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Food intake and body weight modifications following medial hypothalamic hormone implants in female rats
Phyaiolog'y and Behavior, Vot. 12. pp. 8"75--879. Brain Research Publications Inc., 1974. Printed in the U.S.A.
BRIEF COMMUNICATION Food Intake and Body Weight Modifications Following Medial Hypothalamic Hormone Implants in Female Rats RICHARD JANKOWlAK AND J E F F R E Y J. STERN 1
Michigan State Univern~ School o f Medicine and The Universi~ o f Michigan-Dearborn
(Received II October 1973) J A N K O W l A K , R. A N D J. J. STERN. Food intake and body weight modifications following medial hyporhalamic hormone
implants in/emale rats. PHYSIOL. BEHAV. 12 (5) 875-879, 1974. - Two groups of female rats had crystalline estradiol benzoate, progesterone and cholesterol implanted Ln the medici hypothalamus. One group was ovariectomized (OVX) and weighed approximately 300 g. The second group was adrenalectomized and ovaziectomized (ADX-OVX) and weighed approximately 250 $. EB placed in the ventromedial hypothalamus significantly reduced feeding in the OVX animtls; progesterone and cholesterol had no effect. In contrast, progesterone increased feeding in the lean ADX-OVX animals; EB and cholesterol were wathout effect. The most effective progesterone placements were in the dorsal medial hypothalamic nuclei. The data are discussed in terms of EB and progesterone influencing a h y p o t h ~ c set point for body weight. Body weight
Hormones
Ovariectomy
Adrenalectomy
THE FOOD INTAKE of female rats is partially dependent on ovarian hormones. During estrus (vaginal cornification) with its increased estrogen release there is a drop in feeding and reduced body weight [3]. Ovariectomy produces an abrupt rise in eating, exogenous estradiol returns food consumption to control level [20]. During pregnancy and pseudopregnancy rats display feeding changes which are correlated with increased progesterone and decreased estrogen titers [ 16,23 ]. The ability of estradiol and progesterone to affect eating is dependent on the rat's body weight. Estradiol suppresses feeding and body weight in overweight rats only [21 ]. Prolonged estrogenic stimulation does not produce an indefinite weight loss; feeding levels off [19]. Lastly, estradiol does not reduce eating in the lean, immature rat [24]. The weight promoting actions of progesterone are expressed in lean ammals only [18]. Exogenous progesterone does not influence feeding or body weight in ovariectomized (overweight) rats [7]. Adrenal removal prevents the weight gain that typically follows ovariectomy and permits exogenous progesterone to increase feeding
HypothaJamus
overweight condition of their animals, the differential effect of the steroids is not surprising. The present experiment was a replication of Wade and Zucker's study using both lean and overweight spayed rats. METHOD The animals were adult Sprague-Dawley female rats purchased from a commerical breeder (Spartan). Animals were housed in individual cages with food and water available ad lib. The laboratory was on a natural light cycle with the temperature thermostatically maintained at approximately 23 ° C. The females were randomly assigned to one of two groups. Group I (N=10) was ovanectomized (OVX) under pentobarbital anesthesia. The rats in Group 2 (N=I6) were adrenalectomized and ov~riectomized (ADX-OVX) under similarly induced anesthesia. After surgery the ADX-OVX animals were offered 0.5 M NaCI ad lib. One week later all animals had unilateral double wailed cannulas au'ned at the medial hypothalamus (DeGroot: A: 5.4; L: 0.8; V : - 3 . 4 o r - 2 . 6 ) ; again using Nembutal. The outer guides were 21 ga stainless steel hypodermic tubes to the skull with four No. 80 machine screws and dental acrylic resin; the inner removable cannulas were 27 ga tubes bent into right angles. On the day after implantation and continuing throughout the experiment food intake to the nearest 0.1 g was
[IS]. Wade and Zucker [22] report that unilateral implants of e s t r a d i o l benzoate (EB) in the ventromedial-arcuate (VMH-ARC) region of the diencephalon reduce feeding in spayed rats. They also report that progesterone in the same regaon does not significantly influence eating. Given the
' Repnnt requests should be sent to Jeffrey J. Stern, Department of Psychology. University of Michigan Dearborn. Michigan 48128 875
876
J A N K O W I A K AND S T E R N
measured daily at a p p r o x i m a t e l y 8 : 0 0 a.m.; note was t a k e n of spillage. The rats were also weighed to the nearest gram each m o r n i n g . Twenty-five to 35 days after i m p l a n t a t i o n (once feeding became stabilized), the inner cannulas were removed and tapped into u n d i l u t e d crystalline estradiol benzoate, progesterone or cholesterol. T h e tubes were placed in the VMH-ARC at approxamately 9 : 0 0 a.m. and secured to the o u t e r guide with a drop of acrylic resin. Prior to insertion the t u b a were e x a m i n e d u n d e r a microscope to insure t h a t only the l u m e n s c o n t a i n e d h o r m o n e . Animals were given 72 h r of c o n t i n u o u s s t i m u l a t i o n w i t h each agent in a r a n d o m order. O n the t h i r d day o f each t r e a t m e n t , vaginal smears were taken. Twelve days elapsed b e t w e e n t r e a t m e n t s . One week after the e x p e r i m e n t a l period, the ADX-OVX animals were tested to establish the c o m p l e t e n e s s of the adrenalectomies. An animal was considered c o m p l e t e l y ADX if it m e t the following criteria: (1) saline intake reliably greater than 10 ml/day, (2) a rapid weight loss followmg NaC1 withdrawal and, (3) weight recovery with salt replacement. All a n i m a l s discussed, satisfied these conditions. T h e animals were sacrificed with an overdose of sodium p e n t o b ~ b i t a l and perfused pericardially with 0.9% NaC1 followed by neutralized 10% F o r m a l i n . T h e brains were removed and fixed in Formalin. F r o z e n sections cut at 50 ~ m were stained with cresyl violet and e x a m i n e d microscopicaLly to d e t e r m i n e c a n n u l a placements. RESULTS
Ovanectomi:ed .4 nimals Table 1 shows the m e a n b o d y weights and food intakes of the females before (Days I - 5 ) , during (Days 6 - 8 ) and after (Days 9 - 1 2 ) i n t r a h y p o t h a l a m i c stimulation. Estradiol b e n z o a t e significantly decreased feeding (p< 0.01 paired t test) in O V X rats (Table 1). Figure l shows t h a t eating was reduced during the first 2 days of s t i m u l a t i o n ; by Day 3 feeding leveled off. Body weight declined on
TABLE
each of the three days. With EB removal, feeding and body weight r e t u r n e d to baseline (Table I; Fig. I). Of the i0 O V X ammals, 3 s h o w e d a r e d u c t i o n in feeding > 20°:0 with EB; 5 s h o w e d a decrease of I 0 - 2 0 % and 2 rats s h o w e d a decline of 0 - 1 0 % . Figure 2 shows the c a n n u l a p l a c e m e n t s for the animals. Progesterone and cholesterol i m p l a n t e d in the VMH had an insignificant effect o n food intake in O V X rats (Table I). T w o rats s h o w e d a decrease in feeding > 20% with progesterone stimulation, 2 displayed a decrease of I 0 - 2 0 % , while 6 s h o w e d a change of < 10%. Cholesterol modified feeding > I0% in only 2 of I0 cases. The vaginal smears t a k e n on the third day of s t i m u l a t i o n were anestrus in all (30) cases.
A drenatectomZzed-Ovartectom~zed A ntmats Prior to t r e a t m e n t , the ADX-OVX animals weighed m = 254 g and c o n s u m e d m = 22.2 g rat c h o w / d a y (Table 1). EB had an insignificant effect on these measures: of the 16 animals, 1 showed a decrease in feeding > 20%, 4 s h o w e d a decline of 1 0 - 2 0 % , while the remaining 11 displayed feeding changes < 10% (Table I ). Progesterone p r o d u c e d a significant (p< 0.02) increase in eating (Table 1, Fig. 1). During the 72 hours of s t i m u l a t i o n , there was an increase in feeding > 20% for 6 ammals, t 0 - 2 0 % for 5 animals and a change < 10% for 5 rats. Body weight was slightly (NS) elevated during t r e a t m e n t . Figure 2 shows the most effective placements of progesterone to be dorsal to the VMH, withan the b o u n d a r i e s of the dorsal medial h y p o t h a l a m u s (DMH). Cholesterol implants had no effect on either food intake or b o d y weight (Table 1). Again, vaginal smears were anestrus in all cases. DISCUSSION H y p o t h a l a m i c control of feeding is u n d e r s t o o d by some (see [ 1 3 1 ) tO be based o n a Go-No-Go regulatory system. Simply stated, the lateral h y p o t h a l a m u s (LH) is viewed as
1
MEAN FOOD INTAKES AND BODY WEIGHTS IN GRAMS : SEM OF THE FEMALES DURING THE VARIOUS STAGES OF EXPERIMENTATION
OVX Group IN = 10) Body Weight Food Intake
ADX-OVX Group ,N = 16~ Body Weight Food [make
Pretreatment
303 -- ti.5
29.2 : 1.2
254 : S.2
22.2 : 0.08
Treatment Cho| EB Prog
307 _- 11.4 291 = 9.8 310 _- 10.5
2S.6 : 1.1 25.5 : 0.9" 29.1 = 1.0
259 : S.3 "60 : 8.6 .'67 : "L5
21£-" ").S '2.7:[.0 26.6: :.5.-
Post-treatment
30,* = 11.2
2S.9 = l.O
263 : 9 0
21.9:t.0
*p
HYPOTHALAMIC HORMONE IMPLANT AND BODY WEIGHT
OVX
32
877
RATS
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FIG. t. Mean food intakes and body weights for the animals before IDays I-5), during (Days 6-8), and after ('Days 9-12) intrahypothalamic hormone stimulation. The OVX rats were stimulated with El}; the ADX-OVX animals received progesterone stimulation. ~ start of treatment, : termination of treatment. the Go signal: Electrical or chemical stimulation of the LH produces stimulus bound food seeking and. consummatory behavior; bilateral lesions in this region produce (allowing for recovery) aphagm, anorexia and a significant loss of body weight. The weight loss is interpreted as a secondary consequence to diminished eating. The VMH is seen as a core brain receptor, sensitive to oral, gastric and humoral factors [ 17]. Heightened VMH activity inhibits LH activity and feeding. Lesions in the VMH produce hyperphagia and a 2 - 3 fold increase in body weight. Recent findings, however, suggest a lipostatic regulatory. system of hypothalamic feeding control. Under this theory, the hypothalamus (1) establishes a should-be weight value for the ammal. (2) monitors existing body weight lipostatically (levels of adipose tissue are related to body weight [I0] ) and (3) corrects for any discrepancy between the set point and body weight by altering the animal's eating habits
and metabolism. Feeding changes are understood as secondary adjustments to a body weight-set point mismatch. The dynamic hyperphag~a consequent to VMH destruction, for example, is viewed as an adaptive process of weight gain in response to an elevated set point [8]. Once the body signal coincides with the set point, body weight plateaus and food intake returns to near control levels (static phase). LH destruction is understood as the lowering of the hypothalamic set point. Keesey and his group [9,12] have shown that weight loss prior to LH lesiomng eliminates or significantly shortens the postoperative period of aphagla and anorexia. In some cases, LH destruction is actually followed by hyperphagia. They also report that following recovery, LH lesioned male rats maintain a contrast percentage of control weight for as long as 250 days. These data are all consistent with the notion that the hypothalamus establishes a set point about which body weight
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JANKOWIAK AND STERN
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FIG. 2. Modified coronal sections depicung the effecuve EB placements for OVX rats ~top 2 boxes) and progesterone placements for ADX-OVX rats (bottom 2 boxes). Solid figures represent a change in feeding > 20%. Half t-flied figures represent a change of ;) [0<20%. Empty figures indicate a change or" < I0%.
equilibrates. LH d e s t r u c t i o n does not eliminate the Go slgnaJ for feeding but rather lowers the animal's should-be weight vatue, Aphag~a and anorexia are adaptive changes to acheive the lowered body weigl'~t. The d e p e n d e n c y of food intake o n ovarian h o r m o n e s (see i n t r o d u c t i o n ) and the present results are readily
incorporated into the lipostatzc theory of feeding control. Given an overweight rat, estradiol lowers ~,he h y p o t h a l a m i c set point and eating adaptively decreases until coincidence b e t w e e n the set point and body weight is reached. Progesterone tends to razse the b o d y ' s set point and a lean rat will adaptively increase its feeding until the new level has been
H Y P O T H A L A M I C H O R M O N E I M P L A N T AND BODY W E I G H T r e a c h e d . A delicate e s t r o g e n - p r o g e s t a t i o n a l balance is established; any p r o c e d u r e w h i c h shifts the balance t o w a r d estrogen will w i t h i n limits [ 11.18 ] reduce feeding and b o d y weight. A shift toward p r o g e s t e r o n e will increase these measures, aga.m w i t h i n limits. As discussed by R o b e r t s er ai. [ 15 ] the only negative evidence for tins h y p o t h e s i s comes from a report by Hervey et al. [ 7 ] : h y p o p h y s e c t o m i z e d female rats (lean) do n o t gaan weight or increase feeding in response to e x o g e n o u s progesterone. Tins finding implicates the pituitary in the a d j u s t m e n t s m a d e by a rat to a b o d y weight-set p o m t i m b a l a n c e and is w o r t h y of f u r t h e r investigation. T h e present findings clearly maplicate the medial h y p o t h a l a m u s as the region w h i c h m e d i a t e s the actions of ovarian h o r m o n e s o n feeding. " T h e fact t h a t this part of the brain c o n t a i n s receptors o u g h t not to be t o o surprising. In e m b r y o n i c d e v e l o p m e n t this middle part of the CNS is derived from the m o s t dorsal part of the tissue t h a t becomes the CNS. This tussue derives from the same origins as does s k i n . . . " ([131 p. 174]. T h e present findings also suggest two a n a t o m i c a l l y separate h o r m o n e r e c e p t o r sites in the medial h y p o t h a l a m u s : The VMH sensitive to estradiol fin a g r e e m e n t with Wade and Zucker, [221 ) and the DMH sensitive to progesterone. Electrolytic lesions in the DMH p r o d u c e h y p o p h a g i a and weight loss in b o t h weanling and
879
adult rats [ 1,2]. It is possible t h a t the observed h o r m o n a l effects were m e d i a t e d by n o n - h y p o t h a l a m i c sites following leakage into the systemic circulation a n d / o r ventricles. T h e finding that vaginal smears were c o n s i s t e n t l y anestrous strongly argues against the first possibility. Direct m e a s u r e m e n t s of b l o o d h o r m o n e levels, however, are necessary before a definitive rejection can be made. The second possibility seems unlikely because two of the least effective h o r m o n e placem e n t s b o r d e r e d on the third ventricle fFig. 2). Here again, direct tests e.g. using labeled h o r m o n e s , are necessary. Up to n o w we have discussed only feeding changes in response to a set p o i n t - b o d y weight ( c o m p o s i t i o n ) mism a t c h . It appears t h a t the h y p o t h a l a m u s also initiates m e t a b o l i c a d j u s t m e n t s to such a discrepancy. VMH damaged rats tube fed an a m o u n t of food eqtuvalent to shams, show increased b o d y fat levels [ 5 , 6 ] . VMH animals show an increase in carcass fat percentage o n c o n t r o l diets [ 1 4 ] . Finally, Collier [4] has s h o w n t h a t during c o m p l e t e starvation (the set p o i n t has p r e s u m a b l y n o t been altered) active and n o n a c t i v e rats lose weight at precisely the same rate. In o t h e r words, given a set p o i n t - b o d y weight imbalance, the h y p o t h a l a m u s alters b o t h the rat's earing habits and basac m e t a b o l i s m until c o i n c i d e n c e h a s b e e n attained.
REFERENCES i. Bemardis, L. L. and L. A. Frohman. Effect of hypothalamic lesions at different lo~ on development of hyperinsulinemla and obesity in the weanling rat. J. comp. NeuroL 141: 107-118. 1971. 2. Bemaxdis, L. L. Hypopha~a, hypodipsaa and hypoactivity following electrolytic lesions in the dorsomed~l hypothalamic nuclei of mature rats of both sexes. J. Neural Transmisaon 33: 1-10, 1972. 3. Brobeck, J. R., M. Wh~tiand and J. L. Strominger. Variations in regulation of energy exchange associated wath estrus. diestrus and pseudopresmancy in rats. Endocrinology 40: 65-~2, 1947. 4. Collier, G., E. Hirsch and A. I. Leshner. The matabolic cost of activity in activity-naive rats. Physiol. Behav. 3: 881-884, 1972. 5. Han. P. W. Hypothalarnic obesity in rats without hyperphagia. Trans..V.Y. Acad. Sci. Series 11 30: 229-243. 1967. 6. Han. P. W. Obesity in force-fed rats bearing hypothalamic lesions. Pro¢. Soc. exp. Biol. Mea. 127: 1057-1060, t968. 7. Hervey. E. and G. R. Hervey. The relauonship between the effects of ovariectomy and progesterone treatment on body weight and composition in the female rat. J. PhysioL (Lond.) 187: 4 4 - 4 5 , 1966. 8. Heobel, B. G. and P. Teitelbaum. Weight regulation in normal and hypothalamic hyperphagic rats. J. comp. physiol. P~chol. 61: 189-193, 1966. 9. Keesey, R. E. and P. C. Boyle. Effects of quinine adulteration upon body weight of LH-lesioned and intact male rats. J. comp. physiol. Ps'ychol. 84: 38-46, 1973. 10. Kennedy, G. C. The role of depot fat in the hypothalamic control of food intake in the rat. Proc. Royal Soc. Series B 140: 578-592. 1953. 11. Mook, D. G., N. J. Kenney, S. Roberts, A. I. Nussbaum and W. 1. Rodier. Ovartan-adrenal interactions in regulation of body wei.~'at by female rats. J. comp. physiol, t~.chol. 81: 198-211. 1972. 12. Powley, T. L. and R. E. Keesey. Relationship of body weight to the lateral hypothalamic feeding syndrome. Z comp. physiol. P~'chol. 70: 25-36, 1970.
13. Pribram, IC H. Lan&'~Kesof the Brain. Englewood Cliffs, NJ: l%enuce-Hall, Inc. 1971. 14. Remley, N. R., W. P. Brittam and J. D. Sea$o. Absence of finickness in VMH-lesioned non-obese rats. Paper presented at Psychonomic Socaety Meeting, San Antonio, Texas 1970. 15. Roberts, S., N. J. Kenney and D. G. Mook. Overeating induced by progesterone in the ovanectomized, adrenalectomized rat. Horm. Behav. 3: 267-276, 1972. 16. Slonaker, J. R. The effect of copulation, pregnancy, pseudopregnancy and lactation on the voluntary actavity and food consumption of the albino rat. Am. J.. Physiol. 71: 362-394, 1925. 17. Stellar, E. Hunger in man: Comparative and physiological studies. Amer. Psychologrst 22: 105-117, 1967. 18. Stem, J. J., A. L. Porterfield and R. J. K,rupa. Endocrine interactions in the regulation of body weight by female rats. Y. comp. physwL PsychoL in press. 19. Tatxtelin. M. F. and R. A. Gorski. Int'luence of ovartan steroids on the regulation of food intake in the rat. Fedn Proc. 30: 253, 1971. 20. Valenstein, E. S., V. C. Cox and J. W. K'okolewski. Sex differences in hyperpha~a and body weight following hypothalamic damage. Ann. N.Y. Acad. Sci. 157: 1030-1048, 1969. 21. Wade, G. N. and I. Zucker. Hormonal and developmental influences on rat saccharine preferences. J. comp. physiol. Psychol. 69: 291-300, 1969. 22. Wade, G. N. and 1. Zucket. Modulation of food intake and locomotor activity in female rats by diencephalic hormone implants. J. comp. physiol. Prychol. 72: 328-336, 1970. 23. Zucker, I. Hormonal determinants of sex differences in saccharine preference, food intake and body weight. Physiol. Behav. 4: 595-602, 1969. 24. Zucker, I. BOdy weight and age as factors determining estrogen responsiveness in the rat feeding system. Behav. Biol. 7: 527-542, 1972.
BRIEF COMMUNICATION Food Intake and Body Weight Modifications Following Medial Hypothalamic Hormone Implants in Female Rats RICHARD JANKOWlAK AND J E F F R E Y J. STERN 1
Michigan State Univern~ School o f Medicine and The Universi~ o f Michigan-Dearborn
(Received II October 1973) J A N K O W l A K , R. A N D J. J. STERN. Food intake and body weight modifications following medial hyporhalamic hormone
implants in/emale rats. PHYSIOL. BEHAV. 12 (5) 875-879, 1974. - Two groups of female rats had crystalline estradiol benzoate, progesterone and cholesterol implanted Ln the medici hypothalamus. One group was ovariectomized (OVX) and weighed approximately 300 g. The second group was adrenalectomized and ovaziectomized (ADX-OVX) and weighed approximately 250 $. EB placed in the ventromedial hypothalamus significantly reduced feeding in the OVX animtls; progesterone and cholesterol had no effect. In contrast, progesterone increased feeding in the lean ADX-OVX animals; EB and cholesterol were wathout effect. The most effective progesterone placements were in the dorsal medial hypothalamic nuclei. The data are discussed in terms of EB and progesterone influencing a h y p o t h ~ c set point for body weight. Body weight
Hormones
Ovariectomy
Adrenalectomy
THE FOOD INTAKE of female rats is partially dependent on ovarian hormones. During estrus (vaginal cornification) with its increased estrogen release there is a drop in feeding and reduced body weight [3]. Ovariectomy produces an abrupt rise in eating, exogenous estradiol returns food consumption to control level [20]. During pregnancy and pseudopregnancy rats display feeding changes which are correlated with increased progesterone and decreased estrogen titers [ 16,23 ]. The ability of estradiol and progesterone to affect eating is dependent on the rat's body weight. Estradiol suppresses feeding and body weight in overweight rats only [21 ]. Prolonged estrogenic stimulation does not produce an indefinite weight loss; feeding levels off [19]. Lastly, estradiol does not reduce eating in the lean, immature rat [24]. The weight promoting actions of progesterone are expressed in lean ammals only [18]. Exogenous progesterone does not influence feeding or body weight in ovariectomized (overweight) rats [7]. Adrenal removal prevents the weight gain that typically follows ovariectomy and permits exogenous progesterone to increase feeding
HypothaJamus
overweight condition of their animals, the differential effect of the steroids is not surprising. The present experiment was a replication of Wade and Zucker's study using both lean and overweight spayed rats. METHOD The animals were adult Sprague-Dawley female rats purchased from a commerical breeder (Spartan). Animals were housed in individual cages with food and water available ad lib. The laboratory was on a natural light cycle with the temperature thermostatically maintained at approximately 23 ° C. The females were randomly assigned to one of two groups. Group I (N=10) was ovanectomized (OVX) under pentobarbital anesthesia. The rats in Group 2 (N=I6) were adrenalectomized and ov~riectomized (ADX-OVX) under similarly induced anesthesia. After surgery the ADX-OVX animals were offered 0.5 M NaCI ad lib. One week later all animals had unilateral double wailed cannulas au'ned at the medial hypothalamus (DeGroot: A: 5.4; L: 0.8; V : - 3 . 4 o r - 2 . 6 ) ; again using Nembutal. The outer guides were 21 ga stainless steel hypodermic tubes to the skull with four No. 80 machine screws and dental acrylic resin; the inner removable cannulas were 27 ga tubes bent into right angles. On the day after implantation and continuing throughout the experiment food intake to the nearest 0.1 g was
[IS]. Wade and Zucker [22] report that unilateral implants of e s t r a d i o l benzoate (EB) in the ventromedial-arcuate (VMH-ARC) region of the diencephalon reduce feeding in spayed rats. They also report that progesterone in the same regaon does not significantly influence eating. Given the
' Repnnt requests should be sent to Jeffrey J. Stern, Department of Psychology. University of Michigan Dearborn. Michigan 48128 875
876
J A N K O W I A K AND S T E R N
measured daily at a p p r o x i m a t e l y 8 : 0 0 a.m.; note was t a k e n of spillage. The rats were also weighed to the nearest gram each m o r n i n g . Twenty-five to 35 days after i m p l a n t a t i o n (once feeding became stabilized), the inner cannulas were removed and tapped into u n d i l u t e d crystalline estradiol benzoate, progesterone or cholesterol. T h e tubes were placed in the VMH-ARC at approxamately 9 : 0 0 a.m. and secured to the o u t e r guide with a drop of acrylic resin. Prior to insertion the t u b a were e x a m i n e d u n d e r a microscope to insure t h a t only the l u m e n s c o n t a i n e d h o r m o n e . Animals were given 72 h r of c o n t i n u o u s s t i m u l a t i o n w i t h each agent in a r a n d o m order. O n the t h i r d day o f each t r e a t m e n t , vaginal smears were taken. Twelve days elapsed b e t w e e n t r e a t m e n t s . One week after the e x p e r i m e n t a l period, the ADX-OVX animals were tested to establish the c o m p l e t e n e s s of the adrenalectomies. An animal was considered c o m p l e t e l y ADX if it m e t the following criteria: (1) saline intake reliably greater than 10 ml/day, (2) a rapid weight loss followmg NaC1 withdrawal and, (3) weight recovery with salt replacement. All a n i m a l s discussed, satisfied these conditions. T h e animals were sacrificed with an overdose of sodium p e n t o b ~ b i t a l and perfused pericardially with 0.9% NaC1 followed by neutralized 10% F o r m a l i n . T h e brains were removed and fixed in Formalin. F r o z e n sections cut at 50 ~ m were stained with cresyl violet and e x a m i n e d microscopicaLly to d e t e r m i n e c a n n u l a placements. RESULTS
Ovanectomi:ed .4 nimals Table 1 shows the m e a n b o d y weights and food intakes of the females before (Days I - 5 ) , during (Days 6 - 8 ) and after (Days 9 - 1 2 ) i n t r a h y p o t h a l a m i c stimulation. Estradiol b e n z o a t e significantly decreased feeding (p< 0.01 paired t test) in O V X rats (Table 1). Figure l shows t h a t eating was reduced during the first 2 days of s t i m u l a t i o n ; by Day 3 feeding leveled off. Body weight declined on
TABLE
each of the three days. With EB removal, feeding and body weight r e t u r n e d to baseline (Table I; Fig. I). Of the i0 O V X ammals, 3 s h o w e d a r e d u c t i o n in feeding > 20°:0 with EB; 5 s h o w e d a decrease of I 0 - 2 0 % and 2 rats s h o w e d a decline of 0 - 1 0 % . Figure 2 shows the c a n n u l a p l a c e m e n t s for the animals. Progesterone and cholesterol i m p l a n t e d in the VMH had an insignificant effect o n food intake in O V X rats (Table I). T w o rats s h o w e d a decrease in feeding > 20% with progesterone stimulation, 2 displayed a decrease of I 0 - 2 0 % , while 6 s h o w e d a change of < 10%. Cholesterol modified feeding > I0% in only 2 of I0 cases. The vaginal smears t a k e n on the third day of s t i m u l a t i o n were anestrus in all (30) cases.
A drenatectomZzed-Ovartectom~zed A ntmats Prior to t r e a t m e n t , the ADX-OVX animals weighed m = 254 g and c o n s u m e d m = 22.2 g rat c h o w / d a y (Table 1). EB had an insignificant effect on these measures: of the 16 animals, 1 showed a decrease in feeding > 20%, 4 s h o w e d a decline of 1 0 - 2 0 % , while the remaining 11 displayed feeding changes < 10% (Table I ). Progesterone p r o d u c e d a significant (p< 0.02) increase in eating (Table 1, Fig. 1). During the 72 hours of s t i m u l a t i o n , there was an increase in feeding > 20% for 6 ammals, t 0 - 2 0 % for 5 animals and a change < 10% for 5 rats. Body weight was slightly (NS) elevated during t r e a t m e n t . Figure 2 shows the most effective placements of progesterone to be dorsal to the VMH, withan the b o u n d a r i e s of the dorsal medial h y p o t h a l a m u s (DMH). Cholesterol implants had no effect on either food intake or b o d y weight (Table 1). Again, vaginal smears were anestrus in all cases. DISCUSSION H y p o t h a l a m i c control of feeding is u n d e r s t o o d by some (see [ 1 3 1 ) tO be based o n a Go-No-Go regulatory system. Simply stated, the lateral h y p o t h a l a m u s (LH) is viewed as
1
MEAN FOOD INTAKES AND BODY WEIGHTS IN GRAMS : SEM OF THE FEMALES DURING THE VARIOUS STAGES OF EXPERIMENTATION
OVX Group IN = 10) Body Weight Food Intake
ADX-OVX Group ,N = 16~ Body Weight Food [make
Pretreatment
303 -- ti.5
29.2 : 1.2
254 : S.2
22.2 : 0.08
Treatment Cho| EB Prog
307 _- 11.4 291 = 9.8 310 _- 10.5
2S.6 : 1.1 25.5 : 0.9" 29.1 = 1.0
259 : S.3 "60 : 8.6 .'67 : "L5
21£-" ").S '2.7:[.0 26.6: :.5.-
Post-treatment
30,* = 11.2
2S.9 = l.O
263 : 9 0
21.9:t.0
*p
HYPOTHALAMIC HORMONE IMPLANT AND BODY WEIGHT
OVX
32
877
RATS
I
29o)
26
fl 2
I 2
, 4
I 6
o s
28op,,
,o ,2
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4
DAYS
,
6
8
,
,
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DAYS A
.~
,
DX- OV X RATS
-
~
270
"
"
z_
g 25o o 24or~~~~~~
22 2
4
6 8 DAYS
I0 12
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6 8 DAYS
J I0
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FIG. t. Mean food intakes and body weights for the animals before IDays I-5), during (Days 6-8), and after ('Days 9-12) intrahypothalamic hormone stimulation. The OVX rats were stimulated with El}; the ADX-OVX animals received progesterone stimulation. ~ start of treatment, : termination of treatment. the Go signal: Electrical or chemical stimulation of the LH produces stimulus bound food seeking and. consummatory behavior; bilateral lesions in this region produce (allowing for recovery) aphagm, anorexia and a significant loss of body weight. The weight loss is interpreted as a secondary consequence to diminished eating. The VMH is seen as a core brain receptor, sensitive to oral, gastric and humoral factors [ 17]. Heightened VMH activity inhibits LH activity and feeding. Lesions in the VMH produce hyperphagia and a 2 - 3 fold increase in body weight. Recent findings, however, suggest a lipostatic regulatory. system of hypothalamic feeding control. Under this theory, the hypothalamus (1) establishes a should-be weight value for the ammal. (2) monitors existing body weight lipostatically (levels of adipose tissue are related to body weight [I0] ) and (3) corrects for any discrepancy between the set point and body weight by altering the animal's eating habits
and metabolism. Feeding changes are understood as secondary adjustments to a body weight-set point mismatch. The dynamic hyperphag~a consequent to VMH destruction, for example, is viewed as an adaptive process of weight gain in response to an elevated set point [8]. Once the body signal coincides with the set point, body weight plateaus and food intake returns to near control levels (static phase). LH destruction is understood as the lowering of the hypothalamic set point. Keesey and his group [9,12] have shown that weight loss prior to LH lesiomng eliminates or significantly shortens the postoperative period of aphagla and anorexia. In some cases, LH destruction is actually followed by hyperphagia. They also report that following recovery, LH lesioned male rats maintain a contrast percentage of control weight for as long as 250 days. These data are all consistent with the notion that the hypothalamus establishes a set point about which body weight
8"/8
JANKOWIAK AND STERN
0
-I
-2
-3
A 5.25 i
l
I
I
RH
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- I I-
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~\
/I
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=2
-3
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FIG. 2. Modified coronal sections depicung the effecuve EB placements for OVX rats ~top 2 boxes) and progesterone placements for ADX-OVX rats (bottom 2 boxes). Solid figures represent a change in feeding > 20%. Half t-flied figures represent a change of ;) [0<20%. Empty figures indicate a change or" < I0%.
equilibrates. LH d e s t r u c t i o n does not eliminate the Go slgnaJ for feeding but rather lowers the animal's should-be weight vatue, Aphag~a and anorexia are adaptive changes to acheive the lowered body weigl'~t. The d e p e n d e n c y of food intake o n ovarian h o r m o n e s (see i n t r o d u c t i o n ) and the present results are readily
incorporated into the lipostatzc theory of feeding control. Given an overweight rat, estradiol lowers ~,he h y p o t h a l a m i c set point and eating adaptively decreases until coincidence b e t w e e n the set point and body weight is reached. Progesterone tends to razse the b o d y ' s set point and a lean rat will adaptively increase its feeding until the new level has been
H Y P O T H A L A M I C H O R M O N E I M P L A N T AND BODY W E I G H T r e a c h e d . A delicate e s t r o g e n - p r o g e s t a t i o n a l balance is established; any p r o c e d u r e w h i c h shifts the balance t o w a r d estrogen will w i t h i n limits [ 11.18 ] reduce feeding and b o d y weight. A shift toward p r o g e s t e r o n e will increase these measures, aga.m w i t h i n limits. As discussed by R o b e r t s er ai. [ 15 ] the only negative evidence for tins h y p o t h e s i s comes from a report by Hervey et al. [ 7 ] : h y p o p h y s e c t o m i z e d female rats (lean) do n o t gaan weight or increase feeding in response to e x o g e n o u s progesterone. Tins finding implicates the pituitary in the a d j u s t m e n t s m a d e by a rat to a b o d y weight-set p o m t i m b a l a n c e and is w o r t h y of f u r t h e r investigation. T h e present findings clearly maplicate the medial h y p o t h a l a m u s as the region w h i c h m e d i a t e s the actions of ovarian h o r m o n e s o n feeding. " T h e fact t h a t this part of the brain c o n t a i n s receptors o u g h t not to be t o o surprising. In e m b r y o n i c d e v e l o p m e n t this middle part of the CNS is derived from the m o s t dorsal part of the tissue t h a t becomes the CNS. This tussue derives from the same origins as does s k i n . . . " ([131 p. 174]. T h e present findings also suggest two a n a t o m i c a l l y separate h o r m o n e r e c e p t o r sites in the medial h y p o t h a l a m u s : The VMH sensitive to estradiol fin a g r e e m e n t with Wade and Zucker, [221 ) and the DMH sensitive to progesterone. Electrolytic lesions in the DMH p r o d u c e h y p o p h a g i a and weight loss in b o t h weanling and
879
adult rats [ 1,2]. It is possible t h a t the observed h o r m o n a l effects were m e d i a t e d by n o n - h y p o t h a l a m i c sites following leakage into the systemic circulation a n d / o r ventricles. T h e finding that vaginal smears were c o n s i s t e n t l y anestrous strongly argues against the first possibility. Direct m e a s u r e m e n t s of b l o o d h o r m o n e levels, however, are necessary before a definitive rejection can be made. The second possibility seems unlikely because two of the least effective h o r m o n e placem e n t s b o r d e r e d on the third ventricle fFig. 2). Here again, direct tests e.g. using labeled h o r m o n e s , are necessary. Up to n o w we have discussed only feeding changes in response to a set p o i n t - b o d y weight ( c o m p o s i t i o n ) mism a t c h . It appears t h a t the h y p o t h a l a m u s also initiates m e t a b o l i c a d j u s t m e n t s to such a discrepancy. VMH damaged rats tube fed an a m o u n t of food eqtuvalent to shams, show increased b o d y fat levels [ 5 , 6 ] . VMH animals show an increase in carcass fat percentage o n c o n t r o l diets [ 1 4 ] . Finally, Collier [4] has s h o w n t h a t during c o m p l e t e starvation (the set p o i n t has p r e s u m a b l y n o t been altered) active and n o n a c t i v e rats lose weight at precisely the same rate. In o t h e r words, given a set p o i n t - b o d y weight imbalance, the h y p o t h a l a m u s alters b o t h the rat's earing habits and basac m e t a b o l i s m until c o i n c i d e n c e h a s b e e n attained.
REFERENCES i. Bemardis, L. L. and L. A. Frohman. Effect of hypothalamic lesions at different lo~ on development of hyperinsulinemla and obesity in the weanling rat. J. comp. NeuroL 141: 107-118. 1971. 2. Bemaxdis, L. L. Hypopha~a, hypodipsaa and hypoactivity following electrolytic lesions in the dorsomed~l hypothalamic nuclei of mature rats of both sexes. J. Neural Transmisaon 33: 1-10, 1972. 3. Brobeck, J. R., M. Wh~tiand and J. L. Strominger. Variations in regulation of energy exchange associated wath estrus. diestrus and pseudopresmancy in rats. Endocrinology 40: 65-~2, 1947. 4. Collier, G., E. Hirsch and A. I. Leshner. The matabolic cost of activity in activity-naive rats. Physiol. Behav. 3: 881-884, 1972. 5. Han. P. W. Hypothalarnic obesity in rats without hyperphagia. Trans..V.Y. Acad. Sci. Series 11 30: 229-243. 1967. 6. Han. P. W. Obesity in force-fed rats bearing hypothalamic lesions. Pro¢. Soc. exp. Biol. Mea. 127: 1057-1060, t968. 7. Hervey. E. and G. R. Hervey. The relauonship between the effects of ovariectomy and progesterone treatment on body weight and composition in the female rat. J. PhysioL (Lond.) 187: 4 4 - 4 5 , 1966. 8. Heobel, B. G. and P. Teitelbaum. Weight regulation in normal and hypothalamic hyperphagic rats. J. comp. physiol. P~chol. 61: 189-193, 1966. 9. Keesey, R. E. and P. C. Boyle. Effects of quinine adulteration upon body weight of LH-lesioned and intact male rats. J. comp. physiol. Ps'ychol. 84: 38-46, 1973. 10. Kennedy, G. C. The role of depot fat in the hypothalamic control of food intake in the rat. Proc. Royal Soc. Series B 140: 578-592. 1953. 11. Mook, D. G., N. J. Kenney, S. Roberts, A. I. Nussbaum and W. 1. Rodier. Ovartan-adrenal interactions in regulation of body wei.~'at by female rats. J. comp. physiol, t~.chol. 81: 198-211. 1972. 12. Powley, T. L. and R. E. Keesey. Relationship of body weight to the lateral hypothalamic feeding syndrome. Z comp. physiol. P~'chol. 70: 25-36, 1970.
13. Pribram, IC H. Lan&'~Kesof the Brain. Englewood Cliffs, NJ: l%enuce-Hall, Inc. 1971. 14. Remley, N. R., W. P. Brittam and J. D. Sea$o. Absence of finickness in VMH-lesioned non-obese rats. Paper presented at Psychonomic Socaety Meeting, San Antonio, Texas 1970. 15. Roberts, S., N. J. Kenney and D. G. Mook. Overeating induced by progesterone in the ovanectomized, adrenalectomized rat. Horm. Behav. 3: 267-276, 1972. 16. Slonaker, J. R. The effect of copulation, pregnancy, pseudopregnancy and lactation on the voluntary actavity and food consumption of the albino rat. Am. J.. Physiol. 71: 362-394, 1925. 17. Stellar, E. Hunger in man: Comparative and physiological studies. Amer. Psychologrst 22: 105-117, 1967. 18. Stem, J. J., A. L. Porterfield and R. J. K,rupa. Endocrine interactions in the regulation of body weight by female rats. Y. comp. physwL PsychoL in press. 19. Tatxtelin. M. F. and R. A. Gorski. Int'luence of ovartan steroids on the regulation of food intake in the rat. Fedn Proc. 30: 253, 1971. 20. Valenstein, E. S., V. C. Cox and J. W. K'okolewski. Sex differences in hyperpha~a and body weight following hypothalamic damage. Ann. N.Y. Acad. Sci. 157: 1030-1048, 1969. 21. Wade, G. N. and I. Zucker. Hormonal and developmental influences on rat saccharine preferences. J. comp. physiol. Psychol. 69: 291-300, 1969. 22. Wade, G. N. and 1. Zucket. Modulation of food intake and locomotor activity in female rats by diencephalic hormone implants. J. comp. physiol. Prychol. 72: 328-336, 1970. 23. Zucker, I. Hormonal determinants of sex differences in saccharine preference, food intake and body weight. Physiol. Behav. 4: 595-602, 1969. 24. Zucker, I. BOdy weight and age as factors determining estrogen responsiveness in the rat feeding system. Behav. Biol. 7: 527-542, 1972.