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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.

Where Cartesian philosophy distinguishes the perceiving and perceptual mind from the body, phenomenology constitutes the experiential/experiencing body as the subject, giving rise to the affective potential of art. An immersive world of digital connections, smart cities and the Internet of Everything dramatises the centrality of relationship, the intertwining of Self and Other, in the lived environments of human experience. This article addresses the contextual, disciplinary and practical challenges encountered in developing an ambitious interactive public art project embedding SMART technology on the coastal fringes of Cardiff, the capital city of Wales (UK). It examines the processes and problems involved in delivering a stimulating aesthetic experience in and on a complex site, for a complex audience profile. It traces, in particular, the dependence of a multi-disciplinary project team on the theoretical and practical effects of affect in their ongoing effort to produce engaging, provocative, socially inclusive interactive public art, in and through human-centred design techniques.

The cognitive impairments that are often observed in patients with alcohol use disorder (AUD) partially contribute to the extremely low rates of treatment initiation and adherence. Brain acetylcholine receptors (AChR) mediate and modulate cognitive and reward-related behavior, and their distribution can be altered by long-term heavy drinking. Therefore, AChRs are promising pharmacotherapeutic targets for treating the cognitive symptoms of AUD. In the present study, the procognitive efficacy of two AChR agonists, xanomeline and varenicline, were evaluated in group-housed monkeys who self-administered ethanol for more than 1 year. The muscarinic AChR antagonist scopolamine was used to disrupt performance of a serial stimulus discrimination and reversal (SDR) task designed to probe cognitive flexibility, defined as the ability to modify a previously learned behavior in response to a change in reinforcement contingencies. The ability of xanomeline and varenicline to remediate the disruptive effects of scopolamine was compared between socially dominant and subordinate monkeys, with lighter and heavier drinking histories, respectively. We hypothesized that subordinate monkeys would be more sensitive to all three drugs. Scopolamine dose-dependently impaired performance on the serial SDR task in all monkeys at doses lower than those that produced nonspecific impairments (e.g., sedation); its potency did not differ between dominant and subordinate monkeys. However, both AChR agonists were effective in remediating the scopolamine-induced deficit in subordinate monkeys but not in dominant monkeys. These findings suggest xanomeline and varenicline may be effective for enhancing cognitive flexibility in individuals with a history of heavy drinking. SIGNIFICANCE STATEMENT: Procognitive effects of two acetylcholine (ACh) receptor agonists were assessed in group-housed monkeys who had several years' experience drinking ethanol. The muscarinic ACh receptor agonist xanomeline and the nicotinic ACh receptor agonist varenicline reversed a cognitive deficit induced by the muscarinic ACh receptor antagonist scopolamine. However, this effect was observed only in lower-ranking (subordinate) monkeys and not higher-ranking (dominant monkeys). Results suggest that ACh agonists may effectively remediate alcohol-induced cognitive deficits in a subpopulation of those with alcohol use disorder., (Copyright © 2024 by The American Society for Pharmacology and Experimental Therapeutics.)

Drug self-administration and intracranial self-stimulation (ICSS) are two preclinical behavioral procedures used to predict abuse potential of drugs, and abuse-related drug effects in both procedures are thought to depend on increased mesolimbic dopamine (DA) signaling. Drug self-administration and ICSS yield concordant metrics of abuse potential across a diverse range of drug mechanisms of action. The "rate of onset," defined as the velocity with which a drug produces its effect once administered, has also been implicated as a determinant of abuse-related drug effects in self-administration procedures, but this variable has not been systematically examined in ICSS. Accordingly, this study compared ICSS effects produced in rats by three DA transporter inhibitors that have different rates of onset (fastest to slowest: cocaine, WIN-35428, RTI-31) and that produced progressively weaker metrics of abuse potential in a drug self-administration procedure in rhesus monkeys. Additionally, in vivo photometry using the fluorescent DA sensor dLight1.1 targeted to the nucleus accumbens (NAc) was used to assess the time course of extracellular DA levels as a neurochemical correlate of behavioral effects. All three compounds produced ICSS facilitation and increased DA levels assessed by dLight. In both procedures, the rank order of onset rate was cocaine > WIN-35428 > RTI-31; however, in contrast to monkey drug self-administration results, maximum effects did not differ across compounds. These results provide additional evidence that drug-induced increases in DA drive ICSS facilitation in rats and illustrate the utility of both ICSS and photometry to evaluate the time course and magnitude of abuse-related drug effects in rats., (© 2023. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

The κ-opioid receptor (KOR) has emerged as an attractive drug target for pain management without addiction, and biased signaling through particular pathways of KOR may be key to maintaining this benefit while minimizing side-effect liabilities. As for most G protein-coupled receptors (GPCRs), however, the molecular mechanisms of ligand-specific signaling at KOR have remained unclear. To better understand the molecular determinants of KOR signaling bias, we apply structure determination, atomic-level molecular dynamics (MD) simulations, and functional assays. We determine a crystal structure of KOR bound to the G protein-biased agonist nalfurafine, the first approved KOR-targeting drug. We also identify an arrestin-biased KOR agonist, WMS-X600. Using MD simulations of KOR bound to nalfurafine, WMS-X600, and a balanced agonist U50,488, we identify three active-state receptor conformations, including one that appears to favor arrestin signaling over G protein signaling and another that appears to favor G protein signaling over arrestin signaling. These results, combined with mutagenesis validation, provide a molecular explanation of how agonists achieve biased signaling at KOR., (© 2023. The Author(s).)

Background: Because of their implications in several pathological conditions, α4β2* nicotinic acetylcholine receptors (nAChRs) are potential targets for the treatment of nicotine dependence, pain, and many psychiatric and neurodegenerative diseases. However, they exist in various subtypes, and finding selective tools to investigate them has proved challenging. The nicotinic receptor agonist, 5-iodo-A-85380 (5IA), has helped in delineating the function of β2-containing subtypes in vitro; however, much is still unknown about its behavioral effects. Furthermore, its effectiveness on α6-containing subtypes is limited., Aims: To investigate the effects of 5IA on nociception (formalin, hot-plate, and tail-flick tests), locomotion, hypothermia, and conditioned reward after acute and repeated administration, and to examine the potential role of β2 and α6 nAChR subunits in these effects. Lastly, its selectivity for expressed low sensitivity (LS) and high sensitivity (HS) α4β2 receptors is investigated., Results: 5IA dose-dependently induced hypothermia, locomotion suppression, conditioned place preference, and antinociception (only in the formalin test but not in the hot-plate or tail-flick tests). Furthermore, these effects were mediated by β2 but not α6 nicotinic subunits. Finally, we show that 5-iodo-A-85380 potently activates both stoichiometries of α4β2 nAChRs with differential efficacies, being a full agonist on HS α4(2)β2(3) nAChRs, and a partial agonist on LS α4(3)β2(2) nAChRs and α6-containing subtypes as well.

Exposure to stress triggers biological changes throughout the body. Accumulating evidence indicates that alterations in immune system function are associated with the development of stress-associated illnesses such as major depressive disorder and post-traumatic stress disorder, increasing interest in identifying immune markers that provide insight into mental health. Recombination events during T-cell receptor rearrangement and T-cell maturation in the thymus produce circular DNA fragments called T-cell receptor excision circles (TRECs) that can be utilized as indicators of thymic function and numbers of newly emigrating T-cells. Given data suggesting that stress affects thymus function, we examined whether blood levels of TRECs might serve as a quantitative peripheral index of cumulative stress exposure and its physiological correlates. We hypothesized that chronic stress exposure would compromise thymus function and produce corresponding decreases in levels of TRECs. In male mice, exposure to chronic social defeat stress (CSDS) produced thymic involution, adrenal hypertrophy, and decreased levels of TRECs in blood. Extending these studies to humans revealed robust inverse correlations between levels of circulating TRECs and childhood emotional and physical abuse. Cell-type specific analyses also revealed associations between TREC levels and blood cell composition, as well as cell-type specific methylation changes in CD4T + and CD8T + cells. Additionally, TREC levels correlated with epigenetic age acceleration, a common biomarker of stress exposure. Our findings demonstrate alignment between findings in mice and humans and suggest that blood-borne TRECs are a translationally-relevant biomarker that correlates with, and provides insight into, the cumulative physiological and immune-related impacts of stress exposure in mammals., (© 2022. The Author(s).)

Low-nanomolar binding constants were recorded for a series of six 2'-fluoro-(carbamoylpyridinyl)deschloroepibatidine analogues with acetylcholine-binding protein (AChBP). The crystal structures of three complexes with AChBP reveal details of molecular recognition in the orthosteric binding site and imply how the other three ligands bind. Comparisons exploiting AChBP as a surrogate for α4β2 and α7 nicotinic acetylcholine receptors (nAChRs) suggest that the key interactions are conserved. The ligands interact with the same residues as the archetypal nAChR agonist nicotine yet display greater affinity, thereby rationalizing their in vivo activity as potent antagonists of nicotine-induced antinociception. An oxyanion-binding site is formed on the periphery of the AChBP orthosteric site by Lys42, Asp94, Glu170 and Glu210. These residues are highly conserved in the human α4, β2 and α7 nAChR sequences. However, specific sequence differences are discussed that could contribute to nAChR subtype selectivity and in addition may represent a point of allosteric modulation. The ability to engage with this peripheral site may explain, in part, the function of a subset of ligands to act as agonists of α7 nAChR., (open access.)

Brain kappa-opioid receptors (KORs) are implicated in the pathophysiology of depressive and anxiety disorders, stimulating interest in the therapeutic potential of KOR antagonists. Research on KOR function has tended to focus on KOR-expressing neurons and pathways such as the mesocorticolimbic dopamine system. However, KORs are also expressed on non-neuronal cells including microglia, the resident immune cells in the brain. The effects of KOR antagonists on microglia are not understood despite the potential contributions of these cells to overall responsiveness to this class of drugs. Previous work in vitro suggests that KOR activation suppresses proinflammatory signaling mediated by immune cells including microglia. Here, we examined how KOR antagonism affects microglia function in vivo, together with its effects on physiological and behavioral responses to an immune challenge. Pretreatment with the prototypical KOR antagonist JDTic potentiates levels of proinflammatory cytokines (IL-1β, IL-6) in blood following administration of lipopolysaccharide (LPS), an immune-activating agent, without triggering effects on its own. Using magnetic-activated cell sorting (MACs), we found that KOR antagonism potentiates LPS-induced cytokine expression within microglia. This effect is accompanied by potentiation of LPS-induced hyperthermia, although reductions in body weight and locomotion were not affected. Histological analyses confirm that LPS produces visible changes in microglia morphology consistent with activation, but this effect is not altered by KOR antagonism. Considering that inflammation is increasingly implicated in depressive and anxiety disorders, these findings raise the possibility that KOR antagonist actions on microglia may detract from actions on neurons that contribute to their therapeutic potential., (Copyright © 2021 Elsevier Inc. All rights reserved.)

Inconsistent results have been reported for the effects of the mitogen-activating extracellular kinase (MEK) inhibitor α-[amino(4-aminophenyl)thio]methylene-2-(trifluoromethyl)benzeneacetonitrile (SL 327) on ethanol-induced conditioned place preference (EtOH-CPP). Since such inconsistencies may be due to the configurational composition of administered SL 327, the interconvertibility of the geometric isomers of this class of compounds has been investigated. This study provides conditions for determination of configurational composition of this class of compounds by HPLC and by 1 H NMR and reports details of configurational equilibria as a function of medium and time in solution along with solubility data for SL 327 in aqueous DMSO. The results suggest that the apparently inconsistent results reported for CPP-EtOH may be due to the administration of suspension vs. solutions, as well as to different configurational compositions of SL 327., (Copyright © 2021 Elsevier Inc. All rights reserved.)

During 2012-2018, the clandestine manufacture of new psychoactive substances (NPS) designed to circumvent substance control regulations increased exponentially worldwide, with concomitant increase in fatalities. This review focuses on three compound classes identified as synthetic opioids, synthetic amphetamines, and synthetic cannabinoids and highlights the medicinal chemistry precedents utilized by clandestine laboratories to develop new NPS with increased brain penetration, longer duration of action, and greater potency. Chemical approaches to illicit drug abuse treatment options, particularly for opioid use disorder, are also discussed., (© 2020 New York Academy of Sciences.)

Epibatidine is a potent analgetic agent with very high affinity for brain nicotinic acetylcholine receptors (nAChR). We determined the activity profiles of three epibatidine derivatives, RTI-36, RTI-76, and RTI-102, which have affinity for brain nAChR equivalent to that of epibatidine but reduced analgetic activity. RNAs coding for nAChR monomeric subunits and/or concatamers were injected into Xenopus oocytes to obtain receptors of defined subunit composition and stoichiometry. The epibatidine analogs produced protracted activation of high sensitivity (HS) α 4- and α 2-containing receptors with the stoichiometry of 2alpha:3beta subunits but not low sensitivity (LS) receptors with the reverse ratio of alpha and beta subunits. Although not strongly activated by the epibatidine analogs, LS α 4- and α 2-containing receptors were potently desensitized by the epibatidine analogs. In general, the responses of α 4(2) β 2(2) α 5 and β 3 α 4 β 2 α 6 β 2 receptors were similar to those of the HS α 4 β 2 receptors. RTI-36, the analog closest in structure to epibatidine, was the most efficacious of the three compounds, also effectively activating α 7 and α 3 β 4 receptors, albeit with lower potency and less desensitizing effect. Although not the most efficacious agonist, RTI-76 was the most potent desensitizer of α 4- and α 2-containing receptors. RTI-102, a strong partial agonist for HS α 4 β 2 receptors, was effectively an antagonist for LS α 4 β 2 receptors. Our results highlight the importance of subunit stoichiometry and the presence or absence of specific accessory subunits for determining the activity of these drugs on brain nAChR, affecting the interpretation of in vivo studies since in most cases these structural details are not known. SIGNIFICANCE STATEMENT: Epibatidine and related compounds are potent ligands for the high-affinity nicotine receptors of the brain, which are therapeutic targets and mediators of nicotine addiction. Far from being a homogeneous population, these receptors are diverse in subunit composition and vary in subunit stoichiometry. We show the importance of these structural details for drug activity profiles, which present a challenge for the interpretation of in vivo experiments since conventional methods, such as in situ hybridization and immunohistochemistry, cannot illuminate these details., (Copyright © 2020 by The American Society for Pharmacology and Experimental Therapeutics.)

Chronic health problems associated with long-term nicotine use are the leading cause of preventable death in the United States. The use of tobacco products is 3-4 times greater among individuals with cocaine use disorder than that observed in the general population. This may reflect the propensity of nicotine to augment the reinforcing effects of cocaine. However, the mechanism of action of nicotine differs from that of cocaine, which presents a significant challenge for the development of pharmacotherapeutic interventions for the management of nicotine + cocaine polydrug abuse. Bupropion, an FDA-approved smoking cessation aid, has pharmacological actions at both monoamine transporters and nicotinic receptors, suggesting that it may be effective at decreasing nicotine + cocaine coabuse. Here, rhesus monkeys (n = 4) responded for food pellets and, separately, intravenous injections of nicotine, cocaine, or nicotine + cocaine mixtures under a second-order FR2(VR16:S) schedule of reinforcement during 7- to 10-day continuous treatment with saline or bupropion (1.0 and 1.8 mg/kg/hr). Results show that bupropion treatment dose-dependently decreased self-administration of nicotine combined with a low dose of cocaine (0.0032 mg/kg/inj); however, when the dose of cocaine in the mixture was higher (i.e., 0.01 mg/kg/inj), bupropion attenuated self-administration in only a subset of subjects. The effective dosage of bupropion increased responding for cocaine alone, nicotine alone, and for saline injections and significantly increased measures of daily activity. The apparent stimulant-like effects of bupropion at the dosage required to decrease cocaine + nicotine self-administration does not support its clinical use for the management of nicotine + cocaine polydrug abuse. (PsycInfo Database Record (c) 2020 APA, all rights reserved).

Recent studies show that GPCRs rapidly interconvert between multiple states although our ability to interrogate, monitor and visualize them is limited by a relative lack of suitable tools. We previously reported two nanobodies (Nb39 and Nb6) that stabilize distinct ligand- and efficacy-delimited conformations of the kappa opioid receptor. Here, we demonstrate via X-ray crystallography a nanobody-targeted allosteric binding site by which Nb6 stabilizes a ligand-dependent inactive state. As Nb39 stabilizes an active-like state, we show how these two state-dependent nanobodies can provide real-time reporting of ligand stabilized states in cells in situ. Significantly, we demonstrate that chimeric GPCRs can be created with engineered nanobody binding sites to report ligand-stabilized states. Our results provide both insights regarding potential mechanisms for allosterically modulating KOR with nanobodies and a tool for reporting the real-time, in situ dynamic range of GPCR activity.

α-PVP (α-pyrrolidinovalerophenone) and MDPV (3,4-methylenedioxypyrovalerone) are potent abused stimulants that are members of the synthetic cathinone class of drugs. Although these drugs are taken with recreational intent, high doses can lead to unintended adverse effects including agitation, cardiovascular effects, sympathomimetic syndromes, hallucinations, and psychoses. One possible treatment is the use of a vaccine to block or attenuate adverse medical effects. These studies report the preparation of a vaccine that generates high affinity antibodies specific for both drugs and the pharmacological testing of this vaccine in male rats. Alkylation of a hydroxy-α-PVP analog with an appropriate thiol-bearing linker afforded the hapten. When hapten-conjugated carrier protein was mixed with adjuvant, the resulting vaccine stimulated production of antibodies in male Sprague Dawley rats that were found to significantly reduce α-PVP- and MDPV-induced hyperlocomotion as well as to significantly reduce the concentrations of MDPV drugs in critical organs. The novel vaccine produced high affinity antibodies against MDPV, (R)-MDPV, (S)-MDPV, and α-PVP. Cross-reactivity testing against nine structurally similar cathinones showed very limited binding, and no binding to off-target endogenous and exogenous compounds. Antibodies generated by this bi-specific vaccine also significantly shortened the duration of locomotor activity induced by both drugs up to a dose of 5.6 mg/kg in male rats., (Copyright © 2019 Elsevier Ltd. All rights reserved.)

The mechanisms behind relapse to ethanol intake in recovering alcoholics are still unclear. The negative reinforcing effects contributing to ethanol addiction, including relapse, are considered to be partly driven by the κ-opioidergic system. As the κ-opioidergic system interacts with the mesolimbic reward pathway, the aim of the study was to clarify the role of nucleus accumbens shell κ-opioidergic mechanisms in relapse to ethanol intake by using the alcohol deprivation effect (ADE) paradigm. The ADE is defined as a transient increase in voluntary ethanol intake after a forced period of abstinence. Male Long-Evans rats were trained to voluntarily consume 10% (v/v) ethanol solution. Ethanol access and deprivation cycles were initiated after stable ethanol intake baselines had been reached and bilateral guide cannulas had been implanted above the nucleus accumbens shell. One cycle consisted of 10 days of 90 min access to ethanol followed by 6 days of ethanol deprivation. The ADE was measured in the beginning of a new cycle. Rats received JDTic, a selective κ-antagonist, either subcutaneously (10 mg/kg) or intra-accumbally (15 µg/site) or, as a reference substance, systemic naltrexone (0.3 mg/kg) before ethanol re-access, and the effects on the ADE were evaluated. Systemic and intra-accumbal JDTic significantly attenuated the ADE on the first day of ethanol re-access, as did systemic naltrexone. Additionally, naltrexone decreased ethanol intake levels. These results suggest that nucleus accumbens shell κ-opioidergic mechanisms may have a role in mediating relapse to ethanol intake. Additionally, κ-antagonism could be a valuable adjunct in ethanol relapse prevention., (Copyright © 2019 Elsevier B.V. and ECNP. All rights reserved.)

Background: New treatments for stress-related disorders including depression, anxiety, and substance use disorder are greatly needed. Kappa opioid receptors are expressed in the central nervous system, including areas implicated in analgesia and affective state. Although kappa opioid receptor agonists share the antinociceptive effects of mu opioid receptor agonists, they also tend to produce negative affective states. In contrast, selective kappa opioid receptor antagonists have antidepressant- and anxiolytic-like effects, stimulating interest in their therapeutic potential. The prototypical kappa opioid receptor antagonists (e.g., norBNI, JDTic) have an exceptionally long duration of action that complicates their use in humans, particularly in tests to establish safety. This study was designed to test dose- and time-course effects of novel kappa opioid receptor antagonists with the goal of identifying short-acting lead compounds for future medication development., Methods: We screened 2 novel, highly selective kappa opioid receptor antagonists (CYM-52220 and CYM-52288) with oral efficacy in the warm water tail flick assay in rats to determine initial dose and time course effects. For comparison, we tested existing kappa opioid receptor antagonists JDTic and LY-2456302 (also known as CERC-501 or JNJ-67953964)., Results: In the tail flick assay, the rank order of duration of action for the antagonists was LY-2456302 < CYM-52288 < CYM-52220 << JDTic. Furthermore, LY-2456302 blocked the depressive (anhedonia-producing) effects of the kappa opioid receptor agonist U50,488 in the intracranial self-stimulation paradigm, albeit at a higher dose than that needed for analgesic blockade in the tail flick assay., Conclusions: These results suggest that structurally diverse kappa opioid receptor antagonists can have short-acting effects and that LY-2456302 reduces anhedonia as measured in the intracranial self-stimulation test., (© The Author(s) 2019. Published by Oxford University Press on behalf of CINP.)

This study focused on formulating conjugate vaccines targeting oxycodone and heroin for technology transfer, good manufacturing practice (GMP), and clinical evaluation. Lead vaccines used the highly immunogenic carrier protein keyhole limpet hemocyanin (KLH), which poses formulation problems because of its size. To address this barrier to translation, an oxycodone-based hapten conjugated to GMP-grade subunit KLH (OXY-sKLH) and adsorbed on alum adjuvant was studied with regard to carbodiimide coupling reaction time, buffer composition, purification methods for conjugates, conjugate size, state of aggregation, and protein/alum ratio. Vaccine formulations were screened for post-immunization antibody levels and efficacy in reducing oxycodone distribution to the brain in rats. While larger conjugates were more immunogenic, their size prevented characterization of the haptenation ratio by standard analytical methods and sterilization by filtration. To address this issue, conjugation chemistry and vaccine formulation were optimized for maximal efficacy, and conjugate size was measured by dynamic light scattering prior to adsorption to alum. An analogous heroin vaccine (M-sKLH) was also optimized for conjugation chemistry, formulated in alum, and characterized for potency against heroin in rats. Finally, this study found that the efficacy of OXY-sKLH was preserved when co-administered with M-sKLH, supporting the proof of concept for a bivalent vaccine formulation targeting both heroin and oxycodone. This study suggests methods for addressing the unique formulation and characterization challenges posed by conjugating small molecules to sKLH while preserving vaccine efficacy.