Leptin action is modified by an interaction between dietary fat content and ambient temperature

Mice adapted to a high-fat diet are reported to be leptin resistant; however, we previously reported that mice fed a high-fat (HF) diet and housed at 23 degrees C remained sensitive to peripheral leptin and specifically lost body fat. This study tested whether leptin action was impaired by a combination of elevated environmental temperature and a HF diet. Male C57BL/6 mice were adapted to low-fat (LF) or HF diet from 10 days of age and were housed at 27 degrees C from 28 days of age. From 35 days of age, baseline food intake and body weight were recorded for 1 wk and then mice on each diet were infused with 10 microg leptin/day or PBS from an intraperitoneal miniosmotic pump for 13 days. HF-fed mice had a higher energy intake than LF-fed mice and were heavier but not fatter. Serum leptin was lower in PBS-infused HF- than LF-fed mice. Leptin significantly inhibited energy intake of both LF-fed and HF-fed mice, and this was associated with a significant increase in hypothalamic long-form leptin receptors with no change in short-form leptin receptor or brown fat uncoupling protein-1 mRNA expression. Leptin significantly inhibited weight gain in both LF- and HF-fed mice but reduced the percentage of body fat mass only in LF-fed mice. The percentage of lean and fat tissue in HF-fed mice did not change, implying that overall growth had been inhibited. These results suggest that dietary fat modifies the mechanisms responsible for leptin-induced changes in body fat content and that those in HF-fed mice are sensitive to environmental temperature.     

温度和高脂食物对黑线仓鼠代谢产热和体脂累积的影响

能量代谢的适应性调节是小型哺乳动物应对环境季节性变化的主要策略之一。为探讨不同温度下动物在代谢产热能量支出与脂肪累积之间的权衡策略,以成年雄性黑线仓鼠为研究对象开展了3 个实验:实验1 将动物驯化于高脂和低脂食物;实验2 将动物暴露于低温(5℃)和暖温(30℃);实验3 将饲喂高脂食物的动物暴露于低温。以食物平衡法测定摄食量、摄入能和消化率,以开放式氧气分析仪测定代谢产热,以索氏抽提法测定脂肪含量。结果发现,取食高脂食物的黑线仓鼠摄食量显著减少,但脂肪累积显著增加;暖温下摄食量显著减少,但体脂含量显著增加,低温下摄食量显著升高,但体脂含量显著减少;饲喂高脂食物的黑线仓鼠在低温下摄入能显著增加,非颤抖性产热增强,但体脂含量显著降低。结果表明高脂食物对黑线仓鼠体脂累积的影响与环境温度有关,低温诱导脂肪动员,暖温促进脂肪贮存;低温下黑线仓鼠增加能量摄入不能完全补偿用于产热的能量支出,导致脂肪动员增加;暖温下代谢产热降低是脂肪累积的主要因素;与能量摄入相比代谢产热的能量支出在体脂累积的适应性变化中发挥更重要的作用。     

Effects of dietary fat and zinc on adiposity,serum leptin and adipose fatty acid composition in C57BL/6J mice

Zinc (Zn) has been implicated in altered adipose metabolism, insulin resistance and obesity. The objective of this study was to investigate the effects dietary Zn deficiency and supplementation on adiposity, serum leptin and fatty acid composition of adipose triglycerides and phospholipid in C57BL/6J mice fed low-fat (LF) or high-fat (HF) diets for a 16 week period. Weanling C57BL/6J mice were fed LF (16% kcal from soybean oil) or HF (39% kcal from lard and 16% kcal from soybean oil) diets containing 3, 30 or 150 mg Zn/kg diet (ZD = Zn-deficient, ZC = Zn control and ZS = Zn-supplemented, respectively). HF-fed mice had higher fat pad weights and lower adipose Zn concentrations than the LF-fed mice. The ZD and ZS groups had a reduced content of fatty acids in adipose triglycerides compared to the ZC group, suggesting that zinc status may influence fatty acid accumulation in adipose tissue. Serum leptin concentration was positively correlated with body weight and body fat, and negatively correlated with adipose Zn concentration. Dietary fat, but not dietary Zn, altered the fatty acid composition of adipose tissue phospholipid and triglyceride despite differences in Zn status assessed by femur Zn concentrations. The fatty acid profile of adipose triglycerides generally reflected the diets. HF-fed mice had a higher percentage of C20:4 n-6, elevated ratio of n-6/n-3, lower ratio of PUFA/SAT and reduced percentage of total n-3 fatty acids in adipose phospholipid, a fatty acid profile associated with obesity-induced risks for insulin resistance and impaired glucose transport. In summary, the reduced adipose Zn concentrations in HF-fed mice and the negative correlation between serum leptin and adipose Zn concentrations support an interrelationship among obesity, leptin and Zn metabolism.     

Induction of muscle thermogenesis by high-fat diet in mice: association with obesity-resistance

The obesogenic effect of a high-fat (HF) diet is counterbalanced by stimulation of energy expenditure and lipid oxidation in response to a meal. The aim of this study was to reveal whether muscle nonshivering thermogenesis could be stimulated by a HF diet, especially in obesity-resistant A/J compared with obesity-prone C57BL/6J (B/6J) mice. Experiments were performed on male mice born and maintained at 30 degrees C. Four-week-old mice were randomly weaned onto a low-fat (LF) or HF diet for 2 wk. In the A/J LF mice, cold exposure (4 degrees C) resulted in hypothermia, whereas the A/J HF, B/6J LF, and B/6J HF mice were cold tolerant. Cold sensitivity of the A/J LF mice was associated with a relatively low whole body energy expenditure under resting conditions, which was normalized by the HF diet. In both strains, the HF diet induced uncoupling protein-1-mediated thermogenesis, with a stronger induction in A/J mice. Only in A/J mice: 1) the HF diet augmented activation of whole body lipid oxidation by cold; and 2) at 30 degrees C, oxygen consumption, total content, and phosphorylation of AMP-activated protein kinase (AMPK), and AICAR-stimulated palmitate oxidation in soleus muscle was increased by the HF diet in parallel with significantly increased leptinemia. Gene expression data in soleus muscle of the A/J HF mice indicated a shift from carbohydrate to fatty acid oxidation. Our results suggest a role for muscle nonshivering thermogenesis and lipid oxidation in the obesity-resistant phenotype of A/J mice and indicate that a HF diet could induce thermogenesis in oxidative muscle, possibly via the leptin-AMPK axis.     

Deficiency of PTP1B in POMC neurons leads to alterations in energy balance and homeostatic response to cold exposure

The adipose tissue-derived hormone leptin regulates energy balance through catabolic effects on central circuits, including proopiomelanocortin (POMC) neurons. Leptin activation of POMC neurons increases thermogenesis and locomotor activity. Protein tyrosine phosphatase 1B (PTP1B) is an important negative regulator of leptin signaling. POMC neuron-specific deletion of PTP1B in mice results in reduced high-fat diet-induced body weight and adiposity gain due to increased energy expenditure and greater leptin sensitivity. Mice lacking the leptin gene (ob/ob mice) are hypothermic and cold intolerant, whereas leptin delivery to ob/ob mice induces thermogenesis via increased sympathetic activity to brown adipose tissue (BAT). Here, we examined whether POMC PTP1B mediates the thermoregulatory response of CNS leptin signaling by evaluating food intake, body weight, core temperature (T(C)), and spontaneous physical activity (SPA) in response to either exogenous leptin or 4-day cold exposure (4°C) in male POMC-Ptp1b-deficient mice compared with wild-type controls. POMC-Ptp1b(-/-) mice were hypersensitive to leptin-induced food intake and body weight suppression compared with wild types, yet they displayed similar leptin-induced increases in T(C). Interestingly, POMC-Ptp1b(-/-) mice had increased BAT weight and elevated plasma triiodothyronine (T(3)) levels in response to a 4-day cold challenge, as well as reduced SPA 24 h after cold exposure, relative to controls. These data show that PTP1B in POMC neurons plays a role in short-term cold-induced reduction of SPA and may influence cold-induced thermogenesis via enhanced activation of the thyroid axis.     

Serum leptin, energy budget, and thermogenesis in striped hamsters exposed to consecutive decreases in ambient temperatures

Leptin has been found to be a direct participant in the regulation of both energy intake and energy expenditure in small mammals showing seasonal declines in body mass (M(b)) and fat mass, but its roles in an animal exhibiting seasonally increased thermogenesis and unchanged M(b) remain unclear. Serum leptin levels, energy budget, and thermogenesis were measured in striped hamsters exposed to consecutive decreases in ambient temperatures ranging from 23° to -23°C. Cold-exposed hamsters had significant increases in gross energy intake (GEI), the rate of basal metabolism, nonshivering thermogenesis, and activity of cytochrome c oxidase (COX) in brown adipose tissue (BAT), compared with control hamsters, indicating a cold-induced elevation of thermogenesis. Body mass and fat content were decreased in cold-exposed animals, and serum leptin levels were increased in hamsters exposed to temperatures of -8°C and below in inverse proportion to body fat content. Serum leptin levels were positively correlated with GEI and BAT COX activity in cold-exposed hamsters, but no such relationships were observed in control animals. These findings suggest that cold-exposed hamsters increase food consumption to meet the energy requirements for increased BAT thermogenesis. The increases in serum leptin levels are likely involved in increased thermogenesis in hamsters under cold stress. Cold-exposed hamsters may become leptin resistant, which is associated with impaired regulation of food intake. This new natural model of leptin resistance may also provide insight into the dynamic long-term control of energy homeostasis for animals that do not exhibit seasonal decline in M(b).     

Differences in response to corticotropin-releasing factor after short- and long-term consumption of a high-fat diet

We previously reported an exaggerated endocrine and weight loss response to stress in rats fed a high-fat (HF) diet for 5 days. Others report blunted stress-induced anxiety in rats made obese on a HF diet. Experiments described here tested whether sensitivity to stress-related peptides was changed in obese and nonobese HF-fed rats. Third ventricle infusion of corticotropin-releasing factor (CRF) in rats made obese on HF diet (40% kcal fat) produced an exaggerated hypophagia, which is thought to be mediated by CRF(2) receptors. Obese rats responded to a lower dose of CRF for a longer time than rats fed a low-fat (LF) diet (12% kcal fat). CRF-induced release of corticosterone, which is thought to be mediated by CRF(1) receptors, was not exaggerated in obese HF-fed rats. In contrast, rats fed HF diet for 5 days showed the same food intake and corticosterone response to CRF as LF-fed rats. CRF mRNA expression in the paraventricular nucleus of the hypothalamus was stimulated by mild stress (ip saline injection and placement in a novel cage) in LF-fed rats but not in rats fed HF diet for 5 days because of a nonsignificant increase in expression in nonstressed HF-fed rats. In addition, nonstressed levels of urocortin (UCN) I mRNA expression in the Edinger-Westphal nucleus were significantly inhibited in HF-fed rats. These data suggest that rats that have become obese on a HF diet show a change in responsiveness to stress peptides, whereas the increased stress response in nonobese HF-fed rats may be associated with changes in basal CRF and UCN I mRNA expression.     

Overexpression of constitutively active PKG-I protects female, but not male mice from diet-induced obesity

Cyclic guanosine monophosphate (cGMP)-dependent protein kinase I (PKG-I) is a multifunctional protein. The direct effects of PKG-I activation on energy homeostasis and obesity development are not well understood. Herein, we generated transgenic mice with expression of the constitutively active PKG-I in adipose tissue as well as in other tissues. Male and female PKG-I overexpressing mice were fed a low-fat (LF) or high-fat (HF) diet for 16 weeks. HF-fed female PKG-I transgenic mice had decreased body weight gain, lower percentage of body fat, and improved glucose tolerance compared to HF-fed wild-type (WT) controls. In contrast, male transgenic PKG-I mice were not resistant to the development of HF-diet-induced obesity, and exhibited similar levels of adiposity and glucose intolerance as HF-fed WT controls. Furthermore, we found that HF-fed female transgenic PKG-I mice had increased energy expenditure and cold-induced adaptive thermogenesis compared to HF-fed WT controls, which was associated with increased expression of uncoupling protein-1 (UCP1) in brown adipose tissue (BAT). In addition, the rates of lipolysis in white adipose tissue (WAT) were also increased in female transgenic PKG-I mice compared to WT controls due to increased phosphorylation of hormone-sensitive lipase (HSL). However, in male mice, adaptive thermogenesis or WAT lipolysis was similar between transgenic PKG-I mice and WT controls. Together, these data demonstrate sex differences in effects of PKG-I activation on the regulation of adipose tissue function and its contribution to diet induced obesity.     

Photoperiodic regulation in energy intake, thermogenesis and body mass in root voles (Microtus oeconomus)

The present study was designed to examine whether photoperiod alone was effective to induce seasonal regulations in physiology in root voles (Microtus oeconomus) from the Qinghai-Tibetan plateau noted for its extreme cold environment. Root voles were randomly assigned into either long photoperiod (LD; 16L:8D) or short photoperiod (SD; 8L:16D) for 4 weeks at constant temperature (20 degrees C). At the end of acclimation, SD voles showed lower body mass and body fat coupled with higher energy intake than LD voles. SD greatly enhanced thermogenic capacities in root voles, as indicated by elevated basal metabolic rate (BMR), nonshivering thermogenesis (NST), mitochondrial protein content and uncoupling protein-1 (UCP1) content in brown adipose tissue (BAT). Although no variations in serum leptin levels were found between SD and LD voles, serum leptin levels were positively correlated with body mass and body fat mass, and negatively correlated with energy intake and UCP1 content in BAT, respectively. To summarize, SD alone is effective in inducing higher thermogenic capacities and energy intake coupled with lower body mass and body fat mass in root voles. Leptin is potentially involved in the photoperiod induced body mass regulation and thermogenesis in root voles.     

Method of leptin dosing,strain, and group housing influence leptin sensitivity in high-fat-fed weanling mice

High-fat diets are reported to induce resistance to peripherally administered leptin. In an attempt to develop a model of juvenile diet-induced obesity, mice were weaned onto high-fat diet. Male and female, 35-day-old, C57BL/6J high-fat (45% kcal fat) diet-fed mice housed individually on grid floors did not decrease food intake or body weight in response to intraperitoneal (30 microg), lateral ventricle (5 microg), or third ventricle (0.5 microg) injections of leptin. Body weight and fat were significantly reduced by 13-day intraperitoneal infusions of 10 microg leptin/day, which doubled circulating leptin. Leptin infusion also reduced body fat in weanling, high-fat diet-fed NIH Swiss mice. Group housing mice on bedding prevented loss of fat in high-fat diet-fed male and female NIH Swiss and female C57BL/6J mice. These results indicate that peripherally infused leptin reduces fat in part by increasing thermogenesis and that inhibition of food intake in high-fat diet-fed mice requires either chronic activation of central leptin receptors or is independent of receptors that inhibit feeding in response to an acute central injection of leptin.     

Photoperiod and temperature can regulate body mass, serum leptin concentration, and uncoupling protein 1 in Brandt's voles (Lasiopodomys brandtii) and Mongolian gerbils (Meriones unguiculatus)

Environmental factors play an important role in the seasonal adaptation of body mass and thermogenesis in wild small mammals. In this study, we performed a factorial experiment (temperature x photoperiod) in which Brandt's voles and Mongolian gerbils were acclimated to different photoperiods (long photoperiod, 16L : 8D; short photoperiod, 8L : 16D) and temperatures (warm, 23 degrees C; cold, 5 degrees C) to test the hypothesis that photoperiod, temperature, or both together can trigger seasonal changes in serum leptin level, body mass, thermogenesis, and energy intake. Our data demonstrate that Brandt's voles showed a remarkable decrease in body mass in both the cold and a short photoperiod. However, no significant changes in body mass were found for gerbils exposed to similar conditions. The short photoperiod induced a decrease in serum leptin levels for both voles and gerbils that might contribute to an increase in energy intake. Furthermore, the short photoperiod induced an increase of uncoupling protein 1 (UCP1) content for both voles and gerbils, and cold can further enhance the increase in voles. No interactions between photoperiod and temperature were detected for the two species. Brandt's voles can decrease their body mass through changes in energy intake and expenditure, while Mongolian gerbils can keep body mass relatively stable by balancing energy metabolism under winterlike conditions. Leptin was potentially involved in the regulation of body mass and thermogenic capacity for the two species.     

光周期和高脂食物对雌性高山姬鼠能量代谢和产热的影响

为了研究光周期和高脂食物对小型哺乳动物能量代谢和产热的影响,将成年雌性高山姬鼠分别驯化于长光照低脂、高脂食物和短光照低脂、高脂食物条件下,7周后测定动物的体重、能量摄入、产热、身体组成、血清瘦素浓度以及体脂含量等参数。结果发现:1)短光照抑制体重增长,降低血清瘦素浓度,增加非颤抖性产热;2)高脂食物使摄入能减少,消化率和体脂含量提高,但未显著影响体重、基础代谢率、非颤抖性产热和血清瘦素浓度;3)血清瘦素浓度与摄入能不相关,但与体脂重量正相关。结果暗示:短光照下瘦素作用敏感性增加和产热能力增强,可能介导了抵抗高脂食物诱导的肥胖。在野外条件下,高山姬鼠能通过能量代谢和产热的适应性调节避免体重的过度增长,有利于降低捕食风险,增强生存能力。     

Divergent effects of leptin in mice susceptible or resistant to obesity.

Consumption of a high-fat diet decreases hypothalamic neuropeptide Y (NPY) and increases proopiomelanocortin (POMC) and brown adipose uncoupling protein (UCP)-1 mRNA in obesity-resistant SWR/J but not obesity-prone C57Bl/6J mice. Although leptin was elevated in both strains in response to a high-fat diet, its role in the development of diet-induced obesity has remained unclear since insulin and other factors that affect similar tissue targets are altered. Thus, we administered recombinant leptin by subcutaneous infusion to chow-fed mice to mimic the changes in plasma leptin across its broad physiologic range. We observed strain differences in responsiveness to reduced and elevated leptin levels. A reduction in leptin during fasting evoked a greater response in C57Bl/6J mice by decreasing energy expenditure and thyroxin, increasing corticosterone and stimulating food intake and weight gain during refeeding. However, C57Bl/6J mice were less responsive to an increase in leptin in the fed state. Conversely, the leptin-mediated response to fasting was blunted in SWR/J mice, whereas an increase in leptin profoundly reduced food intake and body weight in SWR/J mice fed ad libitum. Sensitivity to fasting in C57Bl/6J mice was associated with higher hypothalamic NPY mRNA and reduced POMC and UCP-1 mRNA expression, while the robust response to high leptin levels in SWR/J mice was associated with suppression of NPY mRNA. These results indicate that differences in leptin responsiveness between strains might occur centrally or peripherally, leading to alteration in the patterns of food intake, thermogenesis and energy storage.     

Npvf: Hypothalamic Biomarker of Ambient Temperature Independent of Nutritional Status

The mechanism by which mice, exposed to the cold, mobilize endogenous or exogenous fuel sources for heat production is unknown. To address this issue we carried out experiments using 3 models of obesity in mice: C57BL/6J+/+ (wild-type B6) mice with variable susceptibility to obesity in response to being fed a high-fat diet (HFD), B6. Ucp1-/- mice with variable diet-induced obesity (DIO) and a deficiency in brown fat thermogenesis and B6. Lep-/- with defects in thermogenesis, fat mobilization and hyperphagia. Mice were exposed to the cold and monitored for changes in food intake and body composition to determine their energy balance phenotype. Upon cold exposure wild-type B6 and Ucp1-/- mice with diet-induced obesity burned endogenous fat in direct proportion to their fat reserves and changes in food intake were inversely related to fat mass, whereas leptin-deficient and lean wild-type B6 mice fed a chow diet depended on increased food intake to fuel thermogenesis. Analysis of gene expression in the hypothalamus to uncover a central regulatory mechanism revealed suppression of the Npvf gene in a manner that depends on the reduced ambient temperature and degree of exposure to the cold, but not on adiposity, leptin levels, food intake or functional brown fat.     

Insulin-like growth factor I mediates high-fat diet-induced adipogenesis in Osborne-Mendel rats

To determine if high-fat (HF) diet-induced changes in adipose tissue cellularity are associated with the presence of paracrine growth factor(s) that alter preadipocyte proliferation, Osborne-Mendel rats were fed either a HF (76% energy) or a low-fat (LF, 12% energy) diet for 85 days. HF-fed rats had greater (P < 0.05) fat pad size, total fat cell number, number of small (30-70 microm) and large (80-140 microm) adipocytes, and percentage of 100- to 140-microm adipocytes compared with LF-fed rats. Preadipocytes in primary cell culture treated with inguinal adipose tissue conditioned medium (ATCM) prepared from HF-fed rats had greater (P < 0.05) proliferation compared with cultures treated with ATCM from LF-fed rats. Proliferative capacity of ATCM prepared from HF-fed rats was attenuated after the stripping of the medium of insulin-like growth factor I using an immunomagnetic bead separation system. These data are consistent with the concept that insulin-like growth factor I is involved in the paracrine regulation of adipogenesis.     

Effects of diet quality on energy budgets and thermogenesis in Brandt's voles

Food quality and availability play an important role in an animal's life history. The aim of this study was to examine the effect of diet quality [high-fiber diet (HF) or low-fiber diet (LF)] on energy budgets and thermogenesis in Brandt's voles (Lasiopodomys (Microtus) brandtii). Dry matter intake and gross energy intake increased and digestibility decreased in HF voles compared with LF voles, while the digestible energy intake was similar for both HF and LF voles. Nonshivering thermogenesis (NST) decreased in HF voles, while LF voles kept stable; no significant differences were detected in basal metabolic rate (BMR), BAT uncoupling protein 1 (UCP1) content and the levels of serum thyroid hormones (T3 and T4) between HF and LF voles. Although there were no differences in body fat content and serum leptin concentrations between HF and LF voles, serum leptin concentrations in HF voles were reduced to nearly half as those seen in LF voles after 4-weeks acclimation. These results support the hypothesis that Brandt's voles can compensate the poor quality diet physiologically by the means of increasing food intake and decreasing thermogenesis.     

Functional evolution of leptin of Ochotona curzoniae in adaptive thermogenesis driven by cold environmental stress

Background

Environmental stress can accelerate the directional selection and evolutionary rate of specific stress-response proteins to bring about new or altered functions, enhancing an organism''s fitness to challenging environments. Plateau pika (Ochotona curzoniae), an endemic and keystone species on Qinghai-Tibetan Plateau, is a high hypoxia and low temperature tolerant mammal with high resting metabolic rate and non-shivering thermogenesis to cope in this harsh plateau environment. Leptin is a key hormone related to how these animals regulate energy homeostasis. Previous molecular evolutionary analysis helped to generate the hypothesis that adaptive evolution of plateau pika leptin may be driven by cold stress.

Methodology/Principal Findings

To test the hypothesis, recombinant pika leptin was first purified. The thermogenic characteristics of C57BL/6J mice injected with pika leptin under warm (23±1°C) and cold (5±1°C) acclimation is investigated. Expression levels of genes regulating adaptive thermogenesis in brown adipose tissue and the hypothalamus are compared between pika leptin and human leptin treatment, suggesting that pika leptin has adaptively and functionally evolved. Our results show that pika leptin regulates energy homeostasis via reduced food intake and increased energy expenditure under both warm and cold conditions. Compared with human leptin, pika leptin demonstrates a superior induced capacity for adaptive thermogenesis, which is reflected in a more enhanced β-oxidation, mitochondrial biogenesis and heat production. Moreover, leptin treatment combined with cold stimulation has a significant synergistic effect on adaptive thermogenesis, more so than is observed with a single cold exposure or single leptin treatment.

Conclusions/Significance

These findings support the hypothesis that cold stress has driven the functional evolution of plateau pika leptin as an ecological adaptation to the Qinghai-Tibetan Plateau.     

Thermogenesis, energy intake and serum leptin in Apodemus chevrieri in Hengduan Mountains region during cold acclimation

Environmental factors play an important role in the seasonal adaptation of body mass and thermogenesis in small, wild mammals. The purpose of the present study was to test the hypothesis that ambient temperature was a cue to trigger the adjustments in body mass, energy intake, and serum leptin level in Apodemus chevrieri during 42 days of cold exposure. Our data demonstrated that cold acclimation induced a decrease in body mass and a significant increase in energy intake in A. chevrieri. Serum leptin levels were positively correlated with body mass and fat mass. These data suggest that A. chevrieri reduced the body mass and increased energy intake and thermogenic capacity under cold acclimation. Further, serum leptin appears to be involved in the energy intake regulation and thermoregulation.     

Effects of cold acclimation on body mass,serum leptin level,energy metabolism and thermognesis in Eothenomys miletus in Hengduan Mountains region

Eothenomys miletus is an important species inhabiting Hengduan mountains region. In order to study adaptive strategy and the role of serum leptin level in response to a 49 d cold exposure, body mass, energy intake, basal metabolic rate (BMR), nonshivering thermogenesis (NST) in E. miletus were measured. During cold exposure (5±1 ^oC), body mass decreased; serum leptin levels decreased significantly and were positively correlated with body mass and fat mass; energy intake, BMR and NST were higher at 5 °C than that of controls. These results suggest that E. miletus enhanced thermogenic capacity and increased maintenance cost during cold acclimation, resulting in increased energy intake. Serum leptin participated in the regulation of energy balance and body mass in E. miletus.