0013-7227/78/1035-1924$02.00/0 Endocrinology Copyright © 1978 by The Endocrine Society
Vol. 103, No. 5 Printed in U.S.A.
Some Effects of Adrenalectomy in the Fatty Rat* Y. YUKIMURA, G. A. BRAY, AND A. R. WOLFSEN Department of Medicine, UCLA School of Medicine, Harbor General Hospital Campus, Torrance, California 90509 ABSTRACT. Function of the pituitary-adrenal axis was examined in lean and genetically obese fatty (Zucker) rats. The diurnal rhythms of plasma corticosterone and plasma ACTH were similar in both groups. The secretion of corticosterone by adrenal glands incubated in vitro with graded doses of ACTH was also comparable in lean and fatty rats. Adrenalectomy reduced food intake and weight gain in the fatty rats to levels similar to those in the lean animals and raised
W
E HAVE previously demonstrated that adrenalectomy would reduce food intake and lower the rate of weight gain in fatty rats to levels approximating those found in lean animals (1). This observation provided a mechanism for the fact that hypophysectomy could prevent the weight gain in fatty rats (2). We suggested that the fatty rat might represent a variety of hypercorticism analogous to that seen in patients with Cushing's disease. The present paper was designed to explore adrenal function in the fatty rat in detail. Two hypotheses were tested. First, we tested the possibility that the fatty rat might have an abnormal pituitary-adrenal axis. Second, we tested the hypothesis that the fatty rat might have an altered responsiveness to exogenous corticosterone or progesterone.
plasma ACTH to the same level. The injection of corticosterone (2 and 10 mg/day) increased food intake more in adrenalectomized fatty rats than in the corresponding lean ones. Progesterone increased food intake similarly in both groups. The enhanced responsiveness to corticosterone could account for many of the metabolical defects in the fatty rat. (Endocrinology 103: 1924, 1978)
tions were performed within 2 weeks after the animals arrived in the laboratory (3). After these operations, drinking water was changed to a 1% solution of NaCl. Procedures
A group of eight lean and eight intact fatty rats, aged 8-10 weeks, were sacrificed between 0900-1000 h and another group between 1600-1700 h. The time between removal from the cage and decapitation was less than 5 sec. Blood was collected in ice-cold tubes, separated in a refrigerated centrifuge, and frozen until assayed. Twenty-three fatty and 24 lean animals were adrenalectomized, and the additional eight pairs were kept as sham-operated controls. The protocol was divided into three phases. For the first 60 days after adrenalectomy, animals were weighed weekly but were otherwise undisturbed. A respiratory viral infection killed three adrenalectomized fatties and nine lean adrenalectomized controls. Food intake was measured Materials and Methods on individual animals between days 60-75 after Animals adrenalectomy. Phase 2 of the experiment began at day 75 with the last measurement of food intake. Forty-eight lean and 47 fatty (Zucker) female Groups of adrenalectomized fatty or lean animals rats were purchased from the Harriet G. Bird Memwere treated with vehicle or progesterone (2 or 10 orial Laboratories, Red Acre Farms, Stow, MA. mg/day for 10 days). This was followed by a period Animals were received at 6-8 weeks of age and of 2 weeks with no treatment. A second 10-day placed on Purina laboratory chow for the duration treatment period with dose levels of 0.5, 2.0, or 10 of the experiment. Adrenalectomy or sham operamg/day corticosterone was then instituted. Phase 3 of the study consisted of the infusion of corticosterone or vehicle iv at 1 or 3 jug/100 g BW/min for Received March 31,1978. Address all correspondence and requests for reprints a 45-min period, using a protocol identical to that to: George A. Bray, M.D., Harbor General Hospital, 1000 of Rotsztejn et al. (4). After the end of the infusion, West Carson Street, Torrance, California 90509. animals were sacrificed for measurement of ACTH * This work is supported in part by NIH Grant AMand corticosterone. 15165. 1924 The Endocrine Society. Downloaded from press.endocrine.org by [${individualUser.displayName}] on 14 November 2015. at 13:16 For personal use only. No other uses without permission. . All rights reserved.
ADRENAL FUNCTION IN FATTY RAT
Methods Plasma corticosterone was measured using a modification of the fluorometric technique described by Glick (5). Plasma ACTH was measured in a RIA using a commercial kit obtained from Amersham/Searle. All samples were run in a single assay. Data on this method has been previously published from this laboratory (6). Glucose was determined by the glucose oxidase method using a Beckman glucose analyzer, and triglycerides were measured with a Boehringer kit. Progesterone and corticosterone were purchased from Sigma Chemical Co. Quartered adrenal glands were incubated using the procedure of Tait (7). Pieces of adrenal were distributed into flasks and incubated for 90 min. Samples of 200 JLJ were withdrawn at 30, 60, and 90 min and assayed for corticosterone. Statistical comparison was done with Student's t test. Statistical significance was taken at a level of probability with P < 0.05.
1925
differences between fatty and lean animals (Table 1). Afternoon corticosterone and ACTH values in 30-week-old sham-operated fatty or lean animals were also not significantly different. The corticosterone values were significantly lower than observed in the 8 to 10-week-old animals, whether fat or lean, but the ACTH values were only significantly lower in the fatty rats. The response of quartered adrenals to ACTH showed no significant difference between fat and lean animals (Fig. 1). Corticosterone secretion in response to ACTH,
Results Adrenal function was assessed by measuring the diurnal rhythm of plasma corticosterone, the response of the quartered adrenal glands to ACTH, and the suppression of ACTH in adrenalectomized animals after infusion of corticosterone. At adrenalectomy, the adrenals of the 8 to 10-week-old fatty rats weighed 48.2 ± 2.0 mg (±SEM) and those of the lean rats weighed 42.5 ± 3.4 mg (P < 0.1). Diurnal variation in plasma corticosterone was identical in the fatty and lean animals (Table 1). Two of the lean and one young fatty rat had undetectably low values of corticosterone at 0900 h, but the mean value for both groups was identical. In the afternoon, the values were higher but again, the two groups were not significantly different. ACTH values showed a similar diurnal rhythm but with no
6.25
6.25 625 ACTH (mU/ml) FIG. 1. Response of the adrenal gland to ACTH in vitro. Quartered adrenal glands from lean or fatty rats were incubated with the indicated doses of ACTH for 90 min. Samples of incubation medium were removed at 30-min intervals, but only the data for the final time period have been plotted. There was no significant difference in concentrations of corticosterone at any time between the groups.
TABLE 1. Plasma corticosterone and ACTH in the morning and afternoon from lean and fatty rats Plasma ACTH (pg/ml)
Plasma corticosterone (/xg/dl)
Group
Age (weeks)
0900 h
1600 h
P
0900 h
1600 h
P
Lean rats
8-10 30
2.4 ± 1.1 (4)
46.8 ± 4.0 (8)° 17.4 ± 1.7 (8)°
Vol. 103, No. 5 Printed in U.S.A.
Some Effects of Adrenalectomy in the Fatty Rat* Y. YUKIMURA, G. A. BRAY, AND A. R. WOLFSEN Department of Medicine, UCLA School of Medicine, Harbor General Hospital Campus, Torrance, California 90509 ABSTRACT. Function of the pituitary-adrenal axis was examined in lean and genetically obese fatty (Zucker) rats. The diurnal rhythms of plasma corticosterone and plasma ACTH were similar in both groups. The secretion of corticosterone by adrenal glands incubated in vitro with graded doses of ACTH was also comparable in lean and fatty rats. Adrenalectomy reduced food intake and weight gain in the fatty rats to levels similar to those in the lean animals and raised
W
E HAVE previously demonstrated that adrenalectomy would reduce food intake and lower the rate of weight gain in fatty rats to levels approximating those found in lean animals (1). This observation provided a mechanism for the fact that hypophysectomy could prevent the weight gain in fatty rats (2). We suggested that the fatty rat might represent a variety of hypercorticism analogous to that seen in patients with Cushing's disease. The present paper was designed to explore adrenal function in the fatty rat in detail. Two hypotheses were tested. First, we tested the possibility that the fatty rat might have an abnormal pituitary-adrenal axis. Second, we tested the hypothesis that the fatty rat might have an altered responsiveness to exogenous corticosterone or progesterone.
plasma ACTH to the same level. The injection of corticosterone (2 and 10 mg/day) increased food intake more in adrenalectomized fatty rats than in the corresponding lean ones. Progesterone increased food intake similarly in both groups. The enhanced responsiveness to corticosterone could account for many of the metabolical defects in the fatty rat. (Endocrinology 103: 1924, 1978)
tions were performed within 2 weeks after the animals arrived in the laboratory (3). After these operations, drinking water was changed to a 1% solution of NaCl. Procedures
A group of eight lean and eight intact fatty rats, aged 8-10 weeks, were sacrificed between 0900-1000 h and another group between 1600-1700 h. The time between removal from the cage and decapitation was less than 5 sec. Blood was collected in ice-cold tubes, separated in a refrigerated centrifuge, and frozen until assayed. Twenty-three fatty and 24 lean animals were adrenalectomized, and the additional eight pairs were kept as sham-operated controls. The protocol was divided into three phases. For the first 60 days after adrenalectomy, animals were weighed weekly but were otherwise undisturbed. A respiratory viral infection killed three adrenalectomized fatties and nine lean adrenalectomized controls. Food intake was measured Materials and Methods on individual animals between days 60-75 after Animals adrenalectomy. Phase 2 of the experiment began at day 75 with the last measurement of food intake. Forty-eight lean and 47 fatty (Zucker) female Groups of adrenalectomized fatty or lean animals rats were purchased from the Harriet G. Bird Memwere treated with vehicle or progesterone (2 or 10 orial Laboratories, Red Acre Farms, Stow, MA. mg/day for 10 days). This was followed by a period Animals were received at 6-8 weeks of age and of 2 weeks with no treatment. A second 10-day placed on Purina laboratory chow for the duration treatment period with dose levels of 0.5, 2.0, or 10 of the experiment. Adrenalectomy or sham operamg/day corticosterone was then instituted. Phase 3 of the study consisted of the infusion of corticosterone or vehicle iv at 1 or 3 jug/100 g BW/min for Received March 31,1978. Address all correspondence and requests for reprints a 45-min period, using a protocol identical to that to: George A. Bray, M.D., Harbor General Hospital, 1000 of Rotsztejn et al. (4). After the end of the infusion, West Carson Street, Torrance, California 90509. animals were sacrificed for measurement of ACTH * This work is supported in part by NIH Grant AMand corticosterone. 15165. 1924 The Endocrine Society. Downloaded from press.endocrine.org by [${individualUser.displayName}] on 14 November 2015. at 13:16 For personal use only. No other uses without permission. . All rights reserved.
ADRENAL FUNCTION IN FATTY RAT
Methods Plasma corticosterone was measured using a modification of the fluorometric technique described by Glick (5). Plasma ACTH was measured in a RIA using a commercial kit obtained from Amersham/Searle. All samples were run in a single assay. Data on this method has been previously published from this laboratory (6). Glucose was determined by the glucose oxidase method using a Beckman glucose analyzer, and triglycerides were measured with a Boehringer kit. Progesterone and corticosterone were purchased from Sigma Chemical Co. Quartered adrenal glands were incubated using the procedure of Tait (7). Pieces of adrenal were distributed into flasks and incubated for 90 min. Samples of 200 JLJ were withdrawn at 30, 60, and 90 min and assayed for corticosterone. Statistical comparison was done with Student's t test. Statistical significance was taken at a level of probability with P < 0.05.
1925
differences between fatty and lean animals (Table 1). Afternoon corticosterone and ACTH values in 30-week-old sham-operated fatty or lean animals were also not significantly different. The corticosterone values were significantly lower than observed in the 8 to 10-week-old animals, whether fat or lean, but the ACTH values were only significantly lower in the fatty rats. The response of quartered adrenals to ACTH showed no significant difference between fat and lean animals (Fig. 1). Corticosterone secretion in response to ACTH,
Results Adrenal function was assessed by measuring the diurnal rhythm of plasma corticosterone, the response of the quartered adrenal glands to ACTH, and the suppression of ACTH in adrenalectomized animals after infusion of corticosterone. At adrenalectomy, the adrenals of the 8 to 10-week-old fatty rats weighed 48.2 ± 2.0 mg (±SEM) and those of the lean rats weighed 42.5 ± 3.4 mg (P < 0.1). Diurnal variation in plasma corticosterone was identical in the fatty and lean animals (Table 1). Two of the lean and one young fatty rat had undetectably low values of corticosterone at 0900 h, but the mean value for both groups was identical. In the afternoon, the values were higher but again, the two groups were not significantly different. ACTH values showed a similar diurnal rhythm but with no
6.25
6.25 625 ACTH (mU/ml) FIG. 1. Response of the adrenal gland to ACTH in vitro. Quartered adrenal glands from lean or fatty rats were incubated with the indicated doses of ACTH for 90 min. Samples of incubation medium were removed at 30-min intervals, but only the data for the final time period have been plotted. There was no significant difference in concentrations of corticosterone at any time between the groups.
TABLE 1. Plasma corticosterone and ACTH in the morning and afternoon from lean and fatty rats Plasma ACTH (pg/ml)
Plasma corticosterone (/xg/dl)
Group
Age (weeks)
0900 h
1600 h
P
0900 h
1600 h
P
Lean rats
8-10 30
2.4 ± 1.1 (4)
46.8 ± 4.0 (8)° 17.4 ± 1.7 (8)°
