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.

Anorexia and weight loss are part of the wasting syndrome of late-stage cancer, are a major cause of morbidity and mortality in cancer, and are thought to be cytokine mediated. Macrophage inhibitory cytokine-1 (MIC-1) is produced by many cancers. Examination of sera from individuals with advanced prostate cancer showed a direct relationship between MIC-1 abundance and cancer-associated weight loss. In mice with xenografted prostate tumors, elevated MIC-1 levels were also associated with marked weight, fat and lean tissue loss that was mediated by decreased food intake and was reversed by administration of antibody to MIC-1. Additionally, normal mice given systemic MIC-1 and transgenic mice overexpressing MIC-1 showed hypophagia and reduced body weight. MIC-1 mediates its effects by central mechanisms that implicate the hypothalamic transforming growth factor-beta receptor II, extracellular signal-regulated kinases 1 and 2, signal transducer and activator of transcription-3, neuropeptide Y and pro-opiomelanocortin. Thus, MIC-1 is a newly defined central regulator of appetite and a potential target for the treatment of both cancer anorexia and weight loss, as well as of obesity.

The relationship between stress and obesity remains elusive. In response to stress, some people lose weight, whereas others gain. Here we report that stress exaggerates diet-induced obesity through a peripheral mechanism in the abdominal white adipose tissue that is mediated by neuropeptide Y (NPY). Stressors such as exposure to cold or aggression lead to the release of NPY from sympathetic nerves, which in turn upregulates NPY and its Y2 receptors (NPY2R) in a glucocorticoid-dependent manner in the abdominal fat. This positive feedback response by NPY leads to the growth of abdominal fat. Release of NPY and activation of NPY2R stimulates fat angiogenesis, macrophage infiltration, and the proliferation and differentiation of new adipocytes, resulting in abdominal obesity and a metabolic syndrome-like condition. NPY, like stress, stimulates mouse and human fat growth, whereas pharmacological inhibition or fat-targeted knockdown of NPY2R is anti-angiogenic and anti-adipogenic, while reducing abdominal obesity and metabolic abnormalities. Thus, manipulations of NPY2R activity within fat tissue offer new ways to remodel fat and treat obesity and metabolic syndrome.

Dietary protein enhances satiety and promotes weight loss, but the mechanisms by which appetite is affected remain unclear. We investigated the role of gut hormones, key regulators of ingestive behavior, in mediating the satiating effects of different macronutrients. In normal-weight and obese human subjects, high-protein intake induced the greatest release of the anorectic hormone peptide YY (PYY) and the most pronounced satiety. Long-term augmentation of dietary protein in mice increased plasma PYY levels, decreased food intake, and reduced adiposity. To directly determine the role of PYY in mediating the satiating effects of protein, we generated Pyy null mice, which were selectively resistant to the satiating and weight-reducing effects of protein and developed marked obesity that was reversed by exogenous PYY treatment. Our findings suggest that modulating the release of endogenous satiety factors, such as PYY, through alteration of specific diet constituents could provide a rational therapy for obesity.

Molecular cloning techniques have recently led to the identification of a growing number of neuropeptide Y-receptor subtypes, suggesting possible subtype-specific involvement in different physiological processes. Here we report the first study which determines and compares the mRNA expression of all four cloned functional Y-receptor subtypes (Y1, Y2, Y4 and Y5) in consecutive sections of the rat brain on a cellular level, using a uniform in situ hybridization technique. Our results demonstrate that Y-receptor subtype mRNA expression is widely distributed throughout the rat brain. Interestingly, coexpression of all four Y-receptors, at different levels, is particularly evident within the limbic system, including the hypothalamus, hippocampus, amygdala, piriform and cingulate cortices and tegmental areas, all of which are heavily involved in behaviour, emotion and homeostatic regulation. Particularly interesting is the demonstration that Y5-receptor mRNA expression always coincides with the presence of Y1-receptor mRNA (although not vice versa), possibly due to the overlapping organization and transcriptional control of their genes. However, it is also clear that several brain nuclei display preferential expression of one or a selective combination of Y-receptor subtype mRNAs. Furthermore, it is evident that there is regionalization of expression within certain loci which express all four receptor subtype mRNAs, particularly within the paraventricular and arcuate hypothalamic nuclei. Our results suggest that some of neuropeptide Y's (NPY) effects may be mediated through one particular subtype, whereas other physiological processes might require the coordinated action of different subtypes within the same or discrete areas.

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(39)), by contrast, Y2 receptors have not been detected on bone. In addition to effects in bone, Y1 receptors have been considered as important regulators of energy homeostasis, consistent with pharmacological evidence from Y receptor agonists and antagonists to stimulate or inhibit feeding (9). Fasting-induced re-feeding is reduced in germ line Y1 receptor knock-out mice (10), and deletion of Y1 receptors in genetically obese ob/ob mice, in which hypothalamic NPY-ergic activity is chronically increased, significantly reduces food intake and body weight (11). Paradoxically, germ line Y1 receptor knock-out mice develop late-onset obesity in the absence of hyperphagia (10,12,13). One hypothesis to reconcile this apparent discrepancy is that hypothalamic and non-hypothalamic Y1 receptors have different effects on energy homeostasis.Given the clear involvement of Y1 receptors in the regulation of energy homeostasis as well as new evidence of a putative role for Y1 receptors on osteoblast-like cells, we investigated the effect of germ line and conditional (adult-onset, hypothalamus-specific) deletion of Y1 receptors in mice. In addition, the potential interaction between Y1 receptor sig-

Neuropeptide Y (NPY) is one of the most abundant neuropeptides in the mammalian nervous system and exhibits a diverse range of important physiological activities, including effects on psychomotor activity, food intake, regulation of central endocrine secretion, and potent vasoactive effects on the cardiovascular system. Two major subtypes of NPY receptor (Y1 and Y2) have been defined by pharmaclgical criteria. We report here the molecular cloning of a cDNA sequence encoding a human NPY receptor and the corrected sequence for a rat homologue. Analysis ofthis sequence confirms that the receptor is a member of the G protein-coupled receptor superfamily. When expressed in Chinese hamster ovary (CHO) or human embryonic kidney (293) cells, the receptor exhibits the characteristic ligand specificity of a Y1 type of NPY receptor. In the 293 cell line, the receptor is coupled to a pertussis toxinsensitive G protein that mediates the inhibition of cyclic AMP accumulation. In the CHO cell line, the receptor is coupled not to the inhibition of adenylate cyclase but rather to the elevation of intraceflular calcium. These results demonstrate that second messenger coupling of the NPY-Y1 receptor is cell type specific, depending on the specific repertoire of G proteins and effector systems present in any cell type.Neuropeptide Y (NPY), a 36-amino acid peptide, is an important regulator in both the central and peripheral nervous systems (1). NPY is highly conserved in primary structure between species, as the sequences of human, rat, rabbit, and guinea pig are identical and differ from the porcine sequence by only a single amino acid (2). NPY also shares close sequence homology and a common tertiary structure with a family of peptides which include peptide YY (PYY) and pancreatic polypeptide (PP) This G protein complex in turn activates a variety of second messenger systems, including a decrease in cyclic AMP and an increase in intracellular calcium (10). However, there are reports of NPY receptors coupled to phosphoinositol metabolism, suggesting the existence ofadditional receptor subtypes and/or multiple functions for the Y1 and Y2 subtypes (6, 11).We report here the molecular cloning of a cDNA sequence encoding a human NPY receptor,* which exhibits the characteristic ligand specificity of a Y1 receptor. When expressed in different cell lines, the receptor couples via pertussis toxin-sensitive G proteins to different second messenger systems.MATERIAL AND METHODS Nucleotide Sequence Determination. Total RNA (3 pug) from rat brain was used as a template to synthesize random primed single-stranded cDNA. The cDNA was used in a polymerase chain reaction (PCR) together with the oligonucleotide primers, which correspond to positions 672-584 and 48-78 in the rat cDNA clone FC5 (12). PCR conditions: 30 cycles at 950C for 1 min, 630C for 2 min, and 720C for 1 min. The reaction product was digested with EcoRI and Pst I, gel purified, and subcloned for sequencing in the Bluescript vector (Stratagene) for confirmation of the seq...