The Neuropeptide Y System Regulates Both Mechanical and Histaminergic Itch.

Itch is a somatosensory modality that serves to alert an organism to harmful elements removable by scratching, such as parasites and chemical irritants. Recently, ablation or silencing of neuropeptide Y (NPY)-expressing spinal interneurons was reported to selectively enhance mechanical itch, whereas chemical itch was unaffected. We examined the effect of activating the NPY/Y1 receptor system on scratch behavior in mice. We found that intrathecal administration of the Y1 agonist [Leu31,Pro34]-NPY (LP-NPY) attenuated itch behavior induced by application of 0.07 g von Frey filament in the nape of the neck compared with saline treatment, indicating that activation of the spinal NPY/Y1 system dampens mechanical itch. However, intrathecal administration of LP-NPY also attenuated chemically induced scratching provoked by intradermal application of histamine or the mast cell degranulator 48/80 (histaminergic itch), and the latter effect could be reversed by administration of the Y1 antagonist BIBO3304. Intrathecal application of the native nonselective agonist NPY also attenuated histamine or 48/80-induced scratching. Our analyses emphasize the importance of including additional quantitative parameters to characterize the full spectrum of itch behavior and show that the NPY/Y1 system dampens both mechanically and chemically induced scratching and hence is shared by the two submodalities of itch.

Dipeptidyl peptidase IV (DPP-IV) inhibition prevents fibrosis in adipose tissue of obese mice.

BACKGROUND: During the development of obesity the expansion of white adipose tissue (WAT) leads to a dysregulation and an excessive remodeling of extracellular matrix (ECM), leading to fibrosis formation. These ECM changes have high impact on WAT physiology and may change obesity progression. Blocking WAT fibrosis may have beneficial effects on the efficacy of diet regimen or therapeutical approaches in obesity. Since dipeptidyl peptidase IV (DPP-IV) inhibitors prevent fibrosis in tissues, such as heart, liver and kidney, the objective of this study was to assess whether vildagliptin, a DPP-IV inhibitor, prevents fibrosis in WAT in a mouse model of obesity, and to investigate the mechanisms underlying this effect. METHODS: We evaluated the inhibitory effect of vildagliptin on fibrosis markers on WAT of high-fat diet (HFD)-induced obese mice and on 3T3-L1 cell line of mouse adipocytes treated with a fibrosis inducer, transforming growth factor beta 1 (TGFß1). RESULTS: Vildagliptin prevents the increase of fibrosis markers in WAT of HFD-fed mice and reduces blood glucose, serum triglycerides, total cholesterol and leptin levels. In the in vitro study, the inhibition of DPP-IV with vildagliptin, neuropeptide Y (NPY) treatment and NPY Y1 receptor activation prevents ECM deposition and fibrosis markers increase induced by TGFß1 treatment. CONCLUSIONS: Vildagliptin prevents fibrosis formation in adipose tissue in obese mice, at least partially through NPY and NPY Y1 receptor activation. GENERAL SIGNIFICANCE: This study highlights the importance of vildagliptin in the treatment of fibrosis that occur in obesity.

Prenatal High Estradiol Exposure Induces Sex-Specific and Dietarily Reversible Insulin Resistance Through Decreased Hypothalamic INSR.

An adverse intrauterine environment may induce adult disease in offspring, but the mechanisms are not well understood. It is reported that fresh embryo transfer (ET) in assisted reproductive technology leads to high maternal estradiol (E2), and prenatal high E2 exposure increases the risk of organ disorders in later life. We found that male newborns and children of fresh ET showed elevated fasting insulin and homeostasis model of assessment for insulin resistance index (HOMA-IR) scores. Male mice with high prenatal estradiol exposure (HE) grew heavier than control mice and developed insulin resistance; they also showed increased food intake, with increased orexigenic hypothalamic neuropeptide Y (NPY) expression. The hypothalamic insulin receptor (INSR) was decreased in male HE mice, associated with elevated promoter methylation. Chronic food restriction (FR) in HE mice reversed insulin resistance and rescued hypothalamic INSR expression by correcting the elevated Insr promoter methylation. Our findings suggest that prenatal exposure to high E2 may induce sex-specific metabolic disorders in later life through epigenetic programming of hypothalamic Insr promoter, and dietary intervention may reverse insulin resistance by remodeling its methylation pattern.

Dietary Macronutrient Composition Affects the Influence of Exogenous Prolactin-Releasing Peptide on Appetite Responses and Hypothalamic Gene Expression in Chickens.

BACKGROUND: The interaction between the effects of exogenous neurotransmitters and dietary composition on appetite regulation in nonmammalian species is unclear. OBJECTIVE: The objective of this study was to determine the effects of exogenous prolactin-releasing peptide (PrRP) and dietary macronutrient composition on food intake regulation in broiler chicks. METHODS: Three isocaloric diets were formulated: high-carbohydrate (HC), high-fat (HF; 60% of ME from lard) and high-protein (HP) diets. In Expt. 1, 4-d-old Hubbard × Cobb-500 chicks fed 1 of the 3 diets since hatch were intracerebroventricularly injected with 0 (vehicle), 3, or 188 pmol PrRP (n = 10). Food intake was measured for 180 min. In Expt. 2, hypothalamic mRNA abundance of appetite-associated factors was measured in hypothalamus samples obtained 1 h postinjection of 0 or 188 pmol PrRP. In Expt. 3, chicks were given free access to all diets before and after intracerebroventricular injection and food intake was measured. RESULTS: Three and 188 pmol PrRP increased (P = 0.0008 and 0.04) HP diet intake, but only 188 pmol PrRP was efficacious at increasing HC (P = 0.0011) and HF (P = 0.01) consumption compared with the vehicle. There was a diet effect on mRNA abundance of all genes (P < 0.05), with greater expression in chicks fed the HF or HP than the HC diet. Whereas neuropeptide Y (NPY) mRNA was similar between vehicle- and PrRP-injected chicks that consumed HP or HF diets, expression was greater (P < 0.05) in PrRP- than vehicle-injected chicks that consumed the HC diet. When chicks had access to all diets, 188 pmol PrRP caused preferential (P < 0.0001) intake of the HP over the HC and HF diets. CONCLUSION: The HP diet enhanced the sensitivity of chicks to the food intake-stimulating effects of PrRP, and PrRP in turn increased preference for the HP diet. Thus, dietary macronutrient composition influences PrRP-mediated food intake, and PrRP in turn affects nutrient intake and transcriptional regulation in chicks.

Activation of Neuropeptide Y Receptors Modulates Retinal Ganglion Cell Physiology and Exerts Neuroprotective Actions In Vitro.

Neuropeptide Y (NPY) is expressed in mammalian retina but the location and potential modulatory effects of NPY receptor activation remain largely unknown. Retinal ganglion cell (RGC) death is a hallmark of several retinal degenerative diseases, particularly glaucoma. Using purified RGCs and ex vivo rat retinal preparations, we have measured RGC intracellular free calcium concentration ([Ca2+]i) and RGC spiking activity, respectively. We found that NPY attenuated the increase in the [Ca2+]i triggered by glutamate mainly via Y1 receptor activation. Moreover, (Leu31, Pro34)-NPY, a Y1/Y5 receptor agonist, increased the initial burst response of OFF-type RGCs, although no effect was observed on RGC spontaneous spiking activity. The Y1 receptor activation was also able to directly modulate RGC responses by attenuating the NMDA-induced increase in RGC spiking activity. These results suggest that Y1 receptor activation, at the level of inner or outer plexiform layers, leads to modulation of RGC receptive field properties. Using in vitro cultures of rat retinal explants exposed to NMDA, we found that NPY pretreatment prevented NMDA-induced cell death. However, in an animal model of retinal ischemia-reperfusion injury, pretreatment with NPY or (Leu31, Pro34)-NPY was not able to prevent apoptosis or rescue RGCs. In conclusion, we found modulatory effects of NPY application that for the first time were detected at the level of RGCs. However, further studies are needed to evaluate whether NPY neuroprotective actions detected in retinal explants can be translated into animal models of retinal degenerative diseases.

Neuropeptide Y Y5-receptor activation on breast cancer cells acts as a paracrine system that stimulates VEGF expression and secretion to promote angiogenesis.

Accumulating data implicate a pathological role for sympathetic neurotransmitters like neuropeptide Y (NPY) in breast cancer progression. Our group and others reported that NPY promotes proliferation and migration in breast cancer cells, however the angiogenic potential of NPY in breast cancer is unknown. Herein we sought to determine if NPY promotes angiogenesis in vitro by increasing vascular endothelial growth factor (VEGF) expression and release from 4T1 breast cancer cells. Western blot analysis revealed that NPY treatment caused a 52 ± 14% increase in VEGF expression in the 4T1 cells compared to non-treated controls. Using selective NPY Y-receptor agonists (Y1R, Y2R and Y5R) we observed an increase in VEGF expression only when cells were treated with Y5R agonist. Congruently, using selective Y1R, Y2R, or Y5R antagonists, NPY-induced increases in VEGF expression in 4T1 cells were attenuated only under Y5R antagonism. Endothelial tube formation assays were conducted using conditioned media (CM) from NPY treated 4T1 cells. Concentration-dependent increases in number of branch points and complete endothelial networks were observed in HUVEC exposed to NPY CM. CM from Y5R agonist treated 4T1 cells caused similar increases in number of branch points and complete endothelial networks. VEGF concentration was quantified in CM (ELISA) from agonist experiments; we observed a 2-fold and 2.5-fold increase in VEGF release from NPY and Y5R agonist treated 4T1 cells respectively. Overall these data highlight a novel mechanism by which NPY may promote breast cancer progression, and further implicate a pathological role of the NPY Y5R.

Opposing roles of corticotropin-releasing factor and neuropeptide Y within the dorsolateral bed nucleus of the stria terminalis in the negative affective component of pain in rats.

Pain is a complex experience composed of sensory and affective components. Although the neural systems of the sensory component of pain have been studied extensively, those of its affective component remain to be determined. In the present study, we examined the effects of corticotropin-releasing factor (CRF) and neuropeptide Y (NPY) injected into the dorsolateral bed nucleus of the stria terminalis (dlBNST) on pain-induced aversion and nociceptive behaviors in rats to examine the roles of these peptides in affective and sensory components of pain, respectively. In vivo microdialysis showed that formalin-evoked pain enhanced the release of CRF in this brain region. Using a conditioned place aversion (CPA) test, we found that intra-dlBNST injection of a CRF1 or CRF2 receptor antagonist suppressed pain-induced aversion. Intra-dlBNST CRF injection induced CPA even in the absence of pain stimulation. On the other hand, intra-dlBNST NPY injection suppressed pain-induced aversion. Coadministration of NPY inhibited CRF-induced CPA. This inhibitory effect of NPY was blocked by coadministration of a Y1 or Y5 receptor antagonist. Furthermore, whole-cell patch-clamp electrophysiology in dlBNST slices revealed that CRF increased neuronal excitability specifically in type II dlBNST neurons, whereas NPY decreased it in these neurons. Excitatory effects of CRF on type II dlBNST neurons were suppressed by NPY. These results have uncovered some of the neuronal mechanisms underlying the affective component of pain by showing opposing roles of intra-dlBNST CRF and NPY in pain-induced aversion and opposing actions of these peptides on neuronal excitability converging on the same target, type II neurons, within the dlBNST.

Modulation of methamphetamine-induced nitric oxide production by neuropeptide Y in the murine striatum.

Methamphetamine (METH) is a potent stimulant that induces both acute and long-lasting neurochemical changes in the brain including neuronal cell loss. Our laboratory demonstrated that the neuropeptide substance P enhances the striatal METH-induced production of nitric oxide (NO). In order to better understand the role of the striatal neuropeptides on the METH-induced production of NO, we used agonists and antagonists of the NPY (Y1R and Y2R) receptors infused via intrastriatal microinjection followed by a bolus of METH (30 mg/kg, ip) and measured 3-NT immunofluorescence, an indirect index of NO production. One striatum received pharmacological agent while the contralateral striatum received aCSF and served as control. NPY receptor agonists dose dependently attenuated the METH-induced production of striatal 3-NT. Conversely, NPY receptor antagonists had the opposite effect. Moreover, METH induced the accumulation of cyclic GMP and activated caspase-3 in approximately 18% of striatal neurons, a phenomenon that was attenuated by pre-treatment with NPY2 receptor agonist. Lastly, METH increased the levels of striatal preproneuropeptide Y mRNA nearly five-fold 16 h after injection as determined by RT-PCR, suggesting increased utilization of the neuropeptide. In conclusion, NPY inhibits the METH-induced production of NO in striatal tissue. Consequently, production of this second messenger induces the accumulation of cyclic GMP and activated caspase-3 in some striatal neurons, an event that may precede the apoptosis of some striatal neurons.

The selective neuropeptide Y Y5 agonist [cPP(1-7),NPY(19-23),Ala31,Aib32,Gln34]hPP differently modulates emotional processes and body weight in the rat.

The neuropeptide Y (NPY) has been suggested to act as a major regulator of emotional processes and body weight. The full spectrum of biological effects of this peptide is mediated by at least four classes of receptors known as the Y(1), Y(2), Y(4), and Y(5) subtypes. However, the respective contribution of each of these receptor subtypes, especially the Y(5) subtype, in emotional processes is still mostly unknown. In the present study, we investigated the effect of long term administration of a selective Y(5) agonist [cPP(1-7),NPY(19-23),Ala(31),Aib(32),Gln(34)]hPP on emotional processes and body weight using two rat models of emotional dysfunctions, the corticosterone (CORT)-induced anxiety model as well as the olfactory bulbectomized (OBX) model of depression and anxiety in Wistar and Sprague-Dawley rats, respectively. The sub-chronic administration of the Y(5) agonist reversed the high levels of locomotion, rearing and grooming in the open field test and the impaired social activity induced by OBX, while increased the percentage of entries and time in the open arm of the elevated plus maze in CORT-treated rats. Furthermore, this Y(5) agonist increased body weight in both strains of control rats. These data further demonstrate that Y(5) receptors are not only involved in the control of body weight but also mediate emotional processing under challenged conditions. Thus, the pharmacotherapeutic administration of a Y(5) agonist could be considered as a potentially novel strategy to alleviate some forms of anxiety and depression in humans.

A role for neuropeptide Y in the gender-specific gastrointestinal, corticosterone and feeding responses to stress.

BACKGROUND AND PURPOSE: Exposure to an acute stress inhibits gastric emptying and stimulates colonic transit via central neuropeptide Y (NPY) pathways; however, peripheral involvement is uncertain. The anxiogenic phenotype of NPY(-/-) mice is gender-dependent, raising the possibility that stress-induced gastrointestinal (GI) responses are female-dominant through NPY. The aim of this study was to determine GI transit rates, corticosterone levels and food intake after acute restraint (AR) or novel environment (NE) stress in male and female NPY(-/-) and WT mice. EXPERIMENTAL APPROACH: Upper gastrointestinal transit (UGIT) (established 30 min after oral gavage) and corticosterone levels were determined under basal or restrained conditions (30 min) and after treatment i.p. with Y(1) antagonist BIBO3304 or Y(2) antagonist BIIE0246. Faecal pellet output (FPO) was established after AR and treatment i.p. with NPY in the NE, as were colonic bead expulsion rates. KEY RESULTS: UGIT and FPO were similar in unrestrained male and female mice. NPY(-/-) females displayed significantly slower UGIT than NPY(-/-) males after AR, but both genders displayed significantly higher FPO and reduced food intake relative to WT counterparts. Peripheral NPY treatment increased bead expulsion time in WT mice. AR male NPY(-/-) mice had higher levels of corticosterone than male WT mice; whilst in AR WT mice, after peripheral Y(1) and Y(2) receptor antagonism in males, and Y(2) antagonism in females, corticosterone was significantly elevated. CONCLUSIONS AND IMPLICATIONS: NPY possesses a role in the gender-dependent susceptibility to stress-induced GI responses. Furthermore, NPY inhibits GI motility through Y(2) receptors and corticosterone release via peripheral Y(1) and Y(2) receptors.

Quantitative analysis of neuropeptide Y receptor association with beta-arrestin2 measured by bimolecular fluorescence complementation.

BACKGROUND AND PURPOSE: beta-Arrestins are critical scaffold proteins that shape spatiotemporal signalling from seven transmembrane domain receptors (7TMRs). Here, we study the association between neuropeptide Y (NPY) receptors and beta-arrestin2, using bimolecular fluorescence complementation (BiFC) to directly report underlying protein-protein interactions. EXPERIMENTAL APPROACH: Y1 receptors were tagged with a C-terminal fragment, Yc, of yellow fluorescent protein (YFP), and beta-arrestin2 fused with the complementary N-terminal fragment, Yn. After Y receptor-beta-arrestin association, YFP fragment refolding to regenerate fluorescence (BiFC) was examined by confocal microscopy in transfected HEK293 cells. Y receptor/beta-arrestin2 BiFC responses were also quantified by automated imaging and granularity analysis. KEY RESULTS: NPY stimulation promoted association between Y1-Yc and beta-arrestin2-Yn, and the specific development of BiFC in intracellular compartments, eliminated when using non-interacting receptor and arrestin mutants. Responses developed irreversibly and were slower than for downstream Y1 receptor-YFP internalization, a consequence of delayed maturation and stability of complemented YFP. However, beta-arrestin2 BiFC measurements delivered appropriate ligand pharmacology for both Y1 and Y2 receptors, and demonstrated higher affinity of Y1 compared to Y2 receptors for beta-arrestin2. Receptor mutagenesis combined with beta-arrestin2 BiFC revealed that alternative arrangements of Ser/Thr residues in the Y1 receptor C tail could support beta-arrestin2 association, and that Y2 receptor-beta-arrestin2 interaction was enhanced by the intracellular loop mutation H155P. CONCLUSIONS AND IMPLICATIONS: The BiFC approach quantifies Y receptor ligand pharmacology focused on the beta-arrestin2 pathway, and provides insight into mechanisms of beta-arrestin2 recruitment by activated and phosphorylated 7TMRs, at the level of protein-protein interaction.

NPY signalling pathway in bone homeostasis: Y1 receptor as a potential drug target.

Neuropeptide (NPY) is a neurotransmitter widely distributed in central and peripheral nervous system that has been implicated in several physiological processes through activation of five different Y receptors: Y1, Y2, Y4, Y5, and y6. NPY system has been extensively studied for the last decades due to its implications in a wide variety of physiological processes. For this purpose a diversity of sophisticated animal models and receptors agonists and antagonists has been developed to better understand its actions throughout body homeostasis. Consequently, NPY and its receptors have recently emerged as a potential regulator of bone homeostasis. This is supported by the demonstration of an increase of bone mass in mice lacking Y1 or Y2 receptor genes. Recent findings revealed Y1 receptor as a potential drug target candidate for prevention and treatment of bone loss. Indeed, it has been demonstrated that osteoblasts express Y1 receptor while no other Y receptor was detected in these cells, implicating Y1 receptor signalling in the local control of bone turnover. In this review, we have summarized the findings obtained from studies on NPY system in skeletogenesis focusing on Y1 receptor.

Relevance of the neuropeptide Y system in the biology of cancer progression.

The peptides pancreatic polypeptide (PP), peptide YY (PYY), and neuropeptide Y (NPY) share a similar structure, known as PP-fold. Within this family of peptides, NPY, a highly conserved 36-aminoacid residue peptide, is involved in the regulation of a wide range of physiological functions, such as food intake and energy metabolism, as well as in the promotion of some remarkable aspects of tumor progression, including cell proliferation, matrix invasion, metastatization, and angiogenesis. NPY exerts its biological effects through five G-protein coupled receptors, named Y1-, Y2-, Y4-, Y5-, and y6-R, which appear associated with different aspects of oncogenesis. Y1-R seems involved in the modulation of cancer cell proliferation, whereas Y2-R activation appears to promote angiogenesis. The development of NPY receptor subtype selective analogs has helped to elucidate the physiological and pathophysiological role and localization of each receptor and may contribute to a better understanding of the receptor-ligand interaction. The NPY system appears to be variously associated with specific tumors, including neural crest-derived tumors, breast and prostate cancers. In addition to NPY, PYY is also able to affect cancer cell growth in a dose-dependent manner and through Y-Rs. In conclusion, peptides of the NPY family and the related receptors play an important role in the progression of different cancer types, with some molecular specificity according to each step of this process. On this basis, future studies may be directed to the implementation of novel diagnostic and therapeutic approaches targeting this system.

Neuropeptide Y suppresses absence seizures in a genetic rat model primarily through effects on Y receptors.

Neuropeptide Y (NPY) potently suppresses absence seizures in a model of genetic generalized epilepsy, genetic absence epilepsy rats of Strasbourg (GAERS). Here we investigated the Y-receptor subtype(s) on which NPY exerts this anti-absence effect. A dual in vivo approach was used: the cumulative duration of seizures was quantified in adult male GAERS in 90-min electroencephalogram recordings following intracerebroventricular (i.c.v.) injection of: (i) subtype-selective agonists of Y1 ([Leu31Pro34]NPY, 2.5 nmol), Y2 (Ac[Leu(28,31)]NPY24-36, 3 nmol), Y5 receptors [hPP1(-17),Ala31,Aib32]NPY, 4 nmol), NPY (3 nmol) or vehicle; and following (ii) i.c.v. injection of antagonists of Y1 (BIBP3226, 20 nmol), Y2 (BIIE0246, 20 nmol) and Y5 (NPY5RA972, 20 nmol) receptors or vehicle, followed by NPY (3 nmol). Injection of the Y1- and Y5-selective agonists resulted in significantly less mean seizure suppression (37.4% and 53.9%, respectively) than NPY (83.2%; P < 0.05), while the Y2 agonist had similar effects to NPY (62.3% suppression, P = 0.57). Food intake was not increased following injection of the Y2 agonist, while significant increases in food intake were seen following NPY and the other Y-subtype agonists. Compared with vehicle, NPY injection suppressed seizures following the Y1 and Y5 antagonists (45.3% and 80.1%, respectively, P < 0.05), but not following the Y2 antagonist (5.1% suppression, P = 0.46). We conclude that NPY Y2 receptors are more important than Y1 and Y5 receptors in mediating the effect of NPY to suppress absence seizures in a genetic rat model. Y2 receptor agonists may represent targets for novel drugs against genetic generalized epilepsies without resulting in appetite stimulation.

Neuropeptide Y-induced enhancement of the evoked release of newly synthesized dopamine in rat striatum: mediation by Y2 receptors.

The purpose of the present study was to determine whether or not activation of neuropeptide Y (NPY) receptors resulted in an enhancement or attenuation of the KCl (50 mM) evoked release of [3H]dopamine newly synthesized from [3H]tyrosine in superfused striatal slices and, if so to identify the NPY receptor subtype mediating the effect. Rat striatal slices were prepared and placed in microsuperfusion chambers and continuously superfused with physiological buffer containing 50 microCi/ml of l-3-5-[3H]tyrosine. Superfusate effluents were collected and analyzed for [3H]dopamine by liquid scintillation spectrometry following amberlite CG50 and alumina chromatography. NPY agonists (NPY and PYY3-36) were added 6 min prior to the addition of KCl, while the Y1, Y2, and Y5 antagonist BIBO3304, BIIE0246 and CGP71683A, respectively were added 6 min prior to the agonists. Continuous superfusion with [3H]tyrosine resulted in the production of [3H]dopamine which reached a steady state at approximately 48 min. Depolarization with KCl resulted in a 2- to 3-fold increase in [3H]dopamine overflow. NPY and PYY3-36 produced a concentration dependent enhancement in the KCl induced increase in newly synthesized [3H]dopamine overflow. The Y2 antagonist BIIE0246 produced an attenuation of both the NPY and PYY3-36 induced enhancement while the Y1 antagonist BIBO3304 and theY5 antagonist CGP71683A failed to alter the NPY or PYY3-36 induced enhancement. These results are consistent with the NPY-Y2 receptor subtype mediating the facilitatory effect.

Neuropeptide Y receptors; antisecretory control of intestinal epithelial function.

This paper reviews the cellular localisation, mechanisms of release and intestinal absorptive actions of neuropeptide Y and its related peptides, peptide YY, pancreatic polypeptide and major fragments NPY(3-36) and PYY(3-36). While NPY is commonly found in inhibitory enteric neurons that can be interneurons, motor neurons or secretomotor-nonvasodilator in nature, its analogue, peptide YY in contrast, is located in neuroendocrine L-cells that predominate in the colorectal mucosa. Peptide YY is released from these cells when nutrients arrive in the small or large bowel, exerting paracrine as well as hormonal actions. Pancreatic polypeptide is found in relatively few, scattered intestinal endocrine cells, the majority of this peptide being produced by, and released from pancreatic islet F-cells in response to food intake. An introduction to the current pharmacology of this family of peptides is provided and the different types of neuropeptide Y (termed Y) receptors, their agonist preferences, antagonism, and preferred signalling pathways, are described. Our current understanding of specific Y receptor localisation within the intestine as determined by immunohistochemistry, is presented as a prelude to an assessment of functional studies that have monitored ion transport across isolated mucosal preparations. It is becoming clear that three Y receptor types are significant functionally in human colon, as well as particular rodent models (e.g. mouse) and these, namely the Y(1), Y(2) and Y(4) receptors, are discussed in detail. Their presence within the basolateral aspect of the epithelial layer (Y(1) and Y(4) receptors) or on enteric neurons (Y(1) and Y(2) receptors) and their activation by endogenous neuropeptide Y, peptide YY (Y(1) and Y(2) receptors) or pancreatic polypeptide (which prefers Y(4) receptors) results consistently in antisecretory/absorptive responses. The recent use of novel mouse knockouts has helped establish loss of specific intestinal functions including Y(1) and Y(2) receptor-mediated absorptive tone in colon mucosa. Progress in this field has been rapid recently, aided by the availability of selective antagonists and mutant mice lacking either one (e.g. Y(4)-/-, for which no antagonists exist at present) or more Y receptor types. It is therefore timely to review this work and present a rational basis for developing stable synthetic Y receptor agonists as novel anti-diarrhoeals.

Design and synthesis of (ant)-agonists that alter appetite and adiposity.

Over the past decade, hypothalamic circuits have been described that impact energy homeostasis in rodents and humans. Our drug development efforts for the treatment of obesity and the metabolic syndrome have largely focused on selected genetic and/or pharmacologically validated pathways. The translation of these pathways into therapeutics for the treatment of obesity will find its first clinical successes over the coming decade. Initial efforts have focused on gaining a better understanding of the relevance of rodent pharmacological and genetic observations for the development of therapeutics for the treatment of human obesity. We pursue pathways defined by the expression of the ghrelin receptor, melanin-concentrating hormone receptors, melanocortin receptors, cannabinoid receptors and neuropeptide Y1 and Y5 receptors. In this review, we will discuss drug development efforts for the treatment of obesity, focused on selective melanocortin 4 receptor agonists and neuropeptide Y1 and Y5 receptor antagonists. These drug development efforts required an in-depth understanding of cell-based observations which drive the development of compound structure-activity relationships. These include understanding of receptor function in selected cell-based backgrounds and early evaluation and validation of ex vivo observations in appropriate in vivo models. In order to develop selective and safe anti-obesity drugs, diverse approaches are needed to increase the likelihood of clinical success, including: (i) developing a detailed understanding of the predictive value of rodent pathways for treatment of human disease; (ii) knowledge of the exact location of targeted receptor subtypes for the clinical indication under study in order to derive a suitable compound profile; (iii) predictive measures of in vivo and/or ex vivo receptor occupancy required to bring about a desired physiological effect; (iv) predictive parameters that outline that the drug-derived effects are safe and mechanism-based; and (v) the refinement of selected compound classes, aimed at their clinical use.

Galanin-neuropeptide Y (NPY) interactions in central cardiovascular control: involvement of the NPY Y receptor subtype.

The interactions between neuropeptide Y (NPY), specifically through NPY Y(1) and Y(2) receptor subtypes, and galanin [GAL(1-29)] have been analysed at the cardiovascular level. The cardiovascular effects of intracisternal coinjections of GAL(1-29) with NPY or NPY Y(1) or Y(2) agonists, as well as quantitative receptor autoradiography of the binding characteristics of NPY Y(1) and Y(2) receptor subtypes in the nucleus of the solitary tract (NTS), in the presence or absence of GAL(1-29), have been investigated. The effects of coinjections of GAL(1-29) and the NPY Y(1) agonist on the expression of c-FOS immunoreactivity in the NTS were also studied. The coinjection of NPY with GAL(1-29) induced a significant vasopressor and tachycardic action with a maximum 40% increase (P < 0.001). The coinjection of the NPY Y(1) agonist and GAL(1-29) induced a similar increase in mean arterial pressure and heart rate as did NPY plus GAL(1-29), actions that were not observed with the NPY Y(2) agonist plus GAL(1-29). GAL(1-29), 3 nm, significantly and substantially (by approximately 40%) decreased NPY Y(1) agonist binding in the NTS. This effect was significantly blocked (P < 0.01) in the presence of the specific galanin antagonist M35. The NPY Y(2) agonist binding was not modified in the presence of GAL(1-29). At the c-FOS level, the coinjection of NPY Y(1) and GAL(1-29) significantly reduced the c-FOS-immunoreactive response induced by either of the two peptides. The present findings suggest the existence of a modulatory antagonistic effect of GAL(1-29) mediated via galanin receptors on the NPY Y(1) receptor subtype and its signalling within the NTS.

Paradoxical effects of NPY in the suprachiasmatic nucleus.

The circadian clock in the suprachiasmatic nucleus (SCN) is synchronized by the 24 h, light : dark cycle, and is reset by photic and non-photic cues. The acute effects of light in the SCN include the increase of mRNA levels of the circadian clock gene Per1 and a dramatic reduction of pineal melatonin. Neuropeptide Y (NPY), which appears to mediate the phase-resetting effects of non-photic stimuli, prevents the ability of light, and stimuli that mimic light, to phase shift the circadian clock when injected into the SCN. The purpose of the present study was to determine if NPY inhibits the ability of light to suppress pineal melatonin. Surprisingly, NPY injected into the SCN of hamsters mimicked the effects of light by suppressing pineal melatonin levels. To confirm that NPY inhibited the effects of light on the induction of Per1 mRNA levels, Per1 mRNA levels in the SCN were measured in these same animals. NPY significantly reduced Per1 mRNA levels induced by the light pulse. The suppression of melatonin by NPY appears to be mediated by the same subtype of NPY receptors in the SCN that mediate the modulation of phase shifts. Injection of Y5 receptor agonists mimicked the effects of NPY on pineal melatonin, while injection of a Y2 agonist did not. Thus, these data are the first to demonstrate the paradoxical effects of NPY within the SCN. NPY mimics the effects of light on pineal melatonin and inhibits the effects of light on the induction of Per1 mRNA.