Alterations in nicotinic receptor alpha5 subunit gene differentially impact early and later stages of cocaine addiction: a translational study in transgenic rats and patients
Introduction
Cocaine use disorder (CUD) is a clinically devastating neuropsychiatric disease without any approved pharmacotherapy (Degenhardt et al., 2011). Relapse to cocaine seeking after a phase of abstinence is the main impediment in the treatment of CUD, with 78 % of patients relapsing within three months after discharge from hospital detoxification (Lima et al., 2019). In such pathological cocaine users, relapse can be precipitated by exposure to cocaine-associated cues, stress, or to cocaine itself (Childress et al., 1993; Jaffe et al., 1989; O’Brien, 1998; Sinha, 2001). Relapse prevention is thus a key target for clinicians in the treatment of substance use disorders in general since it has been associated with the chronicization and the burden of these disorders (Hendershot et al., 2011). Similarly, reinstatement of cocaine-seeking can be elicited in laboratory animals after extinction of cocaine self-administration (SA), by exposure to a conditioned stimulus (CS) previously associated with cocaine intake, stress or a cocaine-priming injection (Crombag and Shaham, 2002; de Wit and Stewart, 1981; Epstein et al., 2006; Fuchs et al., 1998).
Cocaine acts primarily by inhibiting monoamine transporters, such as the dopamine transporter (DAT), leading to increased extracellular dopamine concentration in the nucleus accumbens (NAcc), an effect considered essential for its reinforcing property. However, cocaine is also an inhibitor of nicotinic acetylcholine receptors (nAChRs) (Damaj et al., 1999; Francis et al., 2000), an action that can alter dopamine release in the NAcc (Acevedo-Rodriguez et al., 2014). nAChRs are pentameric ligand-gated ion channels composed of α (α2– α7, α9, α10) and β (β2– β4) subunits that co-assemble according to various combinations with distinct brain localizations and functional properties (Changeux, 2010). nAChRs have been investigated in preclinical models of cocaine addiction. For example, the nAChR antagonist mecamylamine reduces cocaine-SA and prevents cocaine-SA escalation in rodents (Blokhina et al., 2005; Hansen and Mark, 2007; Levin et al., 2000). Additionally, 18-MC, an antagonist of the α3β4 containing (*) nAChRs, decreases cocaine-SA in rats (Glick et al., 1996; Pace et al., 2004), and mice knockout (KO) for the β2 nAChR subunit exhibit decreased cocaine-induced conditioned place preference (Zachariou et al., 2001). Interestingly, mecamylamine also reduces craving for cocaine in CUD subjects (Reid et al., 1999).
Several independent large-scale human genome-wide association studies (GWAS) identified variants in a region on human chromosome 15 encoding the α3, α5 and β4 nAChR subunits that increase the risk for nicotine dependence (Bierut et al., 2007, 2008; Sherva et al., 2008). One of these variants, the non-synonymous coding single nucleotide polymorphism (SNP) rs16969968 found in exon 5 of the α5 gene (α5SNP) that causes an amino acid change D398N in the second intracellular loop of the α5 subunit, has been recently introduced into the genome of the rat, resulting in increased nicotine-SA at high doses and relapse (Forget et al., 2018). This α5SNP is very frequent in the general population and may induce a partial loss of function of the corresponding pentamer (Bierut et al., 2008; Frahm et al., 2011; Kuryatov et al., 2011) without altering the overall expression of the subunit, at least in rats (Forget et al., 2018). It does not modify the electrophysiological function of the receptor when expressed in concatemer in Xenopus oocytes (Prevost et al., 2020), and it lies away from agonists’ binding site, suggesting that it affects other features of the receptor, but its precise mechanism of action is currently unknown (Maskos, 2020). Interestingly, in candidate gene association studies, the α5SNP has also been associated with a lower prevalence of CUD in humans (Grucza et al., 2008; Sherva et al., 2010). In addition, three other SNPs in the α5 nAChR subunit, linked to altered levels of α5 mRNA in humans, have also been associated with CUD (Sherva et al., 2010; Wang et al., 2009). These genetic studies suggest an involvement of the α5 nAChR subunit in cocaine dependence, although the precise mechanisms and stages of CUD implicated have never been further assessed so far.
In fact, multiple behaviors related to CUD may be impacted by alterations in α5*nAChR density or mutations in the α5 nAChR subunit gene, that now need to be identified through longitudinal studies to understand how they contribute to the risk for this pathology. For this, the first goal of the present study was to clarify the role of α5*nAChRs and the impact of the α5SNP, found to confer protection for CUD in humans, on the different stages of an intravenous (IV) cocaine-SA procedure, from first cocaine intake to relapse to cocaine seeking after SA extinction, using transgenic rat lines previously generated in the laboratory (Forget et al., 2018). At the molecular level, since cocaine has been shown to inhibit nAChR-mediated currents in midbrain dopaminergic neurons (Acevedo-Rodriguez et al., 2014), we hypothesized that the association between α5*nAChRs and CUD may be linked to an implication of the α5 subunit in nAChR response to cocaine. Thus, the second objective of the study was to characterize the influence of the α5 subunit on nAChR functional response to cocaine in vitro. Finally, for translational purposes, we took advantage of the information generated by the SA longitudinal study implemented in our preclinical models to i) clarify, in humans, the association of the α5SNP with specific CUD phenotypes, and ii) seek for novel polymorphisms in the CHRNA5/A3/B4 gene cluster, linked to α5 expression levels, in association with these CUD phenotypes.
Section snippets
Experimental model and subject details
Male rats experimentally naive at the start of the study and initially weighing 250–275 g were used for all behavioral experiments. All rats were individually housed in a temperature-controlled environment on a 12-h reverse light/dark cycle (lights off from 08:00 h to 20:00 h) at the start of the experiments and were mildly food restricted (20 g/day/rat).
Wild-type (WT) rats, homozygous α5KO rats and homozygous rats constitutively carrying the rs16969968 SNP (α5SNP rats), on Long Evans
Acquisition of cocaine-SA, dose-response curve, saline substitution and motivation for cocaine-SA in α5SNP, α5KO and WT rats
α5SNP, α5KO and WT rats acquired the food training similarly (continuous reinforcement, no associated cues; Fig. 1A), suggesting an absence of learning or motor deficit in α5SNP and α5KO rats.
Rats were then submitted to chronic IV cocaine-SA. The three groups increased similarly their lever pressing behavior when the ratio progressively increased (FR-1 then FR-2 then FR-3 then FR-4 and finally FR-5; Fig. 1B). However, there was a significant interaction between sessions and genotypes on the
Phenotypes of α5SNP and α5KO rats in cocaine-SA and relapse
We first observed that the presence the α5SNP did not prevent rats from acquiring cocaine-SA, and did not alter the dose-response curve for cocaine-SA. Yet, contrary to WTs, α5SNP rats did not adapt their AL pressing to maintain constant amounts of cocaine intake when the ratio schedule of reinforcement was increased (Fig. 1C and D). This seems in line with previous indications of a protective effect of this polymorphism on cocaine dependence in humans (Grucza et al., 2008; Sherva et al., 2010
Declaration of Competing Interest
RI has received honoraria and support to attend congresses from Indivior and honoraria and consulting fees from Lundbeck. FV has received an honorarium from Lundbeck for a single lecture. FB has received honoraria or research or educational conference grants from Bristol-Myers Squibb, Otsuka, Eli Lilly and Co., Servier, Sanofi Aventis, Lundbeck, AstraZeneca, the European Space Agency and has received peer review research funding from French Ministry of research, Assistance Publique – Hôpitaux
Acknowledgements
Funding: This work was supported by the Institut Pasteur, Centre National de la Recherche ScientifiqueUMR 3571, Fondation pour la Recherche Médicale (SPF20140129365, DPA20140629803andEQU201903007822), Agence Nationale de la Recherche (ANR), Neuroscience "SNP-NIC" and BLANC, LABEX BIO-PSY, FP7 ERANET programme NICO-GENE grant agreement convention ANR no 2010-NEUR-004-01, European Commission FP7 RTD Project HEALTH-2009-Neurocyp.08-202088 grant 242167, French National Cancer Institute grant
References (89)
- et al.
Effects of nicotinic and NMDA receptor channel blockers on intravenous cocaine and nicotine self-administration in mice
Eur. Neuropsychopharmacol.
(2005) - et al.
Dose-response cocaine pharmacokinetics and metabolite profile following intravenous administration and arterial sampling in unanesthetized, freely moving male rats
Neurotoxicol. Teratol.
(1997) - et al.
Acute effects of cocaine on human brain activity and emotion
Neuron
(1997) - et al.
Reduced dopamine response to amphetamine in subjects at ultra-high risk for addiction
Biol. Psychiatry
(2014) - et al.
Second-generation PLINK: rising to the challenge of larger and richer datasets
GigaScience
(2015) - et al.
Using c-fos to study neuronal ensembles in corticostriatal circuitry of addiction
Brain Res.
(2015) - et al.
Mortality among cocaine users: a systematic review of cohort studies
Drug Alcohol Depend.
(2011) - et al.
Modeling cocaine relapse in rodents: behavioral considerations and circuit mechanisms
Prog. Neuropsychopharmacol. Biol. Psychiatry
(2018) - et al.
Granular insular cortex inactivation as a novel therapeutic strategy for nicotine addiction
Biol. Psychiatry
(2010) - et al.
Aversion to nicotine is regulated by the balanced activity of beta4 and alpha5 nicotinic receptor subunits in the medial habenula
Neuron
(2011)
18-Methoxycoronaridine, a non-toxic iboga alkaloid congener: effects on morphine and cocaine self-administration and on mesolimbic dopamine release in rats
Brain Res.
A risk allele for nicotine dependence in CHRNA5 is a protective allele for cocaine dependence
Biol. Psychiatry
Serious suicide attempts in outpatients with multiple substance use disorders
Drug Alcohol Depend.
The nicotinic antagonist mecamylamine preferentially inhibits cocaine vs. Food self-administration in rats
Physiol. Behav.
The role of neurocognitive functioning, substance use variables and the DSM-5 severity scale in cocaine relapse: a prospective study
Drug Alcohol Depend.
The Ultra-High-Risk for psychosis groups: evidence to maintain the status quo
Schizophr. Res.
Novel iboga alkaloid congeners block nicotinic receptors and reduce drug self-administration
Eur. J. Pharmacol.
Circuit changes in motor cortex during motor skill learning
Neuroscience
Assessing contributions of nucleus accumbens shell subregions to reward-seeking behavior
Drug Alcohol Depend.
A nicotine antagonist, mecamylamine, reduces cue-induced cocaine craving in cocaine-dependent subjects
Neuropsychopharmacology
Neural correlates of high and craving during cocaine self-administration using BOLD fMRI
NeuroImage
The distribution of mRNA encoded by a new member of the neuronal nicotinic acetylcholine receptor gene family (alpha 5) in the rat central nervous system
Brain Res.
Nicotine receptor inactivation decreases sensitivity to cocaine
Neuropsychopharmacology
Cocaine inhibition of nicotinic acetylcholine receptors influences dopamine release
Front. Synaptic Neurosci.
Administration of the D1-like dopamine receptor antagonist SCH-23390 into the medial nucleus accumbens shell attenuates cocaine priming-induced reinstatement of drug-seeking behavior in rats
Psychopharmacology
Expression of neuronal nicotinic acetylcholine receptor subunit mRNAs within midbrain dopamine neurons
J. Comp. Neurol.
The landscape of genomic imprinting across diverse adult human tissues
Genome Res.
The CHRNA5/CHRNA3/CHRNB4 nicotinic receptor regulome: genomic architecture, regulatory variants, and clinical associations
Hum. Mutat.
The pattern of c-Fos expression and its refractory period in the brain of rats and monkeys
Front. Cell. Neurosci.
Profound alteration in reward processing due to a human polymorphism in CHRNA5: a role in alcohol dependence and feeding behavior
Neuropsychopharmacology
Novel genes identified in a high-density genome wide association study for nicotine dependence
Hum. Mol. Genet.
Variants in nicotinic receptors and risk for nicotine dependence
Am. J. Psychiatry
Noradrenergic mechanisms in stress and anxiety: I. Preclinical studies
Synapse
Noradrenergic mechanisms in stress and anxiety: II. Clinical studies
Synapse
Interaction between noradrenaline and corticotrophin-releasing factor in the reinstatement of cocaine seeking in the rat
Psychopharmacology
From GWAS to function: using functional genomics to identify the mechanisms underlying complex diseases
Front. Genet.
Nicotine addiction and nicotinic receptors: lessons from genetically modified mice
Nat. Rev. Neurosci.
Cue reactivity and cue reactivity interventions in drug dependence
NIDA Res. Monogr.
QuantiSNP: an Objective Bayes Hidden-Markov Model to detect and accurately map copy number variation using SNP genotyping data
Nucleic Acids Res.
Residuals and Influence in Regression
Cocaine cue-induced dopamine release in recreational cocaine users
Sci. Rep.
Renewal of drug seeking by contextual cues after prolonged extinction in rats
Behav. Neurosci.
Pharmacological characterization of nicotine’s interaction with cocaine and cocaine analogs
J. Pharmacol. Exp. Ther.
Reinstatement of cocaine-reinforced responding in the rat
Psychopharmacology
Cited by (6)
Linking the CHRNA5 SNP to drug abuse liability: From circuitry to cellular mechanisms
2021, NeuropharmacologyCitation Excerpt :In line with these findings, Aroche et al. (2020) identified a protective effect of the ‘A’ allele for crack-cocaine addiction in European-descended patients (Aroche et al., 2020). Furthermore, the progression from initial cocaine use to disordered use appears to be slower amongst individuals carrying at least one copy of the ‘A’ allele (Forget et al., 2020). Chen et al. (2009) identified an association of the ‘G’ allele at CHRNA5 D398N with symptoms of alcohol use disorder in individuals of European descent (X. Chen et al., 2009).
Biomarkers of Relapse in Cocaine Use Disorder: A Narrative Review
2022, Brain SciencesIdentification of the Risk Genes Associated With Vulnerability to Addiction: Major Findings From Transgenic Animals
2022, Frontiers in NeuroscienceDeterminants of Addiction: Neurobiological, Behavioral, Cognitive, and Sociocultural Factors
2022, Determinants of Addiction: Neurobiological, Behavioral, Cognitive, and Sociocultural FactorsCOGNITIVE IMPAIRMENT IN PEOPLE WHO CONSUME BENZOYLMETILECGONINE AND ITS DERIVATIVES: A BIBLIOGRAPHIC REVIEW
2022, Health and Addictions / Salud y DrogasThe α5 Nicotinic Acetylcholine Receptor Subunit Differentially Modulates α4β2<sup>*</sup> and α3β4<sup>*</sup> Receptors
2020, Frontiers in Synaptic Neuroscience
- 1
Equal contribution.