The hypothalamus and other regions within the central nervous system (CNS) link the sensing of nutrients to the control of metabolism and feeding behavior. Here, we report that intracerebroventricular (ICV) administration of the long-chain fatty acid oleic acid markedly inhibits glucose production and food intake. The anorectic effect of oleic acid was independent of leptin and was accompanied by a decrease in the hypothalamic expression of neuropeptide Y. The short-chain fatty acid octanoic acid failed to reproduce the metabolic effects of oleic acid, and ICV coadministration of inhibitors of ATP-sensitive K ؉ channels blunted the effect of oleic acid on glucose production. This is the first demonstration that fatty acids can signal nutrient availability to the CNS, which in turn limits further delivery of nutrients to the circulation. Diabetes 51:271-275, 2002 E xcessive consumption of nutrients with high caloric density is largely responsible for the epidemic of obesity and type 2 diabetes in the western hemisphere and in developing nations (1,2). The association of nutrient excess and obesity with type 2 diabetes is largely mediated via their negative impact on intermediary metabolism and insulin action (1-3). Control of metabolism and food intake in response to nutrients occurs partly at the level of hypothalamic nuclei (3-5). In this regard, macronutrients, such as carbohydrates and lipids, regulate the circulating levels of leptin and insulin (3,6,7), which in turn modulate appetite, energy expenditure, and intermediary metabolism mainly via their hypothalamic receptors (Fig. 1A) (3-5). However, central nervous system (CNS) neurons may also sense nutrients directly via metabolic signaling (8). In this regard, the potential role of CNS lipid metabolism in the control of appetite is supported by the potent anorectic property of fatty acid synthase inhibitors (9,10). Further- FIG. 1. Rapid effects of ICV oleic acid on plasma insulin and glucose levels.A: Hypothesis on the negative feedback mechanisms regulating circulating nutrients. There are two main sources of circulating nutrients: intake and absorption of food (Food) and hepatic production of glucose and lipids (Liver). Increased levels of plasma glucose and lipids regulate hypothalamic efferent pathways via their stimulatory effects on insulin and leptin biosynthesis and secretion. The activation of these central responses leads to decreases in food intake and in hepatic output of glucose and lipids. It is proposed herein that circulating nutrients may also directly signal the nutritional status to the hypothalamus and may therefore play a role in this feedback system. B: Schematic representation of the experimental procedures. Surgical implantation of ICV cannulae was performed on day 1 (ϳ3 weeks before the in vivo study). Full recovery of body weight and food intake was achieved by day 7. Surgical implantation of IV catheters was performed on day 14. Finally, on day 21, ICV infusions were started and blood chemistries, food intake, and/or insulin ac...

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.

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.

IntroductionInsulin resistance is regarded as the main link between obesity and type 2 diabetes mellitus (1, 2). While multiple hypotheses have been proposed to explain this association, recent discoveries on the regulation of feeding behavior and energy expenditure (3-6) have refocused attention on the potential role of hypothalamic centers in the regulation of both energy homeostasis and insulin action (Figure 1) (7-13). In fact, leptin has potent effects on glucose tolerance and insulin action, which appear to be independent of its modulation of feeding behavior (7,9,13). Leptin exerts its actions on food intake and weight gain partly by activation of the melanocortin pathway in the hypothalamus and in other areas within the central nervous system (14). The pivotal role of melanocortinergic neurons in the regulation of energy balance is supported by solid genetic (15, 16) and pharmacologic (17) evidence.To examine whether the activity of the melanocortin pathway in the hypothalamus modulates insulin action in vivo we used the natural agonist, α-melanocyte-stimulating hormone (α-MSH), and a high-affinity antagonist (SHU9119) of the neural melanocortin receptors type 3 and 4 (MCR3, MCR4) (17) to generate bidirectional variations in the activity of this pathway. Body composition and metabolic parameters were assessed using tracer dilution techniques and pancreatic-insulin clamp studies in conscious rats. MethodsExperimental procedures. Thirty-nine male Sprague-Dawley rats (Charles River Laboratories, Wilmington, Massachusetts, USA) were studied ( Figure 2). Rats were housed in individual cages and subjected to a standard light-dark (0600 to 1800 hours/1800 to 0600 hours) cycle. Three weeks before the in vivo study, rats (at ∼10 weeks of age) were equipped with chronic catheters placed in the third cerebral ventricle (18). Rats were anesthetized with intraperitoneal ketamine (Ketaset; 87 mg/kg) and xylazine (Rompun; 11 mg/kg) and fixed in a stereotaxic apparatus with ear bars and a nose piece set at +5.0 mm. A 26-gauge stainless steel guide cannula (Plastics One Inc., Roanoke, Virginia, USA) was chronically implanted into the third ventricle using the following coordinates from bregma: anterior-posterior; +0.2 mm, dorsal-ventral; -9.0 mm, medial-lateral; 0.0 directly on the midsagittal suture. A 28-gauge dummy cannula was inserted to prevent clogging of the guide cannula. The implant is secured to the skull with Caulk Grip dental cement, and the skin is closed over the implant using wound clips. One week before the study, rats (at ∼12 weeks of age) were anesthetized with an intraperitoneal injection of pentobarbital (50 mg/kg body weight), and indwelling catheters were inserted in the right internal jugular vein and in the left carotid artery (18-21). The venous catheter was extended to the level of the right atrium, and the arterial catheter was advanced to the level of the aortic arch. All studies were performed in awake, unstressed, chronically catheterized rats. Histological verification of the intracerebrove...

). Here we examine whether intracerebroventricular (ICV) leptin administration regulates peripheral and hepatic insulin action. Recombinant mouse leptin (n ‫؍‬ 14; 0.02 or 1 g/kg⅐h) or vehicle (n ‫؍‬ 9) were administered ICV for 6 h to conscious rats, and insulin action was determined by insulin (3 milliunits/kg⅐min) clamp and tracer dilution techniques. During physiologic hyperinsulinemia (ϳ65 microunits/ml), the rates of glucose uptake (R d , 20.1 ؎ 0.6 and 23.1 ؎ 0.7 versus 21.7 ؎ 0.6 mg/kg⅐min; p ‫؍‬ NS), glycolysis and glycogen synthesis were similar in rats receiving low-and high-dose leptin versus vehicle. ICV leptin resulted in a 2-3-fold increase in hepatic phosphoenolpyruvate carboxykinase mRNA levels. Glycogenolysis and PEP-gluconeogenesis (2.1 ؎ 0.3 mg/ kg⅐min) contributed similarly to endogenous glucose production (GP) in the vehicle-infused group. However, gluconeogenesis accounted for ϳ80% of GP in both groups receiving ICV leptin, while hepatic glycogenolysis was markedly suppressed (0.7 ؎ 0.3 and 1.2 ؎ 0.3 versus 2.2 ؎ 0.4 mg/kg⅐min, in rats receiving low-and high-dose leptin versus vehicle, respectively; p < 0.01). In summary, short-term ICV leptin administration: 1) failed to affect peripheral insulin action, but 2) induced a striking re-distribution of intrahepatic glucose fluxes. The latter effect largely reproduced that of leptin given systemically at much higher doses. Thus, the regulation of hepatic glucose fluxes by leptin is largely mediated via its central receptors.Leptin, the protein encoded by the ob gene, is an anorectic hormone secreted by adipose cells (1-6). The effects of leptin on food intake are reproduced by its injection directly in the central nervous system, thus suggesting a prominent role of the hypothalamic receptors in mediating its actions (7-12). In the long term, circulating leptin levels correlate with adiposity (3,8,10,13,14) and leptin has been suggested to function as a "biochemical messenger" between fat depots and the hypothalamus (7,10,11,14,15).Nutritional and hormonal factors can also regulate ob gene expression in adipose cells and leptin levels in plasma (3,13,14,16,17). Most important, it has become increasingly evident that leptin plays in turn an important role in the regulation of carbohydrate and lipid metabolism (9, 14, 18 -24). We have recently demonstrated that marked and acute elevations in the plasma leptin concentrations modulate the hepatic gene expression of the gluconeogenic enzyme phosphoenolpyruvate carboxykinase (PEPCK) 1 and the rate of gluconeogenesis (22). It is presently unknown whether the latter metabolic effects of leptin are, at least in part, mediated through its action on hypothalamic receptors.Therefore the primary aim of this study was to examine the metabolic impact of intracerebroventricular (ICV) infusions of leptin on peripheral and hepatic glucose metabolism under basal conditions and in response to physiologic hyperinsulinemia. Furthermore, as recent studies have suggested that a highly selective ␤ 3 -adrenergic recep...

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.

IntroductionInsulin resistance is regarded as the main link between obesity and type 2 diabetes mellitus (1, 2). While multiple hypotheses have been proposed to explain this association, recent discoveries on the regulation of feeding behavior and energy expenditure (3-6) have refocused attention on the potential role of hypothalamic centers in the regulation of both energy homeostasis and insulin action (Figure 1) (7-13). In fact, leptin has potent effects on glucose tolerance and insulin action, which appear to be independent of its modulation of feeding behavior (7,9,13). Leptin exerts its actions on food intake and weight gain partly by activation of the melanocortin pathway in the hypothalamus and in other areas within the central nervous system (14). The pivotal role of melanocortinergic neurons in the regulation of energy balance is supported by solid genetic (15, 16) and pharmacologic (17) evidence.To examine whether the activity of the melanocortin pathway in the hypothalamus modulates insulin action in vivo we used the natural agonist, α-melanocyte-stimulating hormone (α-MSH), and a high-affinity antagonist (SHU9119) of the neural melanocortin receptors type 3 and 4 (MCR3, MCR4) (17) to generate bidirectional variations in the activity of this pathway. Body composition and metabolic parameters were assessed using tracer dilution techniques and pancreatic-insulin clamp studies in conscious rats. MethodsExperimental procedures. Thirty-nine male Sprague-Dawley rats (Charles River Laboratories, Wilmington, Massachusetts, USA) were studied ( Figure 2). Rats were housed in individual cages and subjected to a standard light-dark (0600 to 1800 hours/1800 to 0600 hours) cycle. Three weeks before the in vivo study, rats (at ∼10 weeks of age) were equipped with chronic catheters placed in the third cerebral ventricle (18). Rats were anesthetized with intraperitoneal ketamine (Ketaset; 87 mg/kg) and xylazine (Rompun; 11 mg/kg) and fixed in a stereotaxic apparatus with ear bars and a nose piece set at +5.0 mm. A 26-gauge stainless steel guide cannula (Plastics One Inc., Roanoke, Virginia, USA) was chronically implanted into the third ventricle using the following coordinates from bregma: anterior-posterior; +0.2 mm, dorsal-ventral; -9.0 mm, medial-lateral; 0.0 directly on the midsagittal suture. A 28-gauge dummy cannula was inserted to prevent clogging of the guide cannula. The implant is secured to the skull with Caulk Grip dental cement, and the skin is closed over the implant using wound clips. One week before the study, rats (at ∼12 weeks of age) were anesthetized with an intraperitoneal injection of pentobarbital (50 mg/kg body weight), and indwelling catheters were inserted in the right internal jugular vein and in the left carotid artery (18-21). The venous catheter was extended to the level of the right atrium, and the arterial catheter was advanced to the level of the aortic arch. All studies were performed in awake, unstressed, chronically catheterized rats. Histological verification of the intracerebrove...

Estradiol treatment for 48 h increases the density of alpha 1B-adrenoceptors in the hypothalamus-preoptic area of ovariectomized female rats by five- to six-fold. Present studies tested the hypothesis that estradiol elevation of hypothalamus-preoptic area alpha 1B-adrenoceptor density is correlated with increased levels of mRNA for this receptor. We developed a semiquantitative reverse transcriptase-polymerase chain reaction (RT-PCR) protocol for measuring brain alpha 1b-adrenoceptor mRNA. The primers chosen yielded the predicted 409 base pair PCR product when used to amplify authentic alpha 1b-adrenoceptor cDNA. The identity of the RT-PCR products from rat brain was confirmed by restriction digest analysis and sequencing. Moreover, there was a good correlation between the levels of alpha 1b-adrenoceptor mRNA measured by RT-PCR in liver, whole brain and cerebellum with previous measurements using Northern blots and RNAse protection assays. We then performed RT-PCR on total RNA from hypothalamic-preoptic area tissue taken from ovariectomized control rats and from ovariectomized rats injected once or twice with 2 micrograms of estradiol benzoate at 24 or 24 and 48 h before sacrifice. Exposure to estradiol for either 24 or 48 h significantly increased levels of alpha 1b-adrenoceptor mRNA by 86-110% in the hypothalamus-preoptic area of ovariectomized female rats when compared to oil-treated controls. We also examined whether estradiol regulates alpha 1b-adrenoceptor mRNA in the cortex. Cortical alpha 1b-adrenoceptor mRNA levels were reduced to approximately 20% of control levels when measured 24 h after hormone injection. A similar decrease in cortical alpha 1b-adrenoceptor mRNA was observed 48 h after estrogen administration. In summary, estradiol treatment significantly increases the level of alpha 1b-adrenoceptor mRNA in the hypothalamus-preoptic area, a brain region involved in the control of reproductive function. In the cortex, a brain region with relatively few estrogen receptors, the same estrogen treatment reduces alpha 1b-adrenoceptor mRNA levels.