Decreasing hypothalamic insulin receptors causes hyperphagia and insulin resistance in rats

We investigated the role of hypothalamic insulin signaling in the regulation of energy balance and insulin action in rats through selective decreases in insulin receptor expression in discrete hypothalamic nuclei. We generated an antisense oligodeoxynucleotide directed against the insulin receptor precursor protein and administered this directly into the third cerebral ventricle. Immunostaining of rat brains after 7-day administration of the oligodeoxynucleotide showed a selective decrease of insulin receptor protein within cells in the medial portion of the arcuate nucleus (decreased by approximately 80% as compared to rats treated with a control oligodeoxynucleotide). Insulin receptors in other hypothalamic and extra-hypothalamic areas were not affected. This selective decrease in hypothalamic insulin receptor protein was accompanied by rapid onset of hyperphagia and increased fat mass. During insulin-clamp studies, physiological hyperinsulinemia decreased glucose production by 55% in rats treated with control oligodeoxynucleotides but by only 25% in rats treated with insulin receptor antisense oligodeoxynucleotides. Thus, insulin receptors in discrete areas of the hypothalamus have a physiological role in the control of food intake, fat mass and hepatic action of insulin.     

Caloric compensation in hypothalamic obese rats

The present study explored the influence of pelleted diets on adjustment to caloric dilution in hypothalamic obese rats. Medial hypothalamic lesioned and normal rats were maintained on a high fat diet until the static stage of hyperphagia was reached. They were given three pelleted diets which consisted of undiluted Noyes pellets and Noyes pellets diluted 25 or 50 percent with kaolin. In contrast to previous reports that hypothalamic obese rats do not compensate for caloric dilution of their diet, the obese animals showed as large an increase in food intake on the dilute pelleted diets as did the control animals. These results were interpreted by viewing texture as a dimension of palatability which influences food intake in hypothalamic obese rats.     

Decreased insulin receptor binding in hyperthyroidism

Summary The binding of¹²⁵I-insulin to insulin receptors on circulating mononuclear leukocytes was studied in ten patients with hyperthyroidism and 20 euthyroid normal volunteers. The hyperthyroid patients demonstrated significantly elevated glucose levels following an oral glucose load, despite normal insulin secretion. The infusion of insulin resulted in a delayed hypoglycaemic effect in the hyperthyroid patients; however, the inhibition of the endogenous insulin secretion as indicated by suppression of C-peptide levels was not different from euthyroid control subjects. Insulin binding to monocytes was significantly decreased in the hyperthyroid patients. Scatchard analysis of binding data indicates that a decrease of receptor number rather than receptor affinity seems to be the cause of the lowered insulin binding in hyperthyroid patients with diffuse toxic goitre. The findings of decreased insulin receptor number, mild degree of glucose intolerance despite normal insulin secretion and the delayed hypoglycaemic effect following insulin infusion suggest that peripheral insulin resistance could be involved in the highly complex pathophysiology of glucose intolerance in hyperthyroidism.Abkürzungsverzeichnis T4 thyroxine - T3 triiodothronine - OGTT Oral Glucose Tolerance Test - K_E insulin tracer concentration Presented in part at the 12th Annual Meeting of the European Thyroid Association, Brussels, September 6–10, 1982     

Decreased pancreatic CCK receptor binding and CCK-stimulated amylase release in Zucker obese rats

In Zucker obese rats the response to the effects of CCK on food intake and pancreatic exocrine function are decreased. However, it is unknown whether the decreased responsiveness is due to decreased receptor number and/or sensitivity or abnormal circulating concentrations of CCK. In these experiments percent total binding of 125I-CCK-33 to pancreatic acini from obese rats was one-half that in lean rats when data was expressed on a per microgram DNA basis (19.6 +/- 5.1 vs. 47.4 +/- 11.4, p less than 0.01). In a second experiment while the maximally effective dose of CCK for stimulating amylase secretion from dispersed pancreatic acini was similar in obese and lean rats (10(-10) M), less amylase was secreted in obese rats across the dose range tested (p less than 0.001). In contrast, carbachol had similar potency and efficacy in stimulating amylase release from obese and lean pancreatic acini. The increase of pancreas size by use of a trypsin inhibitor was greater in lean than obese rats (p less than 0.03). In addition, stimulation of amylase release by CCK from obese trypsin inhibitor-treated compared with control obese rats was greater than that from lean trypsin inhibitor-treated compared with control lean rats (p less than 0.002). However, overall, stimulation of amylase secretion by CCK was only 36% of control (p less than 0.001) and by carbachol was only 20% of control (p less than 0.001). Thus, increased size by increased cell number was associated with decreased response per cell.(ABSTRACT TRUNCATED AT 250 WORDS)     

Decreased pancreatic exocrine response to cholecystokinin in Zucker obese rats

Cholecystokinin (CCK), one of the peptides secreted by the gastrointestinal tract during a meal, stimulates release of enzymes into pancreatic juice and is a trophic hormone for the pancreas. Administration of CCK also decreases food intake, and obese rats have been shown to have a higher threshold than lean rats for this apparent effect on satiety. In this study experiments were designed to compare the sensitivity of obese and lean rats to the effects of CCK octapeptide (CCK-8) on pancreatic structure and exocrine function. In both growing and adult Zucker rats DNA content of the pancreas from obese rats was decreased compared with that from lean rats [2.42 +/- 0.21 vs. 3.07 +/- 0.18 mg (P less than 0.01) and 2.46 +/- 0.25 vs. 3.01 +/- 0.19 mg (P less than 0.05), respectively], and in adult obese rats this was accompanied by decreased pancreas size on both absolute weight and percent of body weight bases. In adult obese Bar Harbor mice, although DNA content of the pancreas was also decreased [1.70 +/- 0.10 vs. 2.41 +/- 0.11 mg (P less than 0.01)], pancreas weight was not different (0.30 +/- 0.01 vs. 0.32 +/- 0.01 g). In young rats growth of the pancreas was stimulated by 2 micrograms/kg CCK-8 administered subcutaneously or 100 mg/kg of a trypsin inhibitor administered orally twice daily for 2 wk. Although both treatments increased weight and DNA and protein content of the pancreas, the increases in DNA and protein content were smaller in obese than lean rats, indicating a decreased responsiveness to both trophic agents. Administration of CCK-8 stimulated smaller increases in pancreatic juice volume and amylase release in obese compared with lean rats, indicating decreased pancreatic exocrine function in response to CCK. In adults the CCK-8 dose-response curve for amylase release from dispersed pancreatic acini of obese rats was similar to that of lean rats, indicating normal sensitivity in vitro. Thus, in obese rats and mice DNA content of the pancreas is decreased when compared with that of lean rats and mice, and this is accompanied by decreased in vivo responses to CCK in obese rats.     

Sympathoadrenal function in genetically obese Zucker rats.

The effects of genetic obesity on the actions and alterations of the sympathetic nervous system were studied in 10-12-month-old obese (fa/fa) and lean (Fa/-) Zucker rats. Blood glucose, plasma insulin, epinephrine (E), norepinephrine (NE), and free fatty acids (FFA) concentrations were measured in blood samples taken through a permanent heart catheter before, during, and after exercise or intravenous infusion of E and NE. Baseline plasma FFA and insulin levels were markedly increased in the obese animals. Exercise, i.e., strenuous swimming against a counter current for 15 min, led to reduction of plasma insulin concentrations and an increase of all other blood components in lean Zucker rats. In obese animals, an exaggerated increase of blood glucose and a large suppression of plasma insulin occurred. Plasma FFA levels tended to decline during exercise. Plasma catecholamine patterns in the exercising fatty Zuckers were not different to those of the lean animals. Infusion of E caused an increase of blood glucose and a decrease of plasma insulin concentrations in both groups of animals. The increase in blood glucose in the obese animals was significantly larger compared to the changes in the lean animals. Infusion of NE significantly reduced plasma insulin concentration in obese but not in lean animals. The results revealed that activation of the sympathetic system, expressed as exercise-induced alterations in plasma E and NE levels, is normal in obese Zucker rats. However, postsynaptic receptor effects of catecholamines on glycogenolysis and lipolysis are different in obese and lean animals, which points to permanent changes in adrenoceptor mechanisms on adipocytes, hepatocytes, and muscle cells in obesity.     

Glucose utilization and insulin binding in discrete brain areas of obese rats.

The present study was carried out to determine whether genetically obese Zucker rats present changes in brain glucose utilization and/or insulin binding when compared to their lean counterparts. Glucose utilization in the whole brain, determined by measurement of 2-deoxy(1-3H)glucose-6-phosphate, was significantly lower in obese than in lean Zucker rats. In order to precise the structure involved, we then used quantitative autoradiography methods after either (1-14C) 2-deoxyglucose injection or 125I-insulin incubation. In obese rats, local cerebral glucose utilization (LCGU) was significantly decreased in the external plexiform layer (-37%, p < 0.05), in the lateral hypothalamus (-23%, p < 0.05), and in the basolateral amygdaloid nucleus (-30%, p < 0.05). In contrast, no difference in specific insulin binding was found between the two genotypes in any of the areas studied. These results are consistent with some data showing a decrease of LCGU in hyperinsulinemic rats. All together, these data show perturbations of glucose utilization, particularly in structures linked to the regulation of body weight and food intake in obese Zucker rats.     

Sympathetic hyperresponsiveness to hypothalamic stimulation in young hypertensive rats.

The possible occurrence of central sympathetic dysfunction during development of spontaneous hypertension was studied by recording aortic pressure and sympathetic nerve activity concurrently during electrical stimulation of the posterior hypothalamus in 9-wk-old Kyoto-Wistar rats. Even at this early age, basal levels for both measurements were already elevated significantly in those with spontaneous hypertension. Increases in sympathetic neural firing induced by graded hypothalamic stimulation were always followed by corresponding increases in blood pressure; magnitude of both effects was appreciably larger in spontaneously hypertensive than in normotensive rats, as was the vasodepression caused by blocking autonomic ganglia with pentolinium. By contrast, pressor responses to injected norepinephrine were almost equal thereby suggesting that cardiovascular reactivity was unaltered and that enhanced responsiveness to hypothalamic stimulation was directly due to the concomitant increase in sympathetic nerve activity. Although the exact site from which sympathetic hyperactivity originates was unidentified, our results support the interpretation that sympathetic mechanisms involving the posterior hypothalamus participate in elevating blood pressure during development of spontaneous hypertension in rats.     

Metabolic responses to fasting and refeeding in lean and genetically obese rats

Injection of norepinephrine (NE) (25 micrograms/100 g body wt) caused a similar rise in metabolic rate in lean and obese (fa/fa) Zucker rats, but 3-day fasting suppressed the response in lean rats and enhanced the rise in obese mutants. Triiodothyronine (T3) injection (10 micrograms/100 g body wt) caused a significantly greater rise in oxygen consumption (Vo2) in obese than lean rats, but the response was attenuated by fasting in all animals. The thermic response to a single meal of either mixed composition, carbohydrate, or protein (40 kJ) was much smaller in obese rats than lean, but the response to the mixed nutrient meal was similar for all rats after a 3-day fast. Refeeding 3-day fasted lean rats with a single carbohydrate meal (40 kJ) caused a rise in plasma T3 levels after 3 h and a delayed increase in metabolic rate 24 h later. Injection of NE instead of refeeding caused a similar delayed rise in metabolic rate. Carbohydrate refeeding had no effect on plasma T3 levels or oxygen consumption in 3-day fasted obese Zuckers, but injection of NE did produce a significant increase in metabolic rate after 24 h. Refeeding 3-day fasted rats with protein (40 kJ) caused a rise in oxygen consumption 24 h later in lean animals but had no effect in obese animals. The data from lean Zucker rats confirm previous findings in Sprague-Dawley rats and suggest that the thermic response to refeeding involves a complex interaction between the sympathetic nervous system and thyroid hormones. Obese Zuckers responded normally to NE and T3, indicating that their reduced thermogenesis after food may be due to insensitivity to nutrient availability or an inability to activate the sympathetic nervous system.     

Endogenous glucagon regulation in genetically hyperlipemic obese rats.

Glucagon concentration and regulation were examined in the Zucker rat, in which obesity and hyperlipemia are phenotypic expressions of an autosomal recessive gene. Using littermate animals which are phenotypically thin and normolipemic as controls, we observed reduced basal plasma glucagon levels in the obese lipemic rats. In response to fasting, obese lipemic animals inappropriately demonstrated a further reduction in plasma glucagon concentration. In response to pharmacologic glucagon stimulation (arginine), a subnormal rise in plasma glucagon concentration was observed in the obese, lipemic animals. Glucagon suppressibility with exogenous glucose remained intact. The reduced secretion of glucagon may be a consequence of the abnormal elevation in concentration of plasma insulin, free fatty acids, and glucose, which are characteristic of the obese, lipemic animal. A possible role of glucagon deficiency in the evolution or maintenance of the lipemic state is suggested.     

Decreased hypothalamic aromatization in female rats of true precocious puberty

The true precocious puberty animal model induced by the single dose of danazol was used for investigating the expressions of hypothalamic aromatase in the advanced onset of puberty in rats. The day of vaginal opening and first estrus showed significant advancement in the model rats compared with the normal and vehicle rats (P < 0.01, respectively). The hypothalamic gonadotropin-releasing hormone (GnRH) mRNA expression increased significantly in the model rats compared with that in the normal and vehicle ones (P < 0.01). The levels of aromatase mRNA and protein expressions detected by RT-PCR and Western blot both decreased in the model rats compared with those in the normal and vehicle groups (P < 0.05). The results suggested that the hypothalamic aromatization might diminish in the onset of true precocious puberty of female rats.     

Decreased sensitivity of Zucker obese rats to the putative satiety agent cholecystokinin

In obese rodents increased daily food intake leading to accumulation of adipose tissue is frequently accompanied by increased meal size and loss of the normal diurnal variations in feeding pattern. Increased meal size of obese rats may be due to decreased sensitivity to factors which elicit satiety. We compared Zucker obese and lean rat feeding behavior responses to octapeptide of cholecystokinin (OP-CCK), a peptide shown to decrease meal size in several species. Obese rats were less sensitive than lean rats to OP-CCK (.06, .25 and 1.0 μg/kg/meal) injected before each of four consecutive scheduled meals in the light portion of the diurnal cycle, when obese meal size was larger than lean. However, neither obese nor lean rats responded to injection of the same doses of OP-CCK during meals in the dark, when average meal size was larger than during the light and when average meal size of the obese rats was similar to that of lean rats. In both obese and lean rats injection of OP-CCK affected daily feeding pattern. Obese and lean Zucker rats are less sensitive to OP-CCK when meal size is larger, whether this is due to phase of the diurnal cycle (dark vs. light in both obese and lean rats) or phenotype (obese vs. lean rats in the light).     

Food motivated behavior in genetically obese and hypothalamic-hyperphagic rats and mice

The food-associated behavior of genetically obese Zucker rats, fa/fa and yellow obese mice, aAy, was studied using classic operant procedures. When obese Zucker rats and yellow mice are compared to their lean littermates and lean littermates made obese by electrolytic lesions and chemical lesions, respectively, the naturally obese animals do not display the behavioral patterns associated with rodents made obese by hypothalamic damage. These experiments point out the necessity for careful selection of animal models in studying behaviors associated with regulatory disturbances of normal food intake.     

Responsivity to NPY and melanocortins in obese OLETF rats lacking CCK-A receptors

Otsuka Long-Evans Tokushima Fatty (OLETF) rat lacking CCK-A receptors are hyperphagic and obese. Previous work has demonstrated alterations in neuropeptide Y (NPY) and proopiomelanocortin (POMC) mRNA expression in ad libitum fed OLETF rats compared to lean Long-Evans Tokushima Otsuka (LETO) controls. In order to determine whether alterations in sensitivity to central peptides involved in overall feeding control may contribute to the hyperphagia and obesity in OLETF rats, we assessed OLETF and LETO rats feeding responses to lateral ventricular infusions of NPY (1 and 3.2 nmol), the melanocortin 3/4 agonist MTII (0.1 and 0.32 nmol) and the melanocortin receptor antagonist SHU-9119 (0.25 and 0.5 nmol). At a 3-h time point, NPY increased food intake in both OLETF and LETO rats. OLETF rats were more sensitive, having significant increases at both NPY doses and a greater increase at the higher dose. The melanocortin agonist MTII decreased intake in both LETO and OLETF rats. At the 20-h time point, the magnitude of suppression was greater in OLETF rats. SHU-9119 increased food intake in both groups. OLETF rats were more sensitive with larger relative increase and longer-lasting effects at the lower dose. These results are consistent with demonstrated alterations in neuropeptide gene expression in OLETF rats and indicate that alterations in responsivity to NPY and melanocortin signaling are unlikely to contribute to their hyperphagia and obesity.     

Daily rhythms of feeding in the genetically obese and lean Zucker rats

Feeding patterns were examined in obese (fa/fa) and lean (Fa/-) adult Zucker rats over the light-dark cycle during 14 days. Obese rats eat more than lean rats especially during the dark phase. Light and dark feeding expressed as percentage of 24 hr intake showed no significant differences between the lean and obese groups. The higher food intake in obese rats is mainly caused by larger meals since obese rats ate fewer meals than lean rats. Only for the obese group differences were observed between mean meal size in light and dark phase. There is some indication that the circadian controlled temporal distribution of meals is different in obese rats compared to lean rats since obese rats eat fewer but larger meals during the first half of the dark phase. During this phase meal size increases gradually in the obese rats, suggesting that the circadian influence on feeding motivation is increased.     

Sleep loss alters hypothalamic growth hormone-releasing hormone receptors in rats

Previous experiments suggest that sleep deprivation (SD) is associated with growth hormone-releasing hormone (GHRH) release and that GHRH promotes sleep via intrahypothalamic sites of action. Binding of [His(1), (125)I-Tyr(10), Nle(27)]hGHRH(1-32) amide and GHRH receptor (GHRH-R) mRNA levels were determined in the hypothalamus and pituitary of rats subjected to 8 h of SD and of undisturbed control rats. The characteristics of the hypothalamic GHRH binding sites differed from those of the pituitary. High affinity GHRH binding and GHRH-R mRNA levels decreased by 50% in the hypothalamus of SD rats, whereas there were no alterations in the pituitary. The results demonstrate that GHRH-Rs exist in the hypothalamus and they respond differently to SD than the GHRH-Rs in the pituitary. The SD-induced changes are explained by down-regulation of the hypothalamic GHRH-Rs induced by GHRH release during and after SD.     

Passive avoidance behavior in lean and obese rats with ventromedial hypothalamic lesions

Lean and obese rats with ventromedial hypothalamic lesions performed reliably worse than control animals in the acquisition of a step-down passive avoidance task. However, obese rats performed significantly better than lean VMH animals, which consistently leaped off the platform on the second and succeeding trials. While there were no significant differences between groups in the acquisition of a step-through passive avoidance task, lean and obese rats with VMH lesions took reliably longer than control animals to reach criterion when an identical step-through response had previously been reinforced (punishment-extinction of a one-way conditioned avoidance response). Both lean and obese VMH-damaged rats made more punished approach responses to water than control animals following water-deprivation to 88% of body weight, but only lean VMH rats made a significantly greater number of punished approach responses to liquid food than unoperated animals following food-deprivation to 88% of body weight. The number of punished consummatory responses appeared to be influenced by baseline intake. Among the animals tested in more than one paradigm, there was a significant positive correlation between the number of punished consummatory responses and the number of shocks received during punishment-extinction of the one-way CAR, but no relationship was observed between the performances in either of these and the step-down avoidance paradigm. The impaired passive avoidance behavior by rats with VMH lesions is attributed to both an inability to inhibit a previously reinforced response and a change in response tendencies to aversive stimuli.