Elevated Monoamine Oxidase A Binding During Major Depressive Episodes Is Associated with Greater Severity and Reversed Neurovegetative Symptoms

Inadequate treatment response occurs in approximately 40% of major depressive episodes (MDEs), and one approach to solve this is careful matching of treatment to the specific pathologies of MDE. One such biological abnormality is elevated monoamine oxidase A (MAO-A) levels, which occurs in the prefrontal and anterior cingulate cortex (PFC and ACC) during MDE; however, the subtypes for which this abnormality is most prominent are unknown. We hypothesized that MAO-A levels in the PFC and ACC are most elevated in MDE with greater severity and reversed neurovegetative symptoms (hypersomnia and either hyperphagia or weight gain). MAO-A V_T (an index of MAO-A density) was measured using [¹¹C]harmine positron emission tomography (PET) in 42 subjects with MDEs secondary to major depressive disorder and 37 healthy controls. The effect of severity and reversed neurovegetative symptoms on MAO-A V_T in the PFC and ACC was analyzed using a multivariate analysis of variance (MANOVA). Greater severity and reversed neurovegetative symptoms were associated with elevated MAO-A V_T in the PFC and ACC (MANOVA, severity: F_(2,38)=5.44, p=0.008; reversed neurovegetative symptoms: F_(2,38)=5.13, p=0.01). Increased MAO-A level, when greater severity and reversed neurovegetative symptoms are present, may explain the association of these clinical features with a preferential response to MAO inhibitors, which is especially well-evidenced for reversed neurovegetative symptoms in MDE. As MAO-A creates oxidative stress, facilitates apoptosis, and metabolizes monoamines, therapeutics opposing these processes are predicted to best treat MDE with greater severity and reversed neurovegetative symptoms.     

Relationship of Monoamine Oxidase-A Distribution Volume to Postpartum Depression and Postpartum Crying

Postpartum depression (PPD) has a prevalence rate of 13% and a similarly high proportion of women report a subclinical state of one or more major depressive episode symptoms. The aim was to investigate whether monoamine oxidase-A (MAO-A) V_T, an index of MAO-A density, is increased in the prefrontal and anterior cingulate cortex (PFC and ACC), during PPD or when a PPD spectrum symptom, greater predisposition to crying, is present. MAO-A is an enzyme that increases in density after estrogen decline, and has several functions including creating oxidative stress, influencing apoptosis and monoamine metabolism. Fifty-seven women were recruited including 15 first-onset, antidepressant naive, PPD subjects, 12 postpartum healthy who cry due to sad mood, 15 asymptomatic postpartum healthy women, and 15 healthy women not recently pregnant. Each underwent [¹¹C]-harmine positron emission tomography scanning to measure MAO-A V_T. Both PPD and greater predisposition to crying were associated with greater MAO-A V_T in the PFC and ACC (multivariate analysis of variance (MANOVA), group effect, F_21,135=1.856; p=0.019; mean combined region elevation 21% and 14% in PPD and crying groups, respectively, relative to postpartum asymptomatic). Greater MAO-A V_T in the PFC and ACC represents a new biomarker in PPD, and the PPD symptom of predisposition to crying. Novel strategies for preventing PPD (and some PPD symptoms) may be possible by avoiding environmental conditions that elevate MAO-A level and enhancing conditions that normalize MAO-A level. These findings also argue for clinical trials in PPD with the newer, well-tolerated MAO-A inhibitor antidepressants.     

Increased impulsivity in cocaine dependent subjects independent of antisocial personality disorder and aggression

Several previous studies show a relationship between impulsivity and substance abuse; however, it is unclear whether the increased impulsivity seen in substance dependent groups is specifically related to substance abuse, or if it is due to concomitant antisocial personality disorder (ASPD) or aggression. The issue of whether impulsivity is specifically related to substance abuse is important since it has a bearing on risk factors for development of substance abuse. To determine whether cocaine dependent subjects show increased impulsivity independent of ASPD, the Barratt impulsiveness scale (BIS-11), a delayed reward laboratory measure of impulsivity, and the life history of aggression scale were administered to 49 cocaine dependent subjects and 25 controls. Results showed that cocaine dependent subjects with ASPD were more impulsive and aggressive than controls, but cocaine dependent subjects without ASPD were also more impulsive compared to controls. Controlling for aggression history, cocaine dependent subjects without ASPD continued to have elevated impulsivity as measured by the BIS-11, but not the delayed reward task. This study supports the hypothesis that the increased impulsivity as measured by the BIS-11 in cocaine dependent individuals is not exclusively due to concomitant increases in aggression or ASPD.     

Effects of topiramate on neural responses to alcohol cues in treatment-seeking individuals with alcohol use disorder: preliminary findings from a randomized,placebo-controlled trial

Topiramate, a GABA/glutamate modulator, is efficacious in reducing alcohol consumption, though the mechanisms underlying this effect are not well characterized. This study analyzed functional magnetic resonance imaging (fMRI) data from 22 heavy drinkers enrolled in a 12-week placebo-controlled, randomized clinical trial of topiramate to examine the effects of topiramate on alcohol cue-elicited brain responses, craving, and heavy drinking in individuals with DSM-5 alcohol use disorder. Patients were randomized to receive either topiramate (maximal daily dosage of 200 mg/day) or placebo and were administered an fMRI alcohol cue-reactivity task at baseline (before starting medication) and after 6 weeks of double-blind treatment. Analyses compared the topiramate (n = 12) and placebo (n = 8) groups on (1) the change in brain responses during alcohol cue exposure (vs non-alcohol cues) within five a priori regions of interest related to reward—the bilateral and medial orbitofrontal cortex (OFC) and bilateral ventral striatum (VS) and (2) change in craving and heavy drinking days (HDDs) from baseline and scan 2. Topiramate, relative to placebo, reduced alcohol cue-elicited activation of the left VS, bilateral OFC, and medial OFC, alcohol cue-elicited craving, and HDDs between baseline and 6 weeks of treatment. The reduction in alcohol cue-elicited activation in the medial OFC correlated with reductions in craving, and reduced activation in the right VS, right OFC, and medial OFC correlated with the reduction in HDD. This preliminary study provides evidence that topiramate’s attenuation of alcohol cue-elicited brain activation and craving are key elements of the drug’s neurobiological mechanism of action in reducing heavy drinking.Subject terms: Translational research, Predictive markers     

Treatment for Tobacco Dependence: Effect on Brain Nicotinic Acetylcholine Receptor Density

Cigarette smoking leads to upregulation of brain nicotinic acetylcholine receptors (nAChRs), including the common α₄β₂* nAChR subtype. Although a substantial percentage of smokers receive treatment for tobacco dependence with counseling and/or medication, the effect of a standard course of these treatments on nAChR upregulation has not yet been reported. In the present study, 48 otherwise healthy smokers underwent positron emission tomography (PET) scanning with the radiotracer 2-FA (for labeling α₄β₂* nAChRs) before and after treatment with either cognitive-behavioral therapy, bupropion HCl, or pill placebo. Specific binding volume of distribution (V_S/f_P), a measure proportional to α₄β₂* nAChR density, was determined for regions known to have nAChR upregulation with smoking (prefrontal cortex, brainstem, and cerebellum). In the overall study sample, significant decreases in V_S/f_P were found for the prefrontal cortex, brainstem, and cerebellum of −20 (±35), −25 (±36), and −25 (±31)%, respectively, which represented movement of V_S/f_P values toward values found in non-smokers (mean 58.2% normalization of receptor levels). Participants who quit smoking had significantly greater reductions in V_S/f_P across regions than non-quitters, and correlations were found between reductions in cigarettes per day and decreases in V_S/f_P for brainstem and cerebellum, but there was no between-group effect of treatment type. Thus, smoking reduction and cessation with commonly used treatments (and pill placebo) lead to decreased α₄β₂* nAChR densities across brain regions. Study findings could prove useful in the treatment of smokers by providing encouragement with the knowledge that decreased smoking leads to normalization of specific brain receptors.     

The Relationship between Antisocial and Borderline Features and Aggression in Young Adult Men in Treatment for Substance Use Disorders

There is a large literature documenting that adult men in treatment for substance use disorders perpetrate more aggression than men without substance use disorders. Unfortunately, there is minimal research on aggression among young adult men (i.e., 18–25 years of age) in treatment for substance use. Moreover, although aggression is more likely to occur when individuals are acutely intoxicated by alcohol or drugs, research also suggests that antisocial (ASPD) and borderline (BPD) personality features increase the chances an individual will use aggression. The current study therefore examined the associations between ASPD and BPD features, including specific features that are reflective of impulsivity, and aggression in young adult men in treatment for substance use disorders (N = 79). Controlling for age, education, alcohol and drug use, ASPD features were positively associated with various indicators of aggression (e.g., physical, verbal, attitudinal), whereas BPD features were only associated with physical aggression. However, ASPD and BPD features that were specific to impulsivity were robustly related to indicators of aggression. Findings suggest that substance use treatment should attempt to target ASPD and BPD features in young adult men, which may help reduce aggression after treatment.     

Contributions of the orbitofrontal cortex to impulsive choice: interactions with basal levels of impulsivity, dopamine signalling, and reward-related cues

Rationale  

Individual differences in impulsive decision-making may be critical determinants of vulnerability to impulse control disorders and substance abuse, yet little is known of their biological or behavioural basis. The orbitofrontal cortex (OFC) has been heavily implicated in the regulation of impulsive decision-making. However, lesions of the OFC in rats have both increased and decreased impulsivity in delay-discounting paradigms, where impulsive choice is defined as the selection of small immediate over larger delayed rewards.     

Cocaine Modulation of Frontostriatal Expression of Zif268, D2, and 5-HT2c Receptors in High and Low Impulsive Rats

Impulsivity shares high comorbidity with substance abuse in humans, and high impulsivity (HI) in rats has been identified as a predictive factor for cocaine addiction-like behavior. Despite the evidence that high impulsivity is associated with altered function of corticostriatal networks, the specific neural substrates underlying the increased vulnerability of impulsive individuals to develop cocaine addiction remain unknown. We therefore investigated specific neural correlates of HI within the corticostriatal circuitry and determined how they interact with a protracted history of cocaine self-administration. We used in situ hybridization to map brain expression of two major genes implicated in impulsivity, encoding the dopamine D2 receptor (DA D2R) and the 5-HT2c receptor (5-HT2cR), and an immediate early gene associated with neuronal plasticity, zif268, in groups of rats selected for HI and low impulsivity (LI) on a 5-choice serial reaction time task (5-CSRTT) immediately after 5-CSRTT training, and following 10 or 50 days of cocaine self-administration. HI rats exhibited decreased DA D2R mRNA in the mesolimbic pathway, and increased 5-HT2cR mRNA in the orbitofrontal cortex compared with LI rats. HI rats also showed decreased zif268 mRNA in the ventral and dorsomedial striatum. Cocaine exposure decreased striatal D2R mRNA in both HI and LI rats, decreased 5-HT2cR mRNA differentially in striatal and prefrontal areas between HI and LI rats, and selectively decreased zif268 mRNA in the orbitofrontal and infralimbic cortices of HI animals. These findings implicate novel markers underlying the vulnerability of impulsive rats to cocaine addiction that localize to the OFC, infralimbic cortex, and striatum.     

Cumulative Adversity Sensitizes Neural Response to Acute Stress: Association with Health Symptoms

Cumulative adversity (CA) increases stress sensitivity and risk of adverse health outcomes. However, neural mechanisms underlying these associations in humans remain unclear. To understand neural responses underlying the link between CA and adverse health symptoms, the current study assessed brain activity during stress and neutral-relaxing states in 75 demographically matched, healthy individuals with high, mid, and low CA (25 in each group), and their health symptoms using the Cornell Medical Index. CA was significantly associated with greater adverse health symptoms (P=0.01) in all participants. Functional magnetic resonance imaging results indicated significant associations between CA scores and increased stress-induced activity in the lateral prefrontal cortex, insula, striatum, right amygdala, hippocampus, and temporal regions in all 75 participants (p<0.05, whole-brain corrected). In addition to these regions, the high vs low CA group comparison revealed decreased stress-induced activity in the medial orbitofrontal cortex (OFC) in the high CA group (p<0.01, whole-brain corrected). Specifically, hypoactive medial OFC and hyperactive right hippocampus responses to stress were each significantly associated with greater adverse health symptoms (p<0.01). Furthermore, an inverse correlation was found between activity in the medial OFC and right hippocampus (p=0.01). These results indicate that high CA sensitizes limbic–striatal responses to acute stress and also identifies an important role for stress-related medial OFC and hippocampus responses in the effects of CA on increasing vulnerability to adverse health consequences.     

Longitudinal imaging of metabotropic glutamate 5 receptors during early and extended alcohol abstinence

Chronic alcohol use has important effects on the glutamate system. The metabotropic glutamate 5 (mGlu5) receptor has shown promise in preclinical models as a target to reduce drinking-related behaviors and cue-induced reinstatement, motivating human studies of mGlu5 receptor negative allosteric modulators. The goal of this work was to measure levels of mGlu5 receptor availability with positron emission tomography (PET) imaging using the mGlu5 receptor-specific radiotracer [¹⁸F]FPEB during early and extended alcohol abstinence. Subjects who met DSM-5 criteria for alcohol use disorder (AUD; n = 17) were admitted inpatient for the study duration. [¹⁸F]FPEB PET scans were acquired first during early abstinence (6 ± 4 days after last drink) and a second time during extended abstinence (n = 13; 27 ± 6 days after last drink). A single scan was acquired in healthy controls matched for sex and smoking status (n = 20). [¹⁸F]FPEB total volumes of distribution (V_T) corrected for partial volume effects were measured using equilibrium analysis throughout the brain. A linear mixed model controlling for smoking status and sex identified significantly higher [¹⁸F]FPEB V_T in AUD subjects at early abstinence compared to controls (F_(1,32) = 7.23, p = 0.011). Post-hoc analyses revealed this effect to occur in cortical brain regions. No evidence for significant changes in [¹⁸F]FPEB V_T over time were established. These findings provide human evidence consistent with a robust preclinical literature supporting mGlu5 receptor drugs as pharmacotherapies for AUD.Subject terms: Addiction, Predictive markers     

Markers of Serotonergic Function in the Orbitofrontal Cortex and Dorsal Raphé Nucleus Predict Individual Variation in Spatial-Discrimination Serial Reversal Learning

Dysfunction of the orbitofrontal cortex (OFC) impairs the ability of individuals to flexibly adapt behavior to changing stimulus-reward (S-R) contingencies. Impaired flexibility also results from interventions that alter serotonin (5-HT) and dopamine (DA) transmission in the OFC and dorsomedial striatum (DMS). However, it is unclear whether similar mechanisms underpin naturally occurring variations in behavioral flexibility. In the present study, we used a spatial-discrimination serial reversal procedure to investigate interindividual variability in behavioral flexibility in rats. We show that flexibility on this task is improved following systemic administration of the 5-HT reuptake inhibitor citalopram and by low doses of the DA reuptake inhibitor GBR12909. Rats in the upper quintile of the distribution of perseverative responses during repeated S-R reversals showed significantly reduced levels of the 5-HT metabolite, 5-hydroxy-indoleacetic acid, in the OFC. Additionally, 5-HT_2A receptor binding in the OFC of mid- and high-quintile rats was significantly reduced compared with rats in the low-quintile group. These perturbations were accompanied by an increase in the expression of monoamine oxidase-A (MAO-A) and MAO-B in the lateral OFC and by a decrease in the expression of MAO-A, MAO-B, and tryptophan hydroxylase in the dorsal raphé nucleus of highly perseverative rats. We found no evidence of significant differences in markers of DA and 5-HT function in the DMS or MAO expression in the ventral tegmental area of low- vs high-perseverative rats. These findings indicate that diminished serotonergic tone in the OFC may be an endophenotype that predisposes to behavioral inflexibility and other forms of compulsive behavior.     

Relationship Between Brain Glutamate Levels and Clinical Outcome in Individuals at Ultra High Risk of Psychosis

Alterations in brain glutamate levels may be associated with psychosis risk, but the relationship to clinical outcome in at-risk individuals is unknown. Glutamate concentration was measured in the left thalamus and anterior cingulate cortex (ACC) using 3-Tesla proton magnetic resonance spectroscopy in 75 participants at ultra high risk (UHR) of psychosis and 56 healthy controls. The severity of attenuated positive symptoms and overall functioning were assessed. Measures were repeated in 51 UHR and 33 Control subjects after a mean of 18 months. UHR subjects were allocated to either remission (no longer meeting UHR criteria) or non-remission (meeting UHR or psychosis criteria) status on follow-up assessment. Thalamic glutamate levels at presentation were lower in the UHR non-remission (N=29) compared with the remission group (N=22) (t(49)=3.03; P=0.004), and were associated with an increase in the severity of total positive symptoms over time (r=−0.33; df=47; P=0.02), most notably abnormal thought content (r=−0.442; df=47; P=0.003). In the UHR group, ACC glutamate levels were lower at follow-up compared with baseline (F(80)=4.28; P=0.04). These findings suggest that measures of brain glutamate function may be useful as predictors of clinical outcome in individuals at high risk of psychosis.     

Mice Lacking the Serotonin Htr2B Receptor Gene Present an Antipsychotic-Sensitive Schizophrenic-Like Phenotype

Impulsivity and hyperactivity share common ground with numerous mental disorders, including schizophrenia. Recently, a population-specific serotonin 2B (5-HT_2B) receptor stop codon (ie, HTR2B Q20*) was reported to segregate with severely impulsive individuals, whereas 5-HT_2B mutant (Htr_2B^−/−) mice also showed high impulsivity. Interestingly, in the same cohort, early-onset schizophrenia was more prevalent in HTR2B Q*20 carriers. However, the putative role of 5-HT_2B receptor in the neurobiology of schizophrenia has never been investigated. We assessed the effects of the genetic and the pharmacological ablation of 5-HT_2B receptors in mice subjected to a comprehensive series of behavioral test screenings for schizophrenic-like symptoms and investigated relevant dopaminergic and glutamatergic neurochemical alterations in the cortex and the striatum. Domains related to the positive, negative, and cognitive symptom clusters of schizophrenia were affected in Htr_2B^−/− mice, as shown by deficits in sensorimotor gating, in selective attention, in social interactions, and in learning and memory processes. In addition, Htr_2B^−/− mice presented with enhanced locomotor response to the psychostimulants dizocilpine and amphetamine, and with robust alterations in sleep architecture. Moreover, ablation of 5-HT_2B receptors induced a region-selective decrease of dopamine and glutamate concentrations in the dorsal striatum. Importantly, selected schizophrenic-like phenotypes and endophenotypes were rescued by chronic haloperidol treatment. We report herein that 5-HT_2B receptor deficiency confers a wide spectrum of antipsychotic-sensitive schizophrenic-like behavioral and psychopharmacological phenotypes in mice and provide first evidence for a role of 5-HT_2B receptors in the neurobiology of psychotic disorders.     

Growth hormone responses to GABAB receptor challenge with baclofen and impulsivity in healthy control and personality disorder subjects

Background  

The role of abnormal GABAergic neural transmission in impulsive aggression is not well understood. We have previously shown that central levels of GABA are positively correlated with impulsivity in adult humans with and without personality disorder. An important regulator of GABAergic function is the GABA_B receptor, a presynaptic autoreceptor and heteroreceptor. GABA_B receptor sensitivity may be tested by measuring the growth hormone response to the receptor-agonist baclofen. The purpose of this investigation is to test the hypothesis that dimensional measures of impulsivity and impulsive aggression are negatively correlated with growth hormone response.     

Monoamine Oxidase A is Required for Rapid Dendritic Remodeling in Response to Stress

Background:

Acute stress triggers transient alterations in the synaptic release and metabolism of brain monoamine neurotransmitters. These rapid changes are essential to activate neuroplastic processes aimed at the appraisal of the stressor and enactment of commensurate defensive behaviors. Threat evaluation has been recently associated with the dendritic morphology of pyramidal cells in the orbitofrontal cortex (OFC) and basolateral amygdala (BLA); thus, we examined the rapid effects of restraint stress on anxiety-like behavior and dendritic morphology in the BLA and OFC of mice. Furthermore, we tested whether these processes may be affected by deficiency of monoamine oxidase A (MAO-A), the primary enzyme catalyzing monoamine metabolism.

Methods:

Following a short-term (1–4h) restraint schedule, MAO-A knockout (KO) and wild-type (WT) mice were sacrificed, and histological analyses of dendrites in pyramidal neurons of the BLA and OFC of the animals were performed. Anxiety-like behaviors were examined in a separate cohort of animals subjected to the same experimental conditions.

Results:

In WT mice, short-term restraint stress significantly enhanced anxiety-like responses, as well as a time-dependent proliferation of apical (but not basilar) dendrites of the OFC neurons; conversely, a retraction in BLA dendrites was observed. None of these behavioral and morphological changes were observed in MAO-A KO mice.

Conclusions:

These findings suggest that acute stress induces anxiety-like responses by affecting rapid dendritic remodeling in the pyramidal cells of OFC and BLA; furthermore, our data show that MAO-A and monoamine metabolism are required for these phenomena.     

CFH Variants Affect Structural and Functional Brain Changes and Genetic Risk of Alzheimer's Disease

The immune response is highly active in Alzheimer''s disease (AD). Identification of genetic risk contributed by immune genes to AD may provide essential insight for the prognosis, diagnosis, and treatment of this neurodegenerative disease. In this study, we performed a genetic screening for AD-related top immune genes identified in Europeans in a Chinese cohort, followed by a multiple-stage study focusing on Complement Factor H (CFH) gene. Effects of the risk SNPs on AD-related neuroimaging endophenotypes were evaluated through magnetic resonance imaging scan, and the effects on AD cerebrospinal fluid biomarkers (CSF) and CFH expression changes were measured in aged and AD brain tissues and AD cellular models. Our results showed that the AD-associated top immune genes reported in Europeans (CR1, CD33, CLU, and TREML2) have weak effects in Chinese, whereas CFH showed strong effects. In particular, rs1061170 (P_meta=5.0 × 10⁻⁴) and rs800292 (P_meta=1.3 × 10⁻⁵) showed robust associations with AD, which were confirmed in multiple world-wide sample sets (4317 cases and 16 795 controls). Rs1061170 (P=2.5 × 10⁻³) and rs800292 (P=4.7 × 10⁻⁴) risk-allele carriers have an increased entorhinal thickness in their young age and a higher atrophy rate as the disease progresses. Rs800292 risk-allele carriers have higher CSF tau and Aβ levels and severe cognitive decline. CFH expression level, which was affected by the risk-alleles, was increased in AD brains and cellular models. These comprehensive analyses suggested that CFH is an important immune factor in AD and affects multiple pathological changes in early life and during disease progress.     

Relationship between limbic and cortical 5-HT neurotransmission and acquisition and reversal learning in a go/no-go task in rats

Rationale Specific brain structures have been suggested to be involved in impulsive responding assessed by a variety of operant tasks. Central serotonin (5-HT) function has also been widely implicated in impulsivity; however, little research has addressed the regional aspect of 5-HT roles in different impulsive indices of task performance.Objective We analyzed the relationships between acquisition and reversal learning in a go/no-go task as different behavioral measures of impulsivity and focal concentrations of 5-HT and its metabolites in the brain.Materials and methods Rats administered with parachloroamphetamine (PCA) and vehicle were tested in both acquisition and reversal phases in a go/no-go visual discrimination task. Neurochemical analysis was performed to determine 5-HT concentrations in micropunched brain tissues.Results PCA administration induced regionally 5-HT depletion in the brain and impaired learning performance in both tests. For both tests, significant negative correlations between learning performance and 5-HT and 5-hydroxyindoleacetic acid (5-HIAA) concentrations were observed in the medial prefrontal cortex (mPFC) and amygdala (Amyg). In contrast, significant negative correlations between learning performance and 5-HT and 5-HIAA concentrations were observed for the orbitofrontal cortex (OFC) exclusively in the reversal learning phase.Conclusions The present data indicate that 5-HT neurotransmission to the mPFC and Amyg is involved in inhibitory control over responses to discriminated stimuli associated with the go/no-go paradigm common to both tests. In contrast, 5-HT neurotransmission to the OFC is especially involved in additional processes associated with reversal learning.     

Epigenome-wide association study of alcohol use disorder in five brain regions

Alcohol use disorder (AUD) is closely linked to the brain regions forming the neurocircuitry of addiction. Postmortem human brain tissue enables the direct study of the molecular pathomechanisms of AUD. This study aims to identify these mechanisms by examining differential DNA-methylation between cases with severe AUD (n = 53) and controls (n = 58) using a brain-region-specific approach, in which sample sizes ranged between 46 and 94. Samples of the anterior cingulate cortex (ACC), Brodmann Area 9 (BA9), caudate nucleus (CN), ventral striatum (VS), and putamen (PUT) were investigated. DNA-methylation levels were determined using the Illumina HumanMethylationEPIC Beadchip. Epigenome-wide association analyses were carried out to identify differentially methylated CpG-sites and regions between cases and controls in each brain region. Weighted correlation network analysis (WGCNA), gene-set, and GWAS-enrichment analyses were performed. Two differentially methylated CpG-sites were associated with AUD in the CN, and 18 in VS (q < 0.05). No epigenome-wide significant CpG-sites were found in BA9, ACC, or PUT. Differentially methylated regions associated with AUD case-/control status (q < 0.05) were found in the CN (n = 6), VS (n = 18), and ACC (n = 1). In the VS, the WGCNA-module showing the strongest association with AUD was enriched for immune-related pathways. This study is the first to analyze methylation differences between AUD cases and controls in multiple brain regions and consists of the largest sample to date. Several novel CpG-sites and regions implicated in AUD were identified, providing a first basis to explore epigenetic correlates of AUD.Subject terms: Epigenetics and behaviour, DNA methylation, Addiction     

Association of Gamma-Aminobutyric Acid A Receptor α2 Gene (GABRA2) with Alcohol Use Disorder

Gamma-aminobutyric acid (GABA) is a major inhibitory neurotransmitter in mammalian brain. GABA receptor are involved in a number of complex disorders, including substance abuse. No variants of the commonly studied GABA receptor genes that have been associated with substance dependence have been determined to be functional or pathogenic. To reconcile the conflicting associations with substance dependence traits, we performed a meta-analysis of variants in the GABA_A receptor genes (GABRB2, GABRA6, GABRA1, and GABRG2 on chromosome 5q and GABRA2 on chromosome 4p12) using genotype data from 4739 cases of alcohol, opioid, or methamphetamine dependence and 4924 controls. Then, we combined the data from candidate gene association studies in the literature with two alcohol dependence (AD) samples, including 1691 cases and 1712 controls from the Study of Addiction: Genetics and Environment (SAGE), and 2644 cases and 494 controls from our own study. Using a Bonferroni-corrected threshold of 0.007, we found strong associations between GABRA2 and AD (P=9 × 10⁻⁶ and odds ratio (OR) 95% confidence interval (CI)=1.27 (1.15, 1.4) for rs567926, P=4 × 10⁻⁵ and OR=1.21 (1.1, 1.32) for rs279858), and between GABRG2 and both dependence on alcohol and dependence on heroin (P=0.0005 and OR=1.22 (1.09, 1.37) for rs211014). Significant association was also observed between GABRA6 rs3219151 and AD. The GABRA2 rs279858 association was observed in the SAGE data sets with a combined P of 9 × 10⁻⁶ (OR=1.17 (1.09, 1.26)). When all of these data sets, including our samples, were meta-analyzed, associations of both GABRA2 single-nucleotide polymorphisms remained (for rs567926, P=7 × 10⁻⁵ (OR=1.18 (1.09, 1.29)) in all the studies, and P=8 × 10⁻⁶ (OR=1.25 (1.13, 1.38)) in subjects of European ancestry and for rs279858, P=5 × 10⁻⁶ (OR=1.18 (1.1, 1.26)) in subjects of European ancestry. Findings from this extensive meta-analysis of five GABA_A receptor genes and substance abuse support their involvement (with the best evidence for GABRA2) in the pathogenesis of AD. Further replications with larger samples are warranted.