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In the present study, we examined whether chronic intracerebroventricular (ICV) leptin administration protects against ischemia/reperfusion (I/R)-induced acute kidney injury (AKI). Twelve-week-old male rats were implanted with an ICV cannula into the right lateral ventricle, and 8-10 days after surgery, leptin (0.021 µg/h, n = 8) or saline vehicle (0.5 µL/h, n = 8) was infused via osmotic minipump connected to the ICV cannula for 12 days. On day 8 of leptin or vehicle infusion, rats were submitted to unilateral ischemia/reperfusion (UIR) by clamping the left pedicle for 30 min. To control for leptin-induced reductions in food intake, the vehicle-treated group was pair-fed (UIR-PF) to match the same amount of food consumed by leptin-treated (UIR-Leptin) rats. On the 12th day of leptin or vehicle infusion (fourth day after AKI), single-left kidney glomerular filtration rate (GFR) was measured, blood samples were collected to quantify white blood cells, and kidneys were collected for histological assessment of injury. UIR-Leptin-treated rats showed reduced right and left kidney weights (right: 1,040 ± 24 vs. 1,281 ± 36 mg; left: 1,127 ± 71 vs. 1,707 ± 45 mg, for UIR-Leptin and UIR-PF, respectively). ICV leptin infusion improved GFR (0.50 ± 0.06 vs. 0.13 ± 0.03 mL/min/g kidney wt) and reduced kidney injury scores. ICV leptin treatment also attenuated the reduction in circulating adiponectin levels that was observed in UIR-PF rats and increased the circulating white blood cells count compared with UIR-PF rats (16.3 ± 1.3 vs. 9.8 ± 0.6 k/µL). Therefore, we show that leptin, via its actions on the central nervous system, confers significant protection against major kidney dysfunction and injury in a model of ischemia/reperfusion-induced AKI. NEW & NOTEWORTHY A major new finding of this study is that chronic activation of leptin receptors in the CNS markedly attenuates acute kidney injury and protects against severe renal dysfunction after ischemia/reperfusion, independently of leptin's anorexic effects.

This experiment investigated which hypothalamic nuclei were activated by a dose of leptin that inhibited food intake. Foodnot intake, energy expenditure, respiratory exchange ratio (RER), and intrascapular brown adipose tissue (IBAT) temperature were measured in male and female Sprague Dawley rats for 36 h following an intraperitoneal injection of 0, 50, 200, 500, or 1,000 µg leptin/kg with each rat tested with each dose of leptin in random order. In both males and females, RER and 12-h food intake were inhibited only by 1,000 µg leptin/kg, but there was no effect on energy expenditure or IBAT temperature. At the end of the experiment, phosphorylated signal transducer and activator of transcription 3 (pSTAT3) immunoreactivity was measured 1 h after injection of 0, 50, 500, or 1,000 µg leptin/kg. In male rats, the lowest dose of leptin produced a maximal activation of STAT3 in the Arc and nucleus of the solitary tract (NTS). There was no response in the dorsomedial hypothalamus, but there was a progressive increase in ventromedial nucleus of the hypothalamus (VMH) pSTAT3 with increasing doses of leptin. In female rats, there was no significant change in Arc and pSTAT3 NTS activation was maximal with 500 mg leptin/kg, but only the highest dose of leptin increased VMH pSTAT3. These results suggest that the VMH plays an important role in the energetic response to elevations of circulating leptin but do not exclude the possibility that multiple nuclei provide the appropriate integrated response to hyperleptinemia. NEW & NOTEWORTHY The results of this experiment show that doses of leptin too small to inhibit food intake produce a maximal response to leptin in the arcuate nucleus. By contrast the VMH shows a robust response that correlates with inhibition of food intake. This suggests that the VMH plays an important role in the energetic response to hyperleptinemia.

Previous studies using male rodents showed the adipocyte-derived hormone leptin acts in the brain to regulate cardiovascular function, energy balance, and glucose homeostasis. The importance of sex differences in cardiometabolic responses to leptin, however, is still unclear. We examined potential sex differences in leptin's chronic central nervous system (CNS)-mediated actions on blood pressure (BP), heart rate (HR), appetite, and glucose homeostasis in normal and type 1 diabetic rats. Female and male Sprague-Dawley (SD) rats were instrumented with intracerebroventricular cannulas for continuous 7-day leptin infusion (15 µg/day), and BP and HR were measured by telemetry 24 h/day. At baseline, females had lower mean arterial pressure (MAP) (96 ± 3 vs. 104 ± 4 mmHg, P < 0.05) but higher HR (375 ± 5 vs. 335 ± 5 beats/min, P < 0.05) compared with males. After leptin treatment, we observed similar increases in BP (∼3 mmHg) and HR (∼25 beats/min) in both sexes. Females had significantly lower body weight (BW, 283 ± 2 vs. 417 ± 7 g, P < 0.05) and caloric intake (162 ± 20 vs. 192 ± 9 kcal/kg of body wt, P < 0.05) compared with males, and leptin infusion reduced BW (-10%) and caloric intake (-62%) similarly in both sexes. In rats with streptozotocin-induced diabetes ( n = 5/sex), intracerebroventricular leptin treatment for 7 days completely normalized glucose levels. The same dose of leptin administered intraperitoneally did not alter MAP, HR, glucose levels, or caloric intake in normal or diabetic rats. These results show that leptin's CNS effects on BP, HR, glucose regulation, and energy homeostasis are similar in male and female rats. Therefore, our results provide no evidence for sex differences in leptin's brain-mediated cardiovascular or metabolic actions.

Blood glucose and insulin homeostasis is disrupted during the progression of type 2 diabetes. Insulin levels and action are regulated by both peripheral and central responses that involve the intestine and microbiome. The intestine and its microbiota process nutrients and generate molecules that influence blood glucose and insulin. Peripheral insulin regulation is regulated by gut-segment-dependent nutrient sensing and microbial factors such as short-chain fatty acids and bile acids that engage G-protein-coupled receptors. Innate immune sensing of gut-derived bacterial cell wall components and lipopolysaccharides also alter insulin homeostasis. These bacterial metabolites and postbiotics influence insulin secretion and insulin clearance in part by altering endocrine responses such as glucagon-like peptide-1. Gut-derived bacterial factors can promote inflammation and insulin resistance, but other postbiotics can be insulin sensitizers. In parallel, activation of small intestinal sirtuin 1 increases insulin sensitivity by reversing high fat-induced hypothalamic insulin resistance through a gut-brain neuronal axis, whereas high fat-feeding alters small intestinal microbiome and increases taurochenodeoxycholic acid in the plasma and the dorsal vagal complex to induce insulin resistance. In summary, emerging evidence indicates that intestinal molecular signaling involving nutrient sensing and the host-microbe symbiosis alters insulin homeostasis and action. Gut-derived host endocrine and paracrine factors as well as microbial metabolites act on the liver, pancreas, and the brain, and in parallel on the gut-brain neuronal axis. Understanding common nodes of peripheral and central insulin homeostasis and action may reveal new ways to target the intestinal host-microbe relationship in obesity, metabolic disease, and type 2 diabetes.

Leptin administration into the hindbrain, and specifically the nucleus of the solitary tract, increases phosphorylated signal transducer and activator of transcription 3 (pSTAT3), a marker of leptin receptor activation, in hypothalamic nuclei known to express leptin receptors. The ventromedial nucleus of the hypothalamus (VMH) shows the greatest response, with a threefold increase in pSTAT3. This experiment tested the importance of VMH leptin receptor-expressing neurons in mediating weight loss caused by fourth ventricle (4V) leptin infusion. Male Sprague-Dawley rats received bilateral VMH 75-nL injections of 260 ng/μL of leptin-conjugated saporin (Lep-Sap) or blank-saporin (Blk-Sap). After 23 days they were fitted with 4V infusion cannulas and 1 wk later adapted to housing in a calorimeter before they were infused with 0.9 μg leptin/day for 14 days. There was no effect of VMH Lep-Sap on weight gain or glucose clearance before leptin infusion. Leptin inhibited food intake and respiratory exchange ratio in Blk-Sap but not Lep-Sap rats. Leptin had no effect on energy expenditure or brown adipose tissue temperature of either group. Inguinal and epididymal fat were significantly reduced in leptin-treated Blk-Sap rats, but the response was greatly attenuated in Lep-Sap rats. VMH pSTAT3 was increased in leptin-treated Blk-Sap but not Lep-Sap rats. These results support the concept that leptin-induced weight loss results from an integrated response across different brain areas. They also support previous reports that VMH leptin receptors do not play a significant role in maintaining energy balance in basal conditions but limit weight gain during positive energy balance.

Leptin receptor (LepR) signaling in neurons of the ventromedial nucleus of the hypothalamus (VMH), specifically those expressing steroidogenic factor-1 (SF1), have been proposed to play a key role in controlling energy balance. By crossing LepR-silenced (LepR loxTB ) mice with those expressing SF1-Cre, we unsilenced native LepR specifically in the VMH and tested whether SF1 neurons in the VMH are critical mediators of leptin's effect on energy homeostasis. LepR loxTB × SF1-Cre [knockout (KO)/Tg+] mice were metabolically phenotyped and compared with littermate controls that either expressed or were deficient in LepRs. Leptin-induced phosphorylated STAT3 was present in the VMH of KO/Tg+ mice and absent in other hypothalamic nuclei. VMH leptin signaling did not ameliorate obesity resulting from LepR deficiency in chow-fed mice. There was no change in food intake or energy expenditure when comparing complete LepR-null mice with KO/Tg+ mice, nor did KO/Tg+ mice show improved glucose tolerance. The presence of functional LepRs in the VMH mildly enhanced sensitivity to the pancreatic hormone amylin. When maintained on a high-fat diet (HFD), there was no reduction in diet-induced obesity in KO/Tg+ mice, but KO/Tg+ mice had improved glucose tolerance after 7 wk on an HFD compared with LepR-null mice. We conclude that LepR signaling in the VMH alone is not sufficient to correct metabolic dysfunction observed in LepR-null mice.

We evaluated the contribution of brown adipose tissue (BAT) sympathetic innervation on central leptin-mediated weight loss. In a short- and long-term study, F344BN rats were submitted to either a denervation of interscapular BAT (Denervated) or a sham operation (Sham). Animals from each group received the Ob (Leptin) or green fluorescent protein (GFP; Control) gene through a single injection of recombinant adeno-associated virus delivered centrally. Changes in body weight were recorded for 14 or 35 days, after which adipose tissues and skeletal muscles were weighed. In both studies, hypothalamic phosphorylated STAT3 (P-STAT3) was significantly higher in Sham-Leptin and Denervated-Leptin groups compared with their respective Control groups ( P < 0.01), indicating that leptin signaling was enhanced at the end point. We measured uncoupling protein 1 (UCP1), a marker of BAT thermogenic activity, and found a significant induction in Leptin in Sham animals ( P < 0.001) but not in Denervated animals, demonstrating that BAT UCP1 protein was only induced in Sham rats. Both Sham-Leptin and Denervated-Leptin rats lost ~15% of their initial body weight ( P < 0.001) by day 14 and reached a maximum of 18% body weight loss that stabilized over week 3 of treatment, indicating that sympathetic outflow to BAT is not required for leptin-mediated weight loss. In summary, interscapular BAT (iBAT) denervation did not prevent body weight loss following central leptin gene delivery. The present data show that sympathetic innervation of iBAT is not essential for leptin-induced body weight loss.

We previously reported that low-dose leptin infusions into the third or fourth ventricle that do not affect energy balance when given independently cause rapid weight loss when given simultaneously. Therefore, we tested whether hindbrain leptin enhances the response to forebrain leptin or whether forebrain leptin enhances the response to hindbrain leptin. Rats received fourth-ventricle infusions of saline or 0.01, 0.1, 0.3, or 0.6 μg leptin/day for 13 days. On days 9 and 13 , 0.1 μg leptin was injected into the third ventricle. The injection inhibited food intake for 36 h in saline-infused rats but for 60 h in those infused with 0.6 μg leptin/day. Leptin injection increased intrascapular brown fat temperature in leptin-infused, but not saline-infused, rats. In a separate experiment, rats received third-ventricle infusions of saline or 0.005, 0.01, 0.05, or 0.1 μg leptin/day and fourth-ventricle injections of 1.0 μg leptin on days 9 and 13 Leptin injection inhibited food intake, respiratory exchange ratio, and 14-h food intake in rats infused with saline or the two lowest doses of leptin. There was no effect with higher-dose leptin infusions because food intake, body fat, and lean mass were already inhibited. These data suggest that activation of leptin receptors in the hindbrain enhances the response to third-ventricle leptin, whereas activation of forebrain leptin receptors does not enhance the response to fourth-ventricle leptin, consistent with our previous finding that weight loss in rats treated with fourth-ventricle leptin is associated with indirect activation of hypothalamic STAT3., (Copyright © 2017 the American Physiological Society.)

This study tested whether ganglionic blockade or hepatic vagotomy attenuates the chronic central nervous system (CNS)-mediated antidiabetic and cardiovascular effects of leptin. Male Sprague-Dawley rats were instrumented with telemetry probes and arterial and venous catheters for determination of blood pressure (BP), heart rate (HR), blood sampling, and intravenous (iv) infusions. An intracerebroventricular (ICV) cannula was placed into the brain lateral ventricle for infusion of leptin or vehicle. After control measurements, streptozotocin (STZ) was injected iv (50 mg/kg) to induce diabetes, and 5 days later leptin ( n = 6) or saline vehicle ( n = 5) was infused ICV for 12 days via osmotic pumps. Beginning on day 6 of leptin treatment, the ganglionic blocker hexamethonium (15 mg·kg -1 ·day -1 iv) was infused, while leptin infusion was continued, to assess the role of the autonomic nervous system. Induction of diabetes was associated with increases in blood glucose (98 ± 7 to 350 ± 19 mg/dl), food intake (23 ± 3 to 43 ± 3 g/day), decreases in HR (-70 ± 11 beats/min), polyuria, and increased water consumption, which were all completely normalized by ICV leptin infusion. Although hexamethonium attenuated leptin's effect on HR, it failed to impair leptin's ability to restore euglycemia or to prevent the polyuria or increased water intake in STZ-diabetic rats. We also found that after pretreatment with hexamethonium ( n = 8), ICV leptin infusion, during continued ganglionic blockade, completely normalized blood glucose in diabetic rats. In addition, selective hepatic vagotomy did not attenuate leptin's ability to restore euglycemia in diabetic rats. These results suggest that leptin's powerful chronic CNS antidiabetic actions are mediated primarily via nonautonomic mechanisms., (Copyright © 2017 the American Physiological Society.)

Peripheral coadministration of amylin and leptin produces enhanced suppression of food intake and body weight, but the central nuclei mediating these effects remain unclear. Because each of these peptides controls feeding via actions at the ventral tegmental area (VTA), we tested the hypothesis that the VTA is a site of action for the cooperative effects of leptin and amylin on energy balance control. First, we show that intra-VTA injection of amylin and leptin at doses of each peptide that are effective in reducing food intake and body weight when administered separately produces an enhanced suppression of feeding when administered in combination. We also demonstrate that subthreshold doses of both amylin and leptin cause significant hypophagia and body weight loss when coadministered into the VTA. Additionally, we provide evidence that VTA amylin receptor blockade significantly attenuates the ability of intra-VTA leptin to reduce feeding and body weight gain. Together, these data provide the first evidence that the VTA mediates the interaction of amylin and leptin to cooperatively promote negative energy balance., (Copyright © 2015 the American Physiological Society.)

We previously demonstrated that leptin has powerful central nervous system (CNS)-mediated antidiabetic actions. In this study we tested the importance of melanocortin-4 receptors (MC4Rs) for leptin's ability to suppress food intake, increase blood pressure (BP) and heart rate (HR), and normalize glucose levels in insulin-dependent diabetes. MC4R knockout (MC4R-KO) and control wild-type (WT) rats were implanted with intracerebroventricular (ICV) cannula and BP and HR were measured 24 h/day by telemetry. After 5-day control period, an injection of streptozotocin (50 mg/kg, ip) was used to induce diabetes. Eight days after injection, an osmotic pump was implanted subcutaneously and connected to the ICV cannula to deliver leptin (15 μg/day) for 7 days. At baseline, MC4R-KO rats were hyperphagic and 40% heavier than WT rats. Despite obesity, BP was similar (112 ± 2 vs. 111 ± 2 mmHg) and HR was lower in MC4R-KO rats (320 ± 6 vs. 347 ± 5 beats/min). Induction of diabetes increased food intake (30%) and reduced BP (∼17 mmHg) and HR (∼61 beats/min) in WT rats, while food intake, BP, and HR were reduced by ∼10%, 7 mmHg, and 33 beats/min, respectively, in MC4R-KO rats. Leptin treatment normalized blood glucose (437 ± 10 to 136 ± 18 mg/dl), reduced food intake (40%), and increased HR (+60 beats/min) and BP (+9 mmHg) in WT rats. Only modest changes in blood glucose (367 ± 16 to 326 ± 23 mg/dl), food intake (5%), HR (+16 beats/min) and BP (+4 mmHg) were observed in MC4R-KO rats. These results indicate that intact CNS MC4R signaling is necessary for leptin to exert its chronic antidiabetic, anorexic, and cardiovascular actions., (Copyright © 2015 the American Physiological Society.)

Leptin receptors (ObRs) in the forebrain and hindbrain have been independently recognized as important mediators of leptin responses. We recently used low-dose leptin infusions to show that chronic activation of both hypothalamic and hindbrain ObRs is required to reduce body fat. The objective of the present study was to identify the brain nuclei that are selectively activated in rats that received chronic infusion of leptin in both the forebrain and hindbrain. Either saline or leptin was infused into third and fourth ventricles (0.1 μg/24 h in the third ventricle and 0.6 μg/24 h in the fourth ventricle) of male Sprague-Dawley rats for 6 days using Alzet pumps. Rats infused with leptin into both ventricles (LL rats) showed a significant increase in phosphorylated (p)STAT3 immunoreactivity in the arcuate nucleus, ventromedial hypothalamus, dorsomedial hypothalamus, and posterior hypothalamus compared with other groups. No differences in pSTAT3 immunoreactivity were observed in midbrain or hindbrain nuclei despite a sixfold higher infusion of leptin into the fourth ventricle than the third ventricle. ΔFosB immunoreactivity, a marker of chronic neuronal activation, showed that multiple brain nuclei were chronically activated due to the process of infusion, but only the arcuate nucleus, ventromedial hypothalamus, dorsomedial hypothalamus, and ventral tuberomamillary nucleus showed a significant increase in LL rats compared with other groups. These data demonstrate that low-dose leptin in the hindbrain increases pSTAT3 in areas of the hypothalamus known to respond to leptin, supporting the hypothesis that leptin-induced weight loss requires an integrated response from both the hindbrain and forebrain., (Copyright © 2015 the American Physiological Society.)

Previous studies showed that Src homology-2 tyrosine phosphatase (Shp2) is an important regulator of body weight. In this study, we examined the impact of Shp2 deficiency specifically in proopiomelanocortin (POMC) neurons on metabolic and cardiovascular function and on chronic blood pressure (BP) and metabolic responses to leptin. Mice with Shp2 deleted in POMC neurons (Shp2/Pomc-cre) and control mice (Shp2(flox/flox)) were implanted with telemetry probes and venous catheters for measurement of mean arterial pressure (MAP) and leptin infusion. After at least 5 days of stable control measurements, mice received leptin infusion (2 μg·kg(-1)·day(-1) iv) for 7 days. Compared with Shp2(flox/flox) controls, Shp2/Pomc-cre mice at 22 wk of age were slightly heavier (34 ± 1 vs. 31 ± 1 g) but consumed a similar amount of food (3.9 ± 0.3 vs. 3.8 ± 0.2 g/day). Leptin infusion reduced food intake in Shp2(flox/flox) mice (2.6 ± 0.5 g) and Shp2/Pomc-cre mice (3.2 ± 0.3 g). Despite decreasing food intake, leptin infusion increased MAP in control mice, whereas no significant change in MAP was observed in Shp2/Pomc-cre mice. Leptin infusion also decreased plasma glucose and insulin levels in controls (12 ± 1 to 6 ± 1 μU/ml and 142 ± 12 to 81 ± 8 mg/100 ml) but not in Shp2/Pomc-cre mice. Leptin increased V̇o2 by 16 ± 2% in controls and 7 ± 1% in Shp2/Pomc-cre mice. These results indicate that Shp2 signaling in POMC neurons contributes to the long-term BP and antidiabetic actions of leptin and may play a modest role in normal regulation of body weight., (Copyright © 2014 the American Physiological Society.)

Pontine parabrachial nucleus (PBN) neurons integrate visceral, oral, and other sensory information, playing an integral role in the neural control of feeding. Current experiments probed whether lateral PBN (lPBN) leptin receptor (LepRb) signaling contributes to this function. Intra-lPBN leptin microinjection significantly reduced cumulative chow intake, average meal size, and body weight in rats, independent of effects on locomotor activity or gastric emptying. In contrast to the effects observed following LepRb activation in other nuclei, lPBN LepRb stimulation did not affect progressive ratio responding for sucrose reward or conditioned place preference for a palatable food. Collectively, results suggest that lPBN LepRb activation reduces food intake by modulating the neural processing of meal size/satiation signaling, and highlight the lPBN as a novel site of action for leptin-mediated food intake control., (Copyright © 2014 the American Physiological Society.)

Leptin may reduce pancreatic lipid deposition, which increases with progression of obesity and can impair β-cell function. The insulinotropic effect of glucagon-like peptide-1 (GLP-1) and the efficacy of GLP-1 receptor agonist are reduced associated with impaired β-cell function. In this study, we examined whether leptin could restore the efficacy of exenatide, a GLP-1 receptor agonist, in type 2 diabetes with increased adiposity. We chronically administered leptin (500 μg·kg⁻¹·day⁻¹) and/or exenatide (20 μg·kg⁻¹·day⁻¹) for 2 wk in a mouse model of type 2 diabetes with increased adiposity induced by streptozotocin and high-fat diet (STZ/HFD mice). The STZ/HFD mice exhibited hyperglycemia, overweight, increased pancreatic triglyceride level, and reduced glucose-stimulated insulin secretion (GSIS); moreover, the insulinotropic effect of exenatide was reduced. However, leptin significantly reduced pancreatic triglyceride level, and adding leptin to exenatide (LEP/EX) remarkably enhanced GSIS. These results suggested that the leptin treatment restored the insulinotropic effect of exenatide in the mice. In addition, LEP/EX reduced food intake, body weight, and triglyceride levels in the skeletal muscle and liver, and corrected hyperglycemia to a greater extent than either monotherapy. The pair-feeding experiment indicated that the marked reduction of pancreatic triglyceride level and enhancement of GSIS by LEP/EX occurred via mechanisms other than calorie restriction. These results suggest that leptin treatment may restore the insulinotropic effect of exenatide associated with the reduction of pancreatic lipid deposition in type 2 diabetes with increased adiposity. Combination therapy with leptin and exenatide could be an effective treatment for patients with type 2 diabetes with increased adiposity., (Copyright © 2014 the American Physiological Society.)

Continuous subcutaneous administration of leptin normalizes blood glucose levels in rodent models of Type 1 and Type 2 diabetes independent of changes in food intake, body weight, and plasma insulin. We tested whether an acute intravenous leptin infusion changed blood glucose in normal and diet-induced leptin-resistant rats to determine whether this measure could be used as a marker of leptin sensitivity. Leptin-responsive chow-fed rats and diet-induced leptin-resistant male Sprague-Dawley rats were fitted with thoracic jugular vein catheters. Four days after surgery, conscious rats were infused intravenously with either saline for 32 min, low-dose (LD) leptin (1.9 μg·kg(-1)·min(-1)) followed by high-dose (HD) leptin (3.8 μg·kg(-1)·min(-1)) for 16 min each, or only HD leptin for 16 min. There was no change in blood glucose after an acute intravenous infusion of either LD leptin or HD leptin alone for 16 min. An intravenous infusion of LD followed by HD leptin for 16 min each significantly decreased serum glucose in leptin-responsive rats but not in leptin-resistant rats. Leptin infusions increased serum leptin in all rat groups but had no effect on plasma glucagon or 12-h weight gain and energy intake in any group of rats. These results show that leptin has an acute glucose-lowering effect that reflects the leptin responsiveness of the rat. This effect is consistent across controls and different leptin-resistant rat models, and the acute nonlethal test provides a novel method of testing leptin responsiveness in rats., (Copyright © 2014 the American Physiological Society.)

Maternal high-fat (HF) diet has long-term consequences on the metabolic phenotype of the offspring. Here, we determined the effects of postweaning exercise in offspring of rat dams fed HF diet during gestation and lactation. Pregnant Sprague-Dawley rats were maintained on chow or HF diet throughout gestation and lactation. All pups were weaned onto chow diet on postnatal day (PND) 21. At 4 wk of age, male pups were given free access to running wheels (RW) or remained sedentary (SED) for 3 wk, after which all rats remained sedentary, resulting in four groups: CHOW-SED, CHOW-RW, HF-SED, and HF-RW. Male HF offspring gained more body weight by PND7 compared with CHOW pups and maintained this weight difference through the entire experiment. Three weeks of postweaning exercise did not affect body weight gain in either CHOW or HF offspring, but reduced adiposity in HF offspring. Plasma leptin was decreased at the end of the 3-wk running period in HF-RW rats but was not different from HF-SED 9 wk after the exercise period ended. At 14 wk of age, intracerebroventricular injection of leptin suppressed food intake in CHOW-SED, CHOW-RW, and HF-RW, while it did not affect food intake in HF-SED group. At death, HF-RW rats also had higher leptin-induced phospho-STAT3 level in the arcuate nucleus than HF-SED rats. Both maternal HF diet and postweaning exercise had effects on hypothalamic neuropeptide and receptor mRNA expression in adult offspring. Our data suggest that postweaning exercise improves central leptin sensitivity and signaling in this model.

We previously reported that peripheral leptin infusions in chronically decrebrate rats, in which the forebrain is neurally isolated from the hindbrain, increased body fat and decreased energy expenditure. Any central leptin response in decerebrate rats would depend upon the hindbrain. Here, we tested whether selective activation of hindbrain leptin receptors increased body fat. Fourth ventricle infusion of 0.6 μg leptin/day for 12 days increased body fat by 13% with no increase in food intake. Third ventricle leptin infusions decreased food intake, body fat, and lean tissue with a maximal response at 0.3 μg leptin/day. To test whether hindbrain receptors opposed activity of hypothalamic receptors, rats received peripheral infusions of 40 μg leptin/day and increasing 4th ventricle doses of the leptin receptor antagonist mutein protein. Mutein (3.0 μg/day) reduced body fat in PBS-infused rats to the same level as leptin-infused rats and reduced lean tissue in all rats. Leptin, but not mutein, inhibited food intake. By contrast, 3.0 μg/day mutein in the 3rd ventricle increased food intake and body fat in both PBS- and leptin-infused rats. In basal conditions, hindbrain leptin receptors may antagonize activity of forebrain receptors to protect lean and fat tissue, but there is no evidence for an anabolic role for hindbrain receptors when leptin is elevated. In a dietary study, rats increased energy intake when offered lard and 30% sucrose solution in addition to chow. Peripheral leptin infusion exaggerated the gain in body fat without altering energy intake confirming the potential for leptin to increase adiposity.

Circulating levels of leptin are elevated in individuals suffering from chronic intermittent hypoxia (CIH). Systemic and central administration of leptin elicits increases in sympathetic nervous activity (SNA), arterial pressure (AP), and heart rate (HR), and it attenuates the baroreceptor reflex, cardiovascular responses that are similar to those observed during CIH as a result of activation of chemoreceptors by the systemic hypoxia. Therefore, experiments were done in anesthetized Wistar rats to investigate the effects of leptin in nucleus of the solitary tract (NTS) on AP and HR responses, and renal SNA (RSNA) responses during activation of NTS neurons and the chemoreceptor reflex. Microinjection of leptin (5-100 ng; 20 nl) into caudal NTS pressor sites (l-glutamate; l-Glu; 0.25 M; 10 nl) elicited dose-related increases in AP, HR, and RSNA. Leptin microinjections (5 ng; 20 nl) into these sites potentiated the increase in AP and HR elicited by l-Glu. Additionally, bilateral injections of leptin (5 ng; 100 nl) into NTS potentiated the increase in AP and attenuated the bradycardia to systemic activation of the chemoreflex. In the Zucker obese rat, leptin injections into NTS neither elicited cardiovascular responses nor altered the cardiovascular responses to activation of the chemoreflex. Taken together, these data indicate that leptin exerts a modulatory effect on neuronal circuits within NTS that control cardiovascular responses elicited during the reflex activation of arterial chemoreceptors and suggest that increased AP and SNA observed in individuals with CIH may be due, in part, by leptin's effects on the chemoreflex at the level of NTS.

Leptin regulates energy balance through central circuits that control food intake and energy expenditure, including proopiomelanocortin (POMC) neurons. POMC neuron-specific deletion of protein tyrosine phosphatase 1B (PTP1B) (Ptpn1(loxP/loxP) POMC-Cre), a negative regulator of CNS leptin signaling, results in resistance to diet-induced obesity and improved peripheral leptin sensitivity in mice, thus establishing PTP1B as an important component of POMC neuron regulation of energy balance. POMC neurons are expressed in the pituitary, the arcuate nucleus of the hypothalamus (ARH), and the nucleus of the solitary tract (NTS) in the hindbrain, and it is unknown how each population might contribute to the phenotype of POMC-Ptp1b(-/-) mice. It is also unknown whether improved leptin sensitivity in POMC-Ptp1b(-/-) mice involves altered melanocortin receptor signaling. Therefore, we examined the effects of hindbrain administration (4th ventricle) of leptin (1.5, 3, and 6 μg) or the melanocortin 3/4R agonist melanotan II (0.1 and 0.2 nmol) in POMC-Ptp1b(-/-) (KO) and control PTP1B(fl/fl) (WT) mice on food intake, body weight, spontaneous physical activity (SPA), and core temperature (T(C)). The results show that KO mice were hypersensitive to hindbrain leptin- and MTII-induced food intake and body weight suppression and SPA compared with WT mice. Greater increases in leptin- but not MTII-induced T(C) were also observed in KO vs. WT animals. In addition, KO mice displayed elevated hindbrain and hypothalamic MC4R mRNA expression. These studies are the first to show that hindbrain administration of leptin or a melanocortin receptor agonist alters energy balance in mice likely via participation of hindbrain POMC neurons.

Obesity is associated with alterations in upper airway collapsibility during sleep. Obese, leptin-deficient mice demonstrate blunted ventilatory control, leading us to hypothesize that (1) obesity and leptin deficiency would predispose to worsening neuromechanical upper airway function and that (2) leptin replacement would acutely reverse neuromuscular defects in the absence of weight loss. In age-matched, anesthetized, spontaneously breathing C57BL/6J (BL6) and ob(-)/ob(-) mice, we characterized upper airway pressure-flow dynamics during ramp decreases in nasal pressure (P(N)) to determine the passive expiratory critical pressure (P(CRIT)) and active responses to reductions in P(N), including the percentage of ramps showing inspiratory flow limitation (IFL; frequency), the P(N) threshold at which IFL developed, maximum inspiratory airflow (Vi(max)), and genioglossus electromyographic (EMG(GG)) activity. Elevations in body weight were associated with progressive elevations in P(CRIT) (0.1 ± 0.02 cmH(2)O/g), independent of mouse strain. P(CRIT) was also elevated in ob(-)/ob(-) compared with BL6 mice (1.6 ± 0.1 cmH(2)O), independent of weight. Both obesity and leptin deficiency were associated with significantly higher IFL frequency and P(N) threshold and lower VI(max). Very obese ob(-)/ob(-) mice treated with leptin compared with nontreated mice showed a decrease in IFL frequency (from 63.5 ± 2.9 to 30.0 ± 8.6%) and P(N) threshold (from -0.8 ± 1.1 to -5.6 ± 0.8 cmH(2)O) and increase in VI(max) (from 354.1 ± 25.3 to 659.0 ± 71.8 μl/s). Nevertheless, passive P(CRIT) in leptin-treated mice did not differ significantly from that seen in nontreated ob(-)/ob(-) mice. The findings suggest that weight and leptin deficiency produced defects in upper airway neuromechanical control and that leptin reversed defects in active neuromuscular responses acutely without reducing mechanical loads.

Leptin enhances insulin sensitivity in addition to reducing food intake and body weight. Recently, amylin, a pancreatic β-cell-derived hormone, was shown to restore a weight-reducing effect of leptin in leptin-resistant diet-induced obesity. However, whether amylin improves the effect of leptin on insulin sensitivity in diet-induced obesity is unclear. Diet-induced obese (DIO) mice were infused with either saline (S), leptin (L; 500 μg·kg⁻¹·day⁻¹), amylin (A; 100 μg·kg⁻¹·day⁻¹), or leptin plus amylin (L/A) for 14 days using osmotic minipumps. Food intake, body weight, metabolic parameters, tissue triglyceride content, and AMP-activated protein kinase (AMPK) activity were examined. Pair-feeding and weight-matched calorie restriction experiments were performed to assess the influence of food intake and body weight reduction. Continuous L/A coadministration significantly reduced food intake, increased energy expenditure, and reduced body weight, whereas administration of L or A alone had no effects. L/A coadministration did not affect blood glucose levels during ad libitum feeding but decreased plasma insulin levels significantly (by 48%), suggesting the enhancement of insulin sensitivity. Insulin tolerance test actually showed the increased effect of insulin in L/A-treated mice. In addition, L/A coadministration significantly decreased tissue triglyceride content and increased AMPKα2 activity in skeletal muscle (by 67%). L/A coadministration enhanced insulin sensitivity more than pair-feeding and weight-matched calorie restriction. In conclusion, this study demonstrates the beneficial effect of L/A coadministration on glucose and lipid metabolism in DIO mice, indicating the possible clinical usefulness of L/A coadministration as a new antidiabetic treatment in obesity-associated diabetes.

Maintenance of a 10% or greater reduced body weight results in decreases in the energy cost of low levels of physical activity beyond those attributable to the altered body weight. These changes in nonresting energy expenditure are due mainly to increased skeletal muscle work efficiency following weight loss and are reversed by the administration of the adipocyte-derived hormone leptin. We have also shown previously that the maintenance of a reduced weight is accompanied by a decrease in ratio of glycolytic (phosphofructokinase) to oxidative (cytochrome c oxidase) activity in vastus lateralis muscle that would suggest an increase in the relative expression of the myosin heavy chain I (MHC I) isoform. We performed analyses of vastus lateralis muscle needle biopsy samples to determine whether maintenance of an altered body weight was associated with changes in skeletal muscle metabolic properties as well as mRNA expression of different isoforms of the MHC and sarcoplasmic endoplasmic reticular Ca(2+)-dependent ATPase (SERCA) in subjects studied before weight loss and then again after losing 10% of their initial weight and receiving twice daily injections of either placebo or replacement leptin in a single blind crossover design. We found that the maintenance of a reduced body weight was associated with significant increases in the relative gene expression of MHC I mRNA that was reversed by the administration of leptin as well as an increase in the expression of SERCA2 that was not significantly affected by leptin. Leptin administration also resulted in a significant increase in the expression of the less MHC IIx isoform compared with subjects receiving placebo. These findings are consistent with the leptin-reversible increase in skeletal muscle chemomechanical work efficiency and decrease in the ratio of glycolytic/oxidative enzyme activities observed in subjects following dietary weight loss.

Objective: Evidence supports an antilipotoxic role for leptin in preventing inappropriate peripheral tissue lipid deposition. Obese, leptin-deficient mice develop left ventricular (LV) hypertrophy and myocardial steatosis with increased apoptosis and decreased longevity. Here we investigated the cardiac effects of caloric restriction versus leptin repletion in obese leptin-deficient (ob/ob) mice., Methods: Echocardiography was performed on 7 mo old C57BL/6 wild-type mice (WT) and ob/ob mice fed ad libitum, leptin-repleted (LR-ob/ob), or calorie-restricted (CR-ob/ob) for 4 wk. Ventricular tissue was examined by electron microscopy (EM), triglyceride (TAG) content, oil red O staining, mitochondrial coupling assay, and microarray expression profiling., Results: LR and CR-ob/ob mice showed decreased body and heart weight, and LV wall thickness compared with ad libitum ob/ob mice. LV fractional shortening was decreased in ad libitum ob/ob mice, but restored to WT in LR and CR groups. However, myocardial lipid content by EM and TAG analysis revealed persistent cardiac steatosis in the CR-ob/ob group. Although CR restored mitochondrial coupling to WT levels, PPARα was suppressed and genes associated with oxidative stress and cell death were upregulated in CR-ob/ob animals. In contrast, LR eliminated cardiac steatosis, normalized mitochondrial coupling, and restored PGC1α and PPARα expression, while inducing core genes involved in glycerolipid/free fatty acid (GL/FFA) cycling, a thermogenic pathway that can reduce intracellular lipids., Conclusions: Thus, CR in the absence of leptin fails to normalize cardiac steatosis. GL/FFA cycling may be, at least in part, leptin-dependent and a key pathway that protects the heart from lipid accumulation.

Chronic intermittent hypoxia (IH) during sleep can result from obstructive sleep apnea (OSA), a disorder that is particularly prevalent in obesity. OSA is associated with high levels of circulating leptin, cardiovascular dysfunction, and dyslipidemia. Relationships between leptin and cardiovascular function in OSA and chronic IH are poorly understood. We exposed lean wild-type (WT) and obese leptin-deficient ob/ob mice to IH for 4 wk, with and without leptin infusion, and measured cardiovascular indices including aortic vascular stiffness, endothelial function, cardiac myocyte morphology, and contractile properties. At baseline, ob/ob mice had decreased vascular compliance and endothelial function vs. WT mice. We found that 4 wk of IH decreased vascular compliance and endothelial relaxation responses to acetylcholine in both WT and leptin-deficient ob/ob animals. Recombinant leptin infusion in both strains restored IH-induced vascular abnormalities toward normoxic WT levels. Cardiac myocyte morphology and function were unaltered by IH. Serum cholesterol and triglyceride levels were significantly decreased by leptin treatment in IH mice, as was hepatic stearoyl-Coenzyme A desaturase 1 expression. Taken together, these data suggest that restoring normal leptin signaling can reduce vascular stiffness, increase endothelial relaxation, and correct dyslipidemia associated with IH.

In the fetus, leptin in the circulation increases at late gestation and likely influences fetal organ development. Increased surfactant by leptin was previously demonstrated in vitro using fetal lung explant. We hypothesized that leptin treatment given to fetal sheep and pregnant mice might increase surfactant synthesis in the fetal lung in vivo. At 122-124 days gestational age (term: 150 days), fetal sheep were injected with 5 mg of leptin or vehicle using ultrasound guidance. Three and a half days after injection, preterm lambs were delivered, and lung function was studied during 30-min ventilation, followed by pulmonary surfactant components analyses. Pregnant A/J mice were given 30 or 300 mg of leptin or vehicle by intraperitoneal injection according to five study protocols with different doses, number of treatments, and gestational ages to treat. Surfactant components were analyzed in fetal lung 24 h after the last maternal treatment. Leptin injection given to fetal sheep increased fetal body weight. Control and leptin-treated groups were similar in lung function (preterm newborn lamb), surfactant components pool sizes (lamb and fetal mice), and expression of genes related to surfactant synthesis in the lung (fetal mice). Likewise, saturated phosphatidylcholine and phospholipid were normal in mice lungs with absence of circulating leptin (ob/ob mice) at all ages. These studies coincided in findings that neither exogenously given leptin nor deficiency of leptin influenced fetal lung maturation or surfactant pool sizes in vivo. Furthermore, the key genes critically required for surfactant synthesis were not affected by leptin treatment.

During lactation, female small mammals frequently reduce their fat reserves to very low levels. The function of this reduction is unclear, as calculations suggest that the contribution of the withdrawn energy from fat to the total energy balance of lactation is trivial. An alternative hypothesis is that reducing fat leads to a reduction in circulating adipokines, such as leptin, that play a role in stimulating the hyperphagia of lactation. We investigated the role of circulating leptin in lactation by repleting leptin levels using miniosmotic pumps during the last 7 days of lactation in Brandt's voles (Lasiopodomys brandtii), a model small wild mammal we have extensively studied in the context of lactation energy demands. Repletion of leptin resulted in a dose-dependent reduction of body mass and food intake in lactating voles. Comparisons to nonreproducing individuals suggests that the reduced leptin in lactation, due to reduced fat stores, may account for ∼16% of the lactational hyperphagia. Reduced leptin in lactation may, in part, cause lactational hyperphagia via stimulatory effects on hypothalamic orexigenic neuropeptides (neuropeptide Y and agouti-related peptide) and inhibition of the anorexigenic neuropeptide (proopiomelanocortin). These effects were reversed by the experimental repletion of leptin. There was no significant effect of leptin treatment on daily energy expenditure, milk production or pup growth, but leptin repletion did result in a reversal of the suppression of uncoupling protein-1 levels in brown adipose tissue, indicating an additional role for reducing body fat and leptin during peak lacation.

In sheep, central leptin infusion reduces food intake and increases energy expenditure in adipose tissue and skeletal muscle. The mechanisms for these peripheral effects of central leptin in sheep are not known but, on the basis of rodent studies, may involve AMPK. In mice, central leptin acutely increases both skeletal muscle AMPK activation and glucose uptake. Thus, to investigate whether these effects exist in higher-order mammals, ovariectomized Corriedale ewes (n = 4 per group) received a continuous lateral ventricular infusion (60 μl/h) of either leptin (50 μg/h) or artificial cerebrospinal fluid (aCSF; CON) for 8 days. Tritiated glucose (3-(3)H-glucose) was infused intravenously for calculation of whole body glucose turnover during both acute (6 h) and chronic (7-8 days) leptin/aCSF infusion. Muscle biopsies were also obtained. Leptin infusion reduced (P < 0.05) food intake and body weight, and it also increased plasma epinephrine concentration at 6 h and 7 days, suggesting increased sympathetic nerve activity. Despite this, and in contrast to rodent studies, central leptin infusion did not increase skeletal muscle AMPKα Thr(172) phosphorylation or ACCβ Ser(221) phosphorylation. Surprisingly, the glucose rate of appearance (glucose Ra) and rate of disappearance (glucose Rd) were reduced by both acute and chronic leptin infusion. Direct infusion of the AMPK activator 5-aminoimidazole-4-carboxyamide-ribonucleoside (AICAR) into the femoral artery increased skeletal muscle AMPK phosphorylation. In conclusion, although central leptin infusion in sheep caused the predicted reduction in food intake and increases plasma epinephrine concentration, it had no effect on AMPK activation in skeletal muscle and actually reduced glucose disposal. This suggests that there are species differences in the peripheral responses to central leptin infusion.

Fasting increases c-Fos expression in neuropeptide Y (NPY) neurons of the hypothalamic arcuate nucleus (ARC) in lean, but not in hyperleptinemic mice with late-onset obesity (LOO). Although obesity is associated with leptin resistance, we hypothesized that under fasting conditions, leptin sensitivity might be restored and that hyperleptinemia may counteract the neuronal response to fasting. We investigated whether the reduced fasting response of ARC neurons in LOO is paralleled by an increase in leptin sensitivity, as measured by leptin-induced STAT-3 phosphorylation. To assess leptin's role in the modulation of the fasting-induced ARC activation, we investigated c-Fos responses and hormone and metabolite levels in hyperleptinemic diet-induced obese (DIO) and in leptin-deficient ob/ob mice. Leptin induced a stronger STAT-3 phosphorylation in fasted LOO and lean mice than in ad libitum-fed animals. Similar to LOO, hyperleptinemic DIO mice showed no c-Fos response after fasting, while ob/ob mice showed a stronger response than lean control mice. Mimicking hyperleptinemia by repeated leptin injections in lean mice during fasting attenuated the fasting-induced c-Fos expression. Our findings indicate that high leptin levels prevent the fasting-induced activation of ARC neurons in mice. Moreover, leptin sensitivity is dynamic in obese subjects and depends on the feeding status. During short-term increases in leptin sensitivity, e.g., during fasting, leptin signaling appears to be effective, even in hyperleptinemic obesity. As reflected by the blockade of the fasting-induced ARC activation, fasting seems to interfere with the responsiveness of the ARC to signals related to the status of energy intake.

Leptin serum concentration in early life is an important factor for adequate future development of the offspring. Previously, we demonstrated that hyperleptinemia on lactation programmed for hyperleptinemia, central leptin resistance with lower expression of the long form of leptin receptor at hypothalamus, and higher medullary catecholamine levels with cardiovascular consequences at adulthood. The central objective of this study was to determine the direct effect of leptin on adrenal medullary function of adult rats that were leptin treated during lactation. Adrenal morphology was also accessed. Recombinant murine leptin was injected in the pups during the first 10 days of life (group L, leptin-programmed) or at adulthood during 6 days (group LC). The controls of both experiments received saline (groups C and CC). Both treatments resulted in hyperleptinemia at 150 days old (+78% and 2-fold increase, respectively; P < 0.05). Programmed animals showed hypertrophy of adrenal and higher adrenal catecholamine content at 150 days old (3-fold increase, P < 0.05), and no changes were observed in the LC group. However, LC rats had lower adrenal content of tyrosine hydroxylase (-17%, P < 0.05). Leptin-programmed rats had a lower response to leptin in vitro stimulation (-22%, P < 0.05) and lower expression of key proteins of the leptin signaling pathway, leptin receptor and janus tyrosine kinase 2 in the medullas (-61% and -29%, respectively, P < 0.05). However, they presented higher expression of phosphorylated signal transducer and activator of transcription 3 (+2-fold, P < 0.05). Leptin treatment at adulthood did not affect these parameters. The higher catecholamine synthesis and secretion in the leptin-programmed rats observed in our previous study does not seem to be a consequence of the direct effect of leptin on the medullas. We suggest that the hyperleptinemia of the programmed animals increases adrenal medullary function through sympathetic nervous system activation. In conclusion, high leptin levels on lactation program the activity of the sympathoadrenal system at adulthood that may contribute to the development of adult chronic diseases such as hypertension.

Gastric-derived leptin affects satiety and gastrointestinal function via vagal mechanisms and has been shown to interact with the gut hormone cholecystokinin (CCK). CCK selectively inhibits splanchnic sympathetic nerve discharge (SND) and the activity of a subset of presympathetic vasomotor neurons in the rostroventrolateral medulla (RVLM). The present study sought to examine the effects of gastric leptin on arterial pressure (AP), heart rate (HR), SND, and RVLM neuronal activity to determine whether its effects on cardiovascular regulation are dependent on CCK(1) receptors and vagal afferent transmission. To mimic gastric leptin, leptin (15-30 microg/kg) was administered close to the coeliac artery in anesthetized, artificially ventilated Sprague-Dawley rats. Within 5 min, leptin selectively decreased the activity of RVLM neurons also inhibited by CCK (-27 +/- 4%; P < 0.001; n = 15); these inhibitory effects were abolished following administration of the CCK(1) receptor antagonist lorglumide. Leptin significantly decreased AP and HR (-10 +/- 2 mmHg, P < 0.001; and -8 +/- 2 beats/min, P < 0.01; n = 35) compared with saline (-1 +/- 2 mmHg, 3 +/- 2 beats/min; n = 30). In separate experiments, leptin inhibited splanchnic SND compared with saline (-9 +/- 2% vs. 2 +/- 3%, P < 0.01; n = 8). Bilateral cervical vagotomy abolished the sympathoinhibitory, hypotensive, and bradycardic effects of leptin (P < 0.05; n = 6). Our results suggest that gastric leptin may exert acute sympathoinhibitory and cardiovascular effects via vagal transmission and CCK(1) receptor activation and may play a separate role to adipose leptin in short-term cardiovascular regulation.

Leptin is thought to reduce food intake, in part, by increasing sensitivity to satiation signals, including CCK. Leptin action in both forebrain and hindbrain reduces food intake, and forebrain leptin action augments both the anorexic and neuronal activation responses to CCK. Here, we asked whether leptin signaling in hindbrain also enhances these responses to CCK. We found that food intake was strongly inhibited at 30 min after a combination of 4th-intracerebroventricular (4th-icv) leptin injection and intraperitoneal CCK administration, whereas neither hormone affected intake during this period when given alone. Leptin injections targeted directly at the dorsal vagal complex (DVC) similarly enhanced the anorexic response to intraperitoneal CCK. Intra-DVC leptin injection also robustly increased the number of neurons positive for phospho-STAT3 staining in the area surrounding the site of injection, confirming local leptin receptor activation. Conversely, the anorexic response to 4th-icv leptin was completely blocked by IP devazepide, a CCKA-R antagonist, suggesting that hindbrain leptin reduces intake via a mechanism requiring endogenous CCK signaling. We then asked whether hindbrain leptin treatment enhances the dorsomedial hindbrain, hypothalamus, or amygdala c-Fos responses to IP CCK. We found that, in contrast to the effects of forebrain leptin administration, 4th-icv leptin injection had no effect on CCK-induced c-Fos in any structures examined. We conclude that leptin signaling in either forebrain or hindbrain areas can enhance the response to satiation signals and that multiple distinct neural circuits likely contribute to this interaction.

Fasting activates orexigenic neuropeptide Y neurons in the hypothalamic arcuate nucleus (ARC) of mice, which is reversed by 2 h refeeding with standard chow. Here, we investigated the contribution of diet-derived macronutrients and anorectic hormones to the reversal of the fasting-induced ARC activation during 2 h refeeding. Refeeding of 12-h-fasted mice with a cellulose-based, noncaloric mash induced only a small reduction in c-Fos expression. Refeeding with diets, containing carbohydrates, protein, or fat alone reversed it similar to chow; however, this effect depended on the amount of intake. The fasting-induced ARC activation was unchanged by subcutaneously injected amylin, CCK (both 20 microg/kg), insulin (0.2 U/kg and 0.05 U/kg) or leptin (2.6 mg/kg). Insulin and leptin had no effect on c-Fos expression in neuropeptide Y or proopiomelanocortin-containing ARC neurons. Interestingly, CCK but not amylin reduced the ghrelin-induced c-Fos expression in the ARC in ad libitum-fed mice, suggesting that CCK may inhibit orexigenic ARC neurons when acting together with other feeding-related signals. We conclude that all three macronutrients and also non-nutritive, ingestion-dependent signals contribute to an inhibition of orexigenic ARC neurons after refeeding. Similar to the previously demonstrated inhibitory in vivo action of peptide YY, CCK may be a postprandial mediator of ARC inhibition.

Recent evidence suggests that leptin reduces food intake via actions in the brain circuitry of food reward, such as the ventral tegmental area (VTA), as leptin receptors are present in the VTA, and leptin injection in the VTA reduces food intake. In the hypothalamus, leptin-induced anorexia requires signaling via Janus kinase-signal transducer and activator of transcription (Jak-STAT), insulin receptor substrate (IRS)-phosphatidylinositol 3-kinase (PI 3-kinase), and mammalian target of rapamycin (mTOR). In this study, we determined whether leptin activates each of these signal transduction pathways in the VTA and whether these signaling pathways are required for VTA-leptin induced anorexia. Here, we show that pSTAT3-Tyr(705), a marker of leptin activation, was induced in a midbrain region containing the VTA and substantia nigra following either intracerebroventricular leptin or direct administration of leptin to the VTA, but these interventions failed to increase levels of either pAKT-Ser(473) or phospho-p70S6K-Thr(389), markers of IRS-PI 3-kinase and mTOR signaling, respectively. Moreover, the effect of intra-VTA leptin administration to reduce 4- and 20-h food intake and 20-h body weight was blocked by an inhibitor of Jak-2, at a dose that had no effect on food intake or body weight by itself, but not by local inhibition of either PI 3-kinase (LY-294002) or mTOR (rapamycin) in this timeframe. Taken together, these data support the hypothesis that leptin signaling in the VTA is involved in the regulation of energy balance, but, in contrast to the leptin signaling in the hypothalamus, these effects are mediated predominantly via Jak-2 signaling rather than via the IRS-PI 3-kinase or mTOR signaling pathway.

The relationship between thyroid function and leptin has been extensively studied; however, the mechanisms underlying the changes in thyroid hormone economy that occur during caloric deprivation remain elusive. Our goal was to evaluate the thyroid function of rats submitted to 40% food restriction after chronic leptin replacement. Caloric restriction for 25 days led to significantly reduced serum leptin, thyroid-stimulating hormone (TSH), thyroxine (T(4)), and triiodothyronine (T(3)) and increased serum corticosterone, while liver, kidney, and thyroid type I deiodinase (D1) and brown adipose tissue (BAT) type II deiodinase (D2) activities were decreased and hypothalamic D2 was significantly increased. Interestingly, thyroid iodide uptake was unchanged by caloric restriction, but thyroperoxidase (TPO) activity was significantly reduced. Leptin replacement for the last 10 days of caloric restriction normalized serum leptin and TSH levels, but serum T(4) and T(3) levels and thyroid D1 and TPO activities were not reestablished. Also, a negative effect of leptin administration on Na(+)-I(-) symporter function was detected. Liver and kidney D1 and hypothalamic and BAT D2 were normalized by leptin, while pituitary D2 was significantly decreased. In conclusion, a tissue-specific modulation of deiodinases might be implicated in the normalization of thyroid function during leptin replacement in food-restricted rats. Although leptin restores the hypothalamus-pituitary axis during food restriction, it exerts a direct negative effect on the thyroid gland; thus normalization of serum thyroid hormones might depend on changes in deiodinase activities and the long-term thyroid stimulation by TSH to counterbalance the direct negative effects of leptin on the thyroid gland.

Food deprivation stimulates foraging and hoarding and to a much lesser extent, food intake in Siberian hamsters. Leptin, the anorexigenic hormone secreted primarily from adipocytes, may act in the periphery, the brain, or both to inhibit these ingestive behaviors. Therefore, we tested whether leptin given either intracerebroventricularly or intraperitoneally, would block food deprivation-induced increases in food hoarding, foraging, and intake in animals with differing foraging requirements. Hamsters were trained in a running wheel-based food delivery foraging system coupled with simulated burrow housing. We determined the effects of food deprivation and several peripheral doses of leptin on plasma leptin concentrations. Hamsters were then food deprived for 48 h and given leptin (0, 10, 40, or 80 microg ip), and additional hamsters were food deprived for 48 h and given leptin (0, 1.25, 2.5, or 5.0 microg icv). Foraging, food intake, and hoarding were measured postinjection. Food deprivation stimulated food hoarding to a greater degree and duration than food intake. In animals with a foraging requirement, intracerebroventricular leptin almost completely blocked food deprivation-induced increased food hoarding and intake, but increased foraging. Peripheral leptin treatment was most effective in a sedentary control group, completely inhibiting food deprivation-induced increased food hoarding and intake at the two highest doses, and did not affect foraging at any dose. Thus, the ability of leptin to inhibit food deprivation-induced increases in ingestive behaviors differs based on foraging effort (energy expenditure) and the route of administration of leptin administration.

Food availability and quality in desert habitats are spatially and temporally unpredictable, and animals face periods of food shortage. The golden spiny mouse (Acomys russatus) is an omnivorous desert rodent that does not hoard food, requiring it to withstand such periods by physiological means alone. In response to food restriction, plasma leptin concentrations, core body temperature, and energy expenditure of the spiny mouse decrease significantly after 24 h, and most spiny mice are able to maintain their body mass to approximately 85% of ad libitum for a prolonged period of time. Both 1-day food deprivation and long-term food restriction had a significant effect on body mass and plasma leptin concentrations, which decreased significantly with a high correlation, as well as on the orexigenic agouti-related protein, which increased significantly as a result of the 24-h food deprivation; and on neuropeptide Y (NPY), in which the increase was more pronounced under long-term food restriction. Food restriction and food deprivation had no effect, however, on the anorexigenic pro-opiomelanocortin and cocaine and amphetamine-related transcript. Leptin administration to food-restricted spiny mice did not affect food intake or the rate of decrease in body mass, indicating that it cannot overcome the drive to eat when food is scarce. However, it did result in a significant decrease in NPY levels, and the spiny mice spent less time at low body temperatures compared with PBS-treated golden spiny mice. These results show that in food-restricted golden spiny mice, leptin affects thermogenesis, but not food consumption, and suggest that the thermoregulatory effects of leptin are mediated by NPY.

Leptin plays a critical role in the control of energy homeostasis. The sympathetic cardiovascular actions of leptin have emerged as a potential link between obesity and hypertension. We previously demonstrated that in mice, modest obesity induced by 10 wk of a high-fat diet is associated with preservation of leptin ability to increase renal sympathetic nerve activity (SNA) despite the resistance to the metabolic effects of leptin. Here, we examined whether selective leptin resistance exists in mice with late-stage diet-induced obesity (DIO) produced by 20 wk of a high-fat diet. The decrease in food intake and body weight induced by intraperitoneal or intracerebroventricular injection of leptin was significantly attenuated in the DIO mice. Regional SNA responses to intravenous leptin were also attenuated in DIO mice. In contrast, intracerebroventricularly administered leptin caused contrasting effects on regional SNA in DIO mice. Renal SNA response to intracerebroventricular leptin was preserved, whereas lumbar and brown adipose tissue SNA responses were attenuated. Intact renal SNA response to leptin combined with the increased cerebrospinal fluid leptin levels in DIO mice represents a potential mechanism for the adverse cardiovascular consequences of obesity. Lastly, we examined the role of phosphoinositol-3 kinase (PI3K) and melanocortin receptors (MCR) in mediating the preserved renal SNA response to leptin in obesity. Presence of PI3K inhibitor (LY294002) or MC3/4R antagonist (SHU9119) significantly attenuated the renal SNA response to leptin in DIO and agouti obese mice. Our results demonstrate the importance of PI3K and melanocortin receptors in the transduction of leptin-induced renal sympathetic activation in obesity.

This study tested whether leptin restores sympathetic-vagal balance, heart rate (HR) variability, and cardiac baroreflex sensitivity (BRS) in streptozotocin (STZ)-induced diabetes. Sprague-Dawley rats were instrumented with arterial and venous catheters, and a cannula was placed in the lateral ventricle for intracerebroventricular (ICV) leptin infusion. Blood pressure (BP) and HR were monitored by telemetry. BRS and HR variability were estimated by linear regression between HR and BP responses to phenylephrine or sodium nitroprusside and autoregressive spectral analysis. Measurements were made during control period, 7 days after induction of diabetes, and 7 days after ICV leptin infusion. STZ diabetes was associated with hyperglycemia (422 +/- 17 mg/dl) and bradycardia (-79 +/- 4 beats/min). Leptin decreased glucose levels (165 +/- 16 mg/dl) and raised HR to control values (303 +/- 10 to 389 +/- 10 beats/min). Intrinsic HR (IHR) and chronotropic responses to a full-blocking dose of propranolol and atropine were reduced during diabetes (260 +/- 7 vs. 316 +/- 6, -19 +/- 2 vs. -43 +/- 6, and 39 +/- 3 vs. 68 +/- 8 beats/min), and leptin treatment restored these variables to normal (300 +/- 7, -68 +/- 10, and 71 +/- 8 beats/min). Leptin normalized BRS (bradycardia, -2.6 +/- 0.3, -1.7 +/- 0.2, and -3.0 +/- 0.5; and tachycardia, -3.2 +/- 0.4, -1.9 +/- 0.3, and -3.4 +/- 0.3 beats.min(-1).mmHg(-1) for control, diabetes, and leptin) and HR variability (23 +/- 4 to 11 +/- 1.5 ms2). Chronic glucose infusion to maintain hyperglycemia during leptin infusion did not alter the effect of leptin on IHR but abolished the improved BRS. These results show rapid impairment of autonomic nervous system control of HR after the induction of diabetes and that central nervous system actions of leptin can abolish the hyperglycemia as well as the altered IHR and BRS in STZ-induced diabetes.

Energy balance controls the expression of the leptin receptor (Lepr) in the ruminant hypothalamus but whether similar regulation occurs in peripheral tissues is unknown. To address this issue, we measured Lepr expression in the liver and adipose tissue of dairy cows during the transition from late pregnancy (LP) to early lactation (EL). This period is characterized by the development of a profound state of energy insufficiency and is associated with reduced plasma insulin and leptin and with increased plasma growth hormone. Hepatic expression of the short (Lepr-a) and long (Lepr-b) isoforms was 40% higher during EL (8 days postpartum) than LP (30 days prepartum). A similar effect was observed when negative energy balance was induced in nonpregnant, late-lactation dairy cows by food restriction, implicating energy insufficiency as a specific cause in EL. The stimulation of hepatic Lepr expression was reversed after a 48-h period of hyperinsulinemic euglycemia in EL. Changes in hepatic Lepr expression during chronic elevation of plasma leptin in EL or plasma growth hormone in nonpregnant, late-lactation cows did not support a role for these hormones in mediating the effects of energy insufficiency on hepatic Lepr expression. In adipose tissue, Lepr expression was increased 10-fold during the transition from LP to EL. Overall, these data indicate that hypoinsulinemia is partly responsible for the induction of Lepr expression in the liver, and perhaps adipose tissue, of energy-deficient dairy cows.

Acute administration of peptide YY(3-36) [PYY(3-36)] results in a reduction in food intake in several different vertebrates. However, long-term continuous administration of PYY(3-36) causes only a transient reduction in food intake, thus potentially limiting its therapeutic efficacy. We hypothesized that a fall in leptin levels associated with reduced food intake could contribute to the transient anorectic effects of continuous PYY(3-36) infusion and thus that leptin replacement might prolong the anorectic effects of PYY(3-36). Seven-day administration of 100 microg x kg body wt(-1) x day(-1) PYY(3-36) using osmotic minipumps caused a significant reduction in food intake of ad libitum-fed rats, but only for the first 2 days postimplantation. Circulating levels of leptin were reduced 1 day following continuous infusion of PYY(3-36), and combined leptin infusion at a dose of leptin that had no anorectic effects on its own (100 microg x kg body wt(-1) x day(-1)) prolonged the anorectic actions of PYY(3-36) in ad libitum-fed rats for up to 6 days postimplantation and yielded reduced weight gain compared with either peptide alone. The inhibitory effects of 100 microg x kg body wt(-1) x day(-1) PYY(3-36) on food intake were absent in rats refed after a 24-h fast and substantially reduced at a dose of 1,000 microg x kg body wt(-1) x day(-1) PYY(3-36). Leptin replacement was unable to recover the anorectic effects of PYY(3-36) in fasted rats. Our results suggest that an acute fall in leptin levels is not solely responsible for limiting duration of action of chronic PYY(3-36) infusion, yet chronic coadministration of a subanorectic dose of leptin can extend the anorectic effects of PYY(3-36).

Plasma leptin is often elevated in obese individuals, and previous studies have suggested leptin as a factor that links obesity and atherosclerosis. Because macrophages play a key role in atherogenesis and are responsive to leptin, we hypothesized that leptin increases aortic root lesion formation, in part, through macrophage leptin receptor (LepR). Three different bone marrow transplantation studies were conducted in which bone marrow, with or without LepR, was transplanted into lethally irradiated 1) LDL receptor-deficient (LDLR(-/-)) mice with moderate hyperleptinemia due to Western diet (WD) feeding, 2) LDLR(-/-) mice with WD feeding plus pharmacologically induced hyperleptinemia (daily injection of 125 microg leptin), or 3) obese, hyperleptinemic, LepR-deficient LDLR(-/-) (LepR(db/db);LDLR(-/-)) mice. Minor differences in plasma parameters such as cholesterol, triglycerides, and insulin were observed in some groups; however, a consistent trend for the role of LepR on these parameters was not detected. In each of the studies, macrophage LepR expression did not have an effect on aortic root atherosclerotic lesion formation. These results suggest that nonhematopoietic cells may have a more significant role than macrophages in leptin-mediated effects on aortic root lesion formation.

The metabolic syndrome is a group of disorders including obesity, insulin resistance, atherogenic dyslipidemia, hyperglycemia, and hypertension. To date, few animal models have been described to recapitulate the phenotypes of the syndrome. In this study, we generated and characterized two lines of triple-knockout mice that are deficient in either apolipoprotein E (Apoe(-/-)) or low-density lipoprotein receptor (Ldlr(-/-)) and express no leptin (Lep(ob/ob)) or apolipoprotein B-48 but exclusively apolipoprotein B-100 (Apob(100/100)). These two lines are referred to as Apoe triple-knockout-Apoe 3KO (Apoe(-/-)Apob(100/100)Lep(ob/ob)) and Ldlr triple-knockout-Ldlr 3KO (Ldlr(-/-)Apob(100/100)Lep(ob/ob)) mice. Both lines develop obesity, hyperinsulinemia, hyperlipidemia, hypertension, and atherosclerosis. However, only Apoe 3KO mice are hyperglycemic and glucose intolerant and are more obese than Ldlr 3KO mice. To evaluate the utility of these lines as pharmacological models, we treated both with leptin and found that leptin therapy ameliorated most metabolic derangements. Leptin was more effective in improving glucose tolerance in Ldlr 3KO than Apoe 3KO animals. The reduction of plasma cholesterol by leptin in Ldlr 3KO mice can be accounted for by its suppressive effect on food intake. However, in Apoe 3KO mice, leptin further reduced plasma cholesterol independently of its effect on food intake, and this improvement correlated with a smaller plaque lesion area. These effects suggest a direct role of leptin in modulating VLDL levels and, likewise, the lesion areas in VLDL-enriched animals. These two lines of mice represent new models with features of the metabolic syndrome and will be useful in testing therapies targeted for combating the human condition.

Leptin reduces body fat selectively, sparing body protein. Accordingly, during chronic leptin administration, food intake is suppressed, and body weight is reduced until body fat is depleted. Body weight then stabilizes at this fat-depleted nadir, while food intake returns to normal caloric levels, presumably in defense of energy and nutritional homeostasis. This model of leptin treatment offers the opportunity to examine controls of food intake that are independent of leptin's actions, and provides a window for examining the nature of feeding controls in a "fatless" animal. Here we evaluate macronutrient selection during this fat-depleted phase of leptin treatment. Adult, male Sprague-Dawley rats were maintained on standard pelleted rodent chow and given daily lateral ventricular injections of leptin or vehicle solution until body weight reached the nadir point and food intake returned to normal levels. Injections were then continued for 8 days, during which rats self-selected their daily diet from separate sources of carbohydrate, protein, and fat. Macronutrient choice differed profoundly in leptin and control rats. Leptin rats exhibited a dramatic increase in protein intake, whereas controls exhibited a strong carbohydrate preference. Fat intake did not differ between groups at any time during the 8-day test. Despite these dramatic differences in macronutrient selection, total daily caloric intake did not differ between groups except on day 2. Thus controls of food intake related to ongoing metabolic and nutritional requirements may supersede the negative feedback signals related to body fat stores.

The HPA axis is hyperactive under conditions of leptin and insulin resistance as well as after ANG II administration. We hypothesized that a hyperreactivity of the HPA axis to ANG contributes to an impaired glucose utilization in obesity, since leptin resistance and an overactive renin-angiotensin-aldosterone system are features of obesity. Zucker rats were treated with ANG via subcutaneous minipumps (0, 0.9, and 9.0 mug/h; 4 wk). PA axis reactivity and glucose homeostasis were characterized after CRH treatment and during an oral glucose tolerance test (OGTT). The elevated plasma profile of corticosterone after CRH stimulation in saline-treated OZR compared with LZR confirmed that the sensitization of the PA axis depended on leptin resistance. Irrespective of the rat strain, circulating ANG levels and blood pressure were selectively increased after administration of 9 mug/h ANG (high ANG). Only high ANG induced an elevation of the corticosterone and glucose response after CRH stimulation in OZR but did not affect the ACTH secretion. During OGTT, corticosterone and consequently glucose increased in OZR after high ANG, whereas the insulin secretion was decreased. In the adrenal glands of OZR, AT(1A) receptor mRNA levels increased after high ANG. We conclude that the impairment of glucose utilization after ANG stimulation is potentiated in leptin-resistant rats as a result of a hyperreactive PA axis, thereby confirming the functional importance of a dysregulation within the HPA axis in metabolic syndrome or obesity. The ACTH-independent stimulation of corticosterone release and the selective increase of AT(1A) receptor mRNA in the adrenals of OZR indicated a sensitization of adrenals toward ANG, causing a stimulation of the PA axis.

High sucrose (HS) feeding in rats induces hepatic steatosis and plasma dyslipidemia. In previous reports (Huang W, Dedousis N, Bhatt BA, O'Doherty RM. J Biol Chem 279: 21695-21700, 2004; and Huang W, Dedousis N, Bandi A, Lopaschuk GD, O'Doherty RM. Endocrinology 147: 1480-1487, 2006), our laboratory demonstrated a rapid ( approximately 100 min) leptin-induced decrease in liver and plasma VLDL triglycerides (TG) in lean rats, effects that were abolished in obese rats fed a high-fat diet, a model that also presents with hepatic steatosis and plasma dyslipidemia. To further examine the capacity of acute leptin treatment to improve metabolic abnormalities induced by nutrient excess, hepatic leptin action was studied in rats after 5 wk of HS feeding. HS feeding induced hepatic steatosis (TG+80+/-8%; P=0.001), plasma hyperlipidemia (VLDL-TG+102+/-14%; P=0.001), hyperinsulinemia (plasma insulin +67+/-12%; P=0.04), and insulin resistance as measured by homeostasis model assessment (+125+/-20%; P=0.02), without increases in adiposity or plasma leptin concentration compared with standard chow-fed controls. A 120-min infusion of leptin (plasma leptin 13.6+/-0.7 ng/ml) corrected hepatic steatosis (liver TG-29+/-3%; P=0.003) and plasma hyperlipidemia in HS (VLDL-TG-42+/-4%; P=0.001) and increased plasma ketones (+45+/-3%; P=0.006), without altering plasma glucose, insulin, or homeostasis model assessment compared with saline-infused HS controls. In addition, leptin activated liver phosphatidylinositol 3-kinase (+70+/-18%; P=0.01) and protein kinase B (Akt; +90+/-29%; P=0.02), and inhibited acetyl-CoA carboxylase (40+/-7%; P=0.04) in HS, further demonstrating that hepatic leptin action was intact in these animals. We conclude that 1) leptin action on hepatic lipid metabolism remains intact in HS-fed rats, 2) leptin rapidly reverses hepatic steatosis and plasma dyslipidemia induced by sucrose, and 3) the preservation of hepatic leptin action after a HS diet is associated with the maintenance of low adiposity and plasma leptin concentrations.

Immunoreactivity of leptin receptor (Ob-R) has been detected in rat dorsal motor nucleus of the vagus (DMNV). Here, we confirmed the presence of Ob-R immunoreactivity on retrograde-labeled parasympathetic preganglionic neurons in the DMNV of neonatal rats. The present study investigated the effects of leptin on DMNV neurons, including parasympathetic preganglionic neurons, by using whole cell patch-clamp recording technique in brain stem slices of neonatal rats. Leptin (30-300 nM) induced membrane depolarization and hyperpolarization, respectively, in 14 and 15 out of 80 DMNV neurons tested. Both leptin-induced inward and outward currents persisted in the presence of TTX, indicating that leptin affected DNMV neurons postsynaptically. The current-voltage (I-V) curve of leptin-induced inward currents is characterized by negative slope conductance and has an average reversal potential of -90 +/- 3 mV. The reversal potential of the leptin-induced inward current was shifted to a more positive potential level in a high-potassium medium. These results indicate that a decrease in potassium conductance is likely the main ionic mechanism underlying the leptin-induced depolarization. On the other hand, the I-V curve of leptin-induced outward currents is characterized by positive slope conductance and has an average reversal potential of -88 +/- 3 mV, suggesting that an increase in potassium conductance may underlie leptin-induced hyperpolarization. Most of the leptin-responsive DMNV neurons were identified as being parasympathetic preganglionic neurons. These results suggest that the DMNV is one of the central target sites of leptin, and leptin can regulate parasympathetic outflow from the DMNV by directly acting on the parasympathetic preganglionic neurons of the DMNV.

One of the major manifestations of obesity is an increased production of the adipocyte-derived 16-kDa peptide leptin, which acts mainly on hypothalamic leptin receptors. Leptin receptors are widely distributed in various tissues, including the heart. Whereas increased plasma leptin levels have been reported in patients with congestive heart failure, systemic alterations induced by obesity can affect cardiac hypertrophy, and the direct effects of leptin on cardiac structure and function still remain to be determined. We first exposed primary cardiac myocytes from neonatal rats to leptin for 48 h. This resulted in a significant increase in myocyte long-axis length (P < 0.05 at 50 ng/ml) but not in the short-axis width. Leptin induced the rapid phosphorylation of STAT3 and its DNA binding in cardiac myocytes. Administration of a JAK2 inhibitor, AG-490, completely inhibited all of these effects by leptin. Furthermore, we examined the effect of continuous infusion of leptin for 4 wk following myocardial infarction in mice. Echocardiography demonstrated that left ventricular fractional shortening in the leptin-infused group (28.4 +/- 2.8%) was significantly higher than that in the PBS-infused group (18.4 +/- 2.2%) following myocardial infarction. Interestingly, left ventricular diastolic dimension in the leptin-infused group (4.56 +/- 0.12 mm) was also higher than that in the PBS-infused group (4.13 +/- 0.09 mm). These results demonstrate that leptin induces the elongation of cardiac myocytes via a JAK/STAT pathway and chronic leptin infusion causes eccentric dilatation with augmented systolic function after myocardial infarction.

Leptin preserves lean tissue but decreases adipose tissue by increasing lipolysis and/or inhibiting lipogenesis. The sympathetic nervous system (SNS) is a primary regulator of lipolysis, but it is not known if leptin increases norepinephrine turnover (NETO) in white adipose tissue. In this study, we examined the effect of leptin administered either as a chronic physiological dose (40 microg/day for 4 days from ip miniosmotic pumps) or as an acute injection in the third ventricle (1.5 microg injected two times daily for 2 days) on NETO and the size of brown and white fat depots in male Sprague Dawley rats. NETO was determined from the decline in tissue norepinephrine (NE) during 4 h following administration of the NE synthesis inhibitor alpha-methyl-para-tryrosine. The centrally injected leptin-treated animals demonstrated more dramatic reductions in food intake, body weight, and fat pad size and an increase in NETO compared with the peripherally infused animals. Neither route of leptin administration caused a uniform increase in NETO across all fat pads tested, and in both treatment conditions leptin decreased the size of certain fat pads independent of an increase in NETO. Similar discrepancies in white fat NETO were found for rats pair fed to leptin-treated animals. These results demonstrate that leptin acting either centrally or peripherally selectively increases sympathetic outflow to white fat depots and that a leptin-induced change in fat pad weight does not require an increase in NETO.