The neuropeptide Y (NPY) system is a multireceptor/multiligand system consisting of four receptors in humans (hY(1), hY(2), hY(4), hY(5)) and three agonists (NPY, PYY, PP) that activate these receptors with different potency. The relevance of this system in diseases like obesity or cancer, and the different role that each receptor plays influencing different biological processes makes this system suitable for the design of subtype selectivity studies. In this review we focus on the latest findings within the NPY system, we summarize recent mutagenesis studies, structure activity relationship studies, receptor chimera, and selective ligands focusing also on the binding mode of the native agonists.

Accumulating data indicate that sensory nerve derived neuropeptides such as substance P and calcitonin gene related-protein (CGRP) can accelerate the progression of endometriosis via their respective receptors, so can agonists to their respective receptors receptor 1 (NK1R), receptor activity modifying protein 1 (RAMP-1) and calcitonin receptor-like receptor (CRLR). Adrenergic β2 receptor (ADRB2) agonists also can facilitate lesional progression. In contrast, women with endometriosis appear to have depressed vagal activity, concordant with reduced expression of α7 nicotinic acetylcholine receptor (α7nAChR). The roles of these receptors in adenomyosis are completely unknown.

Expression of neuropeptide Y (NPY) receptors in human SK-N-MC neuroblastoma cells was investigated. Reverse transcriptase-polymerase chain reaction (RT-PCR) revealed that Y1, Y4, and Y5 receptors were expressed in these cells. Expression was confirmed by Western blot and immunocytochemistry demonstrated abundant presence of all three receptors on cell membranes. NPY peptide was also expressed in these cells, but other members of the larger peptide family (peptide YY and pancreatic polypeptide) were not expressed. Incubation with U0126, a specific mitogen-activated protein kinase (MAPK) inhibitor, decreased cell number in serum-free medium culture. Since NPY activates MAPK via different subtypes of NPY receptors, results suggest that endogenously expressed NPY may control proliferation of these cells through a paracrine/autocrine mechanism.

Neuropeptide Y (NPY) is a well established anticonvulsant and first-in-class antiepileptic neuropeptide. In this study, the controversial role of NPY1 receptors in epilepsy was reassessed by testing two highly selective NPY1 receptor ligands and a mixed NPY1/NPFF receptor antagonist BIBP3226 in a rat model for limbic seizures. While BIBP3226 significantly attenuated the pilocarpine-induced seizures, neither of the highly selective NPY1 receptor ligands altered the seizure severity. Administration of the NPFF1/NPFF2 receptor antagonist RF9 also significantly attenuated limbic seizure activity. To further prove the involvement of NPFF receptors in these seizure-modulating effects, low and high affinity antagonists for the NPFF receptors were tested. We observed that the low affinity ligand failed to exhibit anticonvulsant properties while the two high affinity ligands significantly attenuated the seizures. Continuous NPFF1 receptor agonist administration also inhibited limbic seizures whereas bolus administration of the NPFF1 receptor agonist was without effect. This suggests that continuous agonist perfusion could result in NPFF1 receptor desensitization and mimic NPFF1 receptor antagonist administration. Our data unveil for the first time the involvement of the NPFF system in the management of limbic seizures.

Neuropeptide Y (NPY) is highly abundant in the brain and is released as a co-transmitter with plasticity-related neurotransmitters such as glutamate, GABA and noradrenaline. Functionally, its release is associated with appetite, anxiety, and stress regulation. NPY acting on Y2 receptors (Y2R), facilitates fear extinction, suggesting a role in associative memory. Here, we explored to what extent NPY action at Y2R contributes to hippocampus-dependent spatial memory and found that dorsal intrahippocampal receptor antagonism improved spatial reference memory acquired in a water maze in rats, without affecting anxiety levels, or spontaneous motor activity. Water maze training resulted in an increase of Y2R, but not Y1R expression in the hippocampus. By contrast, in the prefrontal cortex there was a decrease in Y2R, and an increase of Y1R expression. Our results indicate that neuropeptide Y2R are significantly involved in hippocampus-dependent spatial memory and that receptor expression is dynamically regulated by this learning experience. Effects are consistent with a metaplastic contribution of NPY receptors to cumulative spatial learning.

Here we describe a novel mechanism for plasma membrane insertion of the delta opioid receptor (DOR). In small dorsal root ganglion neurons, only low levels of DORs are present on the cell surface, in contrast to high levels of intracellular DORs mainly associated with vesicles containing calcitonin gene-related peptide (CGRP). Activation of surface DORs caused Ca(2+) release from IP(3)-sensitive stores and Ca(2+) entry, resulting in a slow and long-lasting exocytosis, DOR insertion, and CGRP release. In contrast, membrane depolarization or activation of vanilloid and P2Y(1) receptors induced a rapid DOR insertion. Thus, DOR activation induces a Ca(2+)-dependent insertion of DORs that is coupled to a release of excitatory neuropeptides, suggesting that treatment of inflammatory pain should include blockade of DORs.

Mechanisms underlying long-term sustained weight loss and glycemic normalization after obesity surgery include changes in gut hormone levels, including glucagon-like peptide 1 (GLP-1) and peptide YY (PYY). We demonstrate that two peptide biased agonists (GEP44 and GEP12) of the GLP-1, neuropeptide Y1, and neuropeptide Y2 receptors (GLP-1R, Y1-R, and Y2-R, respectively) elicit Y1-R antagonist-controlled, GLP-1R-dependent stimulation of insulin secretion in both rat and human pancreatic islets, thus revealing the counteracting effects of Y1-R and GLP-1R agonism. These agonists also promote insulin-independent Y1-R-mediated glucose uptake in muscle tissue ex vivo and more profound reductions in food intake and body weight than liraglutide when administered to diet-induced obese rats. Our findings support a role for Y1-R signaling in glucoregulation and highlight the therapeutic potential of simultaneous receptor targeting to achieve long-term benefits for millions of patients.

Neuropeptide Y (NPY) prominently inhibits epileptic seizures in different animal models. The NPY receptors mediating this effect remain controversial partially due to lack of highly selective agonists and antagonists. To circumvent this problem, we used various NPY receptor knockout mice with the same genetic background and explored anti-epileptic action of NPY in vitro and in vivo. In Y2 (Y2-/-) and Y5 (Y5-/-) receptor knockouts, NPY partially inhibited 0 Mg2+-induced epileptiform activity in hippocampal slices. In contrast, in double knockouts (Y2Y5-/-), NPY had no effect, suggesting that in the hippocampus in vitro both receptors mediate anti-epileptiform action of NPY in an additive manner. Systemic kainate induced more severe seizures in Y5-/- and Y2Y5-/-, but not in Y2-/- mice, as compared to wild-type mice. Moreover, kainate seizures were aggravated by administration of the Y5 antagonist L-152,804 in wild-type mice. In Y5-/- mice, hippocampal kindling progressed faster, and afterdischarge durations were longer in amygdala, but not in hippocampus, as compared to wild-type controls. Taken together, these data suggest that, in mice, both Y2 and Y5 receptors regulate hippocampal seizures in vitro, while activation of Y5 receptors in extra-hippocampal regions reduces generalized seizures in vivo.

The mammalian circadian clock in the suprachiasmatic nucleus (SCN) is a heterogeneous structure. Two key populations of cells that receive retinal input and are believed to participate in circadian responses to light are cells that contain vasoactive intestinal polypeptide (VIP) and gastrin-releasing peptide (GRP). VIP acts primarily through the VPAC2 receptor, while GRP works primarily through the BB2 receptor. Both VIP and GRP phase shift the circadian clock in a manner similar to light when applied to the SCN, both in vivo and in vitro, indicating that they are sufficient to elicit photic-like phase shifts. However, it is not known if they are necessary signals for light to elicit phase shifts. Here we test the hypothesis that GRP and VIP are necessary signaling components for the photic phase shifting of the hamster circadian clock by examining two antagonists for each of these neuropeptides. The BB2 antagonist PD176252 had no effect on light-induced delays on its own, while the BB2 antagonist RC-3095 had the unexpected effect of significantly potentiating both phase delays and advances. Neither of the VIP antagonists ([d-p-Cl-Phe6, Leu17]-VIP, or PG99-465) altered phase shifting responses to light on their own. When the BB2 antagonist PD176252 and the VPAC2 antagonist PG99-465 were delivered together to the SCN, phase delays were significantly attenuated. These results indicate that photic phase shifting requires participation of either VIP or GRP; phase shifts to light are only impaired when signalling in both pathways are inhibited. Additionally, the unexpected potentiation of light-induced phase shifts by RC-3095 should be investigated further for potential chronobiotic applications.

Neuropeptide Y (NPY) receptors are present in cardiac membranes. However, its physiological roles in the heart are not clear. The aim of this study was to define the direct effects of pancreatic polypeptide (PP) on atrial dynamics and atrial natriuretic peptide (ANP) release in perfused beating atria. Pancreatic polypeptides, a NPY Y(4) receptor agonist, decreased atrial contractility but was not dose-dependent. The ANP release was stimulated by PP in a dose-dependent manner. GR 23118, a NPY Y(4) receptor agonist, also increased the ANP release and the potency was greater than PP. In contrast, peptide YY (3-36) (PYY), an NPY Y(2) receptor agonist, suppressed the release of ANP with positive inotropy. NPY, an agonist for Y(1, 2, 5) receptor, did not cause any significant changes. The pretreatment of NPY (18-36), an antagonist for NPY Y(3) receptor, markedly attenuated the stimulation of ANP release by PP but did not affect the suppression of ANP release by PYY. BIIE0246, an antagonist for NPY Y(2) receptor, attenuated the suppression of ANP release by PYY. The responsiveness of atrial contractility to PP or PYY was not affected by either of the antagonists. These results suggest that NPY Y(4) and Y(2) receptor differently regulate the release of atrial ANP.

Female Aedes aegypti mosquitoes are the principal vector for dengue fever, causing 50-100 million infections per year, transmitted between human and mosquito by blood feeding. Ae. aegypti host-seeking behavior is known to be inhibited for three days following a blood meal by a hemolymph-borne humoral factor. Head Peptide-I is a candidate peptide mediating this suppression, but the mechanism by which this peptide alters mosquito behavior and the receptor through which it signals are unknown.

Developmental exposure to selective serotonin reuptake inhibitor (SSRI) increases the risk of Autism Spectrum Disorder (ASD), however, the underlying neurobiology of this effect is not fully understood. Here we used the socially monogamous prairie vole as a translational model of developmental SSRI exposure. Paired female prairie voles (n = 20) were treated with 5 mg/kg subcutaneous fluoxetine (FLX) or saline (SAL) daily from birth of the second litter until the day of birth of the 4th litter. This design created three cohorts of FLX exposure: postnatal exposure in litter 2, both prenatal and postnatal exposure in litter 3, and prenatal exposure in litter 4. Post-weaning, subjects underwent behavioral testing to detect changes in sociality, repetitive behavior, pair-bond formation, and anxiety-like behavior. Quantitative receptor autoradiography was performed for oxytocin, vasopressin 1a, and serotonin 1a receptor density in a subset of brains. We observed increased anxiety-like behavior and reduced sociality in developmentally FLX exposed adults. FLX exposure decreased oxytocin receptor binding in the nucleus accumbens core and central amygdala, and vasopressin 1a receptor binding in the medial amygdala. FLX exposure did not affect serotonin 1A receptor binding in any areas examined. Changes to oxytocin and vasopressin receptors may underlie the behavioral changes observed and have translational implications for the mechanism of the increased risk of ASD subsequent to prenatal SSRI exposure.

The vertebrate gene family for neuropeptide Y (NPY) receptors expanded by duplication of the chromosome carrying the ancestral Y1-Y2-Y5 gene triplet. After loss of some duplicates, the ancestral jawed vertebrate had seven receptor subtypes forming the Y1 (including Y1, Y4, Y6, Y8), Y2 (including Y2, Y7) and Y5 (only Y5) subfamilies. Lampreys are considered to have experienced the same chromosome duplications as gnathostomes and should also be expected to have multiple receptor genes. However, previously only a Y4-like and a Y5 receptor have been cloned and characterized. Here we report the cloning and characterization of two additional receptors from the sea lamprey Petromyzon marinus. Sequence phylogeny alone could not with certainty assign their identity, but based on synteny comparisons of P. marinus and the Arctic lamprey, Lethenteron camtschaticum, with jawed vertebrates, the two receptors most likely are Y1 and Y2. Both receptors were expressed in human HEK293 cells and inositol phosphate assays were performed to determine the response to the three native lamprey peptides NPY, PYY and PMY. The three peptides have similar potencies in the nanomolar range for Y1. No obvious response to the three peptides was detected for Y2. Synteny analysis supports identification of the previously cloned receptor as Y4. No additional NPY receptor genes could be identified in the presently available lamprey genome assemblies. Thus, four NPY-family receptors have been identified in lampreys, orthologs of the same subtypes as in humans (Y1, Y2, Y4 and Y5), whereas many other vertebrate lineages have retained additional ancestral subtypes.

The hypothalamic neuropeptide 26RFa is the most recently identified member of the RFamide peptide family, and this 26RFa signaling system has been shown to be implicated in regulating a variety of physiological processes. In zebrafish，26RFa and two putative receptors, DrGPR103A and DrGPR103B, have been in silico identified, and in vivo data derived from overexpression and loss of function mutation experiments suggest the 26RFa signaling system plays an important role in the hypothalamic regulation of sleep. However, the biochemical and pharmacological information on DrGPR103A/B receptors is still unknown. Here, after cloning of cDNAs of two putative 26RFa receptor genes, DrGPR103A and B, from the total RNA of zebrafish whole body, functional assays demonstrated that both receptors were activated by synthetic zebrafish 26RFa neuropeptide, leading to a significant increase in CRE-driven luciferase activity and intracellular Ca2+ mobilization in a Gαq inhibitor- and Gαi/o inhibitor-sensitive manner. Upon activation by 26RFa, DrGPR103A and B evoked ERK1/2 phosphorylation and underwent internalization. Further functional determination also revealed that zebrafish kisspeptin-1 exhibited a slight potency for activating both DrGPR103A and B, and vice versa, zebrafish 26RFa also showed some activity at zebrafish GPR54A and B. Our findings provided evidence that zebrafish GPR103A and B are two functional Gαq- and Gαi/o-dually coupled receptors for 26RFa, enabling the further elucidation of the endocrinological roles of zebrafish 26RFa signaling system in the regulation of physiological activities.

Morphine suppresses a number of immune parameters, such as natural killer (NK) cell activity and lymphocyte proliferation, by acting through mu-opioid receptors in the central nervous system. Prior studies have implicated the sympathetic nervous system in mediating the immunomodulatory effects of acute morphine treatment. However, the peripheral mechanism whereby morphine inhibits NK cell activity is not fully understood. The aim of the present study was to investigate the role of the sympathetic transmitter neuropeptide Y (NPY) in mediating morphine-induced immune alterations. The results showed that administration of the selective NPY Y1 receptor antagonist BIBP3226 blocked morphine's effect on splenic NK activity but did not attenuate the suppression splenocyte proliferative responses to Con-A or LPS. Furthermore, intravenous NPY administration produced a dose-dependent inhibition of splenic NK activity but did not suppress lymphocyte proliferation. Recent studies from our laboratory have demonstrated that morphine modulates NK activity through a central mechanism that requires the activation of dopamine D1 receptors in the nucleus accumbens. Results from the present study showed that microinjection of the D1 receptor agonist SKF-38393 into the nucleus accumbens shell induced a suppression of NK activity that was reversed by BIBP3226. Collectively, these findings demonstrate that NPY Y1 receptors mediate morphine's suppressive effect on NK activity and further suggest that opioid-induced increases in nucleus accumbens D1 receptor activation inhibit splenic NK activity via NPY released from the sympathetic nervous system.

A radioiodinated analogue of neuropeptide FF, [125I][D. Tyr1,(NMe) Phe3]neuropeptide FF, was used as a selective probe to label neuropeptide FF receptors in the rat spinal cord. Following neonatal capsaicin treatment, dorsal rhizotomy or sciatic nerve section, the distribution and possible alterations of spinal cord specific [125I][D.Tyr1,(NMe)Phe3]neuropeptide FF binding sites were evaluated using in vitro quantitative receptor autoradiography. In normal rats, the highest densities of sites were observed in the superficial layers of the dorsal horn (laminae I-II) whereas moderate to low amounts of labelling were seen in the deeper (III-VI) laminae, around the central canal, and in the ventral horn. Capsaicin-treated rats showed a bilateral decrease (47%) in [125I][D.Tyr1,(NMe)Phe3]neuropeptide FF binding in all spinal areas. Unilateral sciatic nerve section and unilateral dorsal rhizotomy induced significant depletions (15-27%) in [125I][D.Tyr1,(NMe)Phe3]neuropeptide FF labelling in the ipsilateral dorsal horn. These results suggest that a proportion of neuropeptide FF receptors is located on primary afferent terminals of the dorsal horn and could thus play a role in the modulation of nociceptive transmission.

Neuropeptide FF1 and FF2 receptors (NPFF1-R and NPFF2-R), and their endogenous ligand NPFF, are one of only several systems responsible for mediating opioid-induced hyperalgesia, tolerance, and dependence. Currently, no small molecules displaying good affinity or selectivity for either subtype have been reported, to decipher the role of NPFF2-R as it relates to opioid-mediated analgesia, for further exploration of NPFF1-R, or for medication development for either subtype. We report the first nonpeptide small molecule scaffold for NPFF1,2-R, the guanidino-piperidines, and SAR studies resulting in the discovery of a NPFF1 agonist (7b, K(i) = 487 ± 117 nM), a NPFF1 antagonist (46, K(i) = 81 ± 17 nM), and a NPFF2 partial antagonist (53a, K(i) = 30 ± 5 nM), which serve as leads for the development of pharmacological probes and potential therapeutic agents. Testing of 46 alone was without effect in the mouse 48 °C warm-water tail-withdrawal test, but pretreatment with 46 prevented NPFF-induced hyperalgesia.

Neuropeptide Y and peptide YY are important central and peripheral modulators of cardiovascular and neuroendocrine functions, that act through multiple receptor subtypes, Y1 through Y5. A neuropeptide Y-binding site of the Y2 type was characterized by ligand-binding studies in isolated nerve terminals from the rat neurohypophysis. Functionally, neuropeptide Y and peptide YY dose-dependently triggered arginine 8-vasopressin and oxytocin release from perfused isolated terminals, and potentiated the arginine-8-vasopressin release induced by depolarization. Osmotic stimulation by salt loading of rats for two and seven days caused a more than three-fold increase in the neuropeptide Y content of the nerve endings. However, the Y2 receptor expression and arginine-8-vasopressin content declined, showing that the neuropeptide Y system is dynamic and suggesting that it plays a physiological role in salt and water homeostasis. Two sets of observations suggest the arginine-8-vasopressin release by neuropeptide Y may not be explained by neuropeptide Y effects on intracellular Ca2+. First, absence of Ca2+ from the perfusion medium did not affect the arginine-8-vasopressin release, and secondly neuropeptide Y did not change intraterminal Ca2+ concentrations. Pretreatment with pertussis toxin blocked arginine-8-vasopressin secretion by neuropeptide Y, suggesting activation of Gi or Go heterotrimeric G-proteins are required for secretion. It is concluded, that the nerve endings of the neurohypophysis contain a complete neuropeptide Y system with ligand and receptors. Neuropeptide Y may act in an autocrine fashion via activation of Y2 neuropeptide Y receptors to stimulate the release of vasopressin and oxytocin via a Gi/Go dependent secretory mechanism.

Caloric restriction is an anti-aging intervention known to extend lifespan in several experimental models, at least in part, by stimulating autophagy. Caloric restriction increases neuropeptide Y (NPY) in the hypothalamus and plasma ghrelin, a peripheral gut hormone that acts in hypothalamus to modulate energy homeostasis. NPY and ghrelin have been shown to be neuroprotective in different brain areas and to induce several physiological modifications similar to those induced by caloric restriction. However, the effect of NPY and ghrelin in autophagy in cortical neurons is currently not known. Using a cell culture of rat cortical neurons we investigate the involvement of NPY and ghrelin in caloric restriction-induced autophagy. We observed that a caloric restriction mimetic cell culture medium stimulates autophagy in rat cortical neurons and NPY or ghrelin receptor antagonists blocked this effect. On the other hand, exogenous NPY or ghrelin stimulate autophagy in rat cortical neurons. Moreover, NPY mediates the stimulatory effect of ghrelin on autophagy in rat cortical neurons. Since autophagy impairment occurs in aging and age-related neurodegenerative diseases, NPY and ghrelin synergistic effect on autophagy stimulation may suggest a new strategy to delay aging process.

Neuropeptide Y (NPY) is a pleiotropic peptide, which is involved in many biological mechanisms important in regulation of cell growth and survival. The aim of this study was a comprehensive analysis of the NPY system in prostate pathology.