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Background and Purpose: The cannabinoid (CB1) receptor is among the most abundant G protein‐coupled receptors in brain. Allosteric ligands bind to a different site on receptors than the orthosteric ligand can have effects that are unique to the allosteric ligand and modulate orthosteric ligand activity. We propose a unified mathematical model describing the interaction effects of the allosteric ligand Org27569 and the orthosteric agonist CP55940 on CB1 receptor. Experimental Approach: A ternary complex model was constructed, which incorporated kinetic properties to describe the time course of effects of Org27569 and CP55940 reported in the literature: (i) enhanced receptor binding of CP55940, (ii) reduced internalisation and (iii), time‐dependent modulation of cAMP. Underlying mechanisms of time‐dependent modulation by Org27569 were evaluated by simulation. Key Results: A hypothetical transitional state of CP55940–CB1–Org27569, which can internalise but cannot inhibit cAMP, was shown to be necessary and was sufficient to describe the allosteric modulation by Org27569, prior to receptors adopting an inactive conformation. The model indicated that the formation of this transitional CP55940–CB1–Org27569 state and final inactive CP55940–CB1–Org27569 state contributes to the enhanced CP55940 binding. The inactive CP55940–CB1–Org27569 cannot internalise or inhibit cAMP, leading to reduced internalisation and cessation of cAMP inhibition. Conclusions and Implications: In conclusion, a kinetic mathematical model for CB1 receptor allosteric modulation was developed. However, a standard ternary complex model was not sufficient to capture the data and a hypothetical transitional state was required to describe the allosteric modulation properties of Org27569. [ABSTRACT FROM AUTHOR]

Background and Purpose: N‐methyl‐D‐aspartate receptors (NMDARs) play a critical role in synaptic plasticity, and mutations in human genes encoding NMDAR subunits have been described in individuals with various neuropsychiatric disorders. Compounds with a positive allosteric effect are thought to compensate for reduced receptor function. Experimental Approach: We have used whole‐cell patch‐clamp electrophysiology on recombinant rat NMDARs and human variants found in individuals with neuropsychiatric disorders, in combination with in silico modelling, to explore the site of action of novel epipregnanolone‐based NMDAR modulators. Key Results: Analysis of the action of 4‐(20‐oxo‐5β‐pregnan‐3β‐yl) butanoic acid (EPA‐But) at the NMDAR indicates that the effect of this steroid with a "bent" structure is different from that of cholesterol and oxysterols and shares a disuse‐dependent mechanism of NMDAR potentiation with the "planar" steroid 20‐oxo‐pregn‐5‐en‐3β‐yl sulfate (PE‐S). The potentiating effects of EPA‐But and PE‐S are additive. Alanine scan mutagenesis identified residues that reduce the potentiating effect of EPA‐But. No correlation was found between the effects of EPA‐But and PE‐S at mutated receptors that were less sensitive to either steroid. The relative degree of potentiation induced by the two steroids also differed in human NMDARs carrying rare variants of hGluN1 or hGluN2B subunits found in individuals with neuropsychiatric disorders, including intellectual disability, epilepsy, developmental delay, and autism spectrum disorder. Conclusion and Implications: Our results show novel sites of action for pregnanolones at the NMDAR and provide an opportunity for the development of new therapeutic neurosteroid‐based ligands to treat diseases associated with glutamatergic system hypofunction. [ABSTRACT FROM AUTHOR]

Glucagon-like peptide-1 (GLP-1) receptors belong to the pharmaceutically important Class B family of GPCRs and are involved in many biologically significant signalling pathways. Its incretin peptide ligand GLP-1 analogues are effective treatments for Type 2 diabetes. Although developing non-peptide low MW drugs targeting GLP-1 receptors remains elusive, considerable progress has been made in discovering non-peptide agonists and positive allosteric modulators (PAMs) of GLP-1 receptors with demonstrated efficacy. Many of these compounds induce biased signalling in GLP-1 receptor-mediated functional pathways. High-quality structures of GLP-1 receptors in both inactive and active states have been reported, revealing detailed molecular interactions between GLP-1 receptors and non-peptide agonists or PAMs. These progresses raise the exciting possibility of developing non-peptide drugs of GLP-1 receptors as alternative treatments for Type 2 diabetes. The insight into the interactions between the receptor and the non-peptide ligand is also useful for developing non-peptide ligands targeting other Class B GPCRs. LINKED ARTICLES: This article is part of a themed issue on GLP1 receptor ligands (BJP 75th Anniversary). To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v179.4/issuetoc. [ABSTRACT FROM AUTHOR]

Background and Purpose: Protease-activated receptor 1 (PAR1) is a GPCR expressed in several skin cell types, including keratinocyte and dermal fibroblast. PAR1 activation plays a crucial role in the process of skin wound healing such as thrombosis, inflammation, proliferation and tissue repair. In the present study, we identified a novel positive allosteric modulator of PAR1, GB83, and investigated its effect on skin wound healing.Experimental Approach: The enhancement of PAR1 activity by GB83 was measured using Fluo-4 calcium assay. In silico docking analysis of GB83 in PAR1 was performed using dock ligands method (CDOCKER) with CHARMm force field. Effects of GB83 on cell viability and gene expression were observed using MTS assay and quantitative real-time PCRs, respectively. SKH-1 hairless mice were used to investigate the wound healing effect of GB83.Key Results: We demonstrated that GB83 did not activate PAR1 by itself but strongly enhanced PAR1 activation by thrombin and PAR1-activating peptide (AP). In silico docking analysis revealed that GB83 can bind to the PAR1 binding site of vorapaxar. GB83 significantly promoted PAR1-mediated cell viability and migration. In addition, the enhancement of PAR1 activity by GB83 strongly increased gene expression of TGF-β, fibronectin and type I collagen in vitro and promoted skin wound healing in vivo.Conclusion and Implications: Our results revealed that GB83 is the first positive allosteric modulator of PAR1 and it can be a useful pharmacological tool for studying PAR1 and a potential therapeutic agent for skin wound healing. [ABSTRACT FROM AUTHOR]

Background and Purpose Strong implications in major neurological diseases make the neuronal α4 β2 nicotinic ACh receptor ( nAChR) a highly interesting drug target. In this study, we present a detailed electrophysiological characterization of NS9283, a potent positive allosteric modulator acting selectively at 3 α:2 β stoichiometry of α2* and α4* nAChRs. Experimental Approach The whole-cell patch-clamp technique equipped with an ultra-fast drug application system was used to perform electrophysiological characterization of NS9283 modulatory actions on human α4 β2 nAChRs stably expressed in HEK293 cells ( HEK293-h α4 β2). Key Results NS9283 was demonstrated to increase the potency of ACh-evoked currents in HEK293-h α4 β2 cells by left-shifting the concentration-response curve ∼60-fold. Interestingly, this modulation did not significantly alter maximal efficacy levels of ACh. Further, NS9283 did not affect the rate of desensitization of ACh-evoked currents, was incapable of reactivating desensitized receptors and only moderately slowed recovery from desensitization. However, NS9283 strongly decreased the rate of deactivation kinetics and also modestly decreased the rate of activation. This resulted in a left-shift of the ACh window current of ( α4)3( β2)2 nAChRs in the presence of NS9283. Conclusions and Implications This study demonstrates that NS9283 increases responsiveness of human ( α4)3( β2)2 nAChR to ACh with no change in maximum efficacy. We propose that this potentiation is due to a significant slowing of deactivation kinetics. In summary, the mechanism of action of NS9283 bears high resemblance to that of benzodiazepines at the GABAA receptor and to our knowledge, NS9283 constitutes the first nAChR compound of this class. [ABSTRACT FROM AUTHOR]

Background and Purpose The activation of α9α10 nicotinic cholinergic receptors ( nAChRs) present at the synapse between efferent olivocochlear fibres and cochlear hair cells can prevent acoustic trauma. Hence, pharmacological potentiators of these receptors could be useful therapeutically. In this work, we characterize ascorbic acid as a positive modulator of recombinant α9α10 nAChRs. Experimental Approach ACh-evoked responses were analysed under two-electrode voltage-clamp recordings in Xenopus laevis oocytes injected with α9 and α10 cRNAs. Key Results Ascorbic acid potentiated ACh responses in X. laevis oocytes expressing α9α10 (but not α4β2 or α7) nAChRs, in a concentration-dependent manner, with an effective concentration range of 1-30 mM. The compound did not affect the receptor's current-voltage profile nor its apparent affinity for ACh, but it significantly enhanced the maximal evoked currents (percentage of ACh maximal response, 240 ± 20%). This effect was specific for the L form of reduced ascorbic acid. Substitution of the extracellular cysteine residues present in loop C of the ACh binding site did not affect the potentiation. Ascorbic acid turned into a partial agonist of α9α10 nAChRs bearing a point mutation at the pore domain of the channel ( TM2 V13′T mutant). A positive allosteric mechanism of action rather than an antioxidant effect of ascorbic acid is proposed. Conclusions and Implications The present work describes one of the few agents that activates or potentiates α9α10 nAChRs and leads to new avenues for designing drugs with potential therapeutic use in inner ear disorders. [ABSTRACT FROM AUTHOR]

Background and purpose:As baclofen is active in patients with anxiety disorders, GABAB receptors have been implicated in the modulation of anxiety. To avoid the side effects of baclofen, allosteric enhancers of GABAB receptors have been studied to provide an alternative therapeutic avenue for modulation of GABAB receptors. The aim of this study was to characterize derivatives of (R,S)-5,7-di-tert-butyl-3-hydroxy-3-trifluoromethyl-3H-benzofuran-2-one (rac-BHFF) as enhancers of GABAB receptors.Experimental approach:Enhancing properties of rac-BHFF were assessed in the Chinese hamster ovary (CHO)-Gα16-hGABAB(1a,2a) cells by Fluorometric Imaging Plate Reader and GTPγ[35S]-binding assays, and in rat hippocampal slices by population spike (PS) recordings. In vivo activities of rac-BHFF were assessed using the loss of righting reflex (LRR) and stress-induced hyperthermia (SIH) models.Key results:In GTPγ[35S]-binding assays, 0.3 μM rac-BHFF or its pure enantiomer (+)-BHFF shifted the GABA concentration–response curve to the left, an effect that resulted in a large increase in both GABA potency (by 15.3- and 87.3-fold) and efficacy (149% and 181%), respectively. In hippocampal slices, rac-BHFF enhanced baclofen-induced inhibition of PS of CA1 pyramidal cells. In an in vivo mechanism-based model in mice, rac-BHFF increased dose-dependently the LRR induced by baclofen with a minimum effective dose of 3 mg kg−1 p.o. rac-BHFF (100 mg kg−1 p.o.) tested alone had no effect on LRR nor on spontaneous locomotor activity, but exhibited anxiolytic-like activity in the SIH model in mice.Conclusions and implications:rac-BHFF derivatives may serve as valuable pharmacological tools to elucidate the pathophysiological roles played by GABAB receptors in the central and peripheral nervous systems.British Journal of Pharmacology (2008) 154, 797–811; doi:10.1038/bjp.2008.135; published online 21 April 2008 [ABSTRACT FROM AUTHOR]

Linked Article: This article is a Commentary on Kalinichev M, Palea S, Haddouk H, Royer-Urios I, Guilloteau V, Lluel P, Schneider M, Saporito M and Poli S (2014). ADX71441, a novel, potent and selective positive allosteric modulator of the GABAB receptor, shows efficacy in rodent models of overactive bladder. Br J Pharmacol 171: 995-1006. doi: 10.1111/bph.12517. [ABSTRACT FROM AUTHOR]