High-fat diet feeding disrupts the coupling of thermoregulation to energy homeostasis.

OBJECTIVE: Preserving core body temperature across a wide range of ambient temperatures requires adaptive changes of thermogenesis that must be offset by corresponding changes of energy intake if body fat stores are also to be preserved. Among neurons implicated in the integration of thermoregulation with energy homeostasis are those that express both neuropeptide Y (NPY) and agouti-related protein (AgRP) (referred to herein as AgRP neurons). Specifically, cold-induced activation of AgRP neurons was recently shown to be required for cold exposure to increase food intake in mice. Here, we investigated how consuming a high-fat diet (HFD) impacts various adaptive responses to cold exposure as well as the responsiveness of AgRP neurons to cold. METHODS: To test this, we used immunohistochemistry, in vivo fiber photometry and indirect calorimetry for continuous measures of core temperature, energy expenditure, and energy intake in both chow- and HFD-fed mice housed at different ambient temperatures. RESULTS: We show that while both core temperature and the thermogenic response to cold are maintained normally in HFD-fed mice, the increase of energy intake needed to preserve body fat stores is blunted, resulting in weight loss. Using both immunohistochemistry and in vivo fiber photometry, we show that although cold-induced AgRP neuron activation is detected regardless of diet, the number of cold-responsive neurons appears to be blunted in HFD-fed mice. CONCLUSIONS: We conclude that HFD-feeding disrupts the integration of systems governing thermoregulation and energy homeostasis that protect body fat mass during cold exposure.

Warm Responsive Neurons in the Hypothalamic Preoptic Area are Potent Regulators of Glucose Homeostasis in Male Mice.

When mammals are exposed to a warm environment, overheating is prevented by activation of "warm-responsive" neurons (WRNs) in the hypothalamic preoptic area (POA) that reduce thermogenesis while promoting heat dissipation. Heat exposure also impairs glucose tolerance, but whether this also results from activation of POA WRNs is unknown. To address this question, we sought in the current work to determine if glucose intolerance induced by heat exposure can be attributed to activation of a specific subset of WRNs that express pituitary adenylate cyclase-activating peptide (ie, POAPacap neurons). We report that when mice are exposed to an ambient temperature sufficiently warm to activate POAPacap neurons, the expected reduction of energy expenditure is associated with glucose intolerance, and that these responses are recapitulated by chemogenetic POAPacap neuron activation. Because heat-induced glucose intolerance was not blocked by chemogenetic inhibition of POAPacap neurons, we conclude that POAPacap neuron activation is sufficient, but not required, to explain the impairment of glucose tolerance elicited by heat exposure.

Leptin receptor neurons in the dorsomedial hypothalamus regulate diurnal patterns of feeding, locomotion, and metabolism.

The brain plays an essential role in driving daily rhythms of behavior and metabolism in harmony with environmental light-dark cycles. Within the brain, the dorsomedial hypothalamic nucleus (DMH) has been implicated in the integrative circadian control of feeding and energy homeostasis, but the underlying cell types are unknown. Here, we identify a role for DMH leptin receptor-expressing (DMHLepR) neurons in this integrative control. Using a viral approach, we show that silencing neurotransmission in DMHLepR neurons in adult mice not only increases body weight and adiposity but also phase-advances diurnal rhythms of feeding and metabolism into the light cycle and abolishes the normal increase in dark-cycle locomotor activity characteristic of nocturnal rodents. Finally, DMHLepR-silenced mice fail to entrain to a restrictive change in food availability. Together, these findings identify DMHLepR neurons as critical determinants of the daily time of feeding and associated metabolic rhythms.

Cold-induced hyperphagia requires AgRP neuron activation in mice.

To maintain energy homeostasis during cold exposure, the increased energy demands of thermogenesis must be counterbalanced by increased energy intake. To investigate the neurobiological mechanisms underlying this cold-induced hyperphagia, we asked whether agouti-related peptide (AgRP) neurons are activated when animals are placed in a cold environment and, if so, whether this response is required for the associated hyperphagia. We report that AgRP neuron activation occurs rapidly upon acute cold exposure, as do increases of both energy expenditure and energy intake, suggesting the mere perception of cold is sufficient to engage each of these responses. We further report that silencing of AgRP neurons selectively blocks the effect of cold exposure to increase food intake but has no effect on energy expenditure. Together, these findings establish a physiologically important role for AgRP neurons in the hyperphagic response to cold exposure.

Leptin regulation of core body temperature involves mechanisms independent of the thyroid axis.

The ability to maintain core temperature within a narrow range despite rapid and dramatic changes in environmental temperature is essential for the survival of free-living mammals, and growing evidence implicates an important role for the hormone leptin. Given that thyroid hormone plays a major role in thermogenesis and that circulating thyroid hormone levels are reduced in leptin-deficient states (an effect partially restored by leptin replacement), we sought to determine the extent to which leptin's role in thermogenesis is mediated by raising thyroid hormone levels. To this end, we 1) quantified the effect of physiological leptin replacement on circulating levels of thyroid hormone in leptin-deficient ob/ob mice, and 2) determined if the effect of leptin to prevent the fall in core temperature in these animals during cold exposure is mimicked by administration of a physiological replacement dose of triiodothyronine (T3). We report that, as with leptin, normalization of circulating T3 levels is sufficient both to increase energy expenditure, respiratory quotient, and ambulatory activity and to reduce torpor in ob/ob mice. Yet, unlike leptin, infusing T3 at a dose that normalizes plasma T3 levels fails to prevent the fall of core temperature during mild cold exposure. Because thermal conductance (e.g., heat loss to the environment) was reduced by administration of leptin but not T3, leptin regulation of heat dissipation is implicated as playing a uniquely important role in thermoregulation. Together, these findings identify a key role in thermoregulation for leptin-mediated suppression of thermal conduction via a mechanism that is independent of the thyroid axis.

Androgen receptor deficiency in monocytes/macrophages does not alter adiposity or glucose homeostasis in male mice.

Androgen deprivation in men leads to increased adiposity, but the mechanisms underlying androgen regulation of fat mass have not been fully defined. Androgen receptor (AR) is expressed in monocytes/macrophages, which are resident in key metabolic tissues and influence energy metabolism in surrounding cells. Male mice bearing a cell-specific knockout of the AR in monocytes/macrophages (M-ARKO) were generated to determine whether selective loss of androgen signaling in these cells would lead to altered body composition. Wild-type (WT) and M-ARKO mice (12-22 weeks of age, n = 12 per group) were maintained on a regular chow diet for 8 weeks and then switched to a high-fat diet for 8 additional weeks. At baseline and on both the regular chow and high-fat diets, no differences in lean mass or fat mass were observed between groups. Consistent with the absence of differential body weight or adiposity, no differences in food intake (3.0 ± 0.5 g per day for WT mice vs 2.8 ± 0.4 g per day for M-ARKO mice) or total energy expenditure (0.6 ± 0.1 Kcal h-1 for WT mice vs 0.5 ± 0.1 Kcal h-1 for M-ARKO mice) were evident between groups during high-fat feeding. Liver weight was greater in M-ARKO than that in WT mice (1.5 ± 0.1 g vs 1.3 ± 0.0 g, respectively, P = 0.02). Finally, M-ARKO mice did not exhibit impairments in glucose tolerance or insulin sensitivity relative to WT mice at any study time point. In aggregate, these findings suggest that AR signaling specifically in monocytes/macrophages does not contribute to the regulation of systemic energy balance, adiposity, or insulin sensitivity in male mice.

Metabolically distinct weight loss by 10,12 CLA and caloric restriction highlight the importance of subcutaneous white adipose tissue for glucose homeostasis in mice.

BACKGROUND: Widely used as a weight loss supplement, trans-10,cis-12 conjugated linoleic acid (10,12 CLA) promotes fat loss in obese mice and humans, but has also been associated with insulin resistance. OBJECTIVE: We therefore sought to directly compare weight loss by 10,12 CLA versus caloric restriction (CR, 15-25%), an acceptable healthy method of weight loss, to determine how 10,12 CLA-mediated weight loss fails to improve glucose metabolism. METHODS: Obese mice with characteristics of human metabolic syndrome were either supplemented with 10,12 CLA or subjected to CR to promote weight loss. Metabolic endpoints such as energy expenditure, glucose and insulin tolerance testing, and trunk fat distribution were measured. RESULTS: By design, 10,12 CLA and CR caused equivalent weight loss, with greater fat loss by 10,12 CLA accompanied by increased energy expenditure, reduced respiratory quotient, increased fat oxidation, accumulation of alternatively activated macrophages, and browning of subcutaneous white adipose tissue (WAT). Moreover, 10,12 CLA-supplemented mice better defended their body temperature against a cold challenge. However, 10,12 CLA concurrently induced the detrimental loss of subcutaneous WAT without reducing visceral WAT, promoted reduced plasma and WAT adipokine levels, worsened hepatic steatosis, and failed to improve glucose metabolism. Obese mice undergoing CR were protected from subcutaneous-specific fat loss, had improved hepatic steatosis, and subsequently showed the expected improvements in WAT adipokines, glucose metabolism and WAT inflammation. CONCLUSIONS: These results suggest that 10,12 CLA mediates the preferential loss of subcutaneous fat that likely contributes to hepatic steatosis and maintained insulin resistance, despite significant weight loss and WAT browning in mice. Collectively, we have shown that weight loss due to 10,12 CLA supplementation or CR results in dramatically different metabolic phenotypes, with the latter promoting a healthier form of weight loss.

Physiological role for leptin in the control of thermal conductance.

OBJECTIVE: To investigate the role played by leptin in thermoregulation, we studied the effects of physiological leptin replacement in leptin-deficient ob/ob mice on determinants of energy balance, thermogenesis and heat retention under 3 different ambient temperatures. METHODS: The effects of housing at 14 °C, 22 °C or 30 °C on core temperature (telemetry), energy expenditure (respirometry), thermal conductance, body composition, energy intake, and locomotor activity (beam breaks) were measured in ob/ob mice implanted subcutaneously with osmotic minipumps at a dose designed to deliver a physiological replacement dose of leptin or its vehicle-control. RESULTS: As expected, the hypothermic phenotype of ob/ob mice was partially rescued by administration of leptin at a dose that restores plasma levels into the physiological range. This effect of leptin was not due to increased energy expenditure, as cold exposure markedly and equivalently stimulated energy expenditure and induced activation of brown adipose tissue irrespective of leptin treatment. Instead, the effect of physiological leptin replacement to raise core body temperature of cold-exposed ob/ob mice was associated with reduced thermal conductance, implying a physiological role for leptin in heat conservation. Finally, both leptin- and vehicle-treated ob/ob mice failed to match energy intake to expenditure during cold exposure, resulting in weight loss. CONCLUSIONS: The physiological effect of leptin to reduce thermal conductance contributes to maintenance of core body temperature under sub-thermoneutral conditions.

Chronic CNS oxytocin signaling preferentially induces fat loss in high-fat diet-fed rats by enhancing satiety responses and increasing lipid utilization.

Based largely on a number of short-term administration studies, growing evidence suggests that central oxytocin is important in the regulation of energy balance. The goal of the current work is to determine whether long-term third ventricular (3V) infusion of oxytocin into the central nervous system (CNS) is effective for obesity prevention and/or treatment in rat models. We found that chronic 3V oxytocin infusion between 21 and 26 days by osmotic minipumps both reduced weight gain associated with the progression of high-fat diet (HFD)-induced obesity and elicited a sustained reduction of fat mass with no decrease of lean mass in rats with established diet-induced obesity. We further demonstrated that these chronic oxytocin effects result from 1) maintenance of energy expenditure at preintervention levels despite ongoing weight loss, 2) a reduction in respiratory quotient, consistent with increased fat oxidation, and 3) an enhanced satiety response to cholecystokinin-8 and associated decrease of meal size. These weight-reducing effects persisted for approximately 10 days after termination of 3V oxytocin administration and occurred independently of whether sucrose was added to the HFD. We conclude that long-term 3V administration of oxytocin to rats can both prevent and treat diet-induced obesity.

In vivo structure-function studies of human hepatic lipase: the catalytic function rescues the lean phenotype of HL-deficient (hl-/-) mice.

The lean body weight phenotype of hepatic lipase (HL)-deficient mice (hl(-/-)) suggests that HL is required for normal weight gain, but the underlying mechanisms are unknown. HL plays a unique role in lipoprotein metabolism performing bridging as well as catalytic functions, either of which could participate in energy homeostasis. To determine if both the catalytic and bridging functions or the catalytic function alone are required for the effect of HL on body weight, we studied (hl(-/-)) mice that transgenically express physiologic levels of human (h)HL (with catalytic and bridging functions) or a catalytically-inactive (ci)HL variant (with bridging function only) in which the catalytic Serine 145 was mutated to Alanine. As expected, HL activity in postheparin plasma was restored to physiologic levels only in hHL-transgenic mice (hl(-/-)hHL). During high-fat diet feeding, hHL-transgenic mice exhibited increased body weight gain and body adiposity relative to hl(-/-)ciHL mice. A similar, albeit less robust effect was observed in female hHL-transgenic relative to hl(-/-)ciHL mice. To delineate the basis for this effect, we determined cumulative food intake and measured energy expenditure using calorimetry. Interestingly, in both genders, food intake was 5-10% higher in hl(-/-)hHL mice relative to hl(-/-)ciHL controls. Similarly, energy expenditure was ~10% lower in HL-transgenic mice after adjusting for differences in total body weight. Our results demonstrate that (1) the catalytic function of HL is required to rescue the lean body weight phenotype of hl(-/-) mice; (2) this effect involves complementary changes in both sides of the energy balance equation; and (3) the bridging function alone is insufficient to rescue the lean phenotype of hl(-/-)ciHL mice.

Leptin signaling is required for adaptive changes in food intake, but not energy expenditure, in response to different thermal conditions.

Survival of free-living animals depends on the ability to maintain core body temperature in the face of rapid and dramatic changes in their thermal environment. If food intake is not adjusted to meet the changing energy demands associated with changes of ambient temperature, a serious challenge to body energy stores can occur. To more fully understand the coupling of thermoregulation to energy homeostasis in normal animals and to investigate the role of the adipose hormone leptin to this process, comprehensive measures of energy homeostasis and core temperature were obtained in leptin-deficient ob/ob mice and their wild-type (WT) littermate controls when housed under cool (14°C), usual (22°C) or ∼ thermoneutral (30°C) conditions. Our findings extend previous evidence that WT mice robustly defend normothermia in response to either a lowering (14°C) or an increase (30°C) of ambient temperature without changes in body weight or body composition. In contrast, leptin-deficient, ob/ob mice fail to defend normothermia at ambient temperatures lower than thermoneutrality and exhibit marked losses of both body fat and lean mass when exposed to cooler environments (14°C). Our findings further demonstrate a strong inverse relationship between ambient temperature and energy expenditure in WT mice, a relationship that is preserved in ob/ob mice. However, thermal conductance analysis indicates defective heat retention in ob/ob mice, irrespective of temperature. While a negative relationship between ambient temperature and energy intake also exists in WT mice, this relationship is disrupted in ob/ob mice. Thus, to meet the thermoregulatory demands of different ambient temperatures, leptin signaling is required for adaptive changes in both energy intake and thermal conductance. A better understanding of the mechanisms coupling thermoregulation to energy homeostasis may lead to the development of new approaches for the treatment of obesity.

Genetic determinants of atherosclerosis, obesity, and energy balance in consomic mice.

Metabolic diseases such as obesity and atherosclerosis result from complex interactions between environmental factors and genetic variants. A panel of chromosome substitution strains (CSSs) was developed to characterize genetic and dietary factors contributing to metabolic diseases and other biological traits and biomedical conditions. Our goal here was to identify quantitative trait loci (QTLs) contributing to obesity, energy expenditure, and atherosclerosis. Parental strains C57BL/6 and A/J together with a panel of 21 CSSs derived from these progenitors were subjected to chronic feeding of rodent chow and atherosclerotic (females) or diabetogenic (males) test diets, and evaluated for a variety of metabolic phenotypes including several traits unique to this report, namely fat pad weights, energy balance, and atherosclerosis. A total of 297 QTLs across 35 traits were discovered, two of which provided significant protection from atherosclerosis, and several dozen QTLs modulated body weight, body composition, and circulating lipid levels in females and males. While several QTLs confirmed previous reports, most QTLs were novel. Finally, we applied the CSS quantitative genetic approach to energy balance, and identified three novel QTLs controlling energy expenditure and one QTL modulating food intake. Overall, we identified many new QTLs and phenotyped several novel traits in this mouse model of diet-induced metabolic diseases.

In uncontrolled diabetes, thyroid hormone and sympathetic activators induce thermogenesis without increasing glucose uptake in brown adipose tissue.

Recent advances in human brown adipose tissue (BAT) imaging technology have renewed interest in the identification of BAT activators for the treatment of obesity and diabetes. In uncontrolled diabetes (uDM), activation of BAT is implicated in glucose lowering mediated by intracerebroventricular (icv) administration of leptin, which normalizes blood glucose levels in streptozotocin (STZ)-induced diabetic rats. The potent effect of icv leptin to increase BAT glucose uptake in STZ-diabetes is accompanied by the return of reduced plasma thyroxine (T4) levels and BAT uncoupling protein-1 (Ucp1) mRNA levels to nondiabetic controls. We therefore sought to determine whether activation of thyroid hormone receptors is sufficient in and of itself to lower blood glucose levels in STZ-diabetes and whether this effect involves activation of BAT. We found that, although systemic administration of the thyroid hormone (TR)ß-selective agonist GC-1 increases energy expenditure and induces further weight loss in STZ-diabetic rats, it neither increased BAT glucose uptake nor attenuated diabetic hyperglycemia. Even when GC-1 was administered in combination with a ß(3)-adrenergic receptor agonist to mimic sympathetic nervous system activation, glucose uptake was not increased in STZ-diabetic rats, nor was blood glucose lowered, yet this intervention potently activated BAT. Similar results were observed in animals treated with active thyroid hormone (T3) instead of GC-1. Taken together, our data suggest that neither returning normal plasma thyroid hormone levels nor BAT activation has any impact on diabetic hyperglycemia, and that in BAT, increases of Ucp1 gene expression and glucose uptake are readily dissociated from one another in this setting.

VASP increases hepatic fatty acid oxidation by activating AMPK in mice.

Activation of AMP-activated protein kinase (AMPK) signaling reduces hepatic steatosis and hepatic insulin resistance; however, its regulatory mechanisms are not fully understood. In this study, we sought to determine whether vasodilator-stimulated phosphoprotein (VASP) signaling improves lipid metabolism in the liver and, if so, whether VASP's effects are mediated by AMPK. We show that disruption of VASP results in significant hepatic steatosis as a result of significant impairment of fatty acid oxidation, VLDL-triglyceride (TG) secretion, and AMPK signaling. Overexpression of VASP in hepatocytes increased AMPK phosphorylation and fatty acid oxidation and reduced hepatocyte TG accumulation; however, these responses were suppressed in the presence of an AMPK inhibitor. Restoration of AMPK phosphorylation by administration of 5-aminoimidazole-4-carboxamide riboside in Vasp(-/-) mice reduced hepatic steatosis and normalized fatty acid oxidation and VLDL-TG secretion. Activation of VASP by the phosphodiesterase-5 inhibitor, sildenafil, in db/db mice reduced hepatic steatosis and increased phosphorylated (p-)AMPK and p-acetyl CoA carboxylase. In Vasp(-/-) mice, however, sildendafil treatment did not increase p-AMPK or reduce hepatic TG content. These studies identify a role of VASP to enhance hepatic fatty acid oxidation by activating AMPK and to promote VLDL-TG secretion from the liver.

Acutely decreased thermoregulatory energy expenditure or decreased activity energy expenditure both acutely reduce food intake in mice.

Despite the suggestion that reduced energy expenditure may be a key contributor to the obesity pandemic, few studies have tested whether acutely reduced energy expenditure is associated with a compensatory reduction in food intake. The homeostatic mechanisms that control food intake and energy expenditure remain controversial and are thought to act over days to weeks. We evaluated food intake in mice using two models of acutely decreased energy expenditure: 1) increasing ambient temperature to thermoneutrality in mice acclimated to standard laboratory temperature or 2) exercise cessation in mice accustomed to wheel running. Increasing ambient temperature (from 21 °C to 28 °C) rapidly decreased energy expenditure, demonstrating that thermoregulatory energy expenditure contributes to both light cycle (40 ± 1%) and dark cycle energy expenditure (15 ± 3%) at normal ambient temperature (21 °C). Reducing thermoregulatory energy expenditure acutely decreased food intake primarily during the light cycle (65 ± 7%), thus conflicting with the delayed compensation model, but did not alter spontaneous activity. Acute exercise cessation decreased energy expenditure only during the dark cycle (14 ± 2% at 21 °C; 21 ± 4% at 28 °C), while food intake was reduced during the dark cycle (0.9 ± 0.1 g) in mice housed at 28 °C, but during the light cycle (0.3 ± 0.1 g) in mice housed at 21 °C. Cumulatively, there was a strong correlation between the change in daily energy expenditure and the change in daily food intake (R(2) = 0.51, p<0.01). We conclude that acutely decreased energy expenditure decreases food intake suggesting that energy intake is regulated by metabolic signals that respond rapidly and accurately to reduced energy expenditure.

Peripheral oxytocin suppresses food intake and causes weight loss in diet-induced obese rats.

Growing evidence suggests that oxytocin plays an important role in the regulation of energy balance and that central oxytocin administration induces weight loss in diet-induced obese (DIO) animals. To gain a better understanding of how oxytocin mediates these effects, we examined feeding and neuronal responses to oxytocin in animals rendered obese following exposure to either a high-fat (HFD) or low-fat diet (LFD). Our findings demonstrate that peripheral administration of oxytocin dose-dependently reduces food intake and body weight to a similar extent in rats maintained on either diet. Moreover, the effect of oxytocin to induce weight loss remained intact in leptin receptor-deficient Koletsky (fa(k)/fa(k)) rats relative to their lean littermates. To determine whether systemically administered oxytocin activates hindbrain areas that regulate meal size, we measured neuronal c-Fos induction in the nucleus of the solitary tract (NTS) and area postrema (AP). We observed a robust neuronal response to oxytocin in these hindbrain areas that was unexpectedly increased in rats rendered obese on a HFD relative to lean, LFD-fed controls. Finally, we report that repeated daily peripheral administration of oxytocin in DIO animals elicited a sustained reduction of food intake and body weight while preventing the reduction of energy expenditure characteristic of weight-reduced animals. These findings extend recent evidence suggesting that oxytocin circumvents leptin resistance and induces weight-loss in DIO animals through a mechanism involving activation of neurons in the NTS and AP, key hindbrain areas for processing satiety-related inputs.

A novel rodent model that mimics the metabolic sequelae of obese craniopharyngioma patients.

Patients with craniopharyngioma (CP), a tumor located in the pituitary and/or hypothalamus, are susceptible to developing obesity and many metabolic complications. The study aim was to create a rodent model that mimics the complex neuroanatomical and metabolic disturbances commonly seen in obese CP patients. We compared the metabolic phenotype of animals with three distinct types of hypothalamic lesions: 1) destruction of the arcuate nucleus (ARC) induced by monosodium glutamate (MSG), 2) electrolytic lesion of the adjacent ventromedial nucleus (VMN) alone, 3) both the VMN and dorsomedial nucleus (DMN), or a 4) combined medial hypothalamic lesion (CMHL) affecting the VMN, DMN, and the ARC. Only the CMHL model exhibited all key features observed in patients with hypothalamic obesity induced by CP. These features included excessive weight gain due to increased adiposity, increased food intake, and pronounced hyperinsulinemia and hyperleptinemia. Similar to characteristics of patients with CP, CMHL animals exhibited reduced plasma levels of alpha-melanocyte stimulating hormone and reduced ambulatory activity compared with weight-matched controls. Therefore, the CMHL model best mimics the complex metabolic abnormalities observed in obese CP patients compared with lesions to other hypothalamic areas and provides a foundation for future pharmacological approaches to treat obesity in children with hypothalamic damage.

Increased energy expenditure and leptin sensitivity account for low fat mass in myostatin-deficient mice.

Myostatin deficiency causes dramatically increased skeletal muscle mass and reduced fat mass. Previously, myostatin-deficient mice were reported to have unexpectedly low total energy expenditure (EE) after normalizing to body mass, and thus, a metabolic cause for low fat mass was discounted. To clarify how myostatin deficiency affects the control of body fat mass and energy balance, we compared rates of oxygen consumption, body composition, and food intake in young myostatin-deficient mice relative to wild-type (WT) and heterozygous (HET) controls. We report that after adjusting for total body mass using regression analysis, young myostatin-deficient mice display significantly increased EE relative to both WT (+0.81 ± 0.28 kcal/day, P = 0.004) and HET controls (+0.92 ± 0.31 kcal/day, P = 0.005). Since food intake was not different between groups, increased EE likely accounts for the reduced body fat mass (KO: 8.8 ± 1.1% vs. WT: 14.5 ± 1.3%, P = 0.003) and circulating leptin levels (KO: 0.7 ± 0.2 ng/ml vs. WT: 1.9 ± 0.3 ng/ml, P = 0.008). Interestingly, the observed increase in adjusted EE in myostatin-deficient mice occurred despite dramatically reduced ambulatory activity levels (-50% vs. WT, P < 0.05). The absence of hyperphagia together with increased EE in myostatin-deficient mice suggests that increased leptin sensitivity may contribute to their lean phenotype. Indeed, leptin-induced anorexia (KO: -17 ± 1.2% vs. WT: -5 ± 0.3%) and weight loss (KO: -2.2 ± 0.2 g vs. WT: -1.6 ± 0.1, P < 0.05) were increased in myostatin-deficient mice compared with WT controls. We conclude that increased EE, together with increased leptin sensitivity, contributes to low fat mass in mice lacking myostatin.

Identification of a physiological role for leptin in the regulation of ambulatory activity and wheel running in mice.

Mechanisms regulating spontaneous physical activity remain poorly characterized despite evidence of influential genetic and acquired factors. We evaluated ambulatory activity and wheel running in leptin-deficient ob/ob mice and in wild-type mice rendered hypoleptinemic by fasting in both the presence and absence of subcutaneous leptin administration. In ob/ob mice, leptin treatment to plasma levels characteristic of wild-type mice acutely increased both ambulatory activity (by 4,000 ± 200 beam breaks/dark cycle, P < 0.05) and total energy expenditure (TEE; by 0.11 ± 0.01 kcal/h during the dark cycle, P < 0.05) in a dose-dependent manner and acutely increased wheel running (+350%, P < 0.05). Fasting potently increased ambulatory activity and wheel running in wild-type mice (AA: +25%, P < 0.05; wheel running: +80%, P < 0.05), and the effect of fasting was more pronounced in ob/ob mice (AA: +400%, P < 0.05; wheel running: +1,600%, P < 0.05). However, unlike what occurred in ad libitum-fed ob/ob mice, physiological leptin replacement attenuated or prevented fasting-induced increases of ambulatory activity and wheel running in both wild-type and ob/ob mice. Thus, plasma leptin is a physiological regulator of spontaneous physical activity, but the nature of leptin's effect on activity is dependent on food availability.