BEHAVIORALBIOLOGY21, 380--392 (1977)
Some Effects of Ventromedial Hypothalamic Lesions, Adrenalectomies, and Corticosterone Replacement Therapy on Energy Expenditure in Rats la,3 CHRISTOPHER WILSON
Chemistry of Behavior Program, Texas Christian University, Fort Worth, Texas 76129 The present study tested the hypothesis that the decreases in energy expenditure reported following ventromedial hypothalamic (VMH) lesions in rats are a function of the decreases in adrenal function. Five groups of rats received 21 lhr/day sessions in activity wheels and then were subjected to either VMH lesions, adrenalectomies, the respective sham-operations, or no operation. Following surgery, each rat was again given access to an activity wheel for 24 1-hr test sessions. During the test phase the rats received a sequence of drug/no drug conditions which consisted of: (1) no injection; (2) injections of the vehicle substance: or (3) injections of corticosterone in the vehicle substance. Under the no-drug conditions, running levels of the VMH-lesioned and adrenalectomized rats decreased, with the levels of VMH-lesioned rats being more severely depressed. Corticosterone replacement therapy reinstated the activity levels of the adrenalectomized rats, but not those of the VMH rats. The hypothesis that the reductions in activity following adrenalectomy or destruction of the VMH are due to disruption of the same mechanism was not supported.
The interaction between central and peripheral nervous system structures in the production of homeostatically derived behaviors is of prime interest to the physiological psychologist. One major category of such behaviors is those behaviors constituting energy regulation. Certain behaviors are generally conceived of as energy incorporating, others, as energy expending. One obvious energy-incorporating response is food intake, while exercise, in the form of locomotor activity, is conceived of 1 The data presented here are based in part on a dissertation by Christopher Wilson in partial fulfillment of the requirements for the Ph.D. degree at Texas Christian University. This study was supported by a grant from the TCU Research Foundation to Dr. N. R. Remley at Texas Christian University. Requests for reprints should be sent to the author at the Department of Psychology, Arkansas State University, State University, Arkansas 72467. 3 The author wishes to express kind appreciation to Dr. N. R. Remley who directed this research and was the focal point of many fine discussions. Appreciation is also expressed to the author's wife, Linda A. Robinson, who assisted him in every aspect of this study and to Dr. Joseph A. Babitch who was an excellent sounding board. 38O Copyright © 1977 by Academic Press, Inc. All rights of reproduction in any form reserved.
1SSN 0091-6773
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as energy expending. An optimal balance of both input and output mechanisms results in a lean healthy organism. Changes in either or both of the mechanisms produces an animal which becomes either undernourished or grossly obese (Bray and York, 1971). Destruction of the ventromedial hypothalamus (VMH) has produced a variety of behavioral alterations in an animal's ability to regulate its own energy balance (Brittain, 1973; Brobeck et al., 1943; Hetherington and Ranson, 1942; Mayer, 1953). The extent to which these alterations are due to central or peripheral factors is a point of some debate (Brittain, 1973; DeCastro and Balagura, 1975; Hustvedt and Lovo, 1972; Vilberg and Beatty, 1975; York and Bray, 1972). Brittain (1973) proposed that the decrease in an animal's ability to expend energy, as measured by decreases in locomotor activity and free fatty acid mobilization, reported in VMH-lesioned rats may be secondarily due to a reduction in adrenocortical activity. He reported decreases in adrenal weights significantly correlated with the levels of spontaneous running-wheel activity in rats having sustained lesions of the VMH. Studies looking specifically at the role of the adrenal glands in the production of locomotor activity reveal a general decrease in that form of energy expenditure following adrenalectomy (Leshner, 1971; Richter, 1936). Replacement therapy by implantation of adrenal glands (Richter, 1936) or by administration of corticosterone (Leshner, 1971) reinstated the spontaneous activity levels of a majority of the operates. Physiological alterations in energy expenditure have also been reported following adrenalectomy. Shafrir et al. (1960) reported that adrenalectomy resulted in a reduction in the levels of plasma free fatty acids following both in vivo and in vitro adrenergic stimulation of rats' epididymal fat, as compared with intact control subjects. Maeckel and Brodie (1963) reported similar results with epinephrine stimulation of fat pads in vitro from both adrenalectomized and control rats. Maeckel and Brodie (1963) then reinstated normal lipolysis rates in the experimental rats with cortisone stimulation in vivo, but not with cortisone stimulation in vitro. So, both behaviorally and physiologically, it appears that adrenal insufficiency results in a decrease in energy expenditure similar to that seen following lesions of the VMH. Mook et al. (1975) reported differential amounts consumed by rats with both VMH lesions and adrenalectomies as a function of diet. The animals with both manipulations consumed larger amounts of a high-fat or liquid diet as compared with normal lab chow. Morrison's (1968) report that high fat or liquid diets require lower levels of energy expenditure would account for the increase in food intake in the rats given those diets. Anatomically, there is a close association between the central VMH and peripheral adrenal systems. Dunn and Critchlow (1972) reported that neural elements directly involved in the release of ACTH are located
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diffusely throughout the VMH. Tseng and Nickerson (1972) also reported a significant reduction in the number of adenohypophyseal ACTHsecreting cells with VMH lesions in rats. The purpose of this experiment was to test the proposal put forth by Brittain (1973) that the decreases in energy expenditure reported following VMH lesions are a result of a decrease in adrenal activity. Rats were subjected to VMH lesions or adrenalectomies and, subsequently, were given corticosterone replacement therapy. Measures of energy expenditure consisted of running-wheel activity and free fatty acid mobilization rates. It was hypothesized that corticosterone therapy would reinstate the measures of energy output to normal levels in both VMH-lesioned and adrenalectomized rats. METHOD
Subjects. Subjects in this experiment were 58 naive male Holtzman, albino rats, approximately 90 days of age at the beginning of the experiment. The subjects were divided into five groups consisting of 13 animals with bilateral adrenalectomies (ADX), 10 with sham-adrenalectomy operations (SH-ADX), 13 with bilateral VMH lesions (VMH), 12 VMH sham-operated subjects (SH-VMH), and 10 nonoperated control animals (CON). Throughout the experiment the subjects were housed individually and were maintained on an ad lib food and water diet. Apparatus. Activity was measured in Wahman activity wheels, with the number of revolutions being recorded via automatic counters. The wheels were located in a constantly lighted experimental room maintained at 22 + 1°C. Surgery and histology. Each subject was first anesthetized with an intraperitoneal injection of Chlor-Mag-Pent (2.2 cc/kg). Each rat in the VMH and SH-VMH groups was then put into a stereotaxic headholder. Lesions aimed at the VMH were made by passing an anodal direct current through the exposed tip of a No. 2 insect pin. Current parameters were 2.0 mA for 20 sec at the following fiat-head coordinates: 6.7 mm anterior to the interaural line; 0.7 mm on either side of the midsagittal sinus; and 8.5 mm below the surface of the cortex, Sham VMH lesions consisted of lowering the electrode to within 1.0 mm of the VMH and withdrawing it without passing current. Adrenalectomies consisted of cutting the skin and muscle wall bilaterally on the dorsolateral surfaces of the body just posterior to the last rib. The adrenals were then exposed and blunt dissected. Shamadrenalectomy operations consisted of the same procedure with the exception of excision of the adrenal glands. The muscle wall was then sutured together, and the skin was closed with wound clips. Three days per week measures of NaC1 intake were recorded. An adrenalectomy was
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considered successful if the animal's daily intake of a 0.5 M solution of NaC1 was greater than 5 ml. Histology consisted of first anesthetizing each subject with ChlorMag-Pent, exsanguinating it with isotonic saline, and then perfusing it with a 10% neutral buffered Formalin. After being rinsed in water, dehydrated in alcohols, and embedded in celloidin, the brains were sectioned in 40 ~m serial coronal sections and were stained with thionin for microscopic verification of lesion placements. P r o c e d u r e . Upon arrival at the laboratory, rats were given a 1- to 2-week adaptation period, after which acclimation to the running wheels was begun. Each rat was placed in a running wheel for a 1-hr daily session for a total of 21 consecutive days, with the number of revolutions run during each session being recorded. At the conclusion of this training phase the rats were matched and assigned to treatment groups based on the activity of the last two training sessions. At this point, the various surgical manipulations were made. At the time of surgery, two epididymal fat pads were removed from each rat, and the rates of free fatty acid mobilization were determined following the mobilization procedure of Starr et al. (1966) and the colorimetric procedure of Itaya and Ui (1965). Beginning on the third postsurgical day and continuing daily throughout the experiment, each animal was subjected to a sequence of drug/no-drug manipulations. This consisted of either no injection (N) or an injection of corticosterone (Cs) (5 mg/kg body weight) and/or the vehicle substance (V) (0.5 ml/kg body weight). The sequence of manipulations was: N (sessions 1-4); V (sessions 5-8); Cs (sessions 9-12): N (sessions 13-16); V (sessions 17-20); Cs (sessions 21-24); V (sessions 25-28). The CON group received no injections throughout the experiment. Also, during the second Cs manipulation phase, the dose of corticosterone administered to the VMH-lesioned rats was increased to l0 mg/kg body weight. Beginning on the third postsurgical day, each subject was given access to an activity wheel for a l-hr session every other day. The number of revolutions per animal for each of the 1-hr sessions served as a dependent variable. At 50 days following surgery one-half of the animals in each group were injected with corticosterone in the vehicle substance. Subsequently, the animals were anesthetized, and epididymal fat pads were removed for the determination of lipolytic rates. These rats were then subjected to the histological procedures outlined above. At postsurgical Day 58 the remaining subjects were injected with the vehicle substance and were anesthetized, with epididymal fat pads being removed for lipolytic measures. These remaining animals were then subjected to the histological procedure described above. Beginning at the time of surgery and continuing throughout the experiment each rat's body weight was recorded every 2 days. Also, at the time
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of histology the left adrenal gland was removed from each subject in the CON, VMH, SH-VMH, and SH-ADX groups and was weighed.
RESULTS
Data Analysis Data were analyzed using standard parametric procedures (Kirk, 1968), with differences at the P < 0.05 level being reported as statistically significant. Post hoc comparisons were carried out using Newman-Keuls a posteriori procedures. In the ADX group one animal died shortly following surgery, and two animals failed to drink more than 5 ml/day of the sodium chloride solution. In the VMH group, five animals died within two weeks of surgery; three of the rats developed what appeared to be the lateral hypothalamic aphagic syndrome, and one rat developed a severe balance problem and was unable to maintain an upright position. These subjects' data were omitted from all statistical analyses. Total N's for each group were thus: ADX, 10; SH-ADX, 10; SH-VMH, 12; VMH, 7; CON, 10.
Body Weight The mean body weights for all groups over postsurgical Days 1-49 are presented in Fig. 1. As can be seen in this figure the rats comprising the VMH group showed an accelerated rate at which they gained weight. Also, those rats in the two sham-operated and adrenalectomy groups showed depressions in body weight levels below those of the CON group during the postsurgical session. A two-way analysis of variance with repeated measures applied to body weight revealed a significant group effect, F (4, 44) = 7.38, a significant day effect, F (23, 1012) = 154.31, and a significant group x day interaction, F (92, 1012) = 14.03. Post hoc comparisons revealed that, following an initial depression in body weight, t~ o • •
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rats having received VMH lesions recovered and showed an accelerated weight gain. By Day 25 the VMH rats showed body weights reliably higher than on any of the other groups. The SH-ADX, SH-VMH, and ADX groups showed body weights significantly below those of the CON group on Days 25, 27, 47, and 49. The body weights of the SH-ADX, SH-VMH, and ADX rats were not different throughout the study.
Adrenal Weight The mean weights of the left adrenal glands for the SH-ADX, SHVMH, VMH, and CON groups are presented in Table 1. As can be seen, a one-way analysis of variance of these data revealed a significant group effect, F (3, 35) = 7.31. A posteriori procedures revealed a significant reduction in the weights of the left adrenal gland for those rats comprising the VMH group. No other differences were statistically reliable.
Locomotor Activity Figure 2 depicts the mean revolutions per hour for the last two presurgical sessions, combined, and for each postsurgical measurement for all groups in this study. As can be seen in Fig. 2, presurgical running levels for all groups were approximately the same, except for a somewhat higher level for the ADX group. A one-way analysis of variance on these data revealed no differences in the presurgical running levels among any of the five groups. The postsurgical data were divided into two separate phases consisting of one complete drug/no-drug sequence in each phase. Three-factor analyses of variance were used to detect differences in each of the two separate phases. Looking at the first drug/no-drug phase, Experimental Days 1-12, there was an immediate reduction in the running levels of the VMH and ADX rats. While corticosterone therapy failed to affect the locomotor activity of the VMH-lesioned animals, it did produce an increase in the activity of the adrenalectomized rats. The analysis of variance and the post hoc tests applied to the first drug/no-drug phase supported the observations made
TABLE 1 Summary of Mean Adrenal Weights for the SH-VMH. SH-ADX, CON, and VMH Groups Group
SH-VMH SH-ADX CON VMH * P < 0.05.
Mean adrenal weight (mg) 26.14 26.09 26.97 19.06 ~
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above. There was a significant surgical condition effect, F (4, 44) = 11.25, a significant drug condition effect, F (2, 88) = 5.57, and a significant day effect, F (3, 132) = 5.69. Also, the analysis of variance revealed a significant drug condition x day interaction, F (6, 264) = 9.03, a reliable surgical group x drug condition interaction, F (8, 88) = 7.06, and a significant drug condition x surgical group × day interaction, F (24,264) = 11.04. Post hoc analyses showed that, under the no-drug and vehicleinjection conditions, the running levels of the ADX and VMH groups were reliably below those of the three control groups, SH-VMH, SHADX, and CON. Further, the running levels of the ADX subjects were reliably higher than those of the VMH rats on Days 4 and 7-12. Looking at the second drug/no-drug phase, Experimental Days 13-24, Fig. 2 shows progressive reductions in the activity levels of those rats comprising the SH-VMH, SH-ADX, and A D X groups during the no-drug condition. Generally, these groups maintained a stable level of locomotor activity during the vehicle condition and then showed increases during the subsequent corticosterone condition. The different drug/no-drug conditions had no effect on the locomotor activity of the VMH group. The analysis of variance and the post hoc tests largely supported the observations made above. There was a significant group effect, F (4, 44) = 8.90, a reliable drug condition effect, F (2, 88) = 19.54, and a significant day effect, F (3, 132) = 3.78. The drug condition x day interaction was significant, F (6,264) = 30.63, as was the drug condition x day x surgical group triple interaction, F (24, 264) = 19.21. Post hoc analyses revealed no differences among the SH-VMH, SH-ADX, ADX, and CON groups on any particular day of the second drug/no-drug phase. Also, those rats suffering medial basal hypothalamic lesions ran reliably fewer revolutions per hour as compared with all other groups throughout the second half of this study. The drug condition x day x surgical group interaction was
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most evident in the activity levels of the SH-VMH and ADX groups, which showed significant variations within each drug/no-drug condition. During the no-drug phase the SH-VMH and ADX rats showed significant reductions in the levels of running from Days 13 to 15 and 16. Stable levels of performance were maintained during the vehicle-injection condition, with increases during the final drug-injection phase.
Lipolysis Measures Mean free fatty acid mobilization rates for all groups are presented in Table 2. The mobilization rates of the VMH-lesioned rats appear severely depressed, while the adrenalectomized rats assume an intermediate level, below those of the three control groups but above the VMH group. A two-way analysis of variance on the presurgical and postsurgical lipolysis rates revealed a significant group effect, F (8, 40) = 3.48, and a significant pre- versus postsurgical (time) effect, F (1, 40) = 50.32. The group x time interaction was also significant, F (8, 40) = 7.53. Post hoc procedures revealed no differences between any of the groups on the presurgical mobilization measures. A posteriori analyses on the postsurgical data revealed marked differences in the rates of the various groups. VMHlesioned subjects showed reduced lipolysis rates, significantly below those of all other groups, with the exception of the ADX rats having received only the vehicle substance. The mobilization rates of the ADX rats, with or without glucocorticoid therapy, were not different from those of the CON, SH-VMH, or SH-ADX subjects. There were no significant TABLE 2 Summary of Mean Free Fatty Acid Mobilization Rates for Each of the Groups of this Study a Group
Control VMH lesioned Lesion-Cs Lesion-V Sham-Cs Sham-V Adrenalectomized Operate-Cs Operate-V Sham-Cs Sham-V
Mean FFA mobilization rate (txg/g/hr) Presurgical
Postsurgical
1.4192
0.9841
1.5617 1.7475 1.5879 1.3471
0.5038* 0.5000* 1.3782 1.2822
1.4481 1.0535 1.7397 1.1465
0.9925 0.8399 1.5978 1.2700
a Corticosterone is designated by Cs, and the vehicle by V. * P < 0.05 from presurgical score. ** P < 0.05 from control animals.
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differences in the postsurgical mobilization measures for the rats comprising the SH-ADX or SH-VMH groups with or without glucocorticoid therapy. Interestingly, though, the SH-VMH rats given corticosterone therapy just prior to sacrifice demonstrated reliably greater mobilization than did the CON rats.
Histological Results Histological examination revealed destruction of the ventromedial hypothalamus in all subjects of the VMH group (Fig. 3a, b, c). The typical lesion was bordered rostrally by the anterior hypothalamus and caudally by the anterior portion of the posterior hypothalamus. Subjects received damage to the arcuate nucleus and parvocellular region of the paraventricular nucleus. Some damage was sustained by the dorsomedial hypothalamic nucleus, the medial forebrain bundle, and the fornix. The lesions appear extensive and involve portions of the lateral hypothalamus. Since all the animals included in the VMH group were clearly hyperphagic and showed increases in body weight above those of all other groups, the reductions in energy expenditure were attributed to the VMH damage. DISCUSSION
Data from the present experiment indicated that destruction of the VMH or the adrenal glands yields differential results with respect to energy expenditure. These data pointed to both quantitative and qualitative differences in energy output following manipulations of the two systems. A much greater suppression of energy output appeared following brain damage than was evident following adrenalectomy. Also, glucocorticoid therapy was ineffective in altering the output measures of the VMH-lesioned subjects, while glucocorticoid therapy did raise the levels of energy expenditure in the adrenalectomized rats. Essentially, then, it appears that the two mechanisms are dissociable. With respect to body weight, it appears that undergoing any surgical manipulation results in a variation from the body weight levels of the nonoperated (CON) rats. During the final phases of experimentation, the animals comprising the ADX, SH-ADX, and SH-VMH groups showed significant reductions in body weight when compared with the CON group. The reductions in body weight in adrenalectomized rats are consistent with those reported by Leshner (1971), who found that adrenalectomy, with or without corticosterone replacement therapy, resulted in decreases in body weights in male rats. The body weights of the SH-ADX and SH-VMH animals differed from those of the rats comprising the CON group only during specific intervals, Experimental Days 25, 27, 47, and 49. These depressions occurred during or just after glucocorticoid therapy and agree with the report by Leshner (1971) that corticosterone therapy, when given to rats, results in a reduction in body weight. As for the
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A
C
FIG. 3. Lesion placements of subject 82 in group VMH. Lesion placements and extent of damage are typical of all subjects in the VMH group. Coronal sections A, B, and C are serial sections from anterior to posterior.
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rats comprising the VMH group, the increases in body weights above all other groups replicated a large number of reports. It is interesting that, while similar decreases in running activity, an energy-expending task, were reported for both the ADX and VMH rats, opposite effects were seen in the respective alterations in body weights. Access to a running wheel generally produces a decrease in body weight and lean body mass in male rats (Leshner, 1971). If animals run in order to maintain a particular body weight and lean physique, then it appears that very different mechanisms have been disturbed with respect to the two different manipulations. The locomotor activity levels of the various groups are most detrimental to the hypothesis that the peripheral adrenal system can account for the energy expenditure of the VMH-lesioned rats. The recoveries in the running levels of the adrenalectomized rats to levels significantly higher than those of the VMH-lesioned rats during the N and V conditions signified a quantitative difference between the two groups. A qualitative difference between the two groups was signified when corticosterone raised the levels of the ADX rats but had no effect on the running levels of the VMH rats. An anamalous finding was that the rats comprising the SH-VMH group also showed increases in their running levels when administered corticosterone. Of particular interest is the delay between the time the drug was first introduced to the adrenalectomized rats and the time when the increases in running were first recorded, a delay of, at least, 24 hr and, at most, 72 hr, since the animals were only given access to the running wheels every 2 days. Of equal interest is the effect seen following cessation of the drug injections. In the adrenalectomized rats, rather than producing an immediate reduction in running, there was a maintenance of the high level produced with corticosterone stimulation at Day 13, followed by a progressive decline over the next seven sessions. What this means is that the corticosterone is probably not the specific messenger which produced the increase in the running response. Zarrow et al. (1964) supported this contention by reporting a half-life of less than 25 rain for adrenal corticoids in the body. The messenger may be a metabolite of corticosterone or an enzyme directly or indirectly resulting from the glucocorticoid stimulation. An enzyme-induction hypothesis is not unheard of; Wolfe et al. (1976) supported this proposal by looking at the relative number of catecholaminergic receptor sites in the levels of normal and adrenalectomized rats with or without cortisone therapy. They reported a three- to fivefold increase in the number of catecholamine receptors following adrenalectomy, an effect analogous to denervation supersensitivity. With cortisone therapy there was a corresponding decrease in the number of these receptors. Wolfe et al. (1976) concluded that the increase in the number of binding sites " m a y be a compensatory response to the impairments of g l u c o n e o g e n e s i s . . , which occurs after
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adrenalectomy (p. 1347)." An enzyme-induction hypothesis would also account for the report, by Maeckel and Brodie (1963), of an increase in the lipolytic rates of rats given cortisone stimulation in vivo, but not in vitro. Whatever the mechanism, glucocorticoid therapy appears to be ineffective in inducing any changes in the energy expenditure of VMH-lesioned rats. The lipolysis measures reported for the various groups are similar to the running-wheel data. While both adrenalectomies and VMH lesions produced marked reductions in the rates of free fatty acid mobilization, corticosterone stimulation was able to raise the lipolysis rates of the animals comprising the ADX group, but not those of the animals comprising the VMH group. Although the mean presurgical mobilization rates were higher than those recorded postsurgically, these differences were generally not statistically reliable. The anomaly of increased mobilization rates with corticosterone therapy for the rats comprising the sham groups is consistent with the body weight and running-wheel data. Apparently, the exogenous glucocorticoid stimulation produces a semiadditive effect with the endogenous adrenocorticoids, which raises activity and free fatty acid mobilization rates and lowers body weight. The physiological evidence combined with the behavioral measures only strengthens the proposal of separate and independent mechanisms involved in the VMH and adrenal control of energy expenditure. The hypothesis that the reductions in energy expenditure reported following destruction of the medial basal hypothalamus and those seen following adrenalectomy are due to destruction of the same mechanism is not supported. The hypothalamic damage produced a more pronounced effect than did the adrenalectomy: while the VMH lesions virtually abolished running-wheel activity, the adrenalectomy lowered it to an intermediate level, higher than that of the VMH-lesioned rats but significantly below the levels of the control subjects. Furthermore, while a high level of activity was produced with corticosterone stimulation of the adrenalectomized animals, administration of the drug to the rats having sustained brain damage produced no increase in the level of activity. In general, this appears to be the case with the free fatty acid mobilization measures. While no reliable elevation in the mobilization was reported with the drug in the VMH-lesioned rats, corticosterone stimulation of the adrenalectomized rats did elevate the rates of free fatty acid mobilization to a level significantly above those of the VMH-lesioned rats. An alternative hypothesis, proposing that the two systems are independent, receives some support from the data. Although significant reductions in adrenal weights following VMH damage were reported, the fact that corticosterone stimulation failed to produce any increases in the running levels or lipolysis rates of the VMH rats leads to the conclusion that, with respect
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t o e n e r g y e x p e n d i t u r e , t h e r e is little i n t e r a c t i o n b e t w e e n t h e t w o m e c h a nisms.
REFERENCES Bray, G. A., and York, D. A. (1971). Genetically transmitted obesity in rodents. Physiol. Rev. 51, 595-646. Brittain, W. P. (1973). "Developmental Effects of Ventromedial Hypothalamic Lesions on Spontaneous Activity, Quinine Finickiness, and Reactivity to Shock in the Male rat. Unpublished doctoral dissertation, Texas Christian University. Brobeck, J. R., Tepperman, J., and Long, C. N. (1943). Experimental hypothalamic hyperphagia in the albino rat. Yale J. Biol. Med. 15, 831-853. DeCastro, J. M., and Balagura, S. (1975). Ontogeny of meal patterning in rats and its recapitulation during recovery from lateral hypothalamic lesions. J. Comp. Physiol. Psychol. 89, 791-802. Dunn, J., and Critchlow, V. (1972). Electrically stimulated ACTH release in pharmacologically blocked rats: Sites of CRF releasing elements. Fed. Proc. 31, 222. Hetherington, A. W., and Ranson, S. W. (1942). The spontaneous activity and food intake of rats with hypothalamic lesions. Amer. J. Physiol. 136, 609-617. Hustvedt, B. E., and Lovo, A. (1972). Correlation between hyperinsulinemia and hyperphagia in rats with ventromedial hypothalamic lesions. Acta Physiol. Scand. 84, 29-33. Itaya, K., and Ui, M. (1965). Colorimetric determination of free fatty acids in biological fluids. J. Lipid Res. 6, 16-20. Kirk, R. E. (1968). "Experimental Design: Procedures for the Behavioral Sciences." Monterey, Calif.: Brooks/Cole. Leshner, A. I. (1971). The adrenals and the regulatory nature of running-wheel activity. Physiol. Behav. 6, 551-558. Maeckel, R. P., and Brodie, B. B. (1963). Interaction of drugs with the pituitaryadrenocortical system in the production of the fatty liver. Ann. N.Y. Acad. Sci. 104, 1059-1064. Mayer, J. (1953). Decreased activity and energy balance in the hereditary obesity-diabetes syndrome of mice. Science 117, 504-505. Mook, D. G., Fisher, J. C., and Durr, J. C. (1975). Some endocrine influences on hypothalamic hyperphagia. Horm. Behav. 6, 65-79. Morrison, S. D. (1968). The constancy of the energy expended by rats on spontaneous activity, and the distribution of activity between feeding and non-feeding. J. Physiol. (London). 197, 305-323. Richter, C. P. (1936). The spontaneous activity of adrenalectomized rats treated with replacement and other therapy. Endocrinology 20, 657-666. Sha£rir, E., Sussman, K. E., and Steinberg, D. (1960). Role of the pituitary and the adrenals in the mobilization of free fatty acids and lipoproteins. J. Lipid Res. 1, 459-465. Starr, S. E., Crawford, J. D.. and Haessler, H. A. (1966). Dynamics of development of the metabolic and compositional alterations of depot fat in hypothalamic obese rats. Metabolism 15, 39-45. Tseng, M. T., and Nickerson, P. A. (1972). Effects of ventromedial nucleus (VMH) lesions on granules in the ACTH secreting cells. Fed. Proc. 31, 222. Vilberg, T. R., and Beatty, W. W. (1975). Behavioral changes following VMH lesions in rats with controlled insulin levels. Pharmacol. Biochem. Behav. 3, 377-384. Wolfe, B. B., Molinoff, P. B., and Harden, T. K. (1976). Beta adrenergic receptors in rat liver: Effects of adrenalectomy. Proc. Nat. Acad. Sci. USA 73, 1343-1347. York, D. A., and Bray, G. A. (1972). Dependence of hypothalamic obesity on insulin, the pituitary, and the adrenal gland. Endrocrinology 90, 885-894. Zarrow, M. X., Yochlm, J. M., and McCarthy, J. L. (1964). "Experimental Endocrinology: A Sourcebook of Basic Techniques." New York: Academic Press.
Some Effects of Ventromedial Hypothalamic Lesions, Adrenalectomies, and Corticosterone Replacement Therapy on Energy Expenditure in Rats la,3 CHRISTOPHER WILSON
Chemistry of Behavior Program, Texas Christian University, Fort Worth, Texas 76129 The present study tested the hypothesis that the decreases in energy expenditure reported following ventromedial hypothalamic (VMH) lesions in rats are a function of the decreases in adrenal function. Five groups of rats received 21 lhr/day sessions in activity wheels and then were subjected to either VMH lesions, adrenalectomies, the respective sham-operations, or no operation. Following surgery, each rat was again given access to an activity wheel for 24 1-hr test sessions. During the test phase the rats received a sequence of drug/no drug conditions which consisted of: (1) no injection; (2) injections of the vehicle substance: or (3) injections of corticosterone in the vehicle substance. Under the no-drug conditions, running levels of the VMH-lesioned and adrenalectomized rats decreased, with the levels of VMH-lesioned rats being more severely depressed. Corticosterone replacement therapy reinstated the activity levels of the adrenalectomized rats, but not those of the VMH rats. The hypothesis that the reductions in activity following adrenalectomy or destruction of the VMH are due to disruption of the same mechanism was not supported.
The interaction between central and peripheral nervous system structures in the production of homeostatically derived behaviors is of prime interest to the physiological psychologist. One major category of such behaviors is those behaviors constituting energy regulation. Certain behaviors are generally conceived of as energy incorporating, others, as energy expending. One obvious energy-incorporating response is food intake, while exercise, in the form of locomotor activity, is conceived of 1 The data presented here are based in part on a dissertation by Christopher Wilson in partial fulfillment of the requirements for the Ph.D. degree at Texas Christian University. This study was supported by a grant from the TCU Research Foundation to Dr. N. R. Remley at Texas Christian University. Requests for reprints should be sent to the author at the Department of Psychology, Arkansas State University, State University, Arkansas 72467. 3 The author wishes to express kind appreciation to Dr. N. R. Remley who directed this research and was the focal point of many fine discussions. Appreciation is also expressed to the author's wife, Linda A. Robinson, who assisted him in every aspect of this study and to Dr. Joseph A. Babitch who was an excellent sounding board. 38O Copyright © 1977 by Academic Press, Inc. All rights of reproduction in any form reserved.
1SSN 0091-6773
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as energy expending. An optimal balance of both input and output mechanisms results in a lean healthy organism. Changes in either or both of the mechanisms produces an animal which becomes either undernourished or grossly obese (Bray and York, 1971). Destruction of the ventromedial hypothalamus (VMH) has produced a variety of behavioral alterations in an animal's ability to regulate its own energy balance (Brittain, 1973; Brobeck et al., 1943; Hetherington and Ranson, 1942; Mayer, 1953). The extent to which these alterations are due to central or peripheral factors is a point of some debate (Brittain, 1973; DeCastro and Balagura, 1975; Hustvedt and Lovo, 1972; Vilberg and Beatty, 1975; York and Bray, 1972). Brittain (1973) proposed that the decrease in an animal's ability to expend energy, as measured by decreases in locomotor activity and free fatty acid mobilization, reported in VMH-lesioned rats may be secondarily due to a reduction in adrenocortical activity. He reported decreases in adrenal weights significantly correlated with the levels of spontaneous running-wheel activity in rats having sustained lesions of the VMH. Studies looking specifically at the role of the adrenal glands in the production of locomotor activity reveal a general decrease in that form of energy expenditure following adrenalectomy (Leshner, 1971; Richter, 1936). Replacement therapy by implantation of adrenal glands (Richter, 1936) or by administration of corticosterone (Leshner, 1971) reinstated the spontaneous activity levels of a majority of the operates. Physiological alterations in energy expenditure have also been reported following adrenalectomy. Shafrir et al. (1960) reported that adrenalectomy resulted in a reduction in the levels of plasma free fatty acids following both in vivo and in vitro adrenergic stimulation of rats' epididymal fat, as compared with intact control subjects. Maeckel and Brodie (1963) reported similar results with epinephrine stimulation of fat pads in vitro from both adrenalectomized and control rats. Maeckel and Brodie (1963) then reinstated normal lipolysis rates in the experimental rats with cortisone stimulation in vivo, but not with cortisone stimulation in vitro. So, both behaviorally and physiologically, it appears that adrenal insufficiency results in a decrease in energy expenditure similar to that seen following lesions of the VMH. Mook et al. (1975) reported differential amounts consumed by rats with both VMH lesions and adrenalectomies as a function of diet. The animals with both manipulations consumed larger amounts of a high-fat or liquid diet as compared with normal lab chow. Morrison's (1968) report that high fat or liquid diets require lower levels of energy expenditure would account for the increase in food intake in the rats given those diets. Anatomically, there is a close association between the central VMH and peripheral adrenal systems. Dunn and Critchlow (1972) reported that neural elements directly involved in the release of ACTH are located
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diffusely throughout the VMH. Tseng and Nickerson (1972) also reported a significant reduction in the number of adenohypophyseal ACTHsecreting cells with VMH lesions in rats. The purpose of this experiment was to test the proposal put forth by Brittain (1973) that the decreases in energy expenditure reported following VMH lesions are a result of a decrease in adrenal activity. Rats were subjected to VMH lesions or adrenalectomies and, subsequently, were given corticosterone replacement therapy. Measures of energy expenditure consisted of running-wheel activity and free fatty acid mobilization rates. It was hypothesized that corticosterone therapy would reinstate the measures of energy output to normal levels in both VMH-lesioned and adrenalectomized rats. METHOD
Subjects. Subjects in this experiment were 58 naive male Holtzman, albino rats, approximately 90 days of age at the beginning of the experiment. The subjects were divided into five groups consisting of 13 animals with bilateral adrenalectomies (ADX), 10 with sham-adrenalectomy operations (SH-ADX), 13 with bilateral VMH lesions (VMH), 12 VMH sham-operated subjects (SH-VMH), and 10 nonoperated control animals (CON). Throughout the experiment the subjects were housed individually and were maintained on an ad lib food and water diet. Apparatus. Activity was measured in Wahman activity wheels, with the number of revolutions being recorded via automatic counters. The wheels were located in a constantly lighted experimental room maintained at 22 + 1°C. Surgery and histology. Each subject was first anesthetized with an intraperitoneal injection of Chlor-Mag-Pent (2.2 cc/kg). Each rat in the VMH and SH-VMH groups was then put into a stereotaxic headholder. Lesions aimed at the VMH were made by passing an anodal direct current through the exposed tip of a No. 2 insect pin. Current parameters were 2.0 mA for 20 sec at the following fiat-head coordinates: 6.7 mm anterior to the interaural line; 0.7 mm on either side of the midsagittal sinus; and 8.5 mm below the surface of the cortex, Sham VMH lesions consisted of lowering the electrode to within 1.0 mm of the VMH and withdrawing it without passing current. Adrenalectomies consisted of cutting the skin and muscle wall bilaterally on the dorsolateral surfaces of the body just posterior to the last rib. The adrenals were then exposed and blunt dissected. Shamadrenalectomy operations consisted of the same procedure with the exception of excision of the adrenal glands. The muscle wall was then sutured together, and the skin was closed with wound clips. Three days per week measures of NaC1 intake were recorded. An adrenalectomy was
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considered successful if the animal's daily intake of a 0.5 M solution of NaC1 was greater than 5 ml. Histology consisted of first anesthetizing each subject with ChlorMag-Pent, exsanguinating it with isotonic saline, and then perfusing it with a 10% neutral buffered Formalin. After being rinsed in water, dehydrated in alcohols, and embedded in celloidin, the brains were sectioned in 40 ~m serial coronal sections and were stained with thionin for microscopic verification of lesion placements. P r o c e d u r e . Upon arrival at the laboratory, rats were given a 1- to 2-week adaptation period, after which acclimation to the running wheels was begun. Each rat was placed in a running wheel for a 1-hr daily session for a total of 21 consecutive days, with the number of revolutions run during each session being recorded. At the conclusion of this training phase the rats were matched and assigned to treatment groups based on the activity of the last two training sessions. At this point, the various surgical manipulations were made. At the time of surgery, two epididymal fat pads were removed from each rat, and the rates of free fatty acid mobilization were determined following the mobilization procedure of Starr et al. (1966) and the colorimetric procedure of Itaya and Ui (1965). Beginning on the third postsurgical day and continuing daily throughout the experiment, each animal was subjected to a sequence of drug/no-drug manipulations. This consisted of either no injection (N) or an injection of corticosterone (Cs) (5 mg/kg body weight) and/or the vehicle substance (V) (0.5 ml/kg body weight). The sequence of manipulations was: N (sessions 1-4); V (sessions 5-8); Cs (sessions 9-12): N (sessions 13-16); V (sessions 17-20); Cs (sessions 21-24); V (sessions 25-28). The CON group received no injections throughout the experiment. Also, during the second Cs manipulation phase, the dose of corticosterone administered to the VMH-lesioned rats was increased to l0 mg/kg body weight. Beginning on the third postsurgical day, each subject was given access to an activity wheel for a l-hr session every other day. The number of revolutions per animal for each of the 1-hr sessions served as a dependent variable. At 50 days following surgery one-half of the animals in each group were injected with corticosterone in the vehicle substance. Subsequently, the animals were anesthetized, and epididymal fat pads were removed for the determination of lipolytic rates. These rats were then subjected to the histological procedures outlined above. At postsurgical Day 58 the remaining subjects were injected with the vehicle substance and were anesthetized, with epididymal fat pads being removed for lipolytic measures. These remaining animals were then subjected to the histological procedure described above. Beginning at the time of surgery and continuing throughout the experiment each rat's body weight was recorded every 2 days. Also, at the time
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of histology the left adrenal gland was removed from each subject in the CON, VMH, SH-VMH, and SH-ADX groups and was weighed.
RESULTS
Data Analysis Data were analyzed using standard parametric procedures (Kirk, 1968), with differences at the P < 0.05 level being reported as statistically significant. Post hoc comparisons were carried out using Newman-Keuls a posteriori procedures. In the ADX group one animal died shortly following surgery, and two animals failed to drink more than 5 ml/day of the sodium chloride solution. In the VMH group, five animals died within two weeks of surgery; three of the rats developed what appeared to be the lateral hypothalamic aphagic syndrome, and one rat developed a severe balance problem and was unable to maintain an upright position. These subjects' data were omitted from all statistical analyses. Total N's for each group were thus: ADX, 10; SH-ADX, 10; SH-VMH, 12; VMH, 7; CON, 10.
Body Weight The mean body weights for all groups over postsurgical Days 1-49 are presented in Fig. 1. As can be seen in this figure the rats comprising the VMH group showed an accelerated rate at which they gained weight. Also, those rats in the two sham-operated and adrenalectomy groups showed depressions in body weight levels below those of the CON group during the postsurgical session. A two-way analysis of variance with repeated measures applied to body weight revealed a significant group effect, F (4, 44) = 7.38, a significant day effect, F (23, 1012) = 154.31, and a significant group x day interaction, F (92, 1012) = 14.03. Post hoc comparisons revealed that, following an initial depression in body weight, t~ o • •
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rats having received VMH lesions recovered and showed an accelerated weight gain. By Day 25 the VMH rats showed body weights reliably higher than on any of the other groups. The SH-ADX, SH-VMH, and ADX groups showed body weights significantly below those of the CON group on Days 25, 27, 47, and 49. The body weights of the SH-ADX, SH-VMH, and ADX rats were not different throughout the study.
Adrenal Weight The mean weights of the left adrenal glands for the SH-ADX, SHVMH, VMH, and CON groups are presented in Table 1. As can be seen, a one-way analysis of variance of these data revealed a significant group effect, F (3, 35) = 7.31. A posteriori procedures revealed a significant reduction in the weights of the left adrenal gland for those rats comprising the VMH group. No other differences were statistically reliable.
Locomotor Activity Figure 2 depicts the mean revolutions per hour for the last two presurgical sessions, combined, and for each postsurgical measurement for all groups in this study. As can be seen in Fig. 2, presurgical running levels for all groups were approximately the same, except for a somewhat higher level for the ADX group. A one-way analysis of variance on these data revealed no differences in the presurgical running levels among any of the five groups. The postsurgical data were divided into two separate phases consisting of one complete drug/no-drug sequence in each phase. Three-factor analyses of variance were used to detect differences in each of the two separate phases. Looking at the first drug/no-drug phase, Experimental Days 1-12, there was an immediate reduction in the running levels of the VMH and ADX rats. While corticosterone therapy failed to affect the locomotor activity of the VMH-lesioned animals, it did produce an increase in the activity of the adrenalectomized rats. The analysis of variance and the post hoc tests applied to the first drug/no-drug phase supported the observations made
TABLE 1 Summary of Mean Adrenal Weights for the SH-VMH. SH-ADX, CON, and VMH Groups Group
SH-VMH SH-ADX CON VMH * P < 0.05.
Mean adrenal weight (mg) 26.14 26.09 26.97 19.06 ~
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above. There was a significant surgical condition effect, F (4, 44) = 11.25, a significant drug condition effect, F (2, 88) = 5.57, and a significant day effect, F (3, 132) = 5.69. Also, the analysis of variance revealed a significant drug condition x day interaction, F (6, 264) = 9.03, a reliable surgical group x drug condition interaction, F (8, 88) = 7.06, and a significant drug condition x surgical group × day interaction, F (24,264) = 11.04. Post hoc analyses showed that, under the no-drug and vehicleinjection conditions, the running levels of the ADX and VMH groups were reliably below those of the three control groups, SH-VMH, SHADX, and CON. Further, the running levels of the ADX subjects were reliably higher than those of the VMH rats on Days 4 and 7-12. Looking at the second drug/no-drug phase, Experimental Days 13-24, Fig. 2 shows progressive reductions in the activity levels of those rats comprising the SH-VMH, SH-ADX, and A D X groups during the no-drug condition. Generally, these groups maintained a stable level of locomotor activity during the vehicle condition and then showed increases during the subsequent corticosterone condition. The different drug/no-drug conditions had no effect on the locomotor activity of the VMH group. The analysis of variance and the post hoc tests largely supported the observations made above. There was a significant group effect, F (4, 44) = 8.90, a reliable drug condition effect, F (2, 88) = 19.54, and a significant day effect, F (3, 132) = 3.78. The drug condition x day interaction was significant, F (6,264) = 30.63, as was the drug condition x day x surgical group triple interaction, F (24, 264) = 19.21. Post hoc analyses revealed no differences among the SH-VMH, SH-ADX, ADX, and CON groups on any particular day of the second drug/no-drug phase. Also, those rats suffering medial basal hypothalamic lesions ran reliably fewer revolutions per hour as compared with all other groups throughout the second half of this study. The drug condition x day x surgical group interaction was
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most evident in the activity levels of the SH-VMH and ADX groups, which showed significant variations within each drug/no-drug condition. During the no-drug phase the SH-VMH and ADX rats showed significant reductions in the levels of running from Days 13 to 15 and 16. Stable levels of performance were maintained during the vehicle-injection condition, with increases during the final drug-injection phase.
Lipolysis Measures Mean free fatty acid mobilization rates for all groups are presented in Table 2. The mobilization rates of the VMH-lesioned rats appear severely depressed, while the adrenalectomized rats assume an intermediate level, below those of the three control groups but above the VMH group. A two-way analysis of variance on the presurgical and postsurgical lipolysis rates revealed a significant group effect, F (8, 40) = 3.48, and a significant pre- versus postsurgical (time) effect, F (1, 40) = 50.32. The group x time interaction was also significant, F (8, 40) = 7.53. Post hoc procedures revealed no differences between any of the groups on the presurgical mobilization measures. A posteriori analyses on the postsurgical data revealed marked differences in the rates of the various groups. VMHlesioned subjects showed reduced lipolysis rates, significantly below those of all other groups, with the exception of the ADX rats having received only the vehicle substance. The mobilization rates of the ADX rats, with or without glucocorticoid therapy, were not different from those of the CON, SH-VMH, or SH-ADX subjects. There were no significant TABLE 2 Summary of Mean Free Fatty Acid Mobilization Rates for Each of the Groups of this Study a Group
Control VMH lesioned Lesion-Cs Lesion-V Sham-Cs Sham-V Adrenalectomized Operate-Cs Operate-V Sham-Cs Sham-V
Mean FFA mobilization rate (txg/g/hr) Presurgical
Postsurgical
1.4192
0.9841
1.5617 1.7475 1.5879 1.3471
0.5038* 0.5000* 1.3782 1.2822
1.4481 1.0535 1.7397 1.1465
0.9925 0.8399 1.5978 1.2700
a Corticosterone is designated by Cs, and the vehicle by V. * P < 0.05 from presurgical score. ** P < 0.05 from control animals.
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differences in the postsurgical mobilization measures for the rats comprising the SH-ADX or SH-VMH groups with or without glucocorticoid therapy. Interestingly, though, the SH-VMH rats given corticosterone therapy just prior to sacrifice demonstrated reliably greater mobilization than did the CON rats.
Histological Results Histological examination revealed destruction of the ventromedial hypothalamus in all subjects of the VMH group (Fig. 3a, b, c). The typical lesion was bordered rostrally by the anterior hypothalamus and caudally by the anterior portion of the posterior hypothalamus. Subjects received damage to the arcuate nucleus and parvocellular region of the paraventricular nucleus. Some damage was sustained by the dorsomedial hypothalamic nucleus, the medial forebrain bundle, and the fornix. The lesions appear extensive and involve portions of the lateral hypothalamus. Since all the animals included in the VMH group were clearly hyperphagic and showed increases in body weight above those of all other groups, the reductions in energy expenditure were attributed to the VMH damage. DISCUSSION
Data from the present experiment indicated that destruction of the VMH or the adrenal glands yields differential results with respect to energy expenditure. These data pointed to both quantitative and qualitative differences in energy output following manipulations of the two systems. A much greater suppression of energy output appeared following brain damage than was evident following adrenalectomy. Also, glucocorticoid therapy was ineffective in altering the output measures of the VMH-lesioned subjects, while glucocorticoid therapy did raise the levels of energy expenditure in the adrenalectomized rats. Essentially, then, it appears that the two mechanisms are dissociable. With respect to body weight, it appears that undergoing any surgical manipulation results in a variation from the body weight levels of the nonoperated (CON) rats. During the final phases of experimentation, the animals comprising the ADX, SH-ADX, and SH-VMH groups showed significant reductions in body weight when compared with the CON group. The reductions in body weight in adrenalectomized rats are consistent with those reported by Leshner (1971), who found that adrenalectomy, with or without corticosterone replacement therapy, resulted in decreases in body weights in male rats. The body weights of the SH-ADX and SH-VMH animals differed from those of the rats comprising the CON group only during specific intervals, Experimental Days 25, 27, 47, and 49. These depressions occurred during or just after glucocorticoid therapy and agree with the report by Leshner (1971) that corticosterone therapy, when given to rats, results in a reduction in body weight. As for the
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A
C
FIG. 3. Lesion placements of subject 82 in group VMH. Lesion placements and extent of damage are typical of all subjects in the VMH group. Coronal sections A, B, and C are serial sections from anterior to posterior.
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rats comprising the VMH group, the increases in body weights above all other groups replicated a large number of reports. It is interesting that, while similar decreases in running activity, an energy-expending task, were reported for both the ADX and VMH rats, opposite effects were seen in the respective alterations in body weights. Access to a running wheel generally produces a decrease in body weight and lean body mass in male rats (Leshner, 1971). If animals run in order to maintain a particular body weight and lean physique, then it appears that very different mechanisms have been disturbed with respect to the two different manipulations. The locomotor activity levels of the various groups are most detrimental to the hypothesis that the peripheral adrenal system can account for the energy expenditure of the VMH-lesioned rats. The recoveries in the running levels of the adrenalectomized rats to levels significantly higher than those of the VMH-lesioned rats during the N and V conditions signified a quantitative difference between the two groups. A qualitative difference between the two groups was signified when corticosterone raised the levels of the ADX rats but had no effect on the running levels of the VMH rats. An anamalous finding was that the rats comprising the SH-VMH group also showed increases in their running levels when administered corticosterone. Of particular interest is the delay between the time the drug was first introduced to the adrenalectomized rats and the time when the increases in running were first recorded, a delay of, at least, 24 hr and, at most, 72 hr, since the animals were only given access to the running wheels every 2 days. Of equal interest is the effect seen following cessation of the drug injections. In the adrenalectomized rats, rather than producing an immediate reduction in running, there was a maintenance of the high level produced with corticosterone stimulation at Day 13, followed by a progressive decline over the next seven sessions. What this means is that the corticosterone is probably not the specific messenger which produced the increase in the running response. Zarrow et al. (1964) supported this contention by reporting a half-life of less than 25 rain for adrenal corticoids in the body. The messenger may be a metabolite of corticosterone or an enzyme directly or indirectly resulting from the glucocorticoid stimulation. An enzyme-induction hypothesis is not unheard of; Wolfe et al. (1976) supported this proposal by looking at the relative number of catecholaminergic receptor sites in the levels of normal and adrenalectomized rats with or without cortisone therapy. They reported a three- to fivefold increase in the number of catecholamine receptors following adrenalectomy, an effect analogous to denervation supersensitivity. With cortisone therapy there was a corresponding decrease in the number of these receptors. Wolfe et al. (1976) concluded that the increase in the number of binding sites " m a y be a compensatory response to the impairments of g l u c o n e o g e n e s i s . . , which occurs after
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adrenalectomy (p. 1347)." An enzyme-induction hypothesis would also account for the report, by Maeckel and Brodie (1963), of an increase in the lipolytic rates of rats given cortisone stimulation in vivo, but not in vitro. Whatever the mechanism, glucocorticoid therapy appears to be ineffective in inducing any changes in the energy expenditure of VMH-lesioned rats. The lipolysis measures reported for the various groups are similar to the running-wheel data. While both adrenalectomies and VMH lesions produced marked reductions in the rates of free fatty acid mobilization, corticosterone stimulation was able to raise the lipolysis rates of the animals comprising the ADX group, but not those of the animals comprising the VMH group. Although the mean presurgical mobilization rates were higher than those recorded postsurgically, these differences were generally not statistically reliable. The anomaly of increased mobilization rates with corticosterone therapy for the rats comprising the sham groups is consistent with the body weight and running-wheel data. Apparently, the exogenous glucocorticoid stimulation produces a semiadditive effect with the endogenous adrenocorticoids, which raises activity and free fatty acid mobilization rates and lowers body weight. The physiological evidence combined with the behavioral measures only strengthens the proposal of separate and independent mechanisms involved in the VMH and adrenal control of energy expenditure. The hypothesis that the reductions in energy expenditure reported following destruction of the medial basal hypothalamus and those seen following adrenalectomy are due to destruction of the same mechanism is not supported. The hypothalamic damage produced a more pronounced effect than did the adrenalectomy: while the VMH lesions virtually abolished running-wheel activity, the adrenalectomy lowered it to an intermediate level, higher than that of the VMH-lesioned rats but significantly below the levels of the control subjects. Furthermore, while a high level of activity was produced with corticosterone stimulation of the adrenalectomized animals, administration of the drug to the rats having sustained brain damage produced no increase in the level of activity. In general, this appears to be the case with the free fatty acid mobilization measures. While no reliable elevation in the mobilization was reported with the drug in the VMH-lesioned rats, corticosterone stimulation of the adrenalectomized rats did elevate the rates of free fatty acid mobilization to a level significantly above those of the VMH-lesioned rats. An alternative hypothesis, proposing that the two systems are independent, receives some support from the data. Although significant reductions in adrenal weights following VMH damage were reported, the fact that corticosterone stimulation failed to produce any increases in the running levels or lipolysis rates of the VMH rats leads to the conclusion that, with respect
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t o e n e r g y e x p e n d i t u r e , t h e r e is little i n t e r a c t i o n b e t w e e n t h e t w o m e c h a nisms.
REFERENCES Bray, G. A., and York, D. A. (1971). Genetically transmitted obesity in rodents. Physiol. Rev. 51, 595-646. Brittain, W. P. (1973). "Developmental Effects of Ventromedial Hypothalamic Lesions on Spontaneous Activity, Quinine Finickiness, and Reactivity to Shock in the Male rat. Unpublished doctoral dissertation, Texas Christian University. Brobeck, J. R., Tepperman, J., and Long, C. N. (1943). Experimental hypothalamic hyperphagia in the albino rat. Yale J. Biol. Med. 15, 831-853. DeCastro, J. M., and Balagura, S. (1975). Ontogeny of meal patterning in rats and its recapitulation during recovery from lateral hypothalamic lesions. J. Comp. Physiol. Psychol. 89, 791-802. Dunn, J., and Critchlow, V. (1972). Electrically stimulated ACTH release in pharmacologically blocked rats: Sites of CRF releasing elements. Fed. Proc. 31, 222. Hetherington, A. W., and Ranson, S. W. (1942). The spontaneous activity and food intake of rats with hypothalamic lesions. Amer. J. Physiol. 136, 609-617. Hustvedt, B. E., and Lovo, A. (1972). Correlation between hyperinsulinemia and hyperphagia in rats with ventromedial hypothalamic lesions. Acta Physiol. Scand. 84, 29-33. Itaya, K., and Ui, M. (1965). Colorimetric determination of free fatty acids in biological fluids. J. Lipid Res. 6, 16-20. Kirk, R. E. (1968). "Experimental Design: Procedures for the Behavioral Sciences." Monterey, Calif.: Brooks/Cole. Leshner, A. I. (1971). The adrenals and the regulatory nature of running-wheel activity. Physiol. Behav. 6, 551-558. Maeckel, R. P., and Brodie, B. B. (1963). Interaction of drugs with the pituitaryadrenocortical system in the production of the fatty liver. Ann. N.Y. Acad. Sci. 104, 1059-1064. Mayer, J. (1953). Decreased activity and energy balance in the hereditary obesity-diabetes syndrome of mice. Science 117, 504-505. Mook, D. G., Fisher, J. C., and Durr, J. C. (1975). Some endocrine influences on hypothalamic hyperphagia. Horm. Behav. 6, 65-79. Morrison, S. D. (1968). The constancy of the energy expended by rats on spontaneous activity, and the distribution of activity between feeding and non-feeding. J. Physiol. (London). 197, 305-323. Richter, C. P. (1936). The spontaneous activity of adrenalectomized rats treated with replacement and other therapy. Endocrinology 20, 657-666. Sha£rir, E., Sussman, K. E., and Steinberg, D. (1960). Role of the pituitary and the adrenals in the mobilization of free fatty acids and lipoproteins. J. Lipid Res. 1, 459-465. Starr, S. E., Crawford, J. D.. and Haessler, H. A. (1966). Dynamics of development of the metabolic and compositional alterations of depot fat in hypothalamic obese rats. Metabolism 15, 39-45. Tseng, M. T., and Nickerson, P. A. (1972). Effects of ventromedial nucleus (VMH) lesions on granules in the ACTH secreting cells. Fed. Proc. 31, 222. Vilberg, T. R., and Beatty, W. W. (1975). Behavioral changes following VMH lesions in rats with controlled insulin levels. Pharmacol. Biochem. Behav. 3, 377-384. Wolfe, B. B., Molinoff, P. B., and Harden, T. K. (1976). Beta adrenergic receptors in rat liver: Effects of adrenalectomy. Proc. Nat. Acad. Sci. USA 73, 1343-1347. York, D. A., and Bray, G. A. (1972). Dependence of hypothalamic obesity on insulin, the pituitary, and the adrenal gland. Endrocrinology 90, 885-894. Zarrow, M. X., Yochlm, J. M., and McCarthy, J. L. (1964). "Experimental Endocrinology: A Sourcebook of Basic Techniques." New York: Academic Press.
