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...

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...

Leptin decreases visceral fat (VF) and increases peripheral and hepatic insulin action. Here, we generated similar decreases in VF using leptin (Lep), β3-adrenoreceptor agonism (β3), or food restriction (FR) and asked whether insulin action would be equally improved. For 8 days before the in vivo study, Sprague-Dawley rats ( n = 24) were either fed ad libitum [control (Con)], treated with Lep or β3 (CL-316,243) by implanted osmotic mini-pumps, or treated with FR. Total VF was similarly decreased in the latter three groups (Lep, 3.11 ± 0.96 g; β3, 2.87 ± 0.48 g; and FR, 3.54 ± 0.77 g compared with 6.91 ± 1.41 g in Con; P < 0.001) independent of total fat mass (by3H2O) and food intake. Insulin (3 mU ⋅ kg−1 ⋅ min−1) clamp studies were performed to assess hepatic and peripheral insulin sensitivity. Decreased VF resulted in similar and marked improvements in insulin action on glucose production (GP) (Lep, 1.19 ± 0.51; β3, 1.46 ± 0.68; FR, 2.27 ±0.71 compared with 6.06 ± 0.70 mg ⋅ kg−1 ⋅ min−1in Con; P < 0.001). By contrast, reduction in VF by β3 and FR failed to reproduce the stimulation of insulin-mediated glucose uptake (∼60%), glycogen synthesis (∼80%), and glycolysis (∼25%) observed with Lep. We conclude that 1) a moderate decrease in VF uniformly leads to a marked increase in hepatic insulin action, but 2) the effects of leptin on peripheral insulin action are not due to the associated changes in VF or β3 activation.

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