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.     

Physiology: does gut hormone PYY3-36 decrease food intake in rodents?

Batterham et al. report that the gut peptide hormone PYY3-36 decreases food intake and body-weight gain in rodents, a discovery that has been heralded as potentially offering a new therapy for obesity. However, we have been unable to replicate their results. Although the reasons for this discrepancy remain undetermined, an effective anti-obesity drug ultimately must produce its effects across a range of situations. The fact that the findings of Batterham et al. cannot easily be replicated calls into question the potential value of an anti-obesity approach that is based on administration of PYY3-36.     

PYY modulation of cortical and hypothalamic brain areas predicts feeding behaviour in humans

The ability to maintain adequate nutrient intake is critical for survival. Complex interrelated neuronal circuits have developed in the mammalian brain to regulate many aspects of feeding behaviour, from food-seeking to meal termination. The hypothalamus and brainstem are thought to be the principal homeostatic brain areas responsible for regulating body weight. However, in the current 'obesogenic' human environment food intake is largely determined by non-homeostatic factors including cognition, emotion and reward, which are primarily processed in corticolimbic and higher cortical brain regions. Although the pleasure of eating is modulated by satiety and food deprivation increases the reward value of food, there is currently no adequate neurobiological account of this interaction between homeostatic and higher centres in the regulation of food intake in humans. Here we show, using functional magnetic resonance imaging, that peptide YY3-36 (PYY), a physiological gut-derived satiety signal, modulates neural activity within both corticolimbic and higher-cortical areas as well as homeostatic brain regions. Under conditions of high plasma PYY concentrations, mimicking the fed state, changes in neural activity within the caudolateral orbital frontal cortex predict feeding behaviour independently of meal-related sensory experiences. In contrast, in conditions of low levels of PYY, hypothalamic activation predicts food intake. Thus, the presence of a postprandial satiety factor switches food intake regulation from a homeostatic to a hedonic, corticolimbic area. Our studies give insights into the neural networks in humans that respond to a specific satiety signal to regulate food intake. An increased understanding of how such homeostatic and higher brain functions are integrated may pave the way for the development of new treatment strategies for obesity.     

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.     

Anti-nociceptive role of neuropeptide Y in the nucleus accumbens in rats with inflammation, an effect modulated by mu- and kappa-opioid receptors

Recent study in our laboratory showed that neuropeptide Y (NPY) plays an antinociceptive role in the nucleus accumbens (NAc) in intact rats. The present study was performed to further investigate the effect of NPY in nociceptive modulation in the NAc of rats with inflammation, and the possible interaction between NPY and the opioid systems. Experimental inflammation was induced by subcutaneous injection of carrageenan into the left hindpaw of rats. Intra-NAc administration of NPY induced a dose-dependent increase of hindpaw withdrawal latencies (HWLs) to thermal and mechanical stimulations in rats with inflammation. The anti-nociceptive effect of NPY was significantly blocked by subsequent intra-NAc injection of the Y1 receptor antagonist NPY28-36, suggesting an involvement of Y1 receptor in the NPY-induced anti-nociception. Furthermore, intra-NAc administration of the opioid antagonist naloxone significantly antagonized the increased HWLs induced by preceding intra-NAc injection of NPY, suggesting an involvement of the endogenous opioid system in the NPY-induced anti-nociception in the NAc during inflammation. Moreover, the NPY-induced anti-nociception was attenuated by following intra-NAc injection of the μ-opioid antagonist β-funaltrexamine (β-FNA), and κ-opioid antagonist nor-binaltorphimine (nor-BNI), but not by δ-opioid antagonist naltrindole, indicating that μ- and κ-opioid receptors, not δ-opioid receptor, are involved in the NPY-induced anti-nociception in the NAc in rats with inflammation.     

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.     

An anorexic lipid mediator regulated by feeding.

Oleylethanolamide (OEA) is a natural analogue of the endogenous cannabinoid anandamide. Like anandamide, OEA is produced in cells in a stimulus-dependent manner and is rapidly eliminated by enzymatic hydrolysis, suggesting a function in cellular signalling. However, OEA does not activate cannabinoid receptors and its biological functions are still unknown. Here we show that, in rats, food deprivation markedly reduces OEA biosynthesis in the small intestine. Administration of OEA causes a potent and persistent decrease in food intake and gain in body mass. This anorexic effect is behaviourally selective and is associated with the discrete activation of brain regions (the paraventricular hypothalamic nucleus and the nucleus of the solitary tract) involved in the control of satiety. OEA does not affect food intake when injected into the brain ventricles, and its anorexic actions are prevented when peripheral sensory fibres are removed by treatment with capsaicin. These results indicate that OEA is a lipid mediator involved in the peripheral regulation of feeding.     

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.     

Neuropeptide Y functions as a neuroproliferative factor

Neuropeptide Y (NPY) has a number of functions in mammalian physiology. Here we identify a role for NPY in promoting proliferation of postnatal neuronal precursor cells. NPY is synthesized in the postnatal olfactory epithelium by sustentacular cells, previously proposed to function only in structural support. Mice with a targeted deletion of NPY contain half as many dividing olfactory neuronal precursor cells as do controls. Furthermore, NPY-deficient mice develop significantly fewer olfactory neurons by adulthood. NPY acts on multipotent neuronal precursor or basal cells to activate rapidly and transiently the extracellular signal-regulated kinase (ERK)1/2 subgroup of mitogen-activated protein kinases. The NPY Y1 receptor subtype appears to mediate this effect. The ability of NPY to induce neuronal precursor proliferation is mediated by protein kinase C (PKC), indicating an upstream PKC-dependent activation of ERK1/2. These results indicate that NPY may regulate neuronal precursor proliferation in the adult mammal.     

糖皮质激素性骨质疏松大鼠弓状核β-内啡肽免疫反应神经元的变化

目的：观察地塞米松（DEX）致雄性大鼠骨质疏松后,其下丘脑弓状核β-内啡肽（β-endorphin,β-EP）免疫反应神经元的变化。方法：选用4月龄健康雄性SD大鼠16只,采用随机数字表法将大鼠随机分为2组,模型组给予地塞米松磷酸钠注射液以0．1mg／100g体重,肌肉注射,2次／周,连续8周。对照组肌肉注射等量生理盐水,8周后处死取材。通过检测大鼠胫骨上段骨组织形态计量学的变化判断骨代谢的情况;采用免疫组织化学方法观察弓状核β-EP免疫反应神经元的变化。结果：与对照组相比,模型组大鼠骨小梁面积百分数、骨小梁宽度和骨小梁数量明显减小（P〈0．05）,骨小梁分离度明显增大（P〈0．01）;模型组大鼠下丘脑弓状核β-EP免疫反应阳性神经元的数量明显减少（p〈0．05）。结论：弓状核β-EP免疫反应阳性神经元数量的减少可能与糖皮质激素性骨盾疏松的形成具有密切的美系。     

Reduced antinociception and plasma extravasation in mice lacking a neuropeptide Y receptor

Neuropeptide Y (NPY) is believed to exert antinociceptive actions by inhibiting the release of substance P and other 'pain neurotransmitters' in the spinal cord dorsal horn. However, the physiological significance and potential therapeutic value of NPY remain obscure. It is also unclear which receptor subtype(s) are involved. To identify a possible physiological role for the NPY Y1 receptor in pain transmission, we generated NPY Y1 receptor null mutant (Y1-/-) mice by homologous recombination techniques. Here we show that Y1-/- mice develop hyperalgesia to acute thermal, cutaneous and visceral chemical pain, and exhibit mechanical hypersensitivity. Neuropathic pain is increased, and the mice show a complete absence of the pharmacological analgesic effects of NPY. In the periphery, Y1 receptor activation is sufficient and required for substance P release and the subsequent development of neurogenic inflammation and plasma leakage. We conclude that the Y1 receptor is required for central physiological and pharmacological NPY-induced analgesia and that its activation is both sufficient and required for the release of substance P and initiation of neurogenic inflammation.     

Deciphering a neuronal circuit that mediates appetite

Hypothalamic neurons that co-express agouti-related protein (AgRP), neuropeptide?Y and γ-aminobutyric acid (GABA) are known to promote feeding and weight gain by integration of various nutritional, hormonal, and neuronal signals. Ablation of these neurons in mice leads to cessation of feeding that is accompanied by activation of Fos in most regions where they project. Previous experiments have indicated that the ensuing starvation is due to aberrant activation of the parabrachial nucleus (PBN) and it could be prevented by facilitating GABA(A) receptor signalling in the PBN within a critical adaptation period. We speculated that loss of GABA signalling from AgRP-expressing neurons (AgRP neurons) within the PBN results in unopposed excitation of the PBN, which in turn inhibits feeding. However, the source of the excitatory inputs to the PBN was unknown. Here we show that glutamatergic neurons in the nucleus tractus solitarius (NTS) and caudal serotonergic neurons control the excitability of PBN neurons and inhibit feeding. Blockade of serotonin (5-HT(3)) receptor signalling in the NTS by either the chronic administration of ondansetron or the genetic inactivation of Tph2 in caudal serotonergic neurons that project to the NTS protects against starvation when AgRP neurons are ablated. Likewise, genetic inactivation of glutamatergic signalling by the NTS onto N-methyl D-aspartate-type glutamate receptors in the PBN prevents starvation. We also show that suppressing glutamatergic output of the PBN reinstates normal appetite after AgRP neuron ablation, whereas it promotes weight gain without AgRP neuron ablation. Thus we identify the PBN as a hub that integrates signals from several brain regions to bidirectionally modulate feeding and body weight.     

Interaction between neuropeptide Y and noradrenaline on central catecholamine neurons

Despite their widespread occurrence in the central nervous system, interactions between co-localized transmitters and their receptors remain poorly understood. Noradrenergic neurons of the nucleus locus coeruleus contain the peptide co-transmitter neuropeptide Y (refs 1,2). In locus coeruleus cells, stimulation of alpha2-adrenoceptors 3,4 or opioid mu-receptors 5,6 increases a potassium conductance and thereby leads to hyperpolarization and inhibition of spontaneous firing. Coupling between these receptors and the inward rectifying K+ channels involves a pertussis toxin-sensitive GTP-binding protein (Gi or Go)7. Here we investigate whether the neuropeptide Y and alpha2-receptors of locus coeruleus neurons interact with one another. When administered alone, neuropeptide Y reduces the discharge of action potentials, probably by increasing the permeability of the membrane to potassium ions through the activation of a G protein; this effect is reduced in the presence of alpha2-adrenoceptor antagonists. Moreover, the peptide selectively increases the hyperpolarizing effect of alpha2-agonists, but does not enhance responses to opioid mu-agonists. We suggest that noradrenaline and its co-transmitter neuropeptide Y stimulate separate receptors, which influence each other in a specific way.     

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.     

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.     

Influence of magnetic field on nitric oxide and neuropeptide Y in rat adrenal gland

We have investigated the effects of magnetic fields on nitric acid (NO) and neuropeptide Y (NPY) on adrenal glands of the rat using NO nitric acid reductase-spectrophotometry ↭d histochemistry techniques. We found that all cellular layers of the adrenal cortex, including zona glomerulosa, zona fasciculata and zona reticularis were stained by NADPH-d, and some chromaffin cells of the medulla were positive for NPY. Furthermore, magnetic fields increased NO so strongly that high NO levels could be maintained for several hours, as well as some neuroganglion cells in medulla that were double-stained for NPY and NADPH-d. Our data showed that the magnetic field can regulate endocrine and neuroendocrine directly by some action on parenchyma cells, or indirectly by action to NO-ergic, NPY-ergic neurons in the adrenal gland.     

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.     

Rapid development of tolerance to the behavioural actions of cholecystokinin

Cholecystokinin (CCK) acts acutely to inhibit food consumption in fasted rats, mice, sheep, pigs, monkeys and humans. CCK has been proposed as a satiety signal, inducing the behavioural sequence of satiety, or as an aversive internal stimulus, which inhibits food intake by inducing malaise. Reductions in food intake and related exploratory behaviours are initiated by CCK at its peripheral receptor in the gut, which appears to transmit sensory feedback via the vagus nerve to brain regions mediating appetitive behaviours. The therapeutic potential of CCK as an appetite suppressant in obesity syndromes rests on the demonstration of significant, long-lasting body weight reduction. Chronic CCK administration by repeated injections is problematic, since this peptide is rapidly degraded in vivo. We chose the Alzet constant infusion osmotic minipump to investigate possible alterations in body weight and food intake during continuous infusion of CCK. We now report that no change was detected in either body weight or total daily food consumption at any time point during 2 weeks of intraperitoneally (i.p.) infused CCK. The mechanism underlying the lack of chronic CCK effects appears to be a rapid development of behavioural tolerance. Acute challenge doses of CCK which induced satiety-related behaviours in saline-infused rats were ineffective in CCK-infused rats. The behavioural tolerance was apparent within a few hours of minipump implantation. These results provide the first evidence that rapid and reversible tolerance develops to the actions of a gut peptide.     

Stimulation of food intake in rats by centrally administered hypothalamic growth hormone-releasing factor

Hypothalamic growth hormone-releasing factors (GRFs) have been purified recently from human pancreatic (hp) tumours and from rat hypothalamus (rh). GRF peptides have strong homology with peptides of the glucagon, vasoactive intestinal polypeptide and PHI-27 family. Aside from their potent actions on release of somatotropin, no other biological actions of GRFs have been reported. GRF has been localized in neurones bordering the ventromedial hypothalamic nucleus, a region associated frequently with experimental analysis of feeding behaviour. We now report that intracerebroventricularly (i.c.v.)-administered rhGRF and hpGRF(1-40) in doses of 0.2, 2.0 and 20.0 pmol, produced an increase in food intake in hungry rats. This effect seemed to be specific to GRF as i.c.v. injections of a structurally related but physiologically inactive peptide in the same doses had no effect on feeding. In addition, peripheral injections of rhGRF or growth hormone had no effect on food intake, suggesting that the present effects may be mediated centrally. Injections (i.c.v.) of rhGRF (0.2, 2.0 and 20.0 pmol) had no effect on general activity, suggesting that GRF does not produce nonspecific arousal.