ORL1 Research Articles

Nociceptin and the nociceptin receptor in learning and memory.

There are many processes in which the neuropeptide nociceptin/orphanin FQ (N/OFQ or nociceptin) is involved in the brain. The role of nociceptin in learning and memory holds promise in modulating these processes in health and disease in the human brain. This review summarizes the body of research focused on N/OFQ and its specific receptor, the nociceptin receptor (NOP receptor), in learning and memory, and its potential mechanisms of action, in which acetylcholine, NMDA receptor, and noradrenaline may be critical. Finally, the association between NOP receptor and posttraumatic stress disorder (PTSD), a psychiatric disorder with altered fear learning, is examined as one of the potential outcomes resulting from pathological consequences of dysregulation of N/OFQ-NOP receptor in the brain.

Bioinformatics and evolution of vertebrate nociceptin and opioid receptors.

G protein-coupled receptors (GPCRs) are ancestrally related membrane proteins on cells that mediate the pharmacological effect of most drugs and neurotransmitters. GPCRs are the largest group of membrane receptor proteins encoded in the human genome. One of the most famous types of GPCRs is the opioid receptors. Opioid family receptors consist of four closely related proteins expressed in all vertebrate brains and spinal cords examined to date. The three classical types of opioid receptors shown unequivocally to mediate analgesia in animal models and in humans are the mu- (MOR), delta- (DOR), and kappa-(KOR) opioid receptor proteins. The fourth and most recent member of the opioid receptor family discovered is the nociceptin or orphanin FQ receptor (ORL). The role of ORL and its ligands in producing analgesia is not as clear, with both analgesic and hyperalgesic effects reported. All four opioid family receptor genes were cloned from expressed mRNA in a number of vertebrate species, and there are enough sequences presently available to carry out bioinformatic analysis. This chapter presents the results of a comparative analysis of vertebrate opioid receptors using pharmacological studies, bioinformatics, and the latest data from human whole-genome studies. Results confirm our initial hypotheses that the four opioid receptor genes most likely arose by whole-genome duplication, that there is an evolutionary vector of opioid receptor type divergence in sequence and function, and that the hMOR gene shows evidence of positive selection or adaptive evolution in Homo sapiens.

Endogenous nociceptin system involvement in stress responses and anxiety behavior.

The mechanisms underpinning stress-related behavior and dysfunctional events leading to the expression of neuropsychiatric disorders remain incompletely understood. Novel candidates involved in the neuromodulation of stress, mediated both peripherally and centrally, provide opportunities for improved understanding of the neurobiological basis of stress disorders and may represent targets for novel therapeutic development. This chapter provides an overview of the mechanisms by which the opioid-related peptide, nociceptin, regulates the neuroendocrine stress response and stress-related behavior. In our research, we have employed nociceptin receptor antagonists to investigate endogenous nociceptin function in tonic control over stress-induced activity of the hypothalamo-pituitary-adrenal axis. Nociceptin demonstrates a wide range of functions, including modulation of psychological and inflammatory stress responses, modulation of neurotransmitter release, immune homeostasis, in addition to anxiety and cognitive behaviors. Greater appreciation of the complexity of limbic-hypothalamic neuronal networks, together with attention toward gender differences and the roles of steroid hormones, provides an opportunity for deeper understanding of the importance of the nociceptin system in the context of the neurobiology of stress and behavior.

Nociceptin and meiosis during spermatogenesis in postnatal testes.

Phosphorylated Rec8, a key component of cohesin, mediates the association and disassociation, "dynamics," of chromosomes occurring in synaptonemal complex formation, crossover recombination, and sister chromatid cohesion during meiosis in germ cells. Yet, the extrinsic factors triggering meiotic chromosome dynamics remained unclear. In postnatal testes, follicle-stimulating hormone (FSH) acts directly on somatic Sertoli cells to activate gene expression via an intracellular signaling pathway composed of cAMP, cAMP-dependent protein kinase (PKA), and cAMP-response element-binding protein (CREB), and promotes germ cell development and spermatogenesis indirectly. Yet, the paracrine factors mediating the FSH effects to germ cells remained elusive. We have shown that nociceptin, known as a neuropeptide, is upregulated by FSH signaling through cAMP/PKA/CREB pathway in Sertoli cells of postnatal murine testes. Chromatin immunoprecipitation from Sertoli cells demonstrated that CREB phosphorylated at Ser133 associates with prepronociceptin gene encoding nociceptin. Analyses with Sertoli cells and testes revealed that both prepronociceptin mRNA and the nociceptin peptide are induced after FSH signaling is activated. In addition, the nociceptin peptide is induced in testes after 9 days post partum following FSH surge. Thus, our findings may identify nociceptin as a novel paracrine mediator of the FSH effects in the regulation of spermatogenesis; however, very little has known about the functional role of nociceptin in spermatogenesis. We have shown that nociceptin induces Rec8 phosphorylation, triggering chromosome dynamics, during meiosis in spermatocytes of postnatal murine testes. The nociceptin receptor Oprl-1 is exclusively expressed in the plasma membrane of testicular germ cells, mostly spermatocytes. Treatment of testes with nociceptin resulted in a rapid phosphorylation of Rec8. Injection of nociceptin into mice stimulated Rec8 phosphorylation and meiotic chromosome dynamics in testes, whereas injection of nocistatin, a specific inhibitor for nociceptin, abolished them. Therefore, our findings suggest that nociceptin is a novel extrinsic factor that plays a crucial role in the progress of meiosis during spermatogenesis.

Time to first cigarette: A potential clinical screening tool for nicotine dependence and cancer risk

Aims: Opioid peptides are implicated in processes related to reward and aversion; however, the how specific opioid peptides are involved remains unclear. Here, we aim to investigate the role nociceptin in behavioral responding for palatable and aversive tastants. Methods: A microstructural licking analysis of the effects of intracerebroventricular administerednociceptinon total licks, total bouts of licking, and lengths of licking bouts was conducted. Additionally, we compared these licking behaviors in wildtype and nociceptin receptor (NOP) knockout mice, to identify the role of endogenous nociceptin in these processes. We hypothesize that NOP knockout mice will show lower levels of licking behavior under normal conditions and will be irresponsive to the appetitestimulating effects of nociceptin administration. Results: Repeated measures ANOVA analyses show that NOP knockout mice generally emit fewer licks than wildtype mice when given access to sucrose solution. Furthermore, results from post-hoc analyses show nociceptin administration increases total number of licking bouts in wildtype mice in both chow sated and hungry states, an effect thatwas absent inNOPknockoutmice. Similar effects were seen for the non-caloric sweetener sucralose, but to a lesser degree. Hunger tended to increase total bouts of licking for sucrose in all mice, but also increased mean bout length in NOPknockoutmiceonly. Finally, adulterating sucrose solutionwith the bitter aversive tastant quinine produced similarly suppressive effects in both wildtype and NOP receptor knockout mice. Conclusions: Together, these results support our hypothesis, suggesting that endogenous nociceptin contributes to the positive incentive properties of palatable tastants. Further understanding of the contributions of opioid neurotransmission to hedonic processing may aid in developing appropriate treatment strategies for pathologies characterized by aberrant reward seeking and taking behaviors. Financial support: Grant #: DA024635, DA09359, and DA05010.

Inhibition of opioid receptor mediated G-protein activity after chronic administration of kynurenic acid and its derivative without direct binding to opioid receptors.

There is an increasing number of evidence showing analgesic properties of the kynurenic acid (KYNA), and also some studies demonstrate that kynurenine might interact with the opioid system. Therefore in this study, for the first time we investigated the direct binding affinity of KYNA and its structural analog KYNA-1 towards mu, kappa and delta opioid receptor in competition binding experiments applying opioid receptor specific radioligands. The binding affinity measurements were performed in Chinese hamster ovary cell lines overexpressing the corresponding opioid receptor (mu and kappa opioid receptor were rat, delta opioid receptor were mouse sequence). Additionally we also examined the chronic effect of these compounds on mu, kappa and delta opioid receptor and also nociceptin peptide receptor mediated G-protein activity in [(35)S]GTPγS binding assays performed in mouse cortex and striatum membranes. Our results showed that KYNA and KYNA-1 had no affinity towards any of the three classic opioid receptors. On the other hand the compounds significantly decreased opioid and nociceptin receptor mediated G-protein activity or in some cases enhanced the potency of the activating ligand. Moreover, the alterations were receptor and brain region specific. Accordingly, we conclude that KYNA and KYNA-1 do not interact directly with the opioid receptors, but more likely alter the receptor functions intracellularly.

Structure-Based Virtual Screening of the Nociceptin Receptor: Hybrid Docking and Shape-Based Approaches for Improved Hit Identification

The antagonist-bound crystal structure of the nociceptin receptor (NOP), from the opioid receptor family, was recently reported along with those of the other opioid receptors bound to opioid antagonists. We recently reported the first homology model of the ‘active-state’ of the NOP receptor, which when docked with ‘agonist’ ligands showed differences in the TM helices and residues, consistent with GPCR activation after agonist binding. In this study, we explored the use of the active-state NOP homology model for structure-based virtual screening to discover NOP ligands containing new chemical scaffolds. Several NOP agonist and antagonist ligands previously reported are based on a common piperidine scaffold. Given the structure–activity relationships for known NOP ligands, we developed a hybrid method that combines a structure-based and ligand-based approach, utilizing the active-state NOP receptor as well as the pharmacophoric features of known NOP ligands, to identify novel NOP binding scaffolds by virtual screening. Multiple conformations of the NOP active site including the flexible second extracellular loop (EL2) loop were generated by simulated annealing and ranked using enrichment factor (EF) analysis and a ligand–decoy dataset containing known NOP agonist ligands. The enrichment factors were further improved by combining shape-based screening of this ligand–decoy dataset and calculation of consensus scores. This combined structure-based and ligand-based EF analysis yielded higher enrichment factors than the individual methods, suggesting the effectiveness of the hybrid approach. Virtual screening of the CNS Permeable subset of the ZINC database was carried out using the above-mentioned hybrid approach in a tiered fashion utilizing a ligand pharmacophore-based filtering step, followed by structure-based virtual screening using the refined NOP active-state models from the enrichment analysis. Determination of the NOP receptor binding affinity of a selected set of top-scoring hits resulted in identification of several compounds with measurable binding affinity at the NOP receptor, one of which had a new chemotype for NOP receptor binding. The hybrid ligand-based and structure-based methodology demonstrates an effective approach for virtual screening that leverages existing SAR and receptor structure information for identifying novel hits for NOP receptor binding. The refined active-state NOP homology models obtained from the enrichment studies can be further used for structure-based optimization of these new chemotypes to obtain potent and selective NOP receptor ligands for therapeutic development.

Anti-nociception mediated by a κ opioid receptor agonist is blocked by a δ receptor agonist.

BACKGROUND AND PURPOSEThe opioid receptor family comprises four structurally homologous but functionally distinct sub-groups, the μ (MOP), δ (DOP), κ (KOP) and nociceptin (NOP) receptors. As most opioid agonists are selective but not specific, a broad spectrum of behaviours due to activation of different opioid receptors is expected. In this study, we examine whether other opioid receptor systems influenced KOP-mediated antinociception.EXPERIMENTAL APPROACHWe used a tail withdrawal assay in C57Bl/6 mice to assay the antinociceptive effect of systemically administered opioid agonists with varying selectivity at KOP receptors. Pharmacological and genetic approaches were used to analyse the interactions of the other opioid receptors in modulating KOP-mediated antinociception.KEY RESULTSEtorphine, a potent agonist at all four opioid receptors, was not anti-nociceptive in MOP knockout (KO) mice, although etorphine is an efficacious KOP receptor agonist and specific KOP receptor agonists remain analgesic in MOP KO mice. As KOP receptor agonists are aversive, we considered KOP-mediated antinociception might be a form of stress-induced analgesia that is blocked by the anxiolytic effects of DOP receptor agonists. In support of this hypothesis, pretreatment with the DOP antagonist, naltrindole (10 mg·kg−1), unmasked etorphine (3 mg·kg−1) antinociception in MOP KO mice. Further, in wild-type mice, KOP-mediated antinociception by systemic U50,488H (10 mg·kg−1) was blocked by pretreatment with the DOP agonist SNC80 (5 mg·kg−1) and diazepam (1 mg·kg−1).CONCLUSIONS AND IMPLICATIONSSystemic DOP receptor agonists blocked systemic KOP antinociception, and these results identify DOP receptor agonists as potential agents for reversing stress-driven addictive and depressive behaviours mediated through KOP receptor activation.LINKED ARTICLESThis article is part of a themed section on Opioids: New Pathways to Functional Selectivity. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2015.172.issue-2

Exploring LPS-induced sepsis in rats and mice as a model to study potential protective effects of the nociceptin/orphanin FQ system

The nociceptin receptor (NOP) and its ligand nociceptin/orphanin FQ (N/OFQ) have been shown to exert a modulatory effect on immune cells during sepsis. We evaluated the suitability of an experimental lipopolysaccharide (LPS)-induced sepsis model for studying changes in the nociceptin system. C57BL/6 mice BALB/c mice and Wistar rats were inoculated with different doses of LPS with or without a nociceptin receptor antagonist (UFP-101 or SB-612111). In C57BL/6 mice LPS 0.85mg/kg injection produced no septic response, whereas 1.2mg/kg produced a profound response within 5h. In BALB/c mice, LPS 4mg/kg produced no response, whereas 7mg/kg resulted in a profound response within 24h. In Wistar rats LPS 15mg/kg caused no septic response in 6/10 animals, whereas 25mg/kg resulted in marked lethargy before 24h. Splenic interleukin-1β mRNA in BALB/c mice, and serum TNF-α concentrations in Wistar rats increased after LPS injection in a dose-dependent manner, but were undetectable in control animals, indicating that LPS had stimulated an inflammatory reaction. IL-1β and TNF-α concentrations in LPS-treated animals were unaffected by administration of a NOP antagonist. Similarly NOP antagonists had no effect on survival or expression of mRNA for NOP or ppN/OFQ (the N/OFQ precursor) in a variety of tissues. In these animal models, the dose–response curve for LPS was too steep to allow use in survival studies and no changes in the N/OFQ system occurred within 24h. We conclude that LPS-inoculation in rodents is an unsuitable model for studying possible changes in the NOP-N/OFQ system in sepsis.

Neuropeptide FF receptors exhibit direct and anti-opioid effects on mice dorsal raphe nucleus neurons

By using acutely dissociated dorsal raphe nucleus neurons (DRN) from young mice, direct and anti-opioid effects of Neuropeptide FF (NPFF) receptors were measured. The NPFF analog 1DMe (10µM) had no effect on resting Ca2+ channels but reduced the magnitude of Ca2+ transients induced by depolarization in 83.3% neurons tested, of which the inhibition rate is 45.4±2.9%. Pertussis toxin treatment reduced to 18.9% the number of responding neurons and attenuated by 47% the response of 1DMe. In contrast, cholera toxin treatment had no significant effect. Eighteen minute perfusion with 1DMe at a very low 10nM concentration, that did not directly inhibit Ca2+ transients triggered by depolarization in every neuron, attenuated by 78% the inhibitory effect of Nociceptin/orphanin FQ (N/OFQ) on Ca2+ transients, but not that of by serotonin. These results demonstrated for the first time that NPFF receptors on mice DRN inhibit Ca2+ transients induced by depolarization via Gi/o protein and also exhibit a specific anti-opioid activity on nociceptin receptors, and that their specific anti-opioid activity is not a direct consequence of their activity on Ca2+ transients.

Activation of the endogenous nociceptin system by selective nociceptin receptor agonist SCH 221510 produces antitransit and antinociceptive effect: a novel strategy for treatment of diarrhea-predominant IBS.

Diarrhea-predominant irritable bowel syndrome (IBS-D) is a functional gastrointestinal (GI) disorder, defined by the presence of loose stools and abdominal pain. In search for a novel anti-IBS-D therapy, here we investigated the nociceptin receptor (NOP)-dependent effects in the GI tract. A novel potent and selective NOP agonist SCH 221510 was used in the study. The effect of NOP activation on mouse intestinal motility was characterized in vitro and in vivo, in physiological conditions and in animal models of hypermotility and diarrhea. Well-established mouse models of visceral pain were used to characterize the antinociceptive effect of the NOP activation. To provide additional evidence that the endogenous nociceptin system is a relevant target for IBS, NOP expression and nociceptin levels were quantified in serum and colonic biopsies from IBS-D patients. SCH 221510 produced a potent NOP-mediated inhibitory effect on mouse intestinal motility in vitro and in vivo in physiological conditions. The NOP agonist displayed an antidiarrheal and analgesic action after oral administration in animal models mimicking the symptoms of IBS-D. Studies on human samples revealed a strong decrease in endogenous nociceptin system expression in IBS-D patients compared with healthy controls. Collectively, mouse and human data suggest that the endogenous nociceptin system is involved in IBS-D and may become a target for anti-IBS-D treatments using potent and selective synthetic NOP agonists.

Nociceptin system as a target in sepsis?

The nociceptin system comprises the nociceptin receptor (NOP) and the ligand nociceptin/orphanin FQ (N/OFQ) that binds to the receptor. The archetypal role of the system is in pain processing but the NOP receptor is also expressed on immune cells. Activation of the NOP receptor is known to modulate inflammatory responses, such as mast-cell degranulation, neutrophil rolling, vasodilation, increased vascular permeability, adhesion molecule regulation and leucocyte recruitment. As there is a loss of regulation of inflammatory responses during sepsis, the nociceptin system could be a target for therapies aimed at modulating sepsis. This review details the known effects of NOP activation on leucocytes and the vascular endothelium and discusses the most recent human and animal data on the role of the nociceptin system in sepsis.

Discovery of a Novel Series of Orally Active Nociceptin/Orphanin FQ (NOP) Receptor Antagonists Based on a Dihydrospiro(piperidine-4,7′-thieno[2,3-c]pyran) Scaffold

Nociceptin/OFQ (N/OFQ) is a 17 amino acid peptide that is the endogenous ligand for the ORL1/NOP receptor. Nociceptin appears to regulate a host of physiological functions such as biological reactions to stress, anxiety, mood, and drug abuse, in addition to feeding behaviors. To develop tools to study the function of nociceptin and NOP receptor, our research effort sought to identify orally available NOP antagonists. Our effort led to the discovery of a novel chemical series based on the dihydrospiro(piperidine-4,7'-thieno[2,3-c]pyran) scaffold. Herein we show that dihydrospiro(piperidine-4,7'-thieno[2,3-c]pyran)-derived compounds are potent NOP antagonists with high selectivity versus classical opioid receptors (μ, δ, and κ). Moreover, these compounds exhibit sufficient bioavailability to produce a high level of NOP receptor occupancy in the brain following oral administration in rats.

Intrathecal nociceptin displaces a PET ligand for its receptor in the brain at analgesic doses in rhesus monkeys (658.1)

The nociceptin receptor (NOP) has been investigated as a potential therapeutic target for pain control. Nociceptin, the endogenous ligand for NOP, produces analgesia in rodents and primates when given intrathecally (i.t.) (Ko et al. 2006, Xu et al. 1996). Recently NOP radiotracers have been developed for use with Positron Emission Tomography (PET) (Kimura et al. 2011, Hostetler et al. 2013). Although behavioral studies have shown that NOP agonists may be therapeutically useful, it is unknown whether nociceptin given i.t. binds to receptors supraspinally, and if so, which areas in the brain display the greatest ligand‐target interaction. The purpose of this study was to measure the amount of radiotracer displacement in the brain at NOP following an analgesic dose of nociceptin. Two female rhesus monkeys were anesthetized, and underwent PET scans with the NOP radiotracer 11C NOP‐1A. Eight minutes following tracer delivery, the monkey was injected i.t. with either saline or 100 nM nociceptin. Preliminary data show that 100 nM nociceptin delivered into the spinal canal displaces the radiotracer 11C NOP‐1A in a variety of brain nuclei. Prominent regions of displacement include the cortex, striatum, and thalamus. These data suggest that analgesia with NOP agonists can be produced with relatively low receptor occupancy and further supports NOP as a novel drug target for analgesia.Grant Funding Source: Supported by: The Michigan Memorial Pheonix Project and NIDA Traning Grant T32‐DA007281

A novel nociceptin‐1 receptor antagonist produces antidepressant‐ and anxiolytic‐related effects in rodents (656.3)

We utilized a potent, selective, and orally‐bio‐available antagonist with documented engagement with nociceptin receptors (NOP) to assess antidepressant‐ and anxioltyic‐related effects of blockade of NOP receptors. The NOP receptor antagonist, Cpd 1, displayed antidepressant‐like neurochemical and behavioral effects in rats and mice. Serotonin outflow in rat prefrontal cortex was enhanced without affecting norepinephrine or dopamine. Correspondingly Cpd 1 decreased immobility in the forced swim test (FST). Cpd 1 augmented the effects of fluoxetine without changing target occupancies of either drug. Anti‐immobility effects of Cpd 1 were absent in mice without NOP receptors. Cpd 1 also produced anxiolytic‐like effects in some rodent models: decreased stimulus‐elicited freezing in mice and decreased stress‐induced hyperthermia in rats. Cdp 1 was not active in: marble‐burying in mice, novelty‐suppressed feeding in mice, conditioned suppression in rats, punished food‐maintained responding in rats, and the 4‐plate test in mice. Cpd 1 did not disrupte performance in 5‐choice serial reaction time tests or delayed matching‐to‐position in rats. Cpd 1 was mildly wake promoting in rats. These data suggest that Cpd 1 has unique antidepressant‐ and anxiolytic‐related pharmacological effects in rodents.

A novel nociceptin receptor antagonist inhibits feeding behavior in rodents (656.1)

Background: Nociceptin is a 17 amino acid peptide that produces physiological effects through activation of the ORL1 (NOP) receptor. We have identified a novel NOP antagonist (Cpd 1) and found it to have high binding affinity (Ki = 0.11 nM), antagonist potency (Kb = 0.17 nM) and selectivity for NOP. Cpd 1 is orally active, CNS penetrant and potently occupies NOP receptors in vivo. Herein, the effects of Cpd 1 on feeding behavior and metabolism were studied.Methods: Male, lean and dietary‐induced obese (DIO) rats, DIO mice and NOP knockout (KO) mice were used in feeding and indirect calorimetry studies.Results: Consistent with the literature, Cpd 1 reduced food intake in rodents. In lean rats, Cpd 1 inhibited consumption of a highly palatable (high fat/sucrose) diet, reducing the calorie intake to control chow levels. In DIO rats, Cpd 1 inhibited feeding induced by a mild caloric restriction. In DIO mice, Cpd 1 decreased 24 hr free access food intake and decreased the respiratory quotient indicating that metabolism was driven by fat utilization. Fasting‐induced feeding was inhibited by Cpd 1 in 129S6 mice, an effect that was absent in NOP KO mice. Moreover, under vehicle conditions fasting‐induced feeding was reduced in NOP KO mice.Conclusions: These data support that Cpd 1 potently inhibits feeding behavior and stimulates fat utilization in rodents. Moreover, the hypophagic effect of Cpd 1 is dependent on NOP receptors.Grant Funding Source: Eli Lilly and Company

Effects on Inflammation of Newly-Synthesized Hexapeptide with Affinity to Opioid and Nociceptin Receptors

Following the discovery of the N/OFQ/NOP system and its modulatory role in physiological and pathophysiological processes, intensive study has started to find selective NOP ligands with hypothetic therapeutic potential. Among the agonists, a hexapeptide Ac-RYYRWK-NH2 has been identified. It expresses high NOP receptor affinity and selectivity. Its molecule was used as a template, in which Tyr5 was substituted by original β2-tryptophan analogue (S)-2-(1-methyl-1H-indol-3-yl)propionic residue (compound HP3) The new compound activates both NOP and opioid receptors. Having in mind that classical opioids, as well as nociceptin, are involved in modulating pain and inflammation, we examined the anti-inflammatory effect of newly-synthesized peptide HP3 on carrageenan-induced peripheral inflammation, and compared it with that of indomethacin (3 mg/kg). It was found that HP3 in dose 40 μg/kg exerts weaker anti-inflammatory action in the first 180 min of the experiment, but is equally effective with indomethacin 3 mg/kg at the end of the observation. The HP3 effect is due mainly of the activation of opioid receptors.

MT-7716, a novel selective nonpeptidergic NOP receptor agonist, effectively blocks ethanol-induced increase in GABAergic transmission in the rat central amygdala.

The GABAergic system in the central amygdala (CeA) plays a major role in ethanol dependence and the anxiogenic-like response to ethanol withdrawal. A large body of evidence shows that Nociceptin/Orphanin FQ (N/OFQ) regulates ethanol intake and anxiety-like behavior. In the rat, ethanol significantly augments CeA GABA release, whereas N/OFQ diminishes it. Using electrophysiological techniques in an in vitro slice preparation, in this study we investigated the effects of a nonpeptidergic NOP receptor agonist, MT-7716 [(R)-2-3-[1-(Acenaphthen-1-yl)piperidin-4-yl]-2-oxo-2,3-dihydro-1H-benzimidazol-1-yl-N-methylacetamide hydrochloride hydrate], and its interaction with ethanol on GABAergic transmission in CeA slices of naïve rats. We found that MT-7716 dose-dependently (100–1000 nM) diminished evoked GABAA receptor-mediated inhibitory postsynaptic potentials (IPSPs) and increased paired-pulse facilitation (PPF) ratio of these evoked IPSPs, suggesting a presynaptic site of action of the MT-7716 by decreasing GABA release at CeA synapses. The presynaptic action of MT-7716 was also supported by the significant decrease in the frequency of miniature inhibitory postsynaptic currents (mIPSCs) induced by the nociceptin receptor (NOP) agonist. Interestingly, MT-7716 prevented the ethanol-induced augmentation of evoked IPSPs. A putative selective NOP antagonist, [Nphe1]Nociceptin(1–13)NH2, totally prevented the MT-7716-induced inhibition of IPSP amplitudes indicating that MT-7716 exerts its effect through NOPs. These data provide support for an interaction between the nociceptin and GABAergic systems in the CeA and for the anti-alcohol properties of the NOP activation. The development of a synthetic nonpeptidergic NOP receptor agonist such as MT-7716 may represent a useful therapeutic target for alcoholism.

The nociceptin receptor (NOPR) and its interaction with clinically important agonist molecules: a membrane molecular dynamics simulation study.

The nociceptin receptor (NOPR) is an orphan G protein-coupled receptor that contains seven transmembrane helices. NOPR has a distinct mechanism of activation, though it shares a significant homology with other opioid receptors. Previously there have been reports on homology modeling of NOPR and also molecular dynamics simulation studies for a short period. Recently the crystal structure of NOPR was reported. In this study, we analyzed the time dependent behavior of NOPR docked with clinically important agonist molecules such as NOP (natural agonist) peptide and compound 10 (SCH-221510 derivative) using molecular dynamics simulations (MDS) for 100 ns. Molecular dynamics simulations of NOPR-agonist complexes allowed us to refine the system and to also identify stable structures with better binding modes. Structure activity relationships (SAR) for SCH221510 derivatives were investigated and reasons for the activities of these derivatives were determined. Our molecular dynamics trajectory analysis of NOPR-peptide and NOPR-compound 10 complexes found residues to be crucial for binding. Mutagenesis studies on the residues identified from our analysis could prove useful. Our results could also provide useful information in the structure-based drug design of novel and potent agonists targeting NOPR.

Anti-Inflammatory and Antinociceptive Action of an Orally Available Nociceptin Receptor Agonist SCH 221510 in a Mouse Model of Inflammatory Bowel Diseases

The nociceptin receptors (NOPs) are expressed in the gastrointestinal (GI) tract on muscle cell membranes and neurons, as well as the immune cells that infiltrate the mucosa. The involvement of NOPs in the pathophysiology of GI inflammation has been suggested, but due to the lack of selective NOP agonists, it never fully elucidated. Our aim was to characterize the anti-inflammatory and antinociceptive effect of the NOP agonist, SCH 221510 [3-endo-8-[bis(2-methylphenyl)methyl]-3-phenyl-8-azabicyclo [3.2.1]octan-3-ol], as a potential therapeutic strategy in the treatment of inflammatory bowel diseases (IBD). The anti-inflammatory action of SCH 221510 was determined after intraperitoneal, oral, and intracolonic administration of SCH 221510 (0.1-3.0 mg/kg once or twice daily) in mice treated with 2,4,6-trinitrobenzenesulfonic acid (TNBS). Antinociceptive action of SCH 221510 was evaluated in the mouse model of mustard oil (MO)-induced abdominal pain. Relative NOP mRNA expression was assessed in patients with IBD using real-time reverse transcriptase-polymerase chain reaction. We found that the expression of NOP mRNA was significantly decreased in patients with IBD. The administration (0.1 and 1.0 mg/kg i.p. twice daily and 3 mg/kg p.o. twice daily) of SCH 221510 attenuated TNBS colitis in mice. This effect was blocked by a selective NOP antagonist [J-113397 [(±)-1-[(3R*,4R*)-1-(cyclooctylmethyl)-3-(hydroxymethyl)-4-piperidinyl]-3-ethyl-1,3-dihydro-2H-benzimidazol-2-one]]. The intracolonic injections of SCH 221510 did not improve colitis in mice. The antinociceptive effect of SCH 221510 was observed after oral administration of SCH 221510 in MO-induced pain tests in mice with acute colitis. In conclusion, our results show a potent anti-inflammatory and antinociceptive effect upon selective activation of NOP receptors and suggest that the NOP agonist SCH 221510 is a promising drug candidate for future treatment of IBD.