Major Psychiatric Disorders, Substance Use Behaviors, and Longevity.

Importance: Observational studies suggest that major psychiatric disorders and substance use behaviors reduce longevity, making it difficult to disentangle their relationships with aging-related outcomes. Objective: To evaluate the associations between the genetic liabilities for major psychiatric disorders, substance use behaviors (smoking and alcohol consumption), and longevity. Design, Settings, and Participants: This 2-sample mendelian randomization (MR) study assessed associations between psychiatric disorders, substance use behaviors, and longevity using single-variable and multivariable models. Multiomics analyses were performed elucidating transcriptomic underpinnings of the MR associations and identifying potential proteomic therapeutic targets. This study sourced summary-level genome-wide association study (GWAS) data, gene expression, and proteomic data from cohorts of European ancestry. Analyses were performed from May 2022 to November 2023. Exposures: Genetic susceptibility for major depression (n = 500 199), bipolar disorder (n = 413 466), schizophrenia (n = 127 906), problematic alcohol use (n = 435 563), weekly alcohol consumption (n = 666 978), and lifetime smoking index (n = 462 690). Main Outcomes and Measures: The main outcome encompassed aspects of health span, lifespan, and exceptional longevity. Additional outcomes were epigenetic age acceleration (EAA) clocks. Results: Findings from multivariable MR models simultaneously assessing psychiatric disorders and substance use behaviorsm suggest a negative association between smoking and longevity in cohorts of European ancestry (n = 709 709; 431 503 [60.8%] female; ß, -0.33; 95% CI, -0.38 to -0.28; P = 4.59 × 10-34) and with increased EAA (n = 34 449; 18 017 [52.3%] female; eg, PhenoAge: ß, 1.76; 95% CI, 0.72 to 2.79; P = 8.83 × 10-4). Transcriptomic imputation and colocalization identified 249 genes associated with smoking, including 36 novel genes not captured by the original smoking GWAS. Enriched pathways included chromatin remodeling and telomere assembly and maintenance. The transcriptome-wide signature of smoking was inversely associated with longevity, and estimates of individual smoking-associated genes, eg, XRCC3 and PRMT6, aligned with the smoking-longevity MR analyses, suggesting underlying transcriptomic mediators. Cis-instrument MR prioritized brain proteins associated with smoking behavior, including LY6H (ß, 0.02; 95% CI, 0.01 to 0.03; P = 2.37 × 10-6) and RIT2 (ß, 0.02; 95% CI, 0.01 to 0.03; P = 1.05 × 10-5), which had favorable adverse-effect profiles across 367 traits evaluated in phenome-wide MR. Conclusions: The findings suggest that the genetic liability of smoking, but not of psychiatric disorders, is associated with longevity. Transcriptomic associations offer insights into smoking-related pathways, and identified proteomic targets may inform therapeutic development for smoking cessation strategies.

Alcohol use disorder is associated with DNA methylation-based shortening of telomere length and regulated by TESPA1: implications for aging.

Chronic heavy alcohol consumption is associated with increased mortality and morbidity and often leads to premature aging; however, the mechanisms of alcohol-associated cellular aging are not well understood. In this study, we used DNA methylation derived telomere length (DNAmTL) as a novel approach to investigate the role of alcohol use on the aging process. DNAmTL was estimated by 140 cytosine phosphate guanines (CpG) sites in 372 individuals with alcohol use disorder (AUD) and 243 healthy controls (HC) and assessed using various endophenotypes and clinical biomarkers. Validation in an independent sample of DNAmTL on alcohol consumption was performed (N = 4219). Exploratory genome-wide association studies (GWAS) on DNAmTL were also performed to identify genetic variants contributing to DNAmTL shortening. Top GWAS findings were analyzed using in-silico expression quantitative trait loci analyses and related to structural MRI hippocampus volumes of individuals with AUD. DNAmTL was 0.11-kilobases shorter per year in AUD compared to HC after adjustment for age, sex, race, and blood cell composition (p = 4.0 × 10-12). This association was partially attenuated but remained significant after additionally adjusting for BMI, and smoking status (0.06 kilobases shorter per year, p = 0.002). DNAmTL shortening was strongly associated with chronic heavy alcohol use (ps < 0.001), elevated gamma-glutamyl transferase (GGT), and aspartate aminotransferase (AST) (ps < 0.004). Comparison of DNAmTL with PCR-based methods of assessing TL revealed positive correlations (R = 0.3, p = 2.2 × 10-5), highlighting the accuracy of DNAmTL as a biomarker. The GWAS meta-analysis identified a single nucleotide polymorphism (SNP), rs4374022 and 18 imputed ones in Thymocyte Expressed, Positive Selection Associated 1(TESPA1), at the genome-wide level (p = 3.75 × 10-8). The allele C of rs4374022 was associated with DNAmTL shortening, lower hippocampus volume (p < 0.01), and decreased mRNA expression in hippocampus tissue (p = 0.04). Our study demonstrates DNAmTL-related aging acceleration in AUD and suggests a functional role for TESPA1 in regulating DNAmTL length, possibly via the immune system with subsequent biological effects on brain regions negatively affected by alcohol and implicated in aging.

Epigenome-wide association study of alcohol consumption in N = 8161 individuals and relevance to alcohol use disorder pathophysiology: identification of the cystine/glutamate transporter SLC7A11 as a top target.

Alcohol misuse is common in many societies worldwide and is associated with extensive morbidity and mortality, often leading to alcohol use disorders (AUD) and alcohol-related end-organ damage. The underlying mechanisms contributing to the development of AUD are largely unknown; however, growing evidence suggests that alcohol consumption is strongly associated with alterations in DNA methylation. Identification of alcohol-associated methylomic variation might provide novel insights into pathophysiology and novel treatment targets for AUD. Here we performed the largest single-cohort epigenome-wide association study (EWAS) of alcohol consumption to date (N = 8161) and cross-validated findings in AUD populations with relevant endophenotypes, as well as alcohol-related animal models. Results showed 2504 CpGs significantly associated with alcohol consumption (Bonferroni p value < 6.8 × 10-8) with the five leading probes located in SLC7A11 (p = 7.75 × 10-108), JDP2 (p = 1.44 × 10-56), GAS5 (p = 2.71 × 10-47), TRA2B (p = 3.54 × 10-42), and SLC43A1 (p = 1.18 × 10-40). Genes annotated to associated CpG sites are implicated in liver and brain function, the cellular response to alcohol and alcohol-associated diseases, including hypertension and Alzheimer's disease. Two-sample Mendelian randomization confirmed the causal relationship of consumption on AUD risk (inverse variance weighted (IVW) p = 5.37 × 10-09). A methylation-based predictor of alcohol consumption was able to discriminate AUD cases in two independent cohorts (p = 6.32 × 10-38 and p = 5.41 × 10-14). The top EWAS probe cg06690548, located in the cystine/glutamate transporter SLC7A11, was replicated in an independent cohort of AUD and control participants (N = 615) and showed strong hypomethylation in AUD (p < 10-17). Decreased CpG methylation at this probe was consistently associated with clinical measures including increased heavy drinking days (p < 10-4), increased liver function enzymes (GGT (p = 1.03 × 10-21), ALT (p = 1.29 × 10-6), and AST (p = 1.97 × 10-8)) in individuals with AUD. Postmortem brain analyses documented increased SLC7A11 expression in the frontal cortex of individuals with AUD and animal models showed marked increased expression in liver, suggesting a mechanism by which alcohol leads to hypomethylation-induced overexpression of SLC7A11. Taken together, our EWAS discovery sample and subsequent validation of the top probe in AUD suggest a strong role of abnormal glutamate signaling mediated by methylomic variation in SLC7A11. Our data are intriguing given the prominent role of glutamate signaling in brain and liver and might provide an important target for therapeutic intervention.

Smoking is significantly associated with increased risk of COVID-19 and other respiratory infections.

Observational studies suggest smoking, cannabis use, alcohol consumption, and substance use disorders (SUDs) may impact risk for respiratory infections, including coronavirus 2019 (COVID-2019). However, causal inference is challenging due to comorbid substance use. Using summary-level European ancestry data (>1.7 million participants), we performed single-variable and multivariable Mendelian randomization (MR) to evaluate relationships between substance use behaviors, COVID-19 and other respiratory infections. Genetic liability for smoking demonstrated the strongest associations with COVID-19 infection risk, including the risk for very severe respiratory confirmed COVID-19 (odds ratio (OR) = 2.69, 95% CI, 1.42, 5.10, P-value = 0.002), and COVID-19 infections requiring hospitalization (OR = 3.49, 95% CI, 2.23, 5.44, P-value = 3.74 × 10-8); these associations generally remained robust in models accounting for other substance use and cardiometabolic risk factors. Smoking was also strongly associated with increased risk of other respiratory infections, including asthma-related pneumonia/sepsis (OR = 3.64, 95% CI, 2.16, 6.11, P-value = 1.07 × 10-6), chronic lower respiratory diseases (OR = 2.29, 95% CI, 1.80, 2.91, P-value = 1.69 × 10-11), and bacterial pneumonia (OR = 2.14, 95% CI, 1.42, 3.24, P-value = 2.84 × 10-4). We provide strong genetic evidence showing smoking increases the risk for COVID-19 and other respiratory infections even after accounting for other substance use behaviors and cardiometabolic diseases, which suggests that prevention programs aimed at reducing smoking may be important for the COVID-19 pandemic and have substantial public health benefits.

A genetically-informed study disentangling the relationships between tobacco smoking, cannabis use, alcohol consumption, substance use disorders and respiratory infections, including COVID-19.

Background: Observational studies suggest smoking, cannabis use, alcohol consumption, cannabis use, and substance use disorders (SUDs) may play a role in the susceptibility for respiratory infections and disease, including coronavirus 2019 (COVID-2019). However, causal inference is challenging due to comorbid substance use. Methods: Using genome-wide association study data of European ancestry (data from >1.7 million individuals), we performed single-variable and multivariable Mendelian randomization to evaluate relationships between smoking, cannabis use, alcohol consumption, SUDs, and respiratory infections. Results: Genetically predicted lifetime smoking was found to be associated with increased risk for hospitalized COVID-19 (odds ratio (OR)=4.039, 95% CI 2.335-6.985, P-value=5.93×10-7) and very severe hospitalized COVID-19 (OR=3.091, 95% CI, 1.883-5.092, P-value=8.40×10-6). Genetically predicted lifetime smoking was also associated with increased risk pneumoniae (OR=1.589, 95% CI, 1.214-2.078, P-value=7.33×10-4), lower respiratory infections (OR=2.303, 95% CI, 1.713-3.097, P-value=3.40×10-8), and several others. Genetically predicted cannabis use disorder (CUD) was associated with increased bronchitis risk (OR=1.078, 95% CI, 1.020-1.128, P-value=0.007). Conclusions: We provide strong genetic evidence showing smoking increases the risk for respiratory infections and diseases even after accounting for other substance use and abuse. Additionally, we provide find CUD may increase the risk for bronchitis, which taken together, may guide future research SUDs and respiratory outcomes.

Epigenome-wide association study and multi-tissue replication of individuals with alcohol use disorder: evidence for abnormal glucocorticoid signaling pathway gene regulation.

Alcohol use disorder (AUD) is a chronic debilitating disorder with limited treatment options and poorly defined pathophysiology. There are substantial genetic and epigenetic components; however, the underlying mechanisms contributing to AUD remain largely unknown. We conducted the largest DNA methylation epigenome-wide association study (EWAS) analyses currently available for AUD (total N = 625) and employed a top hit replication (N = 4798) using a cross-tissue/cross-phenotypic approach with the goal of identifying novel epigenetic targets relevant to AUD. Results show that a network of differentially methylated regions in glucocorticoid signaling and inflammation-related genes were associated with alcohol use behaviors. A top probe consistently associated across all cohorts was located in the long non-coding RNA growth arrest specific five gene (GAS5) (p < 10-24). GAS5 has been implicated in regulating transcriptional activity of the glucocorticoid receptor and has multiple functions related to apoptosis, immune function and various cancers. Endophenotypic analyses using peripheral cortisol levels and neuroimaging paradigms showed that methylomic variation in GAS5 network-related probes were associated with stress phenotypes. Postmortem brain analyses documented increased GAS5 expression in the amygdala of individuals with AUD. Our data suggest that alcohol use is associated with differential methylation in the glucocorticoid system that might influence stress and inflammatory reactivity and subsequently risk for AUD.

Evaluating the relationship between alcohol consumption, tobacco use, and cardiovascular disease: A multivariable Mendelian randomization study.

BACKGROUND: Alcohol consumption and smoking, 2 major risk factors for cardiovascular disease (CVD), often occur together. The objective of this study is to use a wide range of CVD risk factors and outcomes to evaluate potential total and direct causal roles of alcohol and tobacco use on CVD risk factors and events. METHODS AND FINDINGS: Using large publicly available genome-wide association studies (GWASs) (results from more than 1.2 million combined study participants) of predominantly European ancestry, we conducted 2-sample single-variable Mendelian randomization (SVMR) and multivariable Mendelian randomization (MVMR) to simultaneously assess the independent impact of alcohol consumption and smoking on a wide range of CVD risk factors and outcomes. Multiple sensitivity analyses, including complementary Mendelian randomization (MR) methods, and secondary alcohol consumption and smoking datasets were used. SVMR showed genetic predisposition for alcohol consumption to be associated with CVD risk factors, including high-density lipoprotein cholesterol (HDL-C) (beta 0.40, 95% confidence interval (CI), 0.04-0.47, P value = 1.72 × 10-28), triglycerides (TRG) (beta -0.23, 95% CI, -0.30, -0.15, P value = 4.69 × 10-10), automated systolic blood pressure (BP) measurement (beta 0.11, 95% CI, 0.03-0.18, P value = 4.72 × 10-3), and automated diastolic BP measurement (beta 0.09, 95% CI, 0.03-0.16, P value = 5.24 × 10-3). Conversely, genetically predicted smoking was associated with increased TRG (beta 0.097, 95% CI, 0.014-0.027, P value = 6.59 × 10-12). Alcohol consumption was also associated with increased myocardial infarction (MI) and coronary heart disease (CHD) risks (MI odds ratio (OR) = 1.24, 95% CI, 1.03-1.50, P value = 0.02; CHD OR = 1.21, 95% CI, 1.01-1.45, P value = 0.04); however, its impact was attenuated in MVMR adjusting for smoking. Conversely, alcohol maintained an association with coronary atherosclerosis (OR 1.02, 95% CI, 1.01-1.03, P value = 5.56 × 10-4). In comparison, after adjusting for alcohol consumption, smoking retained its association with several CVD outcomes including MI (OR = 1.84, 95% CI, 1.43, 2.37, P value = 2.0 × 10-6), CHD (OR = 1.64, 95% CI, 1.28-2.09, P value = 8.07 × 10-5), heart failure (HF) (OR = 1.61, 95% CI, 1.32-1.95, P value = 1.9 × 10-6), and large artery atherosclerosis (OR = 2.4, 95% CI, 1.41-4.07, P value = 0.003). Notably, using the FinnGen cohort data, we were able to replicate the association between smoking and several CVD outcomes including MI (OR = 1.77, 95% CI, 1.10-2.84, P value = 0.02), HF (OR = 1.67, 95% CI, 1.14-2.46, P value = 0.008), and peripheral artery disease (PAD) (OR = 2.35, 95% CI, 1.38-4.01, P value = 0.002). The main limitations of this study include possible bias from unmeasured confounders, inability of summary-level MR to investigate a potentially nonlinear relationship between alcohol consumption and CVD risk, and the generalizability of the UK Biobank (UKB) to other populations. CONCLUSIONS: Evaluating the widest range of CVD risk factors and outcomes of any alcohol consumption or smoking MR study to date, we failed to find a cardioprotective impact of genetically predicted alcohol consumption on CVD outcomes. However, alcohol was associated with and increased HDL-C, decreased TRG, and increased BP, which may indicate pathways through impact CVD risk, warranting further study. We found smoking to be a risk factor for many CVDs even after adjusting for alcohol. While future studies incorporating alcohol consumption patterns are necessary, our data suggest causal inference between alcohol, smoking, and CVD risk, further supporting that lifestyle modifications might be able to reduce overall CVD risk.

Educational attainment reduces the risk of suicide attempt among individuals with and without psychiatric disorders independent of cognition: a bidirectional and multivariable Mendelian randomization study with more than 815,000 participants.

Rates of suicidal behavior are increasing in the United States and identifying causal risk factors continues to be a public health priority. Observational literature has shown that educational attainment (EA) and cognitive performance (CP) influence suicide attempt risk; however, the causal nature of these relationships is unknown. Using summary statistics from genome-wide association studies (GWAS) of EA, CP, and suicide attempt risk with > 815,000 combined white participants of European ancestry, we performed multivariable Mendelian randomization (MR) to disentangle the effects of EA and CP on attempted suicide. In single-variable MR (SVMR), EA and CP appeared to reduce suicide attempt risk (EA odds ratio (OR) per standard deviation (SD) increase in EA (4.2 years), 0.524, 95% CI, 0.412-0.666, P = 1.07 × 10-7; CP OR per SD increase in standardized score, 0.714, 95% CI, 0.577-0.885, P = 0.002). Conversely, bidirectional analyses found no effect of a suicide attempt on EA or CP. Using various multivariable MR (MVMR) models, EA seems to be the predominant risk factor for suicide attempt risk with the independent effect (OR, 0.342, 95% CI, 0.206-0.568, P = 1.61 × 10-4), while CP had no effect (OR, 1.182, 95% CI, 0.842-1.659, P = 0.333). In additional MVMR analyses accounting simultaneously for potential behavioral and psychiatric mediators (tobacco smoking; alcohol consumption; and self-reported nerves, tension, anxiety, or depression), the effect of EA was little changed (OR, 0.541, 95% CI, 0.421-0.696, P = 3.33 × 10-6). Consistency of results across complementary MR methods accommodating different assumptions about genetic pleiotropy strengthened causal inference. Our results show that even after accounting for psychiatric disorders and behavioral mediators, EA, but not CP, may causally influence suicide attempt risk among white individuals of European ancestry, which could have important implications for health policy and programs aimed at reducing the increasing rates of suicide. Future work is necessary to examine the EA-suicide relationship populations of different ethnicities.

Adverse Childhood Experiences are Associated with High-Intensity Binge Drinking Behavior in Adulthood and Mediated by Psychiatric Disorders.

AIM: High-intensity binge drinking (HIBD), defined as two or more times the gender-specific binge threshold, is rapidly increasing in the USA; however, the underlying contributing factors are poorly understood. This study investigated the relationship of adverse childhood experiences (ACEs) and HIBD. METHODS: Two independent, cross-sectional samples were analysed: (a) past 12-month drinkers in the National Epidemiological Survey on Alcohol and Related Conditions-III (NESARC-III; n = 25,552) and (b) the National Institute on Alcohol Abuse and Alcoholism (NIAAA) clinical sample (n = 1303). Multinomial logistic regressions were utilized to estimate adjusted odds ratios (AORs) of ACEs on HIBD. Mediation analysis was performed to examine the relationship between the past 12-month psychiatric disorders, ACEs, and HIBD. RESULTS: In the NESARC-III sample, prevalence of ACEs increased across all binge levels with the highest prevalence in extreme HIBD; ACEs were associated with higher odds for HIBD (level II, odds ratio (OR) = 1.2-1.4; P = 0.03-0.001; level III, OR = 1.3-1.9; P < 0.001). Prevalence of DSM-5 diagnoses also increased across all binge levels. Substance use disorders (SUD), mood, personality and post-traumatic stress disorders (PTSD) conferred the highest odds with extreme HIBD (SUD: OR = 21.32; mood: 1.73; personality: 2.84; PTSD: 1.97; all Ps < 0.001). Mediation analyses showed that the association between ACEs and HIBD was fully mediated through SUD (proportion mediated: 70-90%) and partially through other psychiatric disorders (20-80%). In the NIAAA sample, ACEs were 2-5 times more prevalent in extreme HIBD with higher odds (ORs = 3-8, P < 0.001) compared with non-bingers. CONCLUSION: ACEs were associated with significantly increased odds of HIBD and the relationship may be mediated by psychiatric disorders.

Epigenetic aging is accelerated in alcohol use disorder and regulated by genetic variation in APOL2.

To investigate the potential role of alcohol use disorder (AUD) in aging processes, we employed Levine's epigenetic clock (DNAm PhenoAge) to estimate DNA methylation age in 331 individuals with AUD and 201 healthy controls (HC). We evaluated the effects of heavy, chronic alcohol consumption on epigenetic age acceleration (EAA) using clinical biomarkers, including liver function test enzymes (LFTs) and clinical measures. To characterize potential underlying genetic variation contributing to EAA in AUD, we performed genome-wide association studies (GWAS) on EAA, including pathway analyses. We followed up on relevant top findings with in silico expression quantitative trait loci (eQTL) analyses for biological function using the BRAINEAC database. There was a 2.22-year age acceleration in AUD compared to controls after adjusting for gender and blood cell composition (p = 1.85 × 10-5). This association remained significant after adjusting for race, body mass index, and smoking status (1.38 years, p = 0.02). Secondary analyses showed more pronounced EAA in individuals with more severe AUD-associated phenotypes, including elevated gamma-glutamyl transferase (GGT) and alanine aminotransferase (ALT), and higher number of heavy drinking days (all ps < 0.05). The genome-wide meta-analysis of EAA in AUD revealed a significant single nucleotide polymorphism (SNP), rs916264 (p = 5.43 × 10-8), in apolipoprotein L2 (APOL2) at the genome-wide level. The minor allele A of rs916264 was associated with EAA and with increased mRNA expression in hippocampus (p = 0.0015). Our data demonstrate EAA in AUD and suggest that disease severity further accelerates epigenetic aging. EAA was associated with genetic variation in APOL2, suggesting potential novel biological mechanisms for age acceleration in AUD.

Imaging resilience and recovery in alcohol dependence.

BACKGROUND AND AIMS: Resilience and recovery are of increasing importance in the field of alcohol dependence (AD). This paper describes how imaging studies in man can be used to assess the neurobiological correlates of resilience and, if longitudinal, of disease trajectories, progression rates and markers for recovery to inform treatment and prevention options. METHODS: Original papers on recovery and resilience in alcohol addiction and its neurobiological correlates were identified from PubMed and have been analyzed and condensed within a systematic literature review. RESULTS: Findings deriving from functional magnetic resonance imaging (fMRI) and positron emission tomography (PET) studies have identified links between increased resilience and less task-elicited neural activation within the basal ganglia, and benefits of heightened neural pre-frontal cortex (PFC) engagement regarding resilience in a broader sense; namely, resilience against relapse in early abstinence of AD. Furthermore, findings consistently propose at least partial recovery of brain glucose metabolism and executive and general cognitive functioning, as well as structural plasticity effects throughout the brain of alcohol-dependent patients during the course of short-, medium- and long-term abstinence, even when patients only lowered their alcohol consumption to a moderate level. Additionally, specific factors were found that appear to influence these observed brain recovery processes in AD, e.g. genotype-dependent neuronal (re)growth, gender-specific neural recovery effects, critical interfering effects of psychiatric comorbidities, additional smoking or marijuana influences or adolescent alcohol abuse. CONCLUSIONS: Neuroimaging research has uncovered neurobiological markers that appear to be linked to resilience and improved recovery capacities that are furthermore influenced by various factors such as gender or genetics. Consequently, future system-oriented approaches may help to establish a broad neuroscience-based research framework for alcohol dependence.

Human neutrophils can mimic myeloid-derived suppressor cells (PMN-MDSC) and suppress microbead or lectin-induced T cell proliferation through artefactual mechanisms.

We report that human conventional CD15+ neutrophils can be isolated in the peripheral blood mononuclear cell (PBMC) layer during Ficoll gradient separation, and that they can impair T cell proliferation in vitro without concomitant neutrophil activation and killing. This effect was observed in a total of 92 patients with organ transplants, lung cancer or anxiety/depression, and in 18 healthy donors. Although such features are typically associated in the literature with the presence of certain myeloid-derived suppressor cell (PMN-MDSC) populations, we found that commercial centrifuge tubes that contained membranes or gels for PBMC isolation led to up to 70% PBMC contamination by CD15+ neutrophils, with subsequent suppressive effects in certain cellular assays. In particular, the suppressive activity of human MDSC should not be evaluated using lectin or microbead stimulation, whereas assays involving soluble or plate-bound antibodies or MLR are unaffected. We conclude that CD15+ neutrophil contamination, and associated effects on suppressor assays, can lead to significant artefacts in studies of human PMN-MDSC.

Dopamine Transporter Gene Methylation is Associated with Nucleus Accumbens Activation During Reward Processing in Healthy but not Alcohol-Dependent Individuals.

BACKGROUND: Alcohol's reinforcement is mediated by dopamine signaling in the ventral striatum, which is modulated by the dopamine transporter (DAT). We hypothesized that methylomic variation in the DAT gene (DAT1/SLC6A3) affects DAT expression, thus contributing to differences in brain reward circuitry in individuals with alcohol dependence (ALC). METHODS: Blood from 45 recently detoxified ALC and 45 healthy control (HC) individuals was used to assess DNA methylation across 5 functional regions of SLC6A3. Participants completed the monetary incentive delay task in a 3-Tesla magnetic resonance imaging (MRI) scanner. Employing regression models, we examined effects of SLC6A3 methylation on nucleus accumbens (NAc) blood-oxygen-level dependent (BOLD) responses during anticipation of high/low reward/loss. RESULTS: Results showed that decreased methylation of the promoter region of SLC6A3 predicted NAc activation during high loss anticipation (p = 0.028) and low loss anticipation (at trend-level; p = 0.057) in HC but not in individuals with ALC. Specifically, percentage of methylation at 2 CpG sites, located -1,001 and -993 base pairs from the transcription start site, accounted for significant variability in NAc activation in the HC group during high (ps ≤ 0.010) and low (ps ≤ 0.006) loss anticipation. There was no effect on reward anticipation. Furthermore, promoter methylation was positively associated with age, which replicates previous findings. CONCLUSIONS: Our data suggest that methylation in the promoter region of SLC6A3 predicts NAc activation during the anticipation of monetary loss in HCs. However, this effect was not present in the ALC group, suggesting that epigenetic regulation of striatal DAT expression might be disrupted in ALC, which may contribute to previously reported differences in sensitivity to reward and punishment in this population. Alternatively, it is possible that a similar relationship in the ALC group remained undetected possibly due to methodological limitations inherent in functional MRI (e.g., poor spatial resolution, low signal-to-noise ratio) that generally restrict interpretations regarding mechanisms of epigenetic factors involved in group differences in BOLD responses. Future neuroimaging studies are needed to further elucidate the relationship between SLC6A3 methylation and NAc activation in ALC.

Identification of CHRNA5 rare variants in African-American heavy smokers.

BACKGROUND: The common CHRNA5 mis-sense coding single-nucleotide polymorphism (SNP) rs16969968:G>A (D398N) has repeatedly been shown to confer risk for heavy smoking in individuals who carry the 'A' allele (encoding the 398N amino acid). The mis-sense SNP has a minor allele frequency of ∼40% in European-Americans, but only ∼7% in African-Americans (http://www.ncbi.nlm.nih.gov/projects/SNP/). We reasoned that there might be other mis-sense variants among African-Americans that could confer the heavy smoking phenotype (defined here as ≥20 cigarettes per day), perhaps in a manner similar to that of the D398N polymorphism in Europeans. MATERIALS AND METHODS: As such, we resequenced 250 African-American heavy smokers, most of whom were homozygous 'G' at rs16969968:G>A (minor allele frequency of 9.6% within the population). RESULTS: Although many novel coding SNPs were not observed, we report an interesting, although rare (perhaps personal), variant in CHRNA5 that could result in nonsense-mediated decay of the aberrant transcript. CONCLUSION: We conclude that, in African-Americans, variants (common or rare) in genes other than CHRNA5 most likely contribute toward the nicotine-dependent phenotype, either independently or in combination with variants in CHRNA5. The functional significance, on CHRNA5 expression or protein function, of the variants found here should be determined in future studies.

Neural correlates of attentional bias for smoking cues: modulation by variance in the dopamine transporter gene.

Cigarette-dependent smokers automatically and involuntarily orient attention toward smoking cues (SCs). This attentional bias is clinically significant, as it may contribute to relapse. Thus, identifying neural and genetic correlates of attentional bias is critical for improving interventions. Our previous studies show that the dopamine transporter (DAT) SLC6A3 genotype exerts profound effects on limbic responses to SCs. One potential mechanism underlying these effects is greater attentional bias for SCs. Here, we explored associations between attentional bias for SCs and neural responses to SCs among 'sated' smokers genotyped for the SLC6A3 polymorphism. Pseudo-continuous arterial spin-labeled perfusion functional magnetic resonance imaging images were acquired during SC exposure in 35 smokers genotyped for the SLC6A3 variable number of tandem repeats polymorphism (n = 16, 9-repeats; n = 19, 10/10-repeats). Participants completed a visual dot-probe attentional bias task, which contained pictures of smoking and non-smoking pictures, to examine whether genetic variation in DAT influences attentional bias and to investigate relationships between attentional bias and neural responses to SCs. Although attentional bias to smoking pictures was not significantly different between 9-repeats and 10/10-repeats, 9-repeats showed a positive correlation between attentional bias and increased SC-induced brain activity in the amygdala, whereas 10/10-repeats showed an inverse correlation in the medial orbitofrontal cortex (mOFC). In group comparisons, 9-repeats exhibited positive correlations between attentional bias and SCs in the mOFC and amygdala, relative to 10/10-repeats. Findings suggest that genetic variation in the DAT gene influences brain responses associated with attentional bias; thus, providing additional support for a SC-vulnerable endophenotype.

Genetic polymorphisms in the PACAP and PAC1 receptor genes and treatment response to venlafaxine XR in generalized anxiety disorder.

The pituitary adenylate cyclase-activating peptide (PACAP) and its receptor (PAC1) are involved in stress response and anxiety. Genotypes for PACAP/PAC1 were examined for effects on treatment response to venlafaxine XR in generalized anxiety disorder. The Asp54Gly (rs2856966) variant in the PACAP gene was associated with better treatment outcome.

Low frequency genetic variants in the µ-opioid receptor (OPRM1) affect risk for addiction to heroin and cocaine.

The µ-opioid receptor (MOR) binds exogenous and endogenous opioids and is known to mediate the rewarding effects of drugs of abuse. Numerous genetic studies have sought to identify common genetic variation in the gene encoding MOR (OPRM1) that affects risk for drug addiction. The purpose of this study was to examine the contribution of rare coding variants in OPRM1 to the risk for addiction. Rare and low frequency variants were selected using the National Heart Lung and Blood Institute - Exome Sequencing Project (NHLBI-ESP) database, which has screened the exomes of over 6500 individuals. Two SNPs (rs62638690 and rs17174794) were selected for genotyping in 1377 European American individuals addicted to heroin and/or cocaine. Two different SNPs (rs1799971 and rs17174801) were genotyped in 1238 African American individuals addicted to heroin and/or cocaine. Using the minor allele frequencies from the NHLBI-ESP dataset as a comparison group, case-control association analyses were performed. Results revealed an association between rs62638690 and cocaine and heroin addiction in European Americans (p=0.02; 95% C.I. 0.47 [0.24-0.92]). This study suggests a potential role for rare OPRM1 variants in addiction disorders and highlights an area worthy of future study.

Dopamine transporter genotype modulation of neural responses to smoking cues: confirmation in a new cohort.

Previously we demonstrated profound effects of dopamine transporter (DAT) SLC6A3 genotype on limbic responses to smoking cues (SCs). Probands carrying at least one copy of the 9-repeat allele (9-repeat carriers) had greater neural responses to SCs in the anatomically interconnected rostral ventral striatum/medial orbitofrontal cortex (VS/mOFC), compared with homozygotes for the 10-repeat allele (10/10-repeats). To test the reliability of the initial findings, we examined perfusion functional magnetic resonance images acquired during SC exposure in a new cohort of smokers (N=26) who were genotyped for the SLC6A3 polymorphism. In smokers overall, activity was enhanced in the VS/mOFC (t=3.77). Contrasts between allelic groups revealed that 9-repeat carriers had a greater response to SCs in the VS (t=3.12) and mOFC (t=3.19). In separate groups, 9-repeat carriers showed increased activity in the VS (t=5.47) and mOFC (T=4.96), while no increases were observed in 10-repeats. Subjective reports of craving correlated with increased activity in reward-related structures including the extended amygdala, insula and post-central gyrus, and decreased activity in the dorsolateral prefrontal cortex, and were DAT-genotype dependent (r=0.63-0.96). In secondary analyses, we found that The Fagerström Test for Nicotine Dependence scores correlated with enhanced SC-induced perfusion in 10/10-repeats in the insula, mOFC, medial temporal and superior frontal gyri (r=0.50-0.82), while correlations were absent in 9-repeat carriers. Despite heterogeneity introduced by a host of factors, including variance in other genes involved in smoking behavior, we confirm that DAT genotype predicts the direction and location of neural responses to SCs.

DAT genotype modulates brain and behavioral responses elicited by cigarette cues.

We previously demonstrated differential activation of the mesocorticolimbic reward circuitry in response to cigarette cues independent of withdrawal. Despite robust effects, we noted considerable individual variability in brain and subjective responses. As dopamine (DA) is critical for reward and its predictive signals, genetically driven variation in DA transmission may account for the observed differences. Evidence suggests that a variable number of tandem repeats (VNTRs) polymorphism in the DA transporter (DAT) SLC6A3 gene may influence DA transport. Brain and behavioral responses may be enhanced in probands carrying the 9-repeat allele. To test this hypothesis, perfusion fMR images were acquired during cue exposure in 19 smokers genotyped for the 40 bp VNTR polymorphism in the SLC6A3 gene. Contrasts between groups revealed that 9-repeat (9-repeats) had a greater response to smoking (vs nonsmoking) cues than smokers homozygous for the 10-repeat allele (10/10-repeats) bilaterally in the interconnected ventral striatal/pallidal/orbitofrontal cortex regions (VS/VP/OFC). Activity was increased in 9-repeats and decreased in 10/10-repeats in the VS/VP/OFC (p<0.001 for all analyses). Brain activity and craving was strongly correlated in 10/10-repeats in these regions and others (anterior cingulate, parahippocampal gyrus, and insula; r(2)=0.79-0.86, p<0.001 in all regions). Alternatively, there were no significant correlations between brain and behavior in 9-repeats. There were no differences in cigarette dependence, demographics, or resting baseline neural activity between groups. These results provide evidence that genetic variation in the DAT gene contributes to the neural and behavioral responses elicited by smoking cues.

Association analysis of the pituitary adenylate cyclase-activating polypeptide (PACAP/ADCYAP1) gene in bipolar disorder.

BACKGROUND: Linkage studies in bipolar disorder (BPD) suggest that a susceptibility locus exists on chromosome 18p11. The pituitary adenylate cyclase-activating polypeptide/adenylate cyclase-activating polypeptide 1 (pituitary) (PACAP/ADCYAP1) gene maps to this region. PACAP is a neuropeptide involved in neurotransmission in both the peripheral nervous system and central nervous system and is required for catecholamine secretion. Animal models of PACAP mutations show remarkable behavioral defects, including hyperactivity and increased exploratory behavior. OBJECTIVE: In this study we tested the hypothesis that genetic variations in the human PACAP gene contribute to BPD. METHODS: Genotyping of seven single nucleotide polymorphisms (rs1893154; rs2846811; rs8192595; rs2856966; rs928978; rs2231187; rs1610037) was performed in BPD patients (n=570) and healthy controls (n=710). Genotypes and allele frequencies were compared between groups using chi contingency analysis. Linkage disequilibrium between markers was calculated and estimated haplotype frequencies were compared between groups. MAIN RESULTS: There were no significant differences between groups on the allele, genotype or haplotype level for any of the tested single nucleotide polymorphisms. CONCLUSION: Our results provide no evidence of an association of the PACAP gene with BPD in this group of patients and controls. Additional studies are necessary to elucidate the BPD susceptibility locus on chromosome 18p.