.99 1.3  13.1 13.1  -  3.49 3.15  1.71 2.26  (  16.4( 13.5 14.3 13.7 15.6 14.1 14.2 14.4 12.9 17.0 13.6 13.2 12.4  -  Age in Days 55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77 78  Weight in GMS.  Plasma  Cholesterol mg%  Fat mgT,  F/C Ratio Actual Relative  -  166.6  -  .  180.1 182.9 182.7  124.0  177.0  1.4  2.45  117.5  196.5  1.7  2.90  130.0  126.0  0.99  1.71  116.7  289.0  2.50  -  188.9 189.1 185.5 192.1 185.4 193.6  -  202.2 203.3 209.9 207.7 212.1 218.8  16.4 16.9 14.4 82°F 14.7 82°F 15.6 82°F 13.9 74°F _ * 17.8  137.0  215.0  1.6  2.74  127.0  237.0  1.9  3.26  148.5  271.0  1.9  3.26  -  222.0 215.0 207.0  Feed Intake GMS/Animal  17.8 14.5 16.2 17.8  -  -  14.8  TABLE x i DAILY BODY WEIGHT, FEED INTAKE AND PLASMA LIPIDS OF PREPUBERTAL £ RAT. NO. 7 Age Weight in in Days GMS. 30 31 32 33 34 35 36 37 38 39 40 41 42 43 44 45 46 47 48 49 50 51 52 53 54  Plasma Cholesterol Fat mgT, mg%  F/C Ratio Actual Relative  Feed Intake GMS/Animal  Age in Days  18.3 18.3 - ( 15.9( 21.8 16.5 14.6 11.8 11.9 16.2 13.2 16.5 16.2 15.8 13.4 11.2  55 56 57 58 59 60 61 62 63 64 65 66 67 68 69 70 71 72 73 74 75 76 77 78  77.7 86.3  -  -  109.5  -  118.6 124.8 131.4  -  133.3 137.5 138.1 139.6 140.0 141.2 145.6 145.8 149.0 147.2 157.0 155.5 152.8  -  197.0  180.0  0.88  89.0  191.0  2.2  128.0  241.0  1.9  173.0  148.5 168.0  230.0  173.5 244.0  1.3  1.2 1.5  ].:-,.  -  Weight in GMS.  Plasma Cholesterol Fat Ol mgT, mg/o  F/C Ratio Actual Relative  160.0  _  -  • -  170.9 174.4 175.0  -  183.3 182.0 179.5 183.9 185.7 185.0  -  -  -  141.0  262.0  1.9  123.5  222.5  1.8  138.0  159.0  1.2  127.0  237.0  1.9  -  173.4 173.2 190.0 193.6 196.0 196.6  -  198.7 200.0 200.1  Feed Intake GMS/Animal  14.1 16.4 14.3  -  -  13.6 17.8 12.6 17.4 16.3  •-  118.0  205.0  1.7  102.0  206.0  2.0  121.0  174.0  1.4  hot  3.0 12.5 21.4 18.6  -  16.6 17.6  82°F 82°F 82°F 74°F  BIBLIOGRAPHY  185.  BIBLIOGRAPHY 1.  Adlersberg, D., Hormonal influences on the serum l i p i d s . 23: 769 - 789, 1957.  Am.  J.  Med.,  2.  Aftergood, L i l l a , Phd Dissertations, University of Southern 1956.  3.  Aftergood, L i l l a , Deuel, H. J . , J r . , and Roslyn B. A l f i n - S l a t e r . The comparative effe°cts of cottonseed o i l and l a r d on cholesterol levels i n the tissues of the r a t . J . Nut. 62: 129, 1957.  4.  A l f i n - S l a t e r , R. B., M. C. Schotz,. F.. Shimoda, and H. J . 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