Effects of ovariectomy and estradiol on body weight and food intake in gold thioglucose-treated mice

Physiology & Behavior, Vol. 17, pp. 1027-1030. Pergamon Press and Brain Research Publ., 1976. Printed in the U.S.A.

BRIEF COMMUNICATION Effects of Ovariectomy and Estradiol on Body Weight and Food Intake in Gold Thioglucose-Treated Mice 1'2 J E F F R E Y D. BLAUSTEIN, R. THOMAS GENTRY, EDWARD J. ROY AND GEORGE N. WADE

Department o f Psychology, University o f Massachusetts, Amherst, MA 01003 (Received 6 April 1976) BLAUSTEIN, J. D., R. T. GENTRY, E. J. ROY AND G. N. WADE. Effects of ovariectomy and estradiol on body weight and food intake in gold thioglucose.treated mice. PHYSIOL. BEHAV. 17(6) 1027-1030, 1976. - Two experiments investigated the effects of ovariectomy and estradiol replacement on the body weight and food intake of mice that had previously been treated with either gold thioglucose or saline. Ovariectomy and estradiol benzoate injections altered food intake in gold thioglucose-treated mice as much as in saline controls. Ovariectomy increased body weight in saline controls but it was without effect on the body weight of gold thioglucose-treated mice. Food intake

Body weight

Ovariectomy

Estradiol

OVARIAN hormones have striking influences on the eating behavior and body weight regulation of female rats [15,16]. Withdrawal of gonadal steroids by ovariectomy results in a transient hyperphagia and a permanent increase in body weight [10]. These changes can be reversed by treatment with free ( [ 3 ] ; Blaustein, unpublished observations) or esterified forms of estradiol-17~ [10,13] injected systemically. The ventromedial hypothalamus (VMH)-arcuate area might be a site of action since cannula implants of estradiol benzoate (EB) in this region result in a suppression of food intake [2, 6, 17] and body weight [6] and it is known that this is an area of high density of estradiol concentrating cells [ 12]. Although studies on effectiveness of VMH-arcuate implants on suppression of food intake and body weight have been consistent [2, 6, 17], studies on the effectiveness of VMH lesions blocking ovariectomy-induced or estradiol-induced changes in food intake have not. There have been reports of independence of the VMH and gonadal effects [7,14] as well as reports of attenuation of the gonadal effects by prior VMH lesions [2,11 ]. Gold thioglucose (GTG) injections in mice result in a hyperphagia and obesity that is similar to the syndrome which follows lesions of the VMH. It has been hypothesized that injections of GTG cause damage in an area

Gold thioglucose

purported to be involved in glucostatic regulation of eating [9]. The present experiments examined the effects of both withdrawal and replacement of estradiol in mice previously treated with GTG in order to determine if GTG caused damage in areas involved in estrogenic regulation of eating and body weight. Previous research had suggested the possibility of a common substrate for the effects of GTG and ovariectomy on body weight [ 18].

GENERAL METHOD Fifty female mice of the C57 B1/6J strain were purchased from Jackson Laboratories, Inc. (Bar Harbor, ME) and arrived in the laboratory weighing 18 g. All animals were housed individually in Wahmann LC-75 wire-bottom cages with unrestricted access to Purina Laboratory Chow and tap water. Environmental illumination was provided from 0800 to 2000 hours daily and room temperature was maintained at approximately 22 °C. Food intake and body weight were measured weekly to the nearest 0.1 g between 0900 and 1100 hours. Spillage was subtracted in computing weekly food intake. All comparisons are between groups and were evaluated by the Student's t-test. Unless otherwise specified, all

~This research was supported by Research Grant NS-10873 and Research Career Development Award NS-00090 from the National Institute of Neurological Diseases and Stroke, and Training Grant MH 11823 from the National Institutes of Health. We gratefully acknowledge the expert technical assistance of Marilyn Blaustein, Elizabeth Silver and Barbara Van Ryzin. Requests for reprints should be sent to Jeffrey D. Blaustein, Department of Psychology, Tobin Hall, University of Massachusetts, Amherst, Massachusetts, 01003. 2Estradiol benzoate was generouslyprovided by Dr. Preston Perlman, Schering Corp., Bloomfield, New Jersey. 1027

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BLAUSTEIN, GENTRY, ROY AND WADE

comparisons are two-tailed and considered statistically significant if p<0.05. 32

EXPERIMENT

1: G T G A N D O V A R I E C T O M Y

It has been well established that ovariectomy results in a transient hyperphagia and a permanent increase in body weight in rats. The purposes of the first experiment were to document this effect in mice of the C57 Black strain and to determine if prior injections of GTG would alter the hyperphagia and body weight gain. It was expected that if GTG destroyed areas that are involved in estrogenic control of body weight and eating, then GTG-injected mice would show a diminished increase in body weight and food intake after ovariectomy when compared with mice that had previously been injected with only saline. Procedure Part 1. Following one week of baseline recording of food intake and body weight, 36 mice were injected intraperitoneally with GTG (Sigma Chemical Co.; 0.6 mg/g body weight dissolved in 100 mg/ml saline), and 14 mice were injected with saline (SAL). Forty-three days later, each animal was given a supplementary injection of either GTG or saline of the same absolute quantity as the first. Part 2. Seventy days after the first GTG injection, 2 mice in the GTG group that had not gained >30% of their initial body weight were discarded. Of the remaining GTG mice, 29 were ovariectomized (GTG-OVX) and five were sham-operated (GTG-SHAM). Of the SAL group, 10 were ovariectomized (SAL-OVX) and 4 were sham-operated (SAL-SHAM). Ovariectomies were performed under methoxyflurane (Metofane) anesthesia through a single midventral incision. Sham ovariectomies consisted of only the midventral incision without disturbance of the ovaries or uterus. Results Part I. Following ovariectomy (Part 2), 9 animals died from surgical trauma. The data reported are based on the surviving animals, 25 of which had received saline injections. GTG resulted in an increase in body weight of approximately eight grams above saline controls by the time of ovariectomy seventy days after the first GTG injection, t(37) = 7.008, p<0.001 (Fig. 1; Week 0). The hyperphagia induced by GTG was not completely transient. At the time of ovariectomy, food intake and body weight had stabilized but food intake was still approximately 14% above saline injected control mice, t(37) = 4.382, p<0.001 (Fig. 1 ; Week 0). Part 2. Ovariectomy resulted in a transient increase in food intake compared with SHAMS in both GTG and SAL groups (Fig. 1). Food intake was significantly elevated for both groups during the third week postsurgically (GTG-OVX versus GTG-SHAM) t(23) = 2.188, p<0.05, (SAL-OVX versus SAL-SHAM) t(12) = 3.082, p<0.01. The magnitude of the increased food intake compared with SHAM-controls was similar for both the GTG-OVX (0.8 g/day) and the SAL-OVX groups i0.8 g/day). Ovariectomy increased body weight in the SAL-OVX group when compared with the SAL-SHAM group by the third postoperative week, t(12) = 2.586, p<0.05. Body

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FIG. l. Body weight and food intake following ovariectomy (OVX) or sham-ovariectomy (SHAM) in mice that had been injected 10 weeks prior with either gold thioglucose (GTG) or saline (SAL). Arrows indicate the day of surgery. Data are unavailable for the seventh week after surgery. weight then leveled off at this increased value. Ovariectomy had no effect on body weight in GTG-injected mice. At no time was body weight of the GTG-OVX group different from the SHAM controls (Fig. 1). EXPERIMENT

2: E S T R A D I O L

BENZOATE

TREATMENT

Experiment 1 demonstrated that GTG-injected mice show a hyperphagia following ovariectomy that is quantitatively similar to mice that had not been treated with GTG. However, body weight was not increased following ovariectomy in the GTG-OVX group although it was in the SAL-OVX group. The second experiment investigated the effects of estradiol replacement on these two measures. A dose of estradiol benzoate was used that has been shown to be adequate for restoration of sexual behavior when given prior to progesterone in ovariectomized mice [4]. Procedure

Eight weeks after ovariectomy, the GTG-OVX and SAL-OVX groups were subdivided. Eleven of the mice in the GTG-OVX group received daily subcutaneous injection of 1 ug EB dissolved in 0.02 ml sesame oil (GTG-OVX-EB) and 10 received daily injections of the oil vehicle alone (GTG-OVX-OIL). Five of the mice from the SAL-OVX group received daily EB injections (SAL-OVX-EB) and 5 served as oil controls (SAL-OVX-OIL). All injections were continued for 3 weeks.

O V A R I A N A N D GTG I N F L U E N C E S ON BODY W E I G H T

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Results A

Since it was n o t possible to m a t c h groups on food; intake and b o d y weight they were b o t h evaluated by examining the changes f r o m baseline expressed in grams. EB injections resulted in a transient decrease in food intake (Fig. 2). During the first week of t r e a t m e n t , b o t h the GTG-OVX-EB and SAL-OVX-EB groups were statistically different from their respective OIL-injected controls, t(19) = 3.383, p < 0 . 0 1 ; t(8) = 2.225, p < 0 . 0 5 , one-tailed test. During this period of m a x i m a l hypophagia, the f o o d intake of the GTG-OVX-EB group was suppressed 0.9 g/day and the SAL-OVX-EB group was suppressed 0.8 g/day. At no time during the course of the e x p e r i m e n t was the change in b o d y weight f r o m preinjection baseline in the EB-treated groups significantly different from the respective OIL-treated control groups. DISCUSSION

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The results of the first e x p e r i m e n t confirm the effects of GTG injections on the b o d y weight regulation and eating in this strain of mice. Part 2 d e m o n s t r a t e d that o v a r i e c t o m y in mice results in a transient hyperphagia and a p e r m a n e n t elevation in b o d y weight quite similarly to the case in rats. Body weights of mice that had been injected with GTG were not affected by ovariectomy, although the f o o d intake increase was of similar magnitude whether or not mice had been previously treated with GTG. This finding suggests two possible conclusions. The GTG-induced b o d y weight increase and the OVX-induced body weight increase have some cause in c o m m o n , since o v a r i e c t o m y was w i t h o u t additional effect in GTG-treated mice. This is consistent with the findings of Wright and Turner [18] that on increasing b o d y weight gain, GTG was less effective in mice ovariectomized 50 days previously than in non-castrates. It also indicates that under some circumstances, the body weight and f o o d intake changes which follow o v a r i e c t o m y are dissociable, a result that has also been observed in o t h e r syndromes (e.g. diabetes; [ 5 ] ). The second e x p e r i m e n t showed that EB suppresses eating in OVX mice as it does in rats. It is unclear why

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FIG. 2. Body weight and food intake, expressed in grams change from baseline of ovariectomized mice injected daffy with 1 /~g estradiol benzoate (EB) or sesame oil (OIL). Mice had been injected 18 weeks earlier with either gold thioglucose (GTG) or saline (SAL) and ovariectomized 8 weeks prior to start of treatment. Arrows indicate start of hormonal treatment. body weight was unaffected by EB in this study. However, as with ovariectomy, the magnitude of the food intake change was nearly identical whether or not animals had initially received GTG injections. This e x p e r i m e n t is consistent with the n o t i o n that although the effects of GTG and ovariectomy on body weight may share a c o m m o n substrate, the alterations in f o o d intake do not.

REFERENCES

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B L A U S T E I N , G E N T R Y , R O Y A N D WADE 17. Wade, G. N. and I. Zucker. Modulation of food intake and locomotor activity in female rats by diencephalic hormone implants. J. eomp. physiol. Psychol. 72: 328-336, 1970. 18. Wright, P. and C. Turner. Sex differences in body weight following gonadectomy and gold thioglucose injections in mice. PhysioL Behav. 11: 155-159, 1972.