Can Neuropeptide S Be an Indicator for Assessing Anxiety in Psychiatric Disorders?

Neuropeptide S (NPS) is a neuropeptide primarily produced within three brainstem regions including locus coeruleus, trigeminal nerve nucleus, and lateral parabrachial nucleus. NPS is involved in the central regulation of stress, fear, and cognitive integration. NPS is a mediator of behavior, seeking food, and the proliferation of new adipocytes in the setting of obesity. So far, current research of NPS is only limited to animal models; data regarding its functions in humans is still scarce. Animal studies showed that anxiety and appetite might be suppressed by the action of NPS. The discovery of this neuromodulator peptide is effective considering its strong anxiolytic action, which has the potential to be an interesting therapeutic option in treating neuropsychiatric disorders. In this article, we aimed to analyze the pharmaceutical properties of NPS as well as its influence on several neurophysiological aspects-modulation of behavior, association with obesity, as well as its potential application in rehabilitation and treatment of psychiatric disorders.

MRGPRX2 Is the Codeine Receptor of Human Skin Mast Cells: Desensitization through ß-Arrestin and Lack of Correlation with the FcεRI Pathway.

Codeine stimulates skin mast cells and is therefore used in skin tests and as an inducer of experimental itch. MRGPRX2 responds to various drugs, including opioids, to elicit pseudoallergic reactions, but whether it represents the main opiate receptor of skin mast cells remains unknown. By combining a number of approaches, including the silencing of MRGPRX2, we now report that MRGPRX2 is indeed the dominant codeine receptor of dermal mast cells. Activation by codeine displayed profound subject variability and correlated with secretion elicited by compound 48/80 or substance P but not by FcεRI aggregation. Degranulation by codeine was attenuated by stem cell factor, whereas the opposite was found for FcεRI. Compound 48/80 or codeine alone was able to achieve maximum MRGPRX2 activation. MRGPRX2 was rapidly internalized on codeine binding in a ß-arrestin-1‒dependent manner. Codeine-triggered ß-arrestin activation was also established by the Tango assay. Prestimulation with MRGPRX2 agonists (but not C3a or FcεRI aggregation) resulted in refractoriness to further stimulation by the same or another MRGPRX2 ligand (cross desensitization). This was duplicated in a cell line (RBL-MRGPRX2). Collectively, codeine degranulates skin mast cells through MRGPRX2, at which it acts as a balanced ligand. It has yet to be determined whether codeine-induced refractoriness could be exploited to desensitize MRGPRX2 to prevent severe pseudoallergic reactions.

MRGPRX2 signals its importance in cutaneous mast cell biology: Does MRGPRX2 connect mast cells and atopic dermatitis?

The discovery of MRGPRX2 marks an important change in MC biology, explaining non-IgE-mediated clinical phenomena relying on MCs. As receptor for multiple drugs, MRGPRX2 is crucial to drug-induced hypersensitivity. However, not only drugs, but also endogenous mediators like neuropeptides and host defense peptides activate MRGPRX2, suggesting its broad impact in cutaneous pathophysiology. Here, we give a brief overview of MRGPRX2 and its regulation by microenvironmental stimuli, which support MCs and can be altered in skin disorders, and briefly touch on the functional programs elicited by MRGPRX2 ligation. Studies in Mrgprb2-deficient mice (the murine ortholog) help illuminate MRGPRX2's function in health and disease. Recent advances in this model support the long-suspected operational unit between MCs and nerves, with MRGPRX2 being a vital component. Based on the limited evidence for a major contribution of FcεRI/IgE-activated MCs to atopic dermatitis (AD), we develop the hypothesis that MRGPRX2 constitutes the missing link connecting MCs and AD, at least in selected endotypes. Support comes from the multifold changes in the MC-neuronal system of AD skin (eg greater density of MCs and closer connections between MCs and nerves, increased PAR-2/Substance P). We theorize that these deregulations suffice to initiate AD, but external triggers, many of which activating MRGPRX2 themselves (eg Staphylococcus aureus) further feed into the loop. Itch, the most burdensome hallmark of AD, is mostly non-histaminergic but tryptase-dependent, and tryptase is preferentially released upon MRGPRX2 activation. Because MRGPRX2 is a very active research field, some of the existing gaps are likely to be closed soon.

Sociability and extinction of conditioned social fear is affected in neuropeptide S receptor-deficient mice.

Being cautious of unfamiliar conspecifics is adaptive because sick or aggressive conspecifics may jeopardize survival and well-being. However, prolonged or excessive caution, i.e. fear related to social situations, is maladaptive and may result in social anxiety disorder. Some anxiety disorders in humans are associated with polymorphisms of the neuropeptide S receptor (NPSR) gene. In line with this finding, animal studies showed an important role of NPS and NPSR in anxiety and fear. The present study investigated the role of NPSR deficiency in social behavior under non-aversive and aversive conditions. For this, female and male NPSR-deficient mice were tested for (1) sociability and social novelty and (2) acquisition, expression, and extinction of conditioned social fear. The present study revealed very particular effects of the NPSR genotype: Sociability was reduced in female heterozygous NPSR-deficient mice, but was unaffected in males and the other genotypes. Furthermore, the NPSR genotype did not affect the acquisition and expression of conditioned social fear, but its extinction was impaired in heterozygous and facilitated in homozygous NPSR-deficient mice. This indicates that the NPS system plays a role in social behavior under non-aversive and aversive conditions, partly in a sex-dependent manner. The present findings may help to explain social symptoms in anxiety disorders associated with the NPSR genotype.

Motilin: a panoply of communications between the gut, brain, and pancreas.

Introduction: Motilin was first alluded to nearly a century ago. But it remains a rather abstruse peptide, in the shadow of its younger but more lucid 'cousin' ghrelin.Areas covered: The review aimed to bring to the fore multifarious aspects of motilin research with a view to aiding prioritization of future studies on this gastrointestinal peptide.Expert opinion: Growing evidence indicates that rodents (mice, rats, guinea pigs) do not have functional motilin system and, hence, studies in these species are likely to have a minimal translational impact. Both the active peptide and motilin receptor were initially localized to the upper gastrointestinal tract only but more recently - also to the brain (in both humans and other mammals with functional motilin system). Motilin is now indisputably implicated in interdigestive contractile activity of the gastrointestinal tract (in particular, gastric phase III of the migrating motor complex). Beyond this role, evidence is building that there is a cross-talk between motilin system and the brain-pancreas axis, suggesting that motilin exerts not only contractile but also orexigenic and insulin secretagogue actions.

The influence of a new derivate of kisspeptin-10 - Kissorphin (KSO) on the rewarding effects of morphine in the conditioned place preference (CPP) test in male rats.

Kissorphin (KSO) is a new peptide derived from kisspeptin-10. Previous study has indicated that this peptide displays neuropeptide FF (NPFF)-like anti-opioid activity. Herein, we examined the influence of KSO (1; 3, and 10 nmol, intravenously [i.v.]), on the rewarding action of morphine (5 mg/kg, intraperitoneally [i.p.]), using the unbiased design of the conditioned place preference (CPP) paradigm in rats. To test the effect of KSO on the acquisition of morphine-induced CPP, KSO and morphine were co-injected during conditioning with no drugs treatment on the test day. To investigate the effect of KSO on the expression of morphine-induced CPP, morphine alone was given during the conditioning phase (1 × 3 days) and KSO was administered 5 min prior to the placement in the CPP apparatus on the test day. To estimate the influence of KSO on the reinstatement of morphine-induced CPP, KSO was given 5 min before a priming dose of morphine (5 mg/kg, i.p.) on the reinstatement test day. The results show that KSO inhibited the acquisition, expression and reinstatement of morphine-induced CPP. The strongest effect of KSO was observed at the dose of 10 nmol (acquisition and reinstatement) or 1 nmol (expression). KSO given alone, neither induced place preference, nor aversion. Furthermore, the morphine-modulating effects of KSO were markedly antagonized by pretreatment with RF9 (10 nmol, i.v.), the NPFF receptors selective antagonist. Thus, KSO inhibited the morphine-induced CPP mainly by involving specific activation of NPFF receptors. Overall, these data further support the anti-opioid character of KSO.

Antagonistic Effect of Short Neuropeptide F on Allatotropin-Inhibited Feeding Motivation of the Silkworm Larva, Bombyx mori.

Here, we demonstrated an antagonistic effect of short neuropeptide F (sNPF) in modulating feeding motivation in the silkworm Bombyx mori; sNPF reduced the feeding-delaying effects caused by administration of an inhibitory peptide, allatotropin (AT). In situ hybridization and MALDI-TOF MS analysis revealed the presence of three subtypes of sNPFs (sNPF-1, -2, and -3) in the midgut enteroendocrine cells. Ca2+-imaging analyses revealed that three subtypes of sNPF receptors (sNPFRs) (BNGR-A7, -A10, and -A11) showed different affinities with the three subtypes of sNPFs. In addition, sNPF activated its signaling via ERK phosphorylation in the midgut, while mixture of sNPF and AT reduced the phosphorylation level, agreeing with the results of behavioral assay. Together, our current findings suggest that intestinal sNPF positively modulates the feeding motivation by reducing the inhibitory effects by AT within the midgut.

In vitro and in vivo characterization of the bifunctional µ- and δ- opioid receptors ligand MCRT on mouse gastrointestinal motility.

BACKGROUND: Chimeric opioid MCRT was a novel multi-target ligand based on morphiceptin and PFRTic-NH2, and produced potent analgesia (ED50 = 0.03 nmol/mouse) with less upper gastrointestinal dysmotility. In this study, we sought to perform the tests to evaluate the pharmacological effects of MCRT on distal colon motility and defecation function. Moreover, opioid receptor antagonists and neuropeptide FF (NPFF) receptor antagonists were utilized to explore the mechanisms. METHODS: Isolated mouse colon bioassay and colonic bead expulsion were to characterize MCRT-induced inhibition of colonic motility in vitro and in vivo, respectively. Fecal pellet output was to evaluate the defecation function. RESULTS: (1) In vitro, MCRT increased colonic contraction via µ- and δ- opioid receptors (MOR and DOR). (2) In vivo, MCRT delayed colonic bead expulsion (ED50 = 1.1 nmol/mouse) independent of opioid and NPFF receptors. (3) In vivo, MCRT inhibited fecal number (ED50 = 1.43 nmol/mouse) and dry weight (ED50 = 1.63 nmol/mouse), which was mediated by DOR partially but not MOR. CONCLUSIONS: (1) Data indicated that MCRT was less prone to induce gastrointestinal dysmotility at analgesic doses, and provided a possibility for safer opioid analgesic. (2) Based on the mechanism explorations, we speculated on the existence of such an opioid receptor subtype or MOR/DOR heterodimer, which was involved in the central analgesia and the in vitro colonic contractions but not the central colonic dysmotility.

Current understanding of trigeminal ganglion structure and function in headache.

INTRODUCTION: The trigeminal ganglion is unique among the somatosensory ganglia regarding its topography, structure, composition and possibly some functional properties of its cellular components. Being mainly responsible for the sensory innervation of the anterior regions of the head, it is a major target for headache research. One intriguing question is if the trigeminal ganglion is merely a transition site for sensory information from the periphery to the central nervous system, or if intracellular modulatory mechanisms and intercellular signaling are capable of controlling sensory information relevant for the pathophysiology of headaches. METHODS: An online search based on PubMed was made using the keyword "trigeminal ganglion" in combination with "anatomy", "headache", "migraine", "neuropeptides", "receptors" and "signaling". From the relevant literature, further references were selected in view of their relevance for headache mechanisms. The essential information was organized based on location and cell types of the trigeminal ganglion, neuropeptides, receptors for signaling molecules, signaling mechanisms, and their possible relevance for headache generation. RESULTS: The trigeminal ganglion consists of clusters of sensory neurons and their peripheral and central axon processes, which are arranged according to the three trigeminal partitions V1-V3. The neurons are surrounded by satellite glial cells, the axons by Schwann cells. In addition, macrophage-like cells can be found in the trigeminal ganglion. Neurons express various neuropeptides, among which calcitonin gene-related peptide is the most prominent in terms of its prevalence and its role in primary headaches. The classical calcitonin gene-related peptide receptors are expressed in non-calcitonin gene-related peptide neurons and satellite glial cells, although the possibility of a second calcitonin gene-related peptide receptor in calcitonin gene-related peptide neurons remains to be investigated. A variety of other signal molecules like adenosine triphosphate, nitric oxide, cytokines, and neurotrophic factors are released from trigeminal ganglion cells and may act at receptors on adjacent neurons or satellite glial cells. CONCLUSIONS: The trigeminal ganglion may act as an integrative organ. The morphological and functional arrangement of trigeminal ganglion cells suggests that intercellular and possibly also autocrine signaling mechanisms interact with intracellular mechanisms, including gene expression, to modulate sensory information. Receptors and neurotrophic factors delivered to the periphery or the trigeminal brainstem can contribute to peripheral and central sensitization, as in the case of primary headaches. The trigeminal ganglion as a target of drug action outside the blood-brain barrier should therefore be taken into account.

Neuropeptide receptors as potential pharmacological targets for obesity.

Obesity is a chronic multifactorial disease, characterized by an excessive accumulation of adipose tissue. It is usually the result of excessive food intake and/or low energy expenditure. Obesity can be triggered by lifestyle, nutritional, genetic, environmental, hormonal and psychological factors. Several strategies are used to treat obesity, including dietary reeducation, with balanced food intake, increased physical exercise, in order to promote energy expenditure and to overcome the insufficiency in weight reduction by other strategies, and administration of drugs. However, these medications are associated to undesirable side effects, resulting in a high withdrawal rate. Several studies have been focused on the development of compounds that act in the hypothalamic region where the center of the regulation of hunger and satiety is located. Some of them target the activity of endogenous peptides, such as ghrelin pancreatic polypeptide, peptide YY and neuropeptide Y, as well as their receptors. This review addresses the importance of understanding the neuropeptide/peptide hormones and their receptors for the development of novel anti-obesity compounds that may aid in weight reduction as a promising alternative for the treatment of obesity.

Modulation of Drosophila post-feeding physiology and behavior by the neuropeptide leucokinin.

Behavior and physiology are orchestrated by neuropeptides acting as central neuromodulators and circulating hormones. An outstanding question is how these neuropeptides function to coordinate complex and competing behaviors. In Drosophila, the neuropeptide leucokinin (LK) modulates diverse functions, but mechanisms underlying these complex interactions remain poorly understood. As a first step towards understanding these mechanisms, we delineated LK circuitry that governs various aspects of post-feeding physiology and behavior. We found that impaired LK signaling in Lk and Lk receptor (Lkr) mutants affects diverse but coordinated processes, including regulation of stress, water homeostasis, feeding, locomotor activity, and metabolic rate. Next, we sought to define the populations of LK neurons that contribute to the different aspects of this physiology. We find that the calcium activity in abdominal ganglia LK neurons (ABLKs), but not in the two sets of brain neurons, increases specifically following water consumption, suggesting that ABLKs regulate water homeostasis and its associated physiology. To identify targets of LK peptide, we mapped the distribution of Lkr expression, mined a brain single-cell transcriptome dataset for genes coexpressed with Lkr, and identified synaptic partners of LK neurons. Lkr expression in the brain insulin-producing cells (IPCs), gut, renal tubules and chemosensory cells, correlates well with regulatory roles detected in the Lk and Lkr mutants. Furthermore, these mutants and flies with targeted knockdown of Lkr in IPCs displayed altered expression of insulin-like peptides (DILPs) and transcripts in IPCs and increased starvation resistance. Thus, some effects of LK signaling appear to occur via DILP action. Collectively, our data suggest that the three sets of LK neurons have different targets, but modulate the establishment of post-prandial homeostasis by regulating distinct physiological processes and behaviors such as diuresis, metabolism, organismal activity and insulin signaling. These findings provide a platform for investigating feeding-related neuroendocrine regulation of vital behavior and physiology.

Sex-Biased Physiological Roles of NPFF1R, the Canonical Receptor of RFRP-3, in Food Intake and Metabolic Homeostasis Revealed by its Congenital Ablation in mice.

BACKGROUND: RF-amide-related peptide-3 (RFRP-3), the mammalian ortholog of gonadotropin-inhibiting hormone, operates as inhibitory signal for the reproductive axis. Recently, RFRP-3 has been also suggested to stimulate feeding, and therefore might contribute to the control of body weight and its alterations. Yet, characterization of the metabolic actions of RFRP-3 has been so far superficial and mostly pharmacological. Here, we aim to investigate the physiological roles of RFRP-3 signaling in the control of feeding and metabolic homeostasis using a novel mouse model of genetic ablation of its canonical receptor, NPFF1R. METHODS: Food intake, body weight gain and composition, and key metabolic parameters, including glucose tolerance and insulin sensitivity, were monitored in mice with constitutive inactivation of NPFF1R. RESULTS: Congenital elimination of NPFF1R in male mice resulted in changes in feeding patterns, with a decrease in spontaneous food intake and altered responses to leptin and ghrelin: leptin-induced feeding suppression was exaggerated in NPFF1R null mice, whereas orexigenic responses to ghrelin were partially blunted. Concordant with this pro-anorectic phenotype, hypothalamic expression of Pomc was increased in NPFF1R null mice. In contrast, spontaneous feeding and neuropeptide expression remained unaltered in NPFF1R KO female mice. Despite propensity for reduced feeding, ablation of NPFF1R signaling in male mice did not cause overt alterations in body weight (BW) gain or composition, neither it affected BW responses to high fat diet (HFD), total energy expenditure or RQ ratios. Yet, NPFF1R KO males showed a decrease in locomotor activity. Conversely, NPFF1R null female mice tended to be heavier and displayed exaggerated BW increases in response to obesogenic insults, such as HFD or ovariectomy. These were associated to increased fat mass, decreased total energy expenditure in HFD, and unaltered RQ ratios or spontaneous locomotor activity. Finally, lack of NPFF1R signaling worsened the metabolic impact of HFD on glycemic homeostasis in males, as revealed by impaired glucose tolerance and insulin sensitivity, while female mice remained unaffected. CONCLUSION: Our data support a discernible orexigenic role of NPFF1R signaling selectively in males, which might modulate the effects of leptin and ghrelin on food intake. In addition, our study is the first to disclose the sex-biased, deleterious impact of the lack of NPFF1R signaling on body weight and fat composition, energy expenditure, locomotor activity and glucose balance, which exaggerates some of the metabolic consequences of concurrent obesogenic insults, such as HFD, in a sexually dimorphic manner. SUMMARY OF TRANSLATIONAL RELEVANCE: Our data are the first to document the nature and magnitude of the regulatory actions of RFRP-3/NPFF1R signaling in the control of feeding and metabolic homeostasis in a physiological setting. Our results not only suggest an orexigenic action of endogenous RFRP-3, specifically in males, but reveal also the detrimental impact of ablation of NPFF1R signaling on body composition, energy expenditure, locomotor activity or glucose balance, especially when concurrent with other obesogenic insults, as HFD, thereby providing the first evidence for additional metabolic effects of RFRP-3, other that the mere control of feeding. Interestingly, alterations of such key metabolic parameters occurred in a sex-biased manner, with males being more sensitive to deregulation of locomotor activity and glycemic control, while females displayed clearer obesogenic responses and deregulated energy expenditure. While our study cannot discard the possibility of RFRP-3 actions via alternative pathways, such as NPFF2R, our data pave the way for future analyses addressing the eventual contribution of altered RFRP-3/NPFF1R signaling in the development of metabolic alterations (including obesity and its comorbidities), especially in conditions associated to reproductive dysfunction.

RFamide-related peptide-3 promotes alpha TC1 clone 6 cell survival likely via GPR147.

Type 2 diabetes mellitus (T2DM) is closely related to islet alpha cell mass and viability. Several types of RFamide-related peptides (RFRPs) are involved in regulating proliferation and function of islet cells. However, current understanding of the role of RFamide-related peptide-3 (RFRP-3) in pancreatic alpha cells is limited. Therefore, we investigated the expression of the RFRP-3 receptor, G protein-coupled receptor 147 (GPR147), in mouse islets and alpha TC1 clone 6 cells, and evaluated the function of RFRP-3 on alpha cells. We show that GPR147 is expressed in mouse islets and alpha cell lines. In addition, RFRP-3 promotes survival of alpha cells under conditions of hyperglycemia and serum starvation. Mechanistic evidence demonstrates that RFRP-3 activated PI3K/AKT and ERK1/2 signaling cascades and treatment with an antagonist of GPR147, 1-adamantanecarbonyl-Arg-Phe-NH2 (RF9) blocked this function. These findings indicate a novel effect of RFRP-3 in promoting alpha cell survival, likely via GPR147.

A subtype-specific neuropeptide FF receptor antagonist attenuates morphine and nicotine withdrawal syndrome in the rat.

Considerable evidence suggests the Neuropeptide FF (NPFF) and related peptides exert pro-nociceptive and anti-opiate actions, particularly at the supra-spinal level, which may contribute to opiate dependence. The FF1 receptor subtype appears to be primarily responsible for anti-opiate effects. In contrast, stimulation of the FF2 receptor primarily induces pro-opiate effects. AC-262620 is a small molecule, systemically active, selective FF1 receptor antagonist. An initial experiment showed that 10 mg/kg i.p. AC-262620 significantly reduced subsequent naloxone-precipitated somatically expressed withdrawal signs in rats infused s.c. for seven days with 0.3 mg/kg/hr morphine sulfate. A second experiment showed that the same dose of AC-262620 significantly reduced subsequent spontaneous withdrawal signs 23.75 h after termination of seven days s.c. infusion of 0.6 mg/kg/hr morphine sulfate. Chronic nicotine intake may contribute to dependence by overstimulating opiate receptors through release of opiate peptides. By analogy to opiate dependence, it was hypothesized that FF1 receptor activation contributes to nicotine dependence and withdrawal syndrome. AC-262620 significantly reduced somatically expressed withdrawal signs precipitated by the nicotinic antagonist mecamylamine in rats infused for seven days with nicotine bitartrate. Taken together, these findings suggest that NPFF or related neuropeptides contribute to opiate, as well as nicotine, dependence and withdrawal syndrome through the FF1 receptor.

Effect of RFRP-3, the mammalian ortholog of GnIH, on the epididymis of male rats.

RFamide-related peptide-3 (RFRP-3) and its receptor (GPR147) play an important role in reproduction regulation in mammals. To understand the role of RFRP-3 in male reproductive function of epididymis, we first investigated the expression changes in RFRP-3 and its receptor at different stages of development, that is, postnatal day 20 (P20), 40 (P40), 60 (P60) and 80 (P80). Our results showed that fluctuations in the expression of GnIH and GPR147 during postnatal development occurred, and the highest epididymal GnIH and GPR147 expression were both detected in P60. Subsequently, we further investigated the effect of RFRP-3 on the histology, apoptosis and autophagy of the epididymis in vivo. For in vivo study, male rats were treated intratesticularly with different doses of RFRP-3 (control, 0.1 µg, 1 µg, and 10 µg per day) for 7 days. Our results show that RFRP-3 caused dose-dependent histological changes in the epididymal duct, such as a decline in the number of spermatozoa and an increase in degenerated and vacuolated epididymal epithelial cells. Rats treated intratesticularly with RFRP-3 also showed dose-dependent effects on caspase-3 activation and the expression of apoptotic markers (whole caspase-3, cleaved caspase-3 and Bcl-2). However, the expression of autophagy markers (Beclin-1 and Atg5) exhibited a bidirectional, dose-dependent effect. It is concluded that RFRP-3 plays a regulatory role in male rat reproduction, possibly because RFRP-3 mediates the apoptosis and autophagy of the epididymis.

FLP-18 Functions through the G-Protein-Coupled Receptors NPR-1 and NPR-4 to Modulate Reversal Length in Caenorhabditis elegans.

Animal behavior is critically dependent on the activity of neuropeptides. Reversals, one of the most conspicuous behaviors in Caenorhabditis elegans, plays an important role in determining the navigation strategy of the animal. Our experiments on hermaphrodite C. elegans show the involvement of a neuropeptide FLP-18 in modulating reversal length in these hermaphrodites. We show that FLP-18 controls the reversal length by regulating the activity of AVA interneurons through the G-protein-coupled neuropeptide receptors, NPR-4 and NPR-1. We go on to show that the site of action of these receptors is the AVA interneuron for NPR-4 and the ASE sensory neurons for NPR-1. We further show that mutants in the neuropeptide, flp-18, and its receptors show increased reversal lengths. Consistent with the behavioral data, calcium levels in the AVA neuron of freely reversing C. elegans were significantly higher and persisted for longer durations in flp-18, npr-1, npr-4, and npr-1 npr-4 genetic backgrounds compared with wild-type control animals. Finally, we show that increasing FLP-18 levels through genetic and physiological manipulations causes shorter reversal lengths. Together, our analysis suggests that the FLP-18/NPR-1/NPR-4 signaling is a pivotal point in the regulation of reversal length under varied genetic and environmental conditions.SIGNIFICANCE STATEMENT In this study, we elucidate the circuit and molecular machinery required for normal reversal behavior in hermaphrodite Caenorhabditis elegans We delineate the circuit and the neuropeptide receptors required for maintaining reversal length in C. elegans Our work sheds light on the importance of a single neuropeptide, FLP-18, and how change in levels in this one peptide could allow the animal to change the length of its reversal, thereby modulating how the C. elegans explores its environment. We also go on to show that FLP-18 functions to maintain reversal length through the neuropeptide receptors NPR-4 and NPR-1. Our study will allow for a better understanding of the complete repertoire of behaviors shown by freely moving animals as they explore their environment.

Molecular and Physiological Characterization of a Receptor for d-Amino Acid-Containing Neuropeptides.

Neuropeptides in several animals undergo an unusual post-translational modification, the isomerization of an amino acid residue from the l-stereoisomer to the d-stereoisomer. The resulting d-amino acid-containing peptide (DAACP) often displays biological activity higher than that of its all-l-residue analogue, with the d-residue being critical for function in many cases. However, little is known about the full physiological roles played by DAACPs, and few studies have examined the interaction of DAACPs with their cognate receptors. Here, we characterized the signaling of several DAACPs derived from a single neuropeptide prohormone, the Aplysia californica achatin-like neuropeptide precursor (apALNP), at their putative receptor, the achatin-like neuropeptide receptor (apALNR). We first used quantitative polymerase chain reaction and in situ hybridization experiments to demonstrate receptor ( apALNR) expression throughout the central nervous system; on the basis of the expression pattern, we identified novel physiological functions that may be mediated by apALNR. To gain insight into ligand signaling through apALNR, we created a library of native and non-native neuropeptide analogues derived from apALNP (the neuropeptide prohormone) and evaluated them for activity in cells co-transfected with apALNR and the promiscuous Gα subunit Gα-16. Several of these neuropeptide analogues were also evaluated for their ability to induce circuit activity in a well-defined neural network associated with feeding behavior in intact ganglia from Aplysia. Our results reveal the specificity of apALNR and provide strong evidence that this receptor mediates diverse physiological functions throughout the central nervous system. Finally, we show that some native apALNP-derived DAACPs exhibit enhanced stability toward endogenous proteases, suggesting that the d-residues in these DAACPs may increase the peptide lifetime, in addition to influencing receptor specificity, in the nervous system. Ultimately, these studies provide insight into signaling at one of the few known DAACP-specific receptors and advance our understanding of the roles that l- to d-residue isomerization play in neuropeptide signaling.

Function of the natalisin receptor in mating of the oriental fruit fly, Bactrocera dorsalis (Hendel) and testing of peptidomimetics.

Natalisins (NTLs) are conservative neuropeptides, which are only found in arthropods and are documented to regulate reproductive behaviors in insects. In our previous study, we have confirmed that NTLs regulate the reproductive process in an important agricultural pest, Bactrocera dorsalis (Hendel). Hence, in this study, to further confirm the in vivo function of NTL receptor (NTLR) and assess the potential of NTLR as an insecticide target, RNA interference targeting NTLR mRNA was performed. We found that mating frequencies of both males and females were reduced by RNAi-mediated knockdown of the NTLR transcript, while there was no effect on mating duration. Moreover, we functionally expressed the B. dorsalis NTLR in Chinese Hamster Ovary (CHO) cells and was co-transfected with an aequorin reporter to measure ligand activities. A total of 13 biostable multi-Aib analogs were tested for agonistic and antagonistic activities. While most of these NTL analogs did not show strong activity, one analog (NLFQV[Aib]DPFF[Aib]TRamide) had moderate antagonistic activity. Taken together, we provided evidence for the important roles of NTLR in regulating mating frequencies of both male and female in this fly and also provided in vitro data on mimetic analogs that serve as leading structures for the development of agonists and antagonists to disrupt the NTL signaling pathway.

Cloning and mRNA expression of NPB and its effect on hormone secretion of the reproductive cells in the pig.

Neuropeptide B (NPB) is an endogenous ligand for the orphan G protein-coupled receptors NPBWR1 (GPR7) and NPBWR2 (GPR8). Some reports have investigated the role of NPB in the regulation of feeding, energy metabolism and hormone secretion in many species. However, few papers reported the physiological function of NPB in the pig. In this study, we cloned and sequenced the NPB mRNA from a pig, which was found to consist of 123 bases. NPB mRNA expression was detected in central and peripheral tissues by the quantitative fluorescence method. The results showed that NPB mRNA expression was higher in hippocampus, cerebellum, spinal cord, thymus, tonsil, duodenum, cecum, colon, ovary and testis. The distribution of NPB suggested that it may be involved in the regulation of reproductive functions in the pig. Subsequently, the expression and distribution of NPBWR1 and NPBWR2 were found in Leydig cells and ovarian granular cells. We then investigated the direct effect of NPB on pig reproductive cells in vitro. The results showed that different concentrations of NPB (10-12, 10-10, 10-8 and 10-6 M) promoted the secretion of testosterone in Leydig cells in concentration-dependent manner. Different doses of NPB could promote the secretion of progesterone in ovarian granulosa cells in dose-dependent manner. Low concentrations of NPB (10-8 and 10-10 M) promoted estradiol secretion, but high concentrations of NPB (10-6 M) inhibited its secretion. All the results suggested that the NPB/NPBWR1 or NPBWR2 system may play a role in modulating the reproductive activity in the pig.

Recurrent Circuitry for Balancing Sleep Need and Sleep.

Sleep-promoting neurons in the dorsal fan-shaped body (dFB) of Drosophila are integral to sleep homeostasis, but how these cells impose sleep on the organism is unknown. We report that dFB neurons communicate via inhibitory transmitters, including allatostatin-A (AstA), with interneurons connecting the superior arch with the ellipsoid body of the central complex. These "helicon cells" express the galanin receptor homolog AstA-R1, respond to visual input, gate locomotion, and are inhibited by AstA, suggesting that dFB neurons promote rest by suppressing visually guided movement. Sleep changes caused by enhanced or diminished allatostatinergic transmission from dFB neurons and by inhibition or optogenetic stimulation of helicon cells support this notion. Helicon cells provide excitation to R2 neurons of the ellipsoid body, whose activity-dependent plasticity signals rising sleep pressure to the dFB. By virtue of this autoregulatory loop, dFB-mediated inhibition interrupts processes that incur a sleep debt, allowing restorative sleep to rebalance the books. VIDEO ABSTRACT.