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Aims/hypothesis: Roux-en-Y gastric bypass (RYGB) surgery, an effective treatment for morbid obesity, commonly leads to near complete resolution of type 2 diabetes. The underlying mechanisms, however, remain unclear and factors other than weight loss alone may be involved., Methods: To determine whether increased hypothalamic insulin sensitivity after RYGB drives the rapid improvement in glucose metabolism, high-fat-fed rats received either an insulin receptor (IR) antisense vector or a control lentiviral vector that was microinjected into the ventromedial hypothalamus (VMH). Six weeks later, rats underwent RYGB or control gastrointestinal surgery., Results: Four weeks after surgery, weight loss was comparable in RYGB and surgical controls. Nevertheless, only RYGB rats that received the control vector demonstrated both improved hepatic and peripheral insulin sensitivity. Insulin suppressed hepatic glucose production (HGP) by 50% (p < 0.05) with RYGB, whereas the effect of insulin on HGP was completely absent in VMH IR knockdown (IRkd) rats. By contrast, both RYGB groups displayed an identical twofold increase in insulin-stimulated peripheral glucose uptake. The animals that underwent control gastrointestinal surgery failed to show any improvement in either hepatic or peripheral insulin sensitivity; VMH IRkd did not influence the magnitude of insulin resistance., Conclusions/interpretation: Our findings demonstrate that RYGB surgery in high-fat-fed obese rats enhances hepatic and peripheral insulin sensitivity independently of weight loss. The improved hepatic, but not the peripheral, response to insulin is mediated centrally at the level of the VMH. These data provide direct evidence that the metabolic benefits of RYGB surgery are not simply a consequence of weight loss but likely in part involve the central nervous system.

Current studies show that type II glucocorticoid receptor (GR) stimulation during recurring insulin-induced hypoglycemia (RIIH) results in diminished hypoglycemic activation of neurons in discrete CNS metabolic structures, namely the lateral hypothalamic area (LHA), hypothalamic paraventricular (PVH) and dorsomedial (DMH) nuclei, and nucleus of the solitary tract (NTS). The present work utilized immunofluorescence histochemistry to evaluate the reactivity of GR-expressing neurons in characterized hypothalamic, thalamic, and hindbrain metabolic structures to glucoprivation, and to determine if antecedent hypoglycemic stimulation of central GR decreases Fos protein expression by these neurons. Groups of adult male rats were injected subcutaneously with one or four doses of the intermediate-acting insulin, Humulin NPH, on as many days, while controls received diluent only. Rats injected with four doses of insulin were pretreated by intracerebroventricular administration of the selective GR antagonist, CP-475222, or vehicle alone prior to insulin doses 1-3. All animals were sacrificed by trancardial perfusion 2 h after injections on day four of the study. Mean numbers of GR-immunoreactive (-ir) neurons did not differ between groups injected with diluent versus one dose of insulin in each structure evaluated, but were significantly elevated above baseline on the fourth day of RIIH in the LHA and DMH, but not the PVH, VMH, ARC, thalamic paraventricular (PVT), or NTS. Counts of GR-ir-positive neurons in each site were similar between groups treated with CP-475222 or vehicle icv during RIIH. While mean numbers of GR-plus Fos-ir neurons in the PVH, DMH, LHA, and NTS, but not the PVT were significantly elevated after one dose of NPH, this increase was abolished in each site by RIIH. Pharmacological antagonism of central GR during antecedent hypoglycemia prevented RIIH-associated habituation of Fos colabeling of GR-expressing neurons in the PVH, DMH, and LHA. These data show that RIIH increases nuclear immunolabeling for GR in discrete CNS metabolic structures, evidence that recurring metabolic stress may amplify receptor-mediated genomic regulatory function in local neurons. The results also demonstrate that GR-containing neurons in the LHA, DMH, PVH, and NTS react to hypoglycemia by induction of the Fos stimulus-transcription cascade, and that precedent stimulation of central GR is critical for RIIH-associated habituation of this functional response in the former three sites.

Hypoglycemia elicits an integrated array of CNS-mediated counterregulatory responses, including activation of the hypothalamic-pituitary-adrenal axis. The role of antecedent adrenocortical hypersecretion in impaired glucose counterregulation remains controversial. The present studies utilized the selective, nonsteroidal glucocorticoid receptor antagonist, CP-472555, as a pharmacological tool to investigate the hypothesis that hypoglycemic hypercorticosteronemia modulates CNS efferent autonomic and neuroendocrine motor responses to recurring insulin-induced hypoglycemia via glucocorticoid receptor-dependent mechanisms. Groups of adult male rats were injected s.c. with either one or four doses of the intermediate-acting insulin, Humulin neutral protamine Hagedorn (NPH), on as many days, while controls were injected with diluent alone. Animals injected with four doses of insulin were pretreated by i.c.v. administration of graded doses of the glucocorticoid receptor antagonist or vehicle alone prior to the first three doses of insulin. Repeated daily injection of NPH exacerbated hypoglycemia, attenuated patterns of glucagon and epinephrine secretion, and diminished neuronal transcriptional activation in discrete CNS metabolic loci, including the lateral hypothalamic area, dorsomedial hypothalamic nucleus, paraventricular hypothalamic nucleus, and nucleus of the solitary tract. While i.c.v. delivery of 25 or 100 ng doses of CP-472555 did not alter any of these parameters, animals treated with 500 ng exhibited circulating glucose, glucagon, and epinephrine levels that were similar to those in rats injected with one dose of insulin, as well as a reversal of recurring insulin-induced hypoglycemia-associated reductions in Fos immunolabeling in the lateral hypothalamic area, dorsomedial hypothalamic nucleus, and paraventricular hypothalamic nucleus. These results provide unique pharmacological evidence that antecedent activation of central glucocorticoid receptor is required for exacerbation of hypoglycemia during recurring insulin-induced hypoglycemia, and that these receptors mediate modulatory effects of hypoglycemic hypercorticosteronemia on autonomic efferent responses to recurring insulin-induced hypoglycemia. The data also suggest that neurons in central loci characterized here by antagonist-mediated overturn of recurring insulin-induced hypoglycemia-induced decreases in neuronal transcriptional activation may be direct or indirect substrates for this hormonal modulation action.

Recurrent insulin-induced hypoglycemia (RIIH) results in glucose counterregulatory dysfunction in men and male rodents. Intensified hypoglycemia in the latter coincides with diminished neuronal Fos expression in central metabolic regulatory structures, evidence that supports habituation of CNS-mediated compensatory motor outflow during re-exposure to this metabolic stress. In light of the evidence for counterregulatory resistance to precedent hypoglycemia in women, we utilized estradiol-treated ovariectomized (OVX) female rats to examine the hypothesis that this hormone regulates neural adaptability to recurring hypoglycemia. Groups of OVX rats were implanted with subcutaneous silastic capsules containing estradiol benzoate (E) or oil alone, and injected subcutaneously with one or four doses of the intermediate-acting insulin, Humulin NPH, one dose daily, or with diluent alone. Blood glucose levels were not altered by RIIH in E-implanted OVX animals, but were significantly decreased after four versus one insulin injection in the OVX+oil group. Mean numbers of Fos-immunoreactive (ir) neurons in the paraventricular nucleus hypothalamus (PVH), dorsomedial nucleus hypothalamus (DMH), and lateral hypothalamic area (LHA) were higher in both E- versus oil-implanted OVX rats injected with diluent only. Acute hypoglycemia significantly increased mean counts of Fos-ir-positive neurons in the PVH, DMH, and LHA, as well as the nucleus of the solitary tract (NTS) and area postrema (AP) in E- and oil-treated animals to an equivalent extent. OVX+E rats exhibited comparable numbers of Fos-positive neurons in the PVH, DMH, and LHA after one versus four insulin injections, whereas the numbers of labeled neurons in NTS and AP were increased or decreased, respectively, by RIIH. Oil-implanted OVX rats showed significantly diminished numbers of Fos-ir-positive neurons in each neural structure after repeated hypoglycemia. The present data demonstrate that estradiol sustains or enhances neuronal reactivity to recurring hypoglycemia in central metabolic structures, whereas hypoglycemic patterns of Fos expression in each site become habituated during RIIH in the absence of this steroid. The brain sites characterized here by estrogen-dependent maintenance of neuronal genomic reactivity to this substrate fuel imbalance may contain direct and/or indirect cellular targets for hormonal actions that prevent adaptation of CNS-controlled motor responses to this metabolic stress., (Copyright (c) 2006 S. Karger AG, Basel.)