Post-transcriptional regulation and subcellular localization of G-protein γ7 subunit: implications for striatal function and behavioral responses to cocaine.

The striatal D1 dopamine receptor (D1R) and A2a adenosine receptor (A2aR) signaling pathways play important roles in drug-related behaviors. These receptors activate the Golf protein comprised of a specific combination of αolfß2γ7 subunits. During assembly, the γ7 subunit sets the cellular level of the Golf protein. In turn, the amount of Golf protein determines the collective output from both D1R and A2aR signaling pathways. This study shows the Gng7 gene encodes multiple γ7 transcripts differing only in their non-coding regions. In striatum, Transcript 1 is the predominant isoform. Preferentially expressed in the neuropil, Transcript 1 is localized in dendrites where it undergoes post-transcriptional regulation mediated by regulatory elements in its 3' untranslated region that contribute to translational suppression of the γ7 protein. Earlier studies on gene-targeted mice demonstrated loss of γ7 protein disrupts assembly of the Golf protein. In the current study, morphological analysis reveals the loss of the Golf protein is associated with altered dendritic morphology of medium spiny neurons. Finally, behavioral analysis of conditional knockout mice with cell-specific deletion of the γ7 protein in distinct populations of medium spiny neurons reveals differential roles of the Golf protein in mediating behavioral responses to cocaine. Altogether, these findings provide a better understanding of the regulation of γ7 protein expression, its impact on Golf function, and point to a new potential target and mechanisms for treating addiction and related disorders.

Rare GPR37L1 Variants Reveal Potential Association between GPR37L1 and Disorders of Anxiety and Migraine.

GPR37L1 is an orphan receptor that couples through heterotrimeric G-proteins to regulate physiological functions. Since its role in humans is not fully defined, we used an unbiased computational approach to assess the clinical significance of rare G-protein-coupled receptor 37-like 1 (GPR37L1) genetic variants found among 51,289 whole-exome sequences from the DiscovEHR cohort. Rare GPR37L1 coding variants were binned according to predicted pathogenicity and analyzed by sequence kernel association testing to reveal significant associations with disease diagnostic codes for epilepsy and migraine, among others. Since associations do not prove causality, rare GPR37L1 variants were functionally analyzed in SK-N-MC cells to evaluate potential signaling differences and pathogenicity. Notably, receptor variants exhibited varying abilities to reduce cAMP levels, activate mitogen-activated protein kinase (MAPK) signaling, and/or upregulate receptor expression in response to the agonist prosaptide (TX14(A)), as compared with the wild-type receptor. In addition to signaling changes, knock-out (KO) of GPR37L1 or expression of certain rare variants altered cellular cholesterol levels, which were also acutely regulated by administration of the agonist TX14(A) via activation of the MAPK pathway. Finally, to simulate the impact of rare nonsense variants found in the large patient cohort, a KO mouse line lacking Gpr37l1 was generated. Although KO animals did not recapitulate an acute migraine phenotype, the loss of this receptor produced sex-specific changes in anxiety-related disorders often seen in chronic migraineurs. Collectively, these observations define the existence of rare GPR37L1 variants associated with neuropsychiatric conditions in the human population and identify the signaling changes contributing to pathological processes.

Rare GPR37L1 variants reveal potential roles in anxiety and migraine disorders.

GPR37L1 is an orphan receptor that couples through heterotrimeric G-proteins to regulate physiological functions. Since its role in humans is not fully defined, we used an unbiased computational approach to assess the clinical significance of rare GPR37L1 genetic variants found among 51,289 whole exome sequences from the DiscovEHR cohort. Briefly, rare GPR37L1 coding variants were binned according to predicted pathogenicity, and analyzed by Sequence Kernel Association testing to reveal significant associations with disease diagnostic codes for epilepsy and migraine, among others. Since associations do not prove causality, rare GPR37L1 variants were then functionally analyzed in SK-N-MC cells to evaluate potential signaling differences and pathogenicity. Notably, receptor variants exhibited varying abilities to reduce cAMP levels, activate MAPK signaling, and/or upregulate receptor expression in response to the agonist prosaptide (TX14(A)), as compared to the wild-type receptor. In addition to signaling changes, knockout of GPR37L1 or expression of certain rare variants altered cellular cholesterol levels, which were also acutely regulated by administration of the agonist TX14(A) via activation of the MAPK pathway. Finally, to simulate the impact of rare nonsense variants found in the large patient cohort, a knockout (KO) mouse line lacking Gpr37L1 was generated, revealing loss of this receptor produced sex-specific changes implicated in migraine-related disorders. Collectively, these observations define the existence of rare GPR37L1 variants in the human population that are associated with neuropsychiatric conditions and identify the underlying signaling changes that are implicated in the in vivo actions of this receptor in pathological processes leading to anxiety and migraine. SIGNIFICANCE STATEMENT: G-protein coupled receptors (GPCRs) represent a diverse group of membrane receptors that contribute to a wide range of diseases and serve as effective drug targets. However, a number of these receptors have no identified ligands or functions, i.e., orphan receptors. Over the past decade, advances have been made, but there is a need for identifying new strategies to reveal their roles in health and disease. Our results highlight the utility of rare variant analyses of orphan receptors for identifying human disease associations, coupled with functional analyses in relevant cellular and animal systems, to ultimately reveal their roles as novel drug targets for treatment of neurological disorders that lack wide-spread efficacy.

Selective Manipulation of G-Protein γ7 Subunit in Mice Provides New Insights into Striatal Control of Motor Behavior.

Stimulatory coupling of dopamine D1 (D1R) and adenosine A2A receptors (A2AR) to adenylyl cyclase within the striatum is mediated through a specific Gαolfß2γ7 heterotrimer to ultimately modulate motor behaviors. To dissect the individual roles of the Gαolfß2γ7 heterotrimer in different populations of medium spiny neurons (MSNs), we produced and characterized conditional mouse models, in which the Gng7 gene was deleted in either the D1R- or A2AR/D2R-expressing MSNs. We show that conditional loss of γ7 disrupts the cell type-specific assembly of the Gαolfß2γ7 heterotrimer, thereby identifying its circumscribed roles acting downstream of either the D1Rs or A2ARs in coordinating motor behaviors, including in vivo responses to psychostimulants. We reveal that Gαolfß2γ7/cAMP signal in D1R-MSNs does not impact spontaneous and amphetamine-induced locomotor behaviors in male and female mice, while its loss in A2AR/D2R-MSNs results in a hyperlocomotor phenotype and enhanced locomotor response to amphetamine. Additionally, Gαolfß2γ7/cAMP signal in either D1R- or A2AR/D2R-expressing MSNs is not required for the activation of PKA signaling by amphetamine. Finally, we show that Gαolfß2γ7 signaling acting downstream of D1Rs is selectively implicated in the acute locomotor-enhancing effects of morphine. Collectively, these results support the general notion that receptors use specific Gαßγ proteins to direct the fidelity of downstream signaling pathways and to elicit a diverse repertoire of cellular functions. Specifically, these findings highlight the critical role for the γ7 protein in determining the cellular level, and hence, the function of the Gαolfß2γ7 heterotrimer in several disease states associated with dysfunctional striatal signaling.SIGNIFICANCE STATEMENT Dysfunction or imbalance of cAMP signaling in the striatum has been linked to several neurologic and neuropsychiatric disorders, including Parkinson's disease, dystonia, schizophrenia, and drug addiction. By genetically targeting the γ7 subunit in distinct striatal neuronal subpopulations in mice, we demonstrate that the formation and function of the Gαolfß2γ7 heterotrimer, which represents the rate-limiting step for cAMP production in the striatum, is selectively disrupted. Furthermore, we reveal cell type-specific roles for Gαolfß2γ7-mediated cAMP production in the control of spontaneous locomotion as well as behavioral and molecular responses to psychostimulants. Our findings identify the γ7 protein as a novel therapeutic target for disease states associated with dysfunctional striatal cAMP signaling.

NOP receptor antagonism reduces alcohol drinking in male and female rats through mechanisms involving the central amygdala and ventral tegmental area.

BACKGROUND AND PURPOSE: Nociceptin/orphanin FQ (N/OFQ) peptide and its cognate receptor (NOP) are widely expressed in mesolimbic brain regions where they play an important role in modulating reward and motivation. Early evidence suggested that NOP receptor activation attenuates the rewarding effects of drugs of abuse, including alcohol. However, emerging data indicate that NOP receptor blockade also effectively attenuates alcohol drinking and relapse. To advance our understanding of the role of the N/OFQ-NOP receptor system in alcohol abuse, we examined the effect of NOP receptor blockade on voluntary alcohol drinking at the neurocircuitry level. EXPERIMENTAL APPROACH: Using male and female genetically selected alcohol-preferring Marchigian Sardinian (msP) rats, we initially evaluated the effects of the selective NOP receptor antagonist LY2817412 (3, 10, and 30 mg·kg-1 , p.o.) on alcohol consumption in a two-bottle free-choice paradigm. We then microinjected LY2817412 (3 and 6 µg·µl-1 per rat) in the central nucleus of the amygdala (CeA), ventral tegmental area (VTA), and nucleus accumbens (NAc). KEY RESULTS: Peripheral LY2817412 administration dose-dependently and selectively reduced voluntary alcohol intake in male and female msP rats. Central injections of LY2817412 markedly attenuated voluntary alcohol intake in both sexes following administration in the CeA and VTA but not in the NAc. CONCLUSION AND IMPLICATIONS: The present results revealed that the CeA and VTA are neuroanatomical substrates that mediate the effects of NOP receptor antagonism on alcohol consumption. Overall, our findings support the potential of NOP receptor antagonism as a treatment strategy to attenuate alcohol use and addiction.

NOP Receptor Antagonists Decrease Alcohol Drinking in the Dark in C57BL/6J Mice.

BACKGROUND: The nociceptin/orphanin FQ opioid peptide (NOP) receptor and its endogenous ligand N/OFQ have been implicated in the regulation of drug and alcohol use disorders (AUD). In particular, evidence demonstrated that NOP receptor activation blocks reinforcing and motivating effects of alcohol across a range of behavioral measures, including alcohol intake, conditioned place preference, and vulnerability to relapse. METHODS: Here, we show the effects of pharmacological activation and inhibition of NOP receptors on binge-like alcohol consumption, as measured by the "drinking in the dark" (DID) model in C57BL/6J mice. RESULTS: We found that 2 potent and selective NOP agonists AT-202 (0, 0.3, 1, 3 mg/kg) and AT-312 (0, 0.3, 1 mg/kg) did not affect binge alcohol drinking at doses that do not affect locomotor activity. AT-202 also failed to alter DID behavior when administered to mice previously exposed to chronic alcohol treatment with an alcohol-containing liquid diet. Conversely, treatment with either the high affinity NOP receptor antagonist SB-612111 (0, 3, 10, 30 mg/kg) or the selective antagonist LY2817412 (0, 3, 10, 30 mg/kg) decreased binge drinking. SB-612111 was effective at all doses examined, and LY2817412 was effective at 30 mg/kg. Consistently, NOP receptor knockout mice consumed less alcohol compared to wild type. SB-612111 reduced DID and increased sucrose consumption at doses that do not appear to affect locomotor activity. However, the high dose of SB-612111 (30 mg/kg) reduced alcohol intake but failed to inhibit preference in a 2-bottle choice DID model that can assess moderate alcohol intake. CONCLUSIONS: The present results suggest that NOP receptor inhibition rather than activation may represent a valuable approach for treatment of AUD characterized by excessive alcohol consumption such as binge drinking.

Agonist-selective NOP receptor phosphorylation correlates in vitro and in vivo and reveals differential post-activation signaling by chemically diverse agonists.

Agonists of the nociceptin/orphanin FQ opioid peptide (NOP) receptor, a member of the opioid receptor family, are under active investigation as novel analgesics, but their modes of signaling are less well characterized than those of other members of the opioid receptor family. Therefore, we investigated whether different NOP receptor ligands showed differential signaling or functional selectivity at the NOP receptor. Using newly developed phosphosite-specific antibodies to the NOP receptor, we found that agonist-induced NOP receptor phosphorylation occurred primarily at four carboxyl-terminal serine (Ser) and threonine (Thr) residues, namely, Ser346, Ser351, Thr362, and Ser363, and proceeded with a temporal hierarchy, with Ser346 as the first site of phosphorylation. G protein-coupled receptor kinases 2 and 3 (GRK2/3) cooperated during agonist-induced phosphorylation, which, in turn, facilitated NOP receptor desensitization and internalization. A comparison of structurally distinct NOP receptor agonists revealed dissociation in functional efficacies between G protein-dependent signaling and receptor phosphorylation. Furthermore, in NOP-eGFP and NOP-eYFP mice, NOP receptor agonists induced multisite phosphorylation and internalization in a dose-dependent and agonist-selective manner that could be blocked by specific antagonists. Our study provides new tools to study ligand-activated NOP receptor signaling in vitro and in vivo. Differential agonist-selective NOP receptor phosphorylation by chemically diverse NOP receptor agonists suggests that differential signaling by NOP receptor agonists may play a role in NOP receptor ligand pharmacology.

PPARα/CB1 receptor dual ligands as a novel therapy for alcohol use disorder: Evaluation of a novel oleic acid conjugate in preclinical rat models.

Recent studies have demonstrated the utility of drugs modulating the endogenous cannabinoid system to control excessive alcohol intake. Among them, drugs interacting with acylethanolamide receptors including cannabinoid CB1 receptor antagonists/inverse agonists, peroxisome proliferator-activated receptor alpha (PPARα) agonists or peroxisome proliferator-activated receptor gamma (PPARγ) agonists have demonstrated utility in the reduction of alcohol intake in animal models. However, few studies have addressed the potential utility of combining these classes of drugs, especially because of expected safety problems. In the present work we took the advantage of the availability of two novel dual ligands for these receptors, to test the hypothesis that these types of drugs might reproduce and even improve the pharmacological profile of those drugs interacting with single targets. To this end we tested (R)-3-[(4-Benzyl-2-oxooxazolidin-3-yl)methyl]-N-[4-(dodecylcarbamoyl)phenyl]benzamide (NF 10-360), a dual PPARα/γ agonist, and N-[1-(3,4-dihydroxyphenyl)propan-2-yl]oleamide (OLHHA), a dual CB1 receptor antagonist/PPARα agonist, in animal models of alcohol consumption. Both drugs were effective in reducing alcohol intake and alcohol self-administration, being OLHHA a very potent alcohol intake inhibitor (EC50 0.2 mg/kg). OLHHA also reduced self-administration of the opioid oxycodone. OLHHA actions on alcohol self-administration were replicated in alcohol-preferring Marchigian-Sardinian msP rats. Repeated administration of OLHHA did result neither in tolerance nor in toxicological or deleterious metabolic changes in the liver of msP rats. These data support the feasibility of developing novel dual ligands interacting with cannabinoid targets to treat alcohol use disorder in humans.

Influence of neuropathic pain on nicotinic acetylcholine receptor plasticity and behavioral responses to nicotine in rats.

Tobacco smoking is particularly evident in individuals experiencing chronic pain. This complex relationship is poorly understood at both molecular and behavioral levels. Here, we describe experiments aimed at understanding whether a chronic pain state induces neuroadaptations into the brain or peripheral nerves that involve nicotinic acetylcholine receptors (nAChRs) and whether these neuroadaptations directly lead to increased vulnerability to nicotine addiction or to the development of coping strategies to relieve pain symptoms. We found that ligation of the rat L5 spinal nerve led to a dramatic downregulation in the mRNA expression levels of all nAChR subunits examined in dorsal root ganglia and a time-dependent downregulation of discrete subunits, particularly in the cingulate cortex and the amygdala. Spinal nerve ligation and sham-operated rats showed minor or no changes in patterns of acquisition and motivation for nicotine taking. Spinal nerve ligation rats also showed similar vulnerability to nicotine seeking as sham animals when reinstatement was induced by nicotine-associated cues, but failed to reinstate lever pressing when relapse was induced by nicotine priming. Spinal nerve ligation and sham rats were equally sensitive to nicotine-induced anxiety-like behavior and antinociception; however, nicotine produced a potent and long-lasting antiallodynic effect in spinal nerve ligation rats. These results demonstrate that chronic pain leads to plasticity of nAChRs that do not directly facilitate nicotine addictive behaviors. Instead, nicotine potently decreases allodynia, an effect that could lead to increased nicotine consumption in chronic pain subjects.

Analysis of the distribution of spinal NOP receptors in a chronic pain model using NOP-eGFP knock-in mice.

BACKGROUND AND PURPOSE: The nociceptin/orphanin FQ opioid peptide (NOP) receptor system plays a significant role in the regulation of pain. This system functions differently in the spinal cord and brain. The mechanism by which the NOP receptor agonists regulate pain transmission in these regions is not clearly understood. Here, we investigate the peripheral and spinal NOP receptor distribution and antinociceptive effects of intrathecal nociceptin/orphanin FQ (N/OFQ) in chronic neuropathic pain. EXPERIMENTAL APPROACH: We used immunohistochemistry to determine changes in NOP receptor distribution triggered by spinal nerve ligation (SNL) using NOP-eGFP knock-in mice. Antinociceptive effects of intrathecal N/OFQ on SNL-mediated allodynia and heat/cold hyperalgesia were assessed in wild-type mice. KEY RESULTS: NOP-eGFP immunoreactivity was decreased by SNL in the spinal laminae I and II outer, regions that mediate noxious heat stimuli. In contrast, immunoreactivity of NOP-eGFP was unchanged in the ventral border of lamina II inner, which is an important region for the development of allodynia. NOP-eGFP expression was also decreased in a large number of primary afferents in the L4 dorsal root ganglion (DRG) of SNL mice. However, SNL mice showed increased sensitivity, compared to sham animals to the effects of i.t administered N/OFQ with respect to mechanical as well as thermal stimuli. CONCLUSIONS AND IMPLICATIONS: Our findings suggest that the spinal NOP receptor system attenuates injury-induced hyperalgesia by direct inhibition of the projection neurons in the spinal cord that send nociceptive signals to the brain and not by inhibiting presynaptic terminals of DRG neurons in the superficial lamina.

Differential regulation of alcohol taking and seeking by antagonism at α4ß2 and α3ß4 nAChRs.

RATIONALE: Alcoholism is a serious public health problem throughout the world. Current pharmacotherapies for the treatment of this disorder are poorly effective. Preclinical and clinical findings point to nicotinic acetylcholine receptors (nAChRs) as a promising target for the development of novel and effective medications. Assuage Pharmaceuticals, in collaboration with Torrey Pines Institute for Molecular Studies, has discovered a new class of potent and selective α4ß2 nAChR antagonists. OBJECTIVE: Here, it was hypothesized that α4ß2 nAChR antagonism is a viable approach for treatment of alcohol use disorders. RESULTS: When tested in rats, one lead compound, AP-202, attenuated both operant alcohol and nicotine self-administration in a paradigm in which the two reinforcers were concurrently available. The conotoxin TP2212-59, a selective α3ß4 nAChR antagonist, was only effective in reducing nicotine self-administration. AP-202 also reduced alcohol but not food responding when alcohol was presented as the only reinforcer, whereas the commercially available α4ß2 nAChR antagonist dihydro-ß-erythroidine failed to alter alcohol self-administration. AP-202 did not block relapse-like behavior induced by previously alcohol-associated stimuli or yohimbine stress. In a reinstatement paradigm, in which alcohol seeking was triggered by a nicotine challenge, a behavior successfully inhibited by the nonselective nAChR antagonist mecamylamine, AP-202 was not effective, while pretreatment with TP2212-59 abolished nicotine-induced reinstatement of alcohol seeking. CONCLUSIONS: These findings suggest differential roles for α4ß2 and α3ß4 nAChR on alcohol taking and seeking with selective blockade of α4ß2 nAChR being more implicated in modulating alcohol taking while selective blockade of α3ß4 nAChR is involved in nicotine-induced alcohol seeking.

Early life exposure to permethrin: a progressive animal model of Parkinson's disease.

INTRODUCTION: Oxidative stress, alpha-synuclein changes, mitochondrial complex I defects and dopamine loss, observed in the striatum of rats exposed to the pesticide permethrin in early life, could represent neuropathological hallmarks of Parkinson's disease (PD). Nevertheless, an animal model of PD should also fulfill criteria of face and predictive validities. This study was designed to: 1) verify dopaminergic status in the striatum and substantia nigra pars compacta; 2) recognize non-motor symptoms; 3) investigate the time-course development of motor disabilities; 4) assess L-Dopa effectiveness on motor symptoms in rats previously exposed to permethrin in early life. METHODS: The permethrin-treated group received 34mg/kg daily of permethrin from postnatal day 6 to 21, whereas the age-matched control group was administered with the vehicle only. RESULTS: At adolescent age, the permethrin-treated group showed decreased levels of dopamine in the striatum, loss of dopaminergic neurons in the substantia nigra pars compacta and cognitive impairments. Motor coordination defects appeared at adult age (150days old) in permethrin-treated rats on rotarod and beam walking tasks, whereas no differences between the treated and control groups were detected on the foot print task. Predictive validity was evaluated by testing the ability of L-Dopa (5, 10 or 15mg/kg, os) to restore the postural instability in permethrin-treated rats (150days old) tested in a beam walking task. The results revealed full reversal of motor deficits starting from 10mg/kg of L-Dopa. DISCUSSION: The overall results indicate that this animal model replicates the progressive, time-dependent nature of the neurodegenerative process in Parkinson's disease.

A key role for the N/OFQ-NOP receptor system in modulating nicotine taking in a model of nicotine and alcohol co-administration.

Alcohol and nicotine are often co-abused. Although the N/OFQ-NOP receptor system is considered a potential target for development of drug abuse pharmacotherapies, especially for alcoholism, little is known about the role of this system in nicotine dependence. Furthermore, the effect of prior history of nicotine dependence on subsequent nicotine and alcohol taking is understudied. Using an operant co-administration paradigm, in which rats concurrently self-administer nicotine and alcohol, we found that nicotine dependent rats increased nicotine self-administration over time as compared to non-dependent animals, while patterns of alcohol lever pressing did not change between groups. Pretreatment with the potent NOP receptor agonist AT-202 (0.3-3 mg/kg) increased nicotine lever pressing of both dependent and non-dependent groups, whereas the selective antagonist SB612111 (1-10 mg/kg) elicited a clear reduction of nicotine responses, in both dependent and non-dependent rats. In parallel, AT-202 only produced minor changes on alcohol responses and SB612111 reduced alcohol taking at a dose that also reduced locomotor behavior. Results indicate that a history of nicotine dependence affects subsequent nicotine- but not alcohol-maintained responding, and that NOP receptor antagonism, rather than agonism, blocks nicotine self-administration, which strongly suggests a critical role for the endogenous N/OFQ in the modulation of nicotine reinforcement processes.

Knock-In Mice with NOP-eGFP Receptors Identify Receptor Cellular and Regional Localization.

The nociceptin/orphanin FQ (NOP) receptor, the fourth member of the opioid receptor family, is involved in many processes common to the opioid receptors including pain and drug abuse. To better characterize receptor location and trafficking, knock-in mice were created by inserting the gene encoding enhanced green fluorescent protein (eGFP) into the NOP receptor gene (Oprl1) and producing mice expressing a functional NOP-eGFP C-terminal fusion in place of the native NOP receptor. The NOP-eGFP receptor was present in brain of homozygous knock-in animals in concentrations somewhat higher than in wild-type mice and was functional when tested for stimulation of [(35)S]GTPγS binding in vitro and in patch-clamp electrophysiology in dorsal root ganglia (DRG) neurons and hippocampal slices. Inhibition of morphine analgesia was equivalent when tested in knock-in and wild-type mice. Imaging revealed detailed neuroanatomy in brain, spinal cord, and DRG and was generally consistent with in vitro autoradiographic imaging of receptor location. Multicolor immunohistochemistry identified cells coexpressing various spinal cord and DRG cellular markers, as well as coexpression with µ-opioid receptors in DRG and brain regions. Both in tissue slices and primary cultures, the NOP-eGFP receptors appear throughout the cell body and in processes. These knock-in mice have NOP receptors that function both in vitro and in vivo and appear to be an exceptional tool to study receptor neuroanatomy and correlate with NOP receptor function. SIGNIFICANCE STATEMENT: The NOP receptor, the fourth member of the opioid receptor family, is involved in pain, drug abuse, and a number of other CNS processes. The regional and cellular distribution has been difficult to determine due to lack of validated antibodies for immunohistochemical analysis. To provide a new tool for the investigation of receptor localization, we have produced knock-in mice with a fluorescent-tagged NOP receptor in place of the native NOP receptor. These knock-in mice have NOP receptors that function both in vitro and in vivo and have provided a detailed characterization of NOP receptors in brain, spinal cord, and DRG neurons. They appear to be an exceptional tool to study receptor neuroanatomy and correlate with NOP receptor function.

Pharmacological stress is required for the anti-alcohol effect of the α3ß4* nAChR partial agonist AT-1001.

Alcohol and nicotine are often taken together. The mechanisms underlying this frequent co-abuse are not well known. Genetic and pharmacological evidence suggests that the nicotinic acetylcholine receptors (nAChRs) containing the α3 and ß4 subunits play a role in alcohol as well as nicotine addiction. AT-1001 is a high affinity α3ß4 nAChR partial agonist recently found to block nicotine self-administration and relapse-like behavior in rats. Here, to study the involvement of α3ß4 nAChRs in the mechanisms that regulate alcohol abuse we evaluated the effects of AT-1001 on alcohol taking and seeking in Sprague-Dawley rats. AT-1001 reduced operant alcohol self-administration at the highest dose examined (3.0 mg/kg), an effect also observed for food self-administration. A dose of 1.5 mg/kg AT-1001, which had no effect on alcohol or food self-administration, essentially eliminated reinstatement of alcohol seeking induced by yohimbine (0.625 mg/kg) whereas, reinstatement induced by alcohol-associated cues was not altered, nor did AT-1001 induce reinstatement of extinguished self-administration on its own. Finally, AT-1001 showed an anxiolytic activity when measured in the presence or absence of yohimbine stress in the elevated plus maze paradigm. Together, these observations do not support a specific involvement of the α3ß4 nAChR in mediating alcohol reward or cue-induced relapse to alcohol seeking but rather indicate that the α3ß4 nAChR partial agonism may constitute an attractive approach for treating alcohol use disorders exacerbated by elevated stress response.