Chronic reduction of insulin receptors in the ventromedial hypothalamus produces glucose intolerance and islet dysfunction in the absence of weight gain

Insulin is believed to regulate glucose homeostasis mainly via direct effects on the liver, muscle, and adipose tissues. The contribution of insulin's central nervous system effects to disorders of glucose metabolism has received less attention. To evaluate whether postnatal reduction of insulin receptors (IRs) within the ventromedial hypothalamus (VMH), a brain region critical for glucose sensing, contributes to disorders of peripheral glucose metabolism, we microinjected a lentiviral vector expressing an antisense sequence to knockdown IRs or a control lentiviral vector into the VMH of nonobese nondiabetic rats. After 3-4 mo, we assessed 1) glucose tolerance, 2) hepatic insulin sensitivity, and 3) insulin and glucagon secretion, using the glucose clamp technique. Knockdown of IRs locally in the VMH caused glucose intolerance without altering body weight. Increments of plasma insulin during a euglycemic clamp study failed to suppress endogenous glucose production and produced a paradoxical rise in plasma glucagon in the VMH-IR knockdown rats. Unexpectedly, these animals also displayed a 40% reduction (P < 0.05) in insulin secretion in response to an identical hyperglycemic stimulus (～220 mg/dl). Our data demonstrate that chronic suppression of VMH-IR gene expression is sufficient to impair glucose metabolism as well as α-cell and β-cell function in nondiabetic, nonobese rats. These data suggest that insulin resistance within the VMH may be a significant contributor to the development of type 2 diabetes.     

Cardiovascular Vagosympathetic Activity in Rats with Ventromedial Hypothalamic Obesity

Objective: Rats with ventromedial hypothalamic lesion (VMH) are massively obese with endogenous hyperinsulinemia, insulin resistance, low sympathetic activity, and high parasympathetic activity, which are likely to induce hypertension. The goal was to follow in this model the long‐term hemodynamic changes and to investigate the role of autonomic nervous system and insulin resistance in these changes. Research Metho ds and Procedures: Heart rate and blood pressure were monitored for 12 weeks after operation using a telemetric system in VMH and sham rats. Plasma catecholamines and heart β‐adrenoceptors were measured. Glucose tolerance was studied after an intravenous glucose injection and insulin sensitivity during a euglycemic hyperinsulinemic clamp test. Results: A marked bradycardia and only a mild increase in blood pressure occurred in VMH rats compared with sham animals. Response to autonomic‐acting drugs showed an increase in heart vagal tone and responsiveness to a β‐agonist drug. Plasma catecholamine levels were markedly increased, and the density and affinity of heart β‐adrenoceptors were similar in VMH, sham, and control rats. Muscle glucose use was reduced by 1 week after operation in VMH animals. Discussion: These results show the following in this model of massively obese rats with sympathetic impairment: 1) adrenal medulla secretion is increased, probably as a result of hyperinsulinemia and increased vagal activity; 2) cardiac responsiveness to β‐agonist stimulation is increased; and 3) despite these changes and suspected resistance to the vasodilative effect of insulin, blood pressure does not increase. We conclude that high vagal activity may be protective against hypertension associated with obesity.     

A mechanism by which primary or secondary hypothalamic involvement results in the development of insulin-dependent diabetes mellitus (IDDM)

A literature survey and hypothesis is presented in which it is concluded that an intracellular ventromedial hypothalamic (VMH) glucopenia results in a bibrachial response consisting of adenohypophysial release of growth hormone and ACTH as well as sympathetic discharge, both of which act to elevate plasma glucose and remove the VMH glucopenia. This glucopenia may occur as a result of either a deficiency of circulating insulin or alterations in the kinetic properties of the VMH cellular insulin receptor. Two mechanisms for the development of insulin dependent diabetes mellitus (IDDM) are presented: (1) a defect in VMH glucose transport and/or metabolism such that a VMH glucopenia occurs with a subsequent bibrachial response. The result of this is glucose overproduction and a chronic excess glucose stimulus will eventually cause B-cell exhaustion (primary hypothalamic involvement). (2) reduction of the B-cell population by chemical, genetic and/or viral interactions with a consequential insulopenia results in a VMH glucopenia (secondary to a reduced glucose transport into the VMH cells) and causes a bibrachial response. This VMH response may temporarily restore plasma glucose balance but a chronically enhanced counter-regulatory response to maintain this balance will eventually stress the remaining B-cell population and cause further reductions in B-cell numbers (secondary hypothalmic involvement).     

Unacylated ghrelin acts as a potent insulin secretagogue in glucose-stimulated conditions

Acylated and unacylated ghrelin (AG and UAG) are gut hormones that exert pleiotropic actions, including regulation of insulin secretion and glucose metabolism. In this study, we investigated whether AG and UAG differentially regulate portal and systemic insulin levels after a glucose load. We studied the effects of the administration of AG (30 nmol/kg), UAG (3 and 30 nmol/kg), the ghrelin receptor antagonist [D-Lys(3)]GHRP-6 (1 micromol/kg), or various combinations of these compounds on portal and systemic levels of glucose and insulin after an intravenous glucose tolerance test (IVGTT, d-glucose 1 g/kg) in anesthetized fasted Wistar rats. UAG administration potently and dose-dependently enhanced the rise of insulin concentration induced by IVGTT in the portal and, to a lesser extent, the systemic circulation. This UAG-induced effect was completely blocked by the coadministration of exogenous AG at equimolar concentrations. Similarly to UAG, [D-Lys(3)]GHRP-6, alone or in combination with AG and UAG, strongly enhanced the portal insulin response to IVGTT, whereas exogenous AG alone did not exert any further effect. Our data demonstrate that, in glucose-stimulated conditions, exogenous UAG acts as a potent insulin secretagogue, whereas endogenous AG exerts a maximal tonic inhibition on glucose-induced insulin release.     

Effects of stimulation of hypothalamic “feeding areas” on endocrines and metabolism in normal and diabetic monkeys

Electrical stimulation of hypothalamic “feeding areas” (VMH & LHA) through stereotaxically implanted electrodes was carried out in normal and streptozotocinized diabetic conscious male rhesus monkeys. In normal monkeys, the VMH stimulation resulted in a significant decrease in serum insulin and blood glucose while opposite responses were observed following LHA stimulation. Serum growth hormone, FFA and plasma cortisol levels increased significantly on stimulation of LHA and VMH in normal animals. The pattern of blood glucose and serum growth hormone responses to stimulation of “feeding areas” was similar in normal and diabetic animals. The serum insulin, FFA and plasma cortisol were largely unaffected while growth hormone increased significantly by stimulation of these areas in diabetes. The present study indicates that the stimulation of “feeding areas” does not alter the diabetic syndrome significantly nor does diabetes prevent the changes in metabolism seen after stimulation of “feeding areas.”     

NEFA-glucose comodulation model of beta-cell insulin secretion in 24-h multiple-meal test

There is experimental evidence that a source of fatty acids (FAs) that is either exogenous or endogenous is necessary to support normal insulin secretion. Therefore, FAs comodulate the glucose-induced pancreatic insulin secretion. To assess the role of FAs, 16 morbidly obese nondiabetic patients and 6 healthy volunteers were studied. The controls and the obese subjects, before and after diet-induced weight loss, spent 24 h in a calorimetric chamber, where they consumed standardized meals. Hourly blood samples were drawn from a central venous catheter for the measurement of glucose, C-peptide, and nonesterified fatty acid (NEFA) concentrations. Insulin sensitivity was measured (as the M value) by euglycemic hyperinsulinemic clamp. In the present study, we propose a mathematical model in which insulin secretion rate (ISR) is expressed as a function of both plasma glucose and NEFA concentrations. Model parameters, obtained by fitting the individual experimental data of plasma C-peptide concentration, gave an estimated ISR comparable with that obtained by the deconvolution method. To evaluate the performance of the model in an experimental condition in which incretin effect was minimized, previous data on insulin secretion following a butter load and subsequent hyperglycemic clamp were reanalyzed. This model of nutrient-stimulated insulin secretion is the first attempt to represent in a simple way the interaction between glucose and NEFA in the regulation of insulin secretion in the beta-cell and explains, at least in part, the "potentiation factor" used in previous models to account for other control factors different from glucose after either an intravenous infusion of glucose or a mixed meal.     

Hypothalamic insulin signaling is required for inhibition of glucose production

Circulating insulin inhibits endogenous glucose production. Here we report that bidirectional changes in hypothalamic insulin signaling affect glucose production. The infusion of either insulin or a small-molecule insulin mimetic in the third cerebral ventricle suppressed glucose production independent of circulating levels of insulin and of other glucoregulatory hormones. Conversely, central antagonism of insulin signaling impaired the ability of circulating insulin to inhibit glucose production. Finally, third-cerebral-ventricle administration of inhibitors of ATP-sensitive potassium channels, but not of antagonists of the central melanocortin receptors, also blunted the effect of hyperinsulinemia on glucose production. These results reveal a new site of action of insulin on glucose production and suggest that hypothalamic insulin resistance can contribute to hyperglycemia in type 2 diabetes mellitus.     

Separate hypoglycaemic thresholds for gastric acid and pepsin secretion following insulin or posterior hypothalamic stimulation.

The level of hypoglycaemia required to elicit gastric secretion of acid and pepsin was studied in urethane-anaesthetized rats. Hypoglycaemia was induced by intravenous injection of insulin or by electrical stimulation of the posterior hypothalamus. In each case the blood glucose values below which gastric secretion was stimulated were significantly higher for pepsin than for acid secretion. This consistently resulted in the onset of pepsin secretion in advance of the onset of acid secretion. These observations suggest that the production of the different components of the gastric juice was under the influence of either separate hypothalamic glucoreceptors, or a single set of glucoreceptors able to respond selectively to different blood glucose levels.     

Hyperglycemia impairs glucose and insulin regulation of nitric oxide production in glucose-inhibited neurons in the ventromedial hypothalamus

Physiological changes in extracellular glucose, insulin, and leptin regulate glucose-excited (GE) and glucose-inhibited (GI) neurons in the ventromedial hypothalamus (VMH). Nitric oxide (NO) signaling, which is involved in the regulation of food intake and insulin signaling, is altered in obesity and diabetes. We previously showed that glucose and leptin inhibit NO production via the AMP-activated protein kinase (AMPK) pathway, while insulin stimulates NO production via the phosphatidylinositol-3-OH kinase (PI3K) pathway in VMH GI neurons. Hyperglycemia-induced inhibition of AMPK reduces PI3K signaling by activating the mammalian target of rapamycin (mTOR). We hypothesize that hyperglycemia impairs glucose and insulin-regulated NO production in VMH GI neurons. This hypothesis was tested in VMH neurons cultured in hyperglycemic conditions or from streptozotocin-induced type 1 diabetic rats using NO- and membrane potential-sensitive dyes. Neither decreased extracellular glucose from 2.5 to 0.5 mM, nor 5 nM insulin increased NO production in VMH neurons in either experimental condition. Glucose- and insulin-regulated NO production was restored in the presence of the AMPK activator, 5-aminoimidazole-4-carboxamide-1-b-4-ribofuranoside or the mTOR inhibitor rapamycin. Finally, decreased glucose and insulin did not alter membrane potential in VMH neurons cultured in hyperglycemic conditions or from streptozotocin-induced rats. These data suggest that hyperglycemia impairs glucose and insulin regulation of NO production through AMPK inhibition. Furthermore, glucose and insulin signaling pathways interact via the mTOR pathway.     

Autoregulation of Free Radicals via Uncoupling Protein Control in Pancreatic β-Cell Mitochondria

Pancreatic β-cells sense the ambient blood-glucose concentration and secrete insulin to signal other tissues to take up glucose. Mitochondria play a key role in this response as they metabolize nutrients to produce ATP and reactive oxygen species (ROS), both of which are involved in insulin secretion signaling. Based on data available in the literature and previously developed mathematical models, we present a model of glucose-stimulated mitochondrial respiration, ATP synthesis, and ROS production and control in β-cells. The model is consistent with a number of experimental observations reported in the literature. Most notably, it captures the nonlinear rise in the proton leak rate at high membrane potential and the increase in this leak due to uncoupling protein (UCP) activation by ROS. The functional forms used to model ROS production and UCP regulation yield insight into these mechanisms, as many details have not yet been unraveled in the experimental literature. We examine short- and long-term effects of UCP activation inhibition and changes in the mitochondrial density on mitochondrial responses to glucose. Results suggest increasing mitochondrial density while decreasing UCP activity may be an effective way to increase glucose-stimulated insulin secretion while decreasing oxidative stress.     

The effect of ventromedial hypothalamic nuclei destruction on somatostatin and insulin release from the isolated perfused rat pancreas

The effect of electrolytic lesions in the ventromedial hypothalamic nuclei (VMH) on somatostatin and insulin release was studied using the isolated perfused rat pancreas. Obesity gradually developed in the rats after placement of the VMH lesions, and fasting insulin levels determined immediately before the isolation of the pancreas were significantly higher than those in sham-operated controls. In the presence of 4.4 mM glucose, both perfusate somatostatin and insulin responses to arginine were significantly greater than in the controls, suggesting that VMH lesions cause not only hypersecretion of insulin but hypersecretion of somatostatin as well.     

Inhibition of hypothalamic carnitine palmitoyltransferase-1 decreases food intake and glucose production

The enzyme carnitine palmitoyltransferase-1 (CPT1) regulates long-chain fatty acid (LCFA) entry into mitochondria, where the LCFAs undergo beta-oxidation. To investigate the mechanism(s) by which central metabolism of lipids can modulate energy balance, we selectively reduced lipid oxidation in the hypothalamus. We decreased the activity of CPT1 by administering to rats a ribozyme-containing plasmid designed specifically to decrease the expression of this enzyme or by infusing pharmacological inhibitors of its activity into the third cerebral ventricle. Either genetic or biochemical inhibition of hypothalamic CPT1 activity was sufficient to substantially diminish food intake and endogenous glucose production. These results indicated that changes in the rate of lipid oxidation in selective hypothalamic neurons signaled nutrient availability to the hypothalamus, which in turn modulated the exogenous and endogenous inputs of nutrients into the circulation.     

Role of insulin and glucagon in oxytocin effects on glucose metabolism.

Oxytocin (OT) infusion in normal dogs increases plasma insulin and glucagon levels and increases rates of glucose production and uptake. The purpose of this study was to determine whether the effects of OT on glucose metabolism were direct or indirect. The studies were carried out in normal, unanesthetized dogs in which OT infusion was superimposed on infusion of either somatostatin, which suppresses insulin and glucagon secretion, or clonidine, which suppresses insulin secretion only. Infusion of 0.2 microgram/kg/min of somatostatin suppressed basal levels of plasma insulin and glucagon and inhibited the OT-induced rise of these hormones by about 60-80% of that seen with OT alone. The rates of glucose production and uptake by tissues, measured with [6-3H] glucose, were significantly lower than those seen with OT alone, and the rise in glucose clearance was completely inhibited. Clonidine (30 micrograms/kg, sc), given along with an insulin infusion to replace basal levels of insulin, completely prevented the OT-induced rise in plasma insulin and markedly reduced the glucose uptake seen with OT alone, but did not reduce the usual increase in plasma glucose and glucagon levels or glucose production. To determine whether the OT-induced rise in plasma insulin was in response to the concomitant increase in plasma glucose, similar plasma glucose levels were established in normal dogs by a continuous infusion of glucose and an OT infusion was superimposed. OT did not raise plasma glucose levels further, but plasma insulin levels were increased, indicating that OT can stimulate insulin secretion independently of the plasma glucose changes. Studies by others have shown that the addition of OT to pancreatic islets or intact pancreas can stimulate insulin and glucagon secretion, indicating a direct effect. Our studies agree with that and suggest that in vivo, OT raises plasma insulin levels, at least in part, through a direct action on the pancreas. These studies also show that OT increases glucose production by increasing glucagon secretion and, in addition, a direct effect of OT on glucose production is likely. The OT-induced increase in glucose uptake is mediated largely by increased insulin secretion.     

Use of an artificial beta cell in surgery.

Using a Biostator glucose-controlled insulin infusion system to monitor blood glucose during surgery, we have shown that both nondiabetic and diabetic patients have a tendency towards hyperglycemia during surgery. This appears to be due to suppression of endogenous insulin secretion as measured by serum C-peptide levels. Some diabetic patients maintained relatively normal glucose values during surgery when infused with saline alone and not given glucose or insulin, but 2 of 5 were not well controlled by this means. Hyperglycemia in both diabetic and non-diabetic patients was related to the rate of infusion of exogenous glucose. The biostator glucose-controlled insulin infusion system could be used in feedback mode as an apparently safe and effective means of controlling blood glucose during surgery on diabetic patients.     

Glycaemic effects of incretins in Type 1 diabetes mellitus: a concise review, with emphasis on studies in humans

The remission phase of Type 1 diabetes mellitus is associated with substantial recovery of beta-cell function and with marked improvement of endogenous insulin responses to meals in the early months after diagnosis, accompanied by little or no improvement in the insulin response to parenteral glucose, suggesting that the incretin function may be important in glycaemic regulation in this phase of diabetes. Preservation of the insulin response to parenteral glucagon-like peptide-1 (GLP-1), contrasting with lack of stimulation of insulin secretion by the other known incretin gastric inhibitory polypeptide (GIP), prompted studies with exogenous GLP-1 in recent-onset Type 1 diabetes. These studies showed substantial reduction of glycaemic excursions after ingestion of mixed nutrients during intravenous infusion of GLP-1 without administration of insulin, in subjects with a range of endogenous secretion of insulin in response to meals as demonstrated by blood levels of the insulin-connecting peptide (CP). These effects were independent of stimulation of blood levels of CP and were reproduced in volunteers with no endogenous release of CP in response to meals. The glycaemic effects were associated with inhibition of abnormal rises of blood levels of glucagon, and with suppression of endogenous release of human pancreatic polypeptide (HPP), by GLP-1. It was hypothesized that a major component of the glycaemic effect is attributable to the known action of GLP-1 to inhibit gastric emptying and to inhibit glucagon secretion. Studies of the effects of GLP-1 agonists (GLP-1 and exendin-4) given together with established insulin doses before a meal supported the hypothesis. The more prolonged actions of exendin-4 were accompanied by greater and more prolonged reduction of glycaemic effects of ingestion of meals in volunteers with CP-negative Type 1 diabetes mellitus, during intensive insulin therapy, in whom delay of gastric emptying was confirmed by studies of blood levels of acetaminophen ingested with the meals. Side effect-free doses of exendin-4 given together with insulin in volunteers with CP-negative Type 1 diabetes receiving continuing intensive insulin therapy demonstrated the capacity of this combination therapy to normalize blood glucose levels after ingestion of meals that were consistent with the dietary program of the volunteers, without apparent increased risk of hypoglycaemia within a normal between-meals interval. It is suggested that further and more prolonged studies of the use of long-acting GLP-1 agonists as congeners with insulin in Type 1 diabetes mellitus are indicated.     

Glucose, insulin, and leptin signaling pathways modulate nitric oxide synthesis in glucose-inhibited neurons in the ventromedial hypothalamus

Glucose-sensing neurons in the ventromedial hypothalamus (VMH) are involved in the regulation of glucose homeostasis. Glucose-sensing neurons alter their action potential frequency in response to physiological changes in extracellular glucose, insulin, and leptin. Glucose-excited neurons decrease, whereas glucose-inhibited (GI) neurons increase, their action potential frequency when extracellular glucose is reduced. Central nitric oxide (NO) synthesis is regulated by changes in local fuel availability, as well as insulin and leptin. NO is involved in the regulation of food intake and is altered in obesity and diabetes. Thus this study tests the hypothesis that NO synthesis is a site of convergence for glucose, leptin, and insulin signaling in VMH glucose-sensing neurons. With the use of the NO-sensitive dye 4-amino-5-methylamino-2',7'-difluorofluorescein in conjunction with the membrane potential-sensitive dye fluorometric imaging plate reader, we found that glucose and leptin suppress, whereas insulin stimulates neuronal nitric oxide synthase (nNOS)-dependent NO production in cultured VMH GI neurons. The effects of glucose and leptin were mediated by suppression of AMP-activated protein kinase (AMPK). The AMPK activator 5-aminoimidazole-4-carboxamide-1-beta-4-ribofuranoside (AICAR) increased both NO production and neuronal activity in GI neurons. In contrast, the effects of insulin on NO production were blocked by the phosphoinositide 3-kinase inhibitors wortmannin and LY-294002. Furthermore, decreased glucose, insulin, and AICAR increase the phosphorylation of VMH nNOS, whereas leptin decreases it. Finally, VMH neurons express soluble guanylyl cyclase, a downstream mediator of NO signaling. Thus NO may mediate, in part, glucose, leptin, and insulin signaling in VMH glucose-sensing neurons.     

Suppression of gastric acid secretion by intracisternal bombesin does not require the ventromedial hypothalamus

Intracisternal administration of the tetradecapeptide peptide bombesin suppresses gastric acid release. Other studies have shown that the ventromedial hypothalamus (VMH) may have an inhibitory role in gastric regulation. To determine if the inhibition of gastric acid secretion by intracisternally administered bombesin is mediated by the ventromedial hypothalamus, bombesin was injected intracisternally in rats with ventromedial hypothalamic lesions. Neither anterior nor posterior VMH lesions altered the effects of bombesin on gastric acid, concentration, volume, total output, or on serum gastrin. The bombesin-induced rise in gastric pH was very mildly attenuated by both lesions. The previous finding of enhanced gastric acid secretion after anterior VMH lesions was confirmed. The results suggest that the VMH is not crucial in the bombesin-induced inhibition of acid secretion.     

The significance of hyperphagia and diet composition on the metabolism in ventromedial hypothalamic lesioned male rats

The metabolic consequences of ventromedial hypothalamic lesion were studied in a group of aged male rats which were obese and had decreased response to insulin. The effects of hyperphagia and ventromedial hypothalamic lesion per se were separated by comparing experimental animals fed isocalorically with controls and animals fed ad libitum. Ventromedial hypothalamic lesion as such led to increases in the glucose conversion to fatty acid and in lipoprotein lipase activity in adipose tissue. Protein catabolism as reflected by plasma urea levels, was enhanced. The lipoprotein lipase activity in heart tended to be lower after VMH lesion. These metabolic changes were amplified in the VMH lesioned rats fed ad libitum. The liver glycogen content was lowered by VMH lesion, but this effect was abolished by hyperphagia. In parallel experiments the influence of diet composition was studied by feeding similar groups with diet of high fat content. The glucose incorporation in fatty acids was in all groups markedly and similarly inhibited by the high fat diet. The increase in lipoprotein lipase activity in heart and adipose tissue of control rats with high fat intake could not be demonstrated in any of the groups with ventromedial hypothalamic lesion. The plasma urea level in the control group was not affected by the diet, but tended to increase in the ventromedial hypothalamic lesioned groups on high fat intake. These findings demonstrate that the well known metabolic effects of ventromedial hypothalamic lesions are also manifest in obese insulin resistant male rats. Furthermore, the responses to changes in diet composition are different from those of the control rats.     

Indomethacin does not affect endogenous glucose production in type 2 diabetes mellitus.

In healthy subjects, basal endogenous glucose production is partly regulated by paracrine intrahepatic factors. It is currently unknown whether paracrine intrahepatic factors also influence the increased basal endogenous glucose production in patients with type 2 diabetes mellitus. Administration of indomethacin to patients with type 2 diabetes mellitus stimulates endogenous glucose production and inhibits insulin secretion. Our aim was to evaluate whether this stimulatory effect on glucose production is solely attributable to inhibition of insulin secretion. In order to do this, we administered indomethacin to 5 patients with type 2 diabetes during continuous infusion of somatostatin to block endogenous insulin and glucagon secretion and infusion of basal concentrations of insulin and glucagon in a placebo-controlled study. Endogenous glucose production was measured 3 hours after the start of the somatostatin, insulin and glucagon infusion, for 4 hours after administration of placebo/indomethacin, by primed, continuous infusion of [6,6-(2)H(2)] glucose. At the time of administration of placebo or indomethacin, there were no significant differences in plasma glucose concentrations and endogenous glucose production rates between the two experiments (16.4 +/- 2.09 mmol/l vs. 16.6 +/- 1.34 mmol/l and 17.7 +/- 1.05 micromol/kg/min and 17.0 +/- 1.06 micromol/kg/min), control vs. indomethacin). Plasma glucose concentration did not change significantly in the four hours after indomethacin or placebo administration. Endogenous glucose production in both experiments was similar after both placebo and indomethacin. Mean plasma C-peptide concentrations were all below the detection limit of the assay, reflecting adequate suppression of endogenous insulin secretion by somatostatin. There were no differences in plasma concentrations of insulin (76 +/- 5 vs. 74 +/- 4 pmol/l) and glucagon (69 +/- 8 vs. 71 +/- 6 ng/l) between the studies with levels remaining unchanged in both experiments. Plasma concentrations of cortisol, epinephrine, and norepinephrine were similar in the two studies and did not change significantly. We conclude that indomethacin stimulates endogenous glucose production in patients with type 2 diabetes mellitus by inhibition of insulin secretion.