The Effects of Subdiaphragmatic Vagotomy in Rats With Ventromedial Hypothalamic Obesity SHUJI INOUE AND GEORGE A. BRAY Department of Medicine, UCLA School of Medicine, Harbor General Hospital Campus, Torrance, California 90509 vent the rise in acid after vagal stimulation. These results suggest that subdiaphragmatic vagotomy reversed the obesity of VMH lesioned rats primarily by decreasing food intake. However, there was a positive correlation (r = .70) between the level of serum insulin and basal gastric acid in VMH lesioned rats which remained significant when the effects of food intake were held constant (partial correlation coefficient = 0.449). This supports the possibility that ventromedial hypothalamic injury is followed by enhanced vagal activity and that the vagus may play an important part in the hyperinsulinemia of VMH obesity. (Endocrinology 100: 108, 1977)
ABSTRACT. The present study investigated the effects of subdiaphragmatic vagotomy in rats with ventromedial hypothalamic (VMH) lesions and obesity. Vagotomy or sham-vagotomy was performed two weeks after VMH lesions and rats were observed for 4 more weeks. Complete vagotomy reversed the VMH obesity, lowered serum insulin, lowered basal gastric acid and blocked the secretion of gastric acid after stimulation of the cervical vagus. Pair-feeding of VMH-lesioned rats without vagotomy to the food intake of vagotomized animals also reversed the obesity, lowered serum insulin and lowered basal acid secretion, but it did not pre-
H
YPOTHALAMIC obesity results from bilateral injury to the ventromedial region of the hypothalamus. Studies from several laboratories have suggested that this syndrome depends in large part on the hypersecretion of insulin. The studies by Frohman and his collaborators (1), by Han and Frohman (2), and by Hustvedt and L0v0 (3) showed that hyperinsulinemia occurred in the first few days following injury to the ventromedial hypothalamus (VMH) before significant changes in body fat could be demonstrated. In addition, York and Bray (4) and Goldman et at. (5) showed that if the islets of Langerhans were damaged with streptozotocin before producing VMH lesions, obesity and hyperphagia were largely prevented. Earlier, Ridley and Brooks (6) had noted that hypothalamic obesity was associated with gastric hyperacidity, a phenomenon related to vagal overactivity. Powley and Opsahl (7) pursued this observation and demonstrated that interruption of the vagus nerve below the diaphragm could reverse the obesity which follows ventromedial hypothalamic injury.
One interpretation of the studies by Powley and Opsahl was that hypersecretion of insulin was reduced after vagotomy and that the weight loss occurred because the hyperinsulinemia was relieved. Alternatively, it is possible that the weight loss represented reduced food intake secondary to gastrointestinal dysfunction and was not the result of a fall in insulin secretion. To evaluate these possibilities, we performed the following experiment. Materials and Methods Animals Fifty-six female Sprague-Dawley rats weighing between 230 and 270 g were used in this study. They were housed in individual metabolic cages and fed powdered Purina Laboratory chow and tap water ad libitum. The lighting was automatically regulated to provide 12 h of darkness and 12 hours of light (0600 to 1800 h). The room temperature was approximately 21 C. Body weights were measured weekly and food intake was recorded daily during the experiment. Experimental
Received February 26,1976. Supported in part by NIH Grant Nos. Am 18165 and RR 00425.
procedures
Under pentobarbital anesthesia, 50 rats received bilateral electrolytic lesions in the ventromedial hypothalamus (VMH) using coordinates
108
VAGOTOMY AND OBESITY from the DeGroot Atlas (at the Bregma anteriorly; ±0.75 mm lateral to the midsagital line, and 1.0 mm above the base of the skull) (8). Another six rats were subject to sham-VMH operations. Only 37 of the 50 VMH-lesioned rats which gained more than 70 g in two weeks were used in the study. Two weeks later, subdiaphragmatic vagotomy was performed in 25 of these rats according to the method of Snowdon and Epstein (9). A sham-vagotomy was performed in 12 others and in the 6 sham-VMH controls. Four weeks following vagotomy, all groups of rats were bled by cardiac puncture under light ether anesthesia for measurement of serum glucose and insulin. Several days later, while under pentobarbital anesthesia, the cervical vagus was electrically stimulated and the secretion of gastric acid from the cannulated stomach was measured before and after stimulation in order to document the completeness of vagotomy (7). Rats were classified as follows: Group 1, six VMH-lesioned rats with sham-vagotomy which were fed ad libitum; Group 2, twelve vagotomized VMH-lesioned rats which were fed ad libitum. This group was subsequently subdivided into two groups: Group 2a—seven rats with complete vagotomy, and Group 2b—five rats with incomplete vagotomy as determined by measuring the release of gastric acid before and after direct electrical stimulation of the cervical vagus nerves using the method of Powley and Opsahl (7); Group 3, five VMH-lesioned rats with a sham-
109
vagotomy which were pair-fed to the group with complete vagotomy by giving the average daily food intake of the latter group. Group 4 consisted of six VMH-sham operated rats with a sham-vagotomy which were fed ad libitum (control group). Analytical methods The Lee index was used to estimate fatness (10). Serum insulin was measured by a double antibody method using rat insulin standard and labeled pork insulin (11). Serum glucose was measured by a glucose-oxidase method. Statistical comparisons utilized data with Student's t test. Correlation and partial correlation coefficients were determined with the BMDP programs and an IBM 360/90 computer.
Results Figure 1 shows that the VMH-lesioned rats with a sham-vagotomy continued to gain weight throughout the study to reach an average weight of 485 g. The group with incomplete vagotomy lost weight for two weeks after vagotomy and then began to regain weight slowly. The control group reached the same final weight as the group with incomplete vagotomy. The group with VMH lesions and complete vagotomy showed a marked loss of weight during the
500-i Sham-Vagotomy
FIG. 1. Body weight of rats with ventromedial hypothalamic lesions. Each group is plotted as the mean ± SEM. Sham-vagotomy — • —; incomplete vagotomy — • —; complete vagotomy — O—; VMH pairfed to sham-vagotomy — • —; control group (sham lesion-sham vagotomy) — A —.
400-
1 Incomplete ""•"¥ Vagotomy ^-Control
300-
Pair-Fed 2 Complete Vagotomy
200-
VMH
VAGOTOMY
WEEKS
Endo • 1977 Vol 100 • No 1
INOUE AND BRAY
110
TABLE 1. Serum glucose and immunoreactive insulin levels after overnight fast in rats subject to ventromedial hypthalamic (VMH) lesions Operative status Group
VMH
1
+\
Serum glucose (mg/dl)
Leef index
Vagotomy Sham
0.352 ±.011*
ABSTRACT. The present study investigated the effects of subdiaphragmatic vagotomy in rats with ventromedial hypothalamic (VMH) lesions and obesity. Vagotomy or sham-vagotomy was performed two weeks after VMH lesions and rats were observed for 4 more weeks. Complete vagotomy reversed the VMH obesity, lowered serum insulin, lowered basal gastric acid and blocked the secretion of gastric acid after stimulation of the cervical vagus. Pair-feeding of VMH-lesioned rats without vagotomy to the food intake of vagotomized animals also reversed the obesity, lowered serum insulin and lowered basal acid secretion, but it did not pre-
H
YPOTHALAMIC obesity results from bilateral injury to the ventromedial region of the hypothalamus. Studies from several laboratories have suggested that this syndrome depends in large part on the hypersecretion of insulin. The studies by Frohman and his collaborators (1), by Han and Frohman (2), and by Hustvedt and L0v0 (3) showed that hyperinsulinemia occurred in the first few days following injury to the ventromedial hypothalamus (VMH) before significant changes in body fat could be demonstrated. In addition, York and Bray (4) and Goldman et at. (5) showed that if the islets of Langerhans were damaged with streptozotocin before producing VMH lesions, obesity and hyperphagia were largely prevented. Earlier, Ridley and Brooks (6) had noted that hypothalamic obesity was associated with gastric hyperacidity, a phenomenon related to vagal overactivity. Powley and Opsahl (7) pursued this observation and demonstrated that interruption of the vagus nerve below the diaphragm could reverse the obesity which follows ventromedial hypothalamic injury.
One interpretation of the studies by Powley and Opsahl was that hypersecretion of insulin was reduced after vagotomy and that the weight loss occurred because the hyperinsulinemia was relieved. Alternatively, it is possible that the weight loss represented reduced food intake secondary to gastrointestinal dysfunction and was not the result of a fall in insulin secretion. To evaluate these possibilities, we performed the following experiment. Materials and Methods Animals Fifty-six female Sprague-Dawley rats weighing between 230 and 270 g were used in this study. They were housed in individual metabolic cages and fed powdered Purina Laboratory chow and tap water ad libitum. The lighting was automatically regulated to provide 12 h of darkness and 12 hours of light (0600 to 1800 h). The room temperature was approximately 21 C. Body weights were measured weekly and food intake was recorded daily during the experiment. Experimental
Received February 26,1976. Supported in part by NIH Grant Nos. Am 18165 and RR 00425.
procedures
Under pentobarbital anesthesia, 50 rats received bilateral electrolytic lesions in the ventromedial hypothalamus (VMH) using coordinates
108
VAGOTOMY AND OBESITY from the DeGroot Atlas (at the Bregma anteriorly; ±0.75 mm lateral to the midsagital line, and 1.0 mm above the base of the skull) (8). Another six rats were subject to sham-VMH operations. Only 37 of the 50 VMH-lesioned rats which gained more than 70 g in two weeks were used in the study. Two weeks later, subdiaphragmatic vagotomy was performed in 25 of these rats according to the method of Snowdon and Epstein (9). A sham-vagotomy was performed in 12 others and in the 6 sham-VMH controls. Four weeks following vagotomy, all groups of rats were bled by cardiac puncture under light ether anesthesia for measurement of serum glucose and insulin. Several days later, while under pentobarbital anesthesia, the cervical vagus was electrically stimulated and the secretion of gastric acid from the cannulated stomach was measured before and after stimulation in order to document the completeness of vagotomy (7). Rats were classified as follows: Group 1, six VMH-lesioned rats with sham-vagotomy which were fed ad libitum; Group 2, twelve vagotomized VMH-lesioned rats which were fed ad libitum. This group was subsequently subdivided into two groups: Group 2a—seven rats with complete vagotomy, and Group 2b—five rats with incomplete vagotomy as determined by measuring the release of gastric acid before and after direct electrical stimulation of the cervical vagus nerves using the method of Powley and Opsahl (7); Group 3, five VMH-lesioned rats with a sham-
109
vagotomy which were pair-fed to the group with complete vagotomy by giving the average daily food intake of the latter group. Group 4 consisted of six VMH-sham operated rats with a sham-vagotomy which were fed ad libitum (control group). Analytical methods The Lee index was used to estimate fatness (10). Serum insulin was measured by a double antibody method using rat insulin standard and labeled pork insulin (11). Serum glucose was measured by a glucose-oxidase method. Statistical comparisons utilized data with Student's t test. Correlation and partial correlation coefficients were determined with the BMDP programs and an IBM 360/90 computer.
Results Figure 1 shows that the VMH-lesioned rats with a sham-vagotomy continued to gain weight throughout the study to reach an average weight of 485 g. The group with incomplete vagotomy lost weight for two weeks after vagotomy and then began to regain weight slowly. The control group reached the same final weight as the group with incomplete vagotomy. The group with VMH lesions and complete vagotomy showed a marked loss of weight during the
500-i Sham-Vagotomy
FIG. 1. Body weight of rats with ventromedial hypothalamic lesions. Each group is plotted as the mean ± SEM. Sham-vagotomy — • —; incomplete vagotomy — • —; complete vagotomy — O—; VMH pairfed to sham-vagotomy — • —; control group (sham lesion-sham vagotomy) — A —.
400-
1 Incomplete ""•"¥ Vagotomy ^-Control
300-
Pair-Fed 2 Complete Vagotomy
200-
VMH
VAGOTOMY
WEEKS
Endo • 1977 Vol 100 • No 1
INOUE AND BRAY
110
TABLE 1. Serum glucose and immunoreactive insulin levels after overnight fast in rats subject to ventromedial hypthalamic (VMH) lesions Operative status Group
VMH
1
+\
Serum glucose (mg/dl)
Leef index
Vagotomy Sham
0.352 ±.011*
