Leptin-regulated endocannabinoids are involved in maintaining food intake

Leptin is the primary signal through which the hypothalamus senses nutritional state and modulates food intake and energy balance. Leptin reduces food intake by upregulating anorexigenic (appetite-reducing) neuropeptides, such as alpha-melanocyte-stimulating hormone, and downregulating orexigenic (appetite-stimulating) factors, primarily neuropeptide Y. Genetic defects in anorexigenic signalling, such as mutations in the melanocortin-4 (ref. 5) or leptin receptors, cause obesity. However, alternative orexigenic pathways maintain food intake in mice deficient in neuropeptide Y. CB1 cannabinoid receptors and the endocannabinoids anandamide and 2-arachidonoyl glycerol are present in the hypothalamus, and marijuana and anandamide stimulate food intake. Here we show that following temporary food restriction, CB1 receptor knockout mice eat less than their wild-type littermates, and the CB1 antagonist SR141716A reduces food intake in wild-type but not knockout mice. Furthermore, defective leptin signalling is associated with elevated hypothalamic, but not cerebellar, levels of endocannabinoids in obese db/db and ob/ob mice and Zucker rats. Acute leptin treatment of normal rats and ob/ob mice reduces anandamide and 2-arachidonoyl glycerol in the hypothalamus. These findings indicate that endocannabinoids in the hypothalamus may tonically activate CB1 receptors to maintain food intake and form part of the neural circuitry regulated by leptin.     

AMP-kinase regulates food intake by responding to hormonal and nutrient signals in the hypothalamus

Obesity is an epidemic in Western society, and causes rapidly accelerating rates of type 2 diabetes and cardiovascular disease. The evolutionarily conserved serine/threonine kinase, AMP-activated protein kinase (AMPK), functions as a 'fuel gauge' to monitor cellular energy status. We investigated the potential role of AMPK in the hypothalamus in the regulation of food intake. Here we report that AMPK activity is inhibited in arcuate and paraventricular hypothalamus (PVH) by the anorexigenic hormone leptin, and in multiple hypothalamic regions by insulin, high glucose and refeeding. A melanocortin receptor agonist, a potent anorexigen, decreases AMPK activity in PVH, whereas agouti-related protein, an orexigen, increases AMPK activity. Melanocortin receptor signalling is required for leptin and refeeding effects on AMPK in PVH. Dominant negative AMPK expression in the hypothalamus is sufficient to reduce food intake and body weight, whereas constitutively active AMPK increases both. Alterations of hypothalamic AMPK activity augment changes in arcuate neuropeptide expression induced by fasting and feeding. Furthermore, inhibition of hypothalamic AMPK is necessary for leptin's effects on food intake and body weight, as constitutively active AMPK blocks these effects. Thus, hypothalamic AMPK plays a critical role in hormonal and nutrient-derived anorexigenic and orexigenic signals and in energy balance.     

Mice lacking the M3 muscarinic acetylcholine receptor are hypophagic and lean

Members of the muscarinic acetylcholine receptor family (M1-M5) have central roles in the regulation of many fundamental physiological functions. Identifying the specific receptor subtype(s) that mediate the diverse muscarinic actions of acetylcholine is of considerable therapeutic interest, but has proved difficult primarily because of a lack of subtype-selective ligands. Here we show that mice deficient in the M3 muscarinic receptor (M3R-/- mice) display a significant decrease in food intake, reduced body weight and peripheral fat deposits, and very low levels of serum leptin and insulin. Paradoxically, hypothalamic messenger RNA levels of melanin-concentrating hormone (MCH), which are normally upregulated in fasted animals leading to an increase in food intake, are significantly reduced in M3R-/- mice. Intra-cerebroventricular injection studies show that an agouti-related peptide analogue lacked orexigenic (appetite-stimulating) activity in M3R-/- mice. However, M3R-/- mice remained responsive to the orexigenic effects of MCH. Our data indicate that there may be a cholinergic pathway that involves M3-receptor-mediated facilitation of food intake at a site downstream of the hypothalamic leptin/melanocortin system and upstream of the MCH system.     

A role for ghrelin in the central regulation of feeding

Ghrelin is an acylated peptide that stimulates the release of growth hormone from the pituitary. Ghrelin-producing neurons are located in the hypothalamus, whereas ghrelin receptors are expressed in various regions of the brain, which is indicative of central-and as yet undefined-physiological functions. Here we show that ghrelin is involved in the hypothalamic regulation of energy homeostasis. Intracerebroventricular injections of ghrelin strongly stimulated feeding in rats and increased body weight gain. Ghrelin also increased feeding in rats that are genetically deficient in growth hormone. Anti-ghrelin immunoglobulin G robustly suppressed feeding. After intracerebroventricular ghrelin administration, Fos protein, a marker of neuronal activation, was found in regions of primary importance in the regulation of feeding, including neuropeptide Y6 (NPY) neurons and agouti-related protein (AGRP) neurons. Antibodies and antagonists of NPY and AGRP abolished ghrelin-induced feeding. Ghrelin augmented NPY gene expression and blocked leptin-induced feeding reduction, implying that there is a competitive interaction between ghrelin and leptin in feeding regulation. We conclude that ghrelin is a physiological mediator of feeding, and probably has a function in growth regulation by stimulating feeding and release of growth hormone.     

UCP2 mediates ghrelin's action on NPY/AgRP neurons by lowering free radicals

The gut-derived hormone ghrelin exerts its effect on the brain by regulating neuronal activity. Ghrelin-induced feeding behaviour is controlled by arcuate nucleus neurons that co-express neuropeptide Y and agouti-related protein (NPY/AgRP neurons). However, the intracellular mechanisms triggered by ghrelin to alter NPY/AgRP neuronal activity are poorly understood. Here we show that ghrelin initiates robust changes in hypothalamic mitochondrial respiration in mice that are dependent on uncoupling protein 2 (UCP2). Activation of this mitochondrial mechanism is critical for ghrelin-induced mitochondrial proliferation and electric activation of NPY/AgRP neurons, for ghrelin-triggered synaptic plasticity of pro-opiomelanocortin-expressing neurons, and for ghrelin-induced food intake. The UCP2-dependent action of ghrelin on NPY/AgRP neurons is driven by a hypothalamic fatty acid oxidation pathway involving AMPK, CPT1 and free radicals that are scavenged by UCP2. These results reveal a signalling modality connecting mitochondria-mediated effects of G-protein-coupled receptors on neuronal function and associated behaviour.     

Gut hormone PYY(3-36) physiologically inhibits food intake

Food intake is regulated by the hypothalamus, including the melanocortin and neuropeptide Y (NPY) systems in the arcuate nucleus. The NPY Y2 receptor (Y2R), a putative inhibitory presynaptic receptor, is highly expressed on NPY neurons in the arcuate nucleus, which is accessible to peripheral hormones. Peptide YY(3-36) (PYY(3-36)), a Y2R agonist, is released from the gastrointestinal tract postprandially in proportion to the calorie content of a meal. Here we show that peripheral injection of PYY(3-36) in rats inhibits food intake and reduces weight gain. PYY(3-36) also inhibits food intake in mice but not in Y2r-null mice, which suggests that the anorectic effect requires the Y2R. Peripheral administration of PYY(3-36) increases c-Fos immunoreactivity in the arcuate nucleus and decreases hypothalamic Npy messenger RNA. Intra-arcuate injection of PYY(3-36) inhibits food intake. PYY(3-36) also inhibits electrical activity of NPY nerve terminals, thus activating adjacent pro-opiomelanocortin (POMC) neurons. In humans, infusion of normal postprandial concentrations of PYY(3-36) significantly decreases appetite and reduces food intake by 33% over 24 h. Thus, postprandial elevation of PYY(3-36) may act through the arcuate nucleus Y2R to inhibit feeding in a gut-hypothalamic pathway.     

Identification of nesfatin-1 as a satiety molecule in the hypothalamus

The brain hypothalamus contains certain secreted molecules that are important in regulating feeding behaviour. Here we show that nesfatin, corresponding to NEFA/nucleobindin2 (NUCB2), a secreted protein of unknown function, is expressed in the appetite-control hypothalamic nuclei in rats. Intracerebroventricular (i.c.v.) injection of NUCB2 reduces feeding. Rat cerebrospinal fluid contains nesfatin-1, an amino-terminal fragment derived from NUCB2, and its expression is decreased in the hypothalamic paraventricular nucleus under starved conditions. I.c.v. injection of nesfatin-1 decreases food intake in a dose-dependent manner, whereas injection of an antibody neutralizing nesfatin-1 stimulates appetite. In contrast, i.c.v. injection of other possible fragments processed from NUCB2 does not promote satiety, and conversion of NUCB2 to nesfatin-1 is necessary to induce feeding suppression. Chronic i.c.v. injection of nesfatin-1 reduces body weight, whereas rats gain body weight after chronic i.c.v. injection of antisense morpholino oligonucleotide against the gene encoding NUCB2. Nesfatin-1-induced anorexia occurs in Zucker rats with a leptin receptor mutation, and an anti-nesfatin-1 antibody does not block leptin-induced anorexia. In contrast, central injection of alpha-melanocyte-stimulating hormone elevates NUCB2 gene expression in the paraventricular nucleus, and satiety by nesfatin-1 is abolished by an antagonist of the melanocortin-3/4 receptor. We identify nesfatin-1 as a satiety molecule that is associated with melanocortin signalling in the hypothalamus.     

IRS-2 pathways integrate female reproduction and energy homeostasis

Severe dietary restriction, catabolic states and even short-term caloric deprivation impair fertility in mammals. Likewise, obesity is associated with infertile conditions such as polycystic ovary syndrome. The reproductive status of lower organisms such as Caenorhabditis elegans is also modulated by availability of nutrients. Thus, fertility requires the integration of reproductive and metabolic signals. Here we show that deletion of insulin receptor substrate-2 (IRS-2), a component of the insulin/insulin-like growth factor-1 signalling cascade, causes female infertility. Mice lacking IRS-2 have small, anovulatory ovaries with reduced numbers of follicles. Plasma concentrations of luteinizing hormone, prolactin and sex steroids are low in these animals. Pituitaries are decreased in size and contain reduced numbers of gonadotrophs. Females lacking IRS-2 have increased food intake and obesity, despite elevated levels of leptin. Our findings indicate that insulin, together with leptin and other neuropeptides, may modulate hypothalamic control of appetite and reproductive endocrinology. Coupled with findings on the role of insulin-signalling pathways in the regulation of fertility, metabolism and longevity in C. elegans and Drosophila, we have identified an evolutionarily conserved mechanism in mammals that regulates both reproduction and energy homeostasis.     

Leptin activates anorexigenic POMC neurons through a neural network in the arcuate nucleus

The administration of leptin to leptin-deficient humans, and the analogous Lepob/Lepob mice, effectively reduces hyperphagia and obesity. But common obesity is associated with elevated leptin, which suggests that obese humans are resistant to this adipocyte hormone. In addition to regulating long-term energy balance, leptin also rapidly affects neuronal activity. Proopiomelanocortin (POMC) and neuropeptide-Y types of neurons in the arcuate nucleus of the hypothalamus are both principal sites of leptin receptor expression and the source of potent neuropeptide modulators, melanocortins and neuropeptide Y, which exert opposing effects on feeding and metabolism. These neurons are therefore ideal for characterizing leptin action and the mechanism of leptin resistance; however, their diffuse distribution makes them difficult to study. Here we report electrophysiological recordings on POMC neurons, which we identified by targeted expression of green fluorescent protein in transgenic mice. Leptin increases the frequency of action potentials in the anorexigenic POMC neurons by two mechanisms: depolarization through a nonspecific cation channel; and reduced inhibition by local orexigenic neuropeptide-Y/GABA (gamma-aminobutyric acid) neurons. Furthermore, we show that melanocortin peptides have an autoinhibitory effect on this circuit. On the basis of our results, we propose an integrated model of leptin action and neuronal architecture in the arcuate nucleus of the hypothalamus.     

多基因突变小鼠模型与动脉粥样硬化研究

目前己知人类有近 2 0 0 0 0种疾病 ,其发生与发展都与基因受损有着直接或间接的关系 ,其中相当一部分疾病的发病涉及到两个以上的基因功能异常。动脉粥样硬化 (AS)、肥胖、糖尿病、高血压等多基因疑难疾病是目前严重影响人类健康的重大疾病。在AS的发病过程中 ,血脂代谢异常是其重要原因之一。在载脂蛋白E(apoE)通过与低密度脂蛋白受体 (LDLR)和乳糜微粒受体的特异性结合 ,介导血浆脂蛋白的转运与清除 ,在脂质的代谢中起着非常重要的作用。瘦素受体 (OB R)在体内介导瘦素的信号传导 ,调节能量代谢与平衡与肥胖以及血脂代谢有关。通过…     

Partial leptin deficiency and human adiposity.

The adipocyte-derived hormone leptin is crucial for energy homeostasis in mammals; mice and humans without it suffer from a voracious appetite and extreme obesity. The effect on energy balance of variations in plasma leptin above a minimal threshold is uncertain, however, particularly in humans. Here we examine a group of individuals who are genetically partially deficient in leptin, and show that differences in circulating leptin levels within the range found in normal human populations can directly influence the laying down of fat tissue (adiposity).     

卵巢内瘦素受体缺失对卵巢生殖功能和全身糖脂代谢的影响

为探索卵巢内瘦素信号传导缺失对卵巢自身功能和全身糖脂代谢的影响,阐明瘦素对外周生殖系统的直接作用,进行了以下研究:①对瘦素长受体缺失(LR-)的雌性B6.Cg-m / Leprdb小鼠和其同窝出生的有完整LR的雌性野生型C57BU6J小鼠(LR ),在性成熟期后的12周,进行卵巢相互移植,构建实验小鼠的LR基因型组合分别为A(躯体 ,卵巢-)、B[躯体 ,卵巢-)、C(躯体 ,卵巢 )、D(躯体-,卵巢 )、E(躯体-,卵巢-)共5组(n=5),移植后观测2个动情周期取材:②检测各自的全身糖脂代谢指标、卵巢周期和生殖激素;③卵泡刺激素(0.75 IU/g)刺激试验后,用免疫组化和蛋白质印迹(WB)方法,检测卵巢内脂代谢信号蛋白包括Janus激酶(JAK2)、磷酸化的丝裂原激活蛋白激酶(p-ERK)和低密度脂蛋白受体(LDLR)的表达.结果显示:①D、E组与A、B、C 3组比较,体重相差约1倍,性腺周围脂肪垫重量相差约10倍(P<0.01);血葡萄糖、低密度脂蛋白胆固醇(LDL-C)和胰岛素水平相差2-3倍(P<0.01);A、B、C 3组的全身血糖脂指标相似,D和E 2组的高糖脂血症升高程度也相似.(②A、B、C 3组小鼠均恢复正常的动情周期,而D、E 2组则始终处于动情间期;小鼠16周龄时D、E组与A、B、C 3组比较:卵巢重量及E2、P、FSH、LH水平显著下降(P<0.05 J.③瘦素刺激后JAK2在A、B、D和E组的卵巢中表达均呈弱阳性,而C组卵巢内呈强阳性表达;FSH刺激后p-ERK和LDLR在A、B、C 3组卵巢内均呈强阳性表达,而D和E组则表达下降.进而得出结论:①单纯卵巢内瘦素信号缺失并不影响卵巢自身的生殖功能、脂质代谢和全身的糖脂代谢.②瘦素受体全身缺失小鼠的卵巢功能障碍,与卵巢外的全身因素即高糖脂血症和低促性腺激素状态有关.③高糖脂血症降调节卵巢内的瘦素信号蛋白JAK2的表达,诱导小鼠卵巢脂质化和功能障碍;FSH刺激可诱导ERK磷酸化后LDLR的表达增强,缓解小鼠卵巢的脂质化和功能障碍.     

Leptin stimulates fatty-acid oxidation by activating AMP-activated protein kinase.

Leptin is a hormone secreted by adipocytes that plays a pivotal role in regulating food intake, energy expenditure and neuroendocrine function. Leptin stimulates the oxidation of fatty acids and the uptake of glucose, and prevents the accumulation of lipids in nonadipose tissues, which can lead to functional impairments known as "lipotoxicity". The signalling pathways that mediate the metabolic effects of leptin remain undefined. The 5'-AMP-activated protein kinase (AMPK) potently stimulates fatty-acid oxidation in muscle by inhibiting the activity of acetyl coenzyme A carboxylase (ACC). AMPK is a heterotrimeric enzyme that is conserved from yeast to humans and functions as a 'fuel gauge' to monitor the status of cellular energy. Here we show that leptin selectively stimulates phosphorylation and activation of the alpha2 catalytic subunit of AMPK (alpha2 AMPK) in skeletal muscle, thus establishing a previously unknown signalling pathway for leptin. Early activation of AMPK occurs by leptin acting directly on muscle, whereas later activation depends on leptin functioning through the hypothalamic-sympathetic nervous system axis. In parallel with its activation of AMPK, leptin suppresses the activity of ACC, thereby stimulating the oxidation of fatty acids in muscle. Blocking AMPK activation inhibits the phosphorylation of ACC stimulated by leptin. Our data identify AMPK as a principal mediator of the effects of leptin on fatty-acid metabolism in muscle.     

TANK-binding kinase-1 delineates innate and adaptive immune responses to DNA vaccines

Successful vaccines contain not only protective antigen(s) but also an adjuvant component that triggers innate immune activation and is necessary for their optimal immunogenicity. In the case of DNA vaccines, this consists of plasmid DNA; however, the adjuvant element(s) as well as its intra- and inter-cellular innate immune signalling pathway(s) leading to the encoded antigen-specific T- and B-cell responses remain unclear. Here we demonstrate in vivo that TANK-binding kinase 1 (TBK1), a non-canonical IkappaB kinase, mediates the adjuvant effect of DNA vaccines and is essential for its immunogenicity in mice. Plasmid-DNA-activated, TBK1-dependent signalling and the resultant type-I interferon receptor-mediated signalling was required for induction of antigen-specific B and T cells, which occurred even in the absence of innate immune signalling through a well known CpG DNA sensor-Toll-like receptor 9 (TLR9) or Z-DNA binding protein 1 (ZBP1, also known as DAI, which was recently reported as a potential B-form DNA sensor). Moreover, bone-marrow-transfer experiments revealed that TBK1-mediated signalling in haematopoietic cells was critical for the induction of antigen-specific B and CD4(+) T cells, whereas in non-haematopoietic cells TBK1 was required for CD8(+) T-cell induction. These data suggest that TBK1 is a key signalling molecule for DNA-vaccine-induced immunogenicity, by differentially controlling DNA-activated innate immune signalling through haematopoietic and non-haematopoietic cells.     

运动与瘦素的研究进展

瘦素（ leptin）是脂肪组织分泌的一种蛋白质类激素,主要作用在下丘脑,具有调节体脂代谢、能量平衡等功能,瘦素水平的高低对肥胖、2-型糖尿病等疾病起到调节作用。血清瘦素可通过血脑屏障与下丘脑的瘦素受体结合,激活 JAK2-STAT3等信号转导通路,进而改变中枢神经系统中一系列神经肽的表达。通过收集大量近年来关于不同运动形式、运动强度、运动时间等情况对血浆瘦素水平影响的文献,总结分析出一次性运动、非一次性运动对血浆瘦素水平和能量代谢的影响,以及目前研究存在的问题,加深对瘦素功能和机理的分子水平认知,并展望了今后的研究前景。     

Distinct molecular mechanism for initiating TRAF6 signalling

Tumour-necrosis factor (TNF) receptor-associated factor 6 (TRAF6) is the only TRAF family member that participates in signal transduction of both the TNF receptor (TNFR) superfamily and the interleukin-1 receptor (IL-1R)/Toll-like receptor (TLR) superfamily; it is important for adaptive immunity, innate immunity and bone homeostasis. Here we report crystal structures of TRAF6, alone and in complex with TRAF6-binding peptides from CD40 and TRANCE-R (also known as RANK), members of the TNFR superfamily, to gain insight into the mechanism by which TRAF6 mediates several signalling cascades. A 40 degrees difference in the directions of the bound peptides in TRAF6 and TRAF2 shows that there are marked structural differences between receptor recognition by TRAF6 and other TRAFs. The structural determinant of the petide TRAF6 interaction reveals a Pro-X-Glu-X-X-(aromatic/acidic residue) TRAF6-binding motif, which is present not only in CD40 and TRANCE-R but also in the three IRAK adapter kinases for IL-1R/TLR signalling. Cell-permeable peptides with the TRAF6-binding motif inhibit TRAF6 signalling, which indicates their potential as therapeutic modulators. Our studies identify a universal mechanism by which TRAF6 regulates several signalling cascades in adaptive immunity, innate immunity and bone homeostasis.     

中枢NMU抑制食欲作用受黑皮质激素受体途径部分介导

研究中枢神经介素U（NMU）受体2（NMU2R）与黑皮质激素（MC）受体途径（MCR3/4）在调节摄食行为和能量平衡方面的相互作用关系.对禁食或不禁食大鼠脑室注射NMU2R内源性配体NMU和MCR3/4信号途径高亲和力拮抗剂SHU9119,通过测定不同时间点大鼠摄食量和体重变化,探讨中枢NMU2R和黑皮质素受体途径在调节动物摄食行为上的作用及其相互关系.脑室注射NMU对大鼠食欲有显著抑制作用;在同时注射NMU和SHU9119的情况下,NMU2R对大鼠的这种食欲抑制作用会部分受到MCR3/4信号途径变化的影响;同样,对SHU9119前处理大鼠脑内注射NMU,NMU2R抑制食欲的作用也会明显降低,其在摄食方面的作用被部分抑制.结果提示,NMU能够有效的调节摄食行为,而且这种在摄食行为上的调控作用可能部分受MC受体途径介导.     

Dysfunction of lipid sensor GPR120 leads to obesity in both mouse and human

Free fatty acids provide an important energy source as nutrients, and act as signalling molecules in various cellular processes. Several G-protein-coupled receptors have been identified as free-fatty-acid receptors important in physiology as well as in several diseases. GPR120 (also known as O3FAR1) functions as a receptor for unsaturated long-chain free fatty acids and has a critical role in various physiological homeostasis mechanisms such as adipogenesis, regulation of appetite and food preference. Here we show that GPR120-deficient mice fed a high-fat diet develop obesity, glucose intolerance and fatty liver with decreased adipocyte differentiation and lipogenesis and enhanced hepatic lipogenesis. Insulin resistance in such mice is associated with reduced insulin signalling and enhanced inflammation in adipose tissue. In human, we show that GPR120 expression in adipose tissue is significantly higher in obese individuals than in lean controls. GPR120 exon sequencing in obese subjects reveals a deleterious non-synonymous mutation (p.R270H) that inhibits GPR120 signalling activity. Furthermore, the p.R270H variant increases the risk of obesity in European populations. Overall, this study demonstrates that the lipid sensor GPR120 has a key role in sensing dietary fat and, therefore, in the control of energy balance in both humans and rodents.