Association of galanin haplotypes with alcoholism and anxiety in two ethnically distinct populations.
ABSTRACT: The neuropeptide galanin (GAL) is widely expressed in the central nervous system. Animal studies have implicated GAL in alcohol abuse and anxiety: chronic ethanol intake increases hypothalamic GAL mRNA; high levels of stress increase GAL release in the central amygdala. The coding sequence of the galanin gene, GAL, is highly conserved and a functional polymorphism has not yet been found. The aim of our study was, for the first time, to identify GAL haplotypes and investigate associations with alcoholism and anxiety. Seven single-nucleotide polymorphisms (SNPs) spanning GAL were genotyped in 65 controls from five populations: US and Finnish Caucasians, African Americans, Plains and Southwestern Indians. A single haplotype block with little evidence of historical recombination was observed for each population. Four tag SNPs were then genotyped in DSM-III-R lifetime alcoholics and nonalcoholics from two population isolates: 514 Finnish Caucasian men and 331 Plains Indian men and women. Tridimensional Personality Questionnaire harm avoidance (HA) scores, a dimensional measure of anxiety, were obtained. There was a haplotype association with alcoholism in both the Finnish (P=0.001) and Plains Indian (P=0.004) men. The SNPs were also significantly associated. Alcoholics were divided into high and low HA groups (>or= and
Project description:The GABAAalpha2 receptor gene (GABRA2) modulates anxiety and stress response. Three recent association studies implicate GABRA2 in alcoholism, however in these papers both common, opposite-configuration haplotypes in the region distal to intron3 predict risk. We have now replicated the GABRA2 association with alcoholism in 331 Plains Indian men and women and 461 Finnish Caucasian men. Using a dimensional measure of anxiety, harm avoidance (HA), we also found that the association with alcoholism is mediated, or moderated, by anxiety. Nine SNPs were genotyped revealing two haplotype blocks. Within the previously implicated block 2 region, we identified the two common, opposite-configuration risk haplotypes, A and B. Their frequencies differed markedly in Finns and Plains Indians. In both populations, most block 2 SNPs were significantly associated with alcoholism. The associations were due to increased frequencies of both homozygotes in alcoholics, indicating the possibility of alcoholic subtypes with opposite genotypes. Congruently, there was no significant haplotype association. Using HA as an indicator variable for anxiety, we found haplotype linkage to alcoholism with high and low dimensional anxiety, and to HA itself, in both populations. High HA alcoholics had the highest frequency of the more abundant haplotype (A in Finns, B in Plains Indians); low HA alcoholics had the highest frequency of the less abundant haplotype (B in Finns, A in Plains Indians) (Finns: P = 0.007, OR = 2.1, Plains Indians: P = 0.040, OR = 1.9). Non-alcoholics had intermediate frequencies. Our results suggest that within the distal GABRA2 region is a functional locus or loci that may differ between populations but that alters risk for alcoholism via the mediating action of anxiety.
Project description:The chromosome 4 cluster of GABA(A) receptor genes is predominantly expressed in the brain reward circuitry and this chromosomal region has been implicated in linkage scans for alcoholism. Variation in one chromosome 4 gene, GABRA2, has been robustly associated with alcohol use disorders (AUD) although no functional locus has been identified. As HapMap data reveal moderate long-distance linkage disequilibrium across GABRA2 and the adjacent gene, GABRG1, it is possible that the functional locus is in GABRG1. We genotyped 24 SNPs across GABRG1 and GABRA2 in two population isolates: 547 Finnish Caucasian men (266 alcoholics) and 311 community-derived Plains Indian men and women (181 alcoholics). In both the Plains Indians and the Caucasians: (1) the GABRG1 haplotype block(s) did not extend to GABRA2; (2) GABRG1 haplotypes and SNPs were significantly associated with AUD; (3) there was no association between GABRA2 haplotypes and AUD; (4) there were several common (>or=0.05) haplotypes that spanned GABRG1 and GABRA2 (341 kb), three of which were present in both populations: one of these ancestral haplotypes was associated with AUD, the other two were more common in non-alcoholics; this association was determined by GABRG1; (5) in the Finns, three less common (<0.05) extended haplotypes showed an association with AUD that was determined by GABRA2. Our results suggest that there are likely to be independent, complex contributions from both GABRG1 and GABRA2 to alcoholism vulnerability.
Project description:BACKGROUND:Animal and human studies indicate that GABBR1, encoding the GABAB1 receptor subunit, and SLC6A1, encoding the neuronal gamma-aminobutyric acid (GABA) transporter GAT1, play a role in addiction by modulating synaptic GABA. Therefore, variants in these genes might predict risk/resilience for alcoholism. METHODS:This study included 3 populations that differed by ethnicity and alcoholism phenotype: African American (AA) men: 401 treatment-seeking inpatients with single/comorbid diagnoses of alcohol and drug dependence, 193 controls; Finnish Caucasian men: 159 incarcerated alcoholics, half with comorbid antisocial personality disorder, 181 controls; and a community sample of Plains Indian (PI) men and women: 239 alcoholics, 178 controls. Seven GABBR1 tag single nucleotide polymorphisms were genotyped in the AA and Finnish samples; rs29220 was genotyped in the PI for replication. Also, a uniquely African, functional SLC6A1 insertion promoter polymorphism (IND) was genotyped in the AAs. RESULTS:We found a significant and congruent association between GABBR1 rs29220 and alcoholism in all 3 populations. The major genotype (heterozygotes in AAs, Finns) and the major allele in PIs were significantly more common in alcoholics. Moreover, SLC6A1 IND was more abundant in controls, that is, the major genotype predicted alcoholism. An analysis of combined GABBR1 rs29220 and SLC6A1 IND genotypes showed that rs29220 heterozygotes, irrespective of their IND status, had an increased risk for alcoholism, whereas carriers of the IND allele and either rs29220 homozygote were more resilient. CONCLUSIONS:Our results show that with both GABBR1 and SLC6A1, the minor genotypes/alleles were protective against risk for alcoholism. Finally, GABBR1 rs29220 might predict treatment response/adverse effects for baclofen, a GABAB receptor agonist.
Project description:The resting EEG is a dynamic index of cortical activation, cognitive function and consciousness and is therefore an intermediate phenotype for many behaviors in which arousal is implicated such as anxiety and alcoholism. We performed a dense whole genome linkage scan using 3878 unlinked SNPs in a large pedigree derived from a population isolate sample of 328 Plains American Indians. Alpha (8-13 Hz), theta (4-8 Hz) and beta (13-30 Hz) EEG power was heritable (0.58-0.27) and stable over a 2 year period (r = 0.82-0.53). Genetic correlations between frequency bands were high (0.75). Linkage peaks for EEG power in all three frequency bands converged on chromosome 5q13-14 with genome-wide significant LOD scores of 3.5 (empirical p<0.0001) for alpha and beta power. A logical candidate gene, corticotropin releasing hormone-binding protein (CRH-BP), was located at the apex of these convergent linkage peaks. CRH-BP was significantly associated with alpha power in the Plains Indians and also in a replication sample of 188 Caucasians. Moreover, the same SNPs and haplotypes, located within the CRH-BP haplotype block, were also associated with anxiety disorders in the Plains Indians and alcohol use disorders in the Caucasians. CRH-BP modulates CRH which influences cortical and hippocampal EEG activity and is the primary mediator of the neuroendocrine stress response. Our results suggest a likely role for CRH-BP in stress-related alcoholism and highlight the use of the resting EEG as an intermediate phenotype for arousal-related behaviors such as anxiety and addiction.
Project description:The corticotropin-releasing hormone type I receptor (CRHR1) gene has been implicated in the liability for neuropsychiatric disorders, particularly under conditions of stress. On the basis of the hypothesized effects of CRHR1 variation on stress reactivity, measures of adulthood traumatic stress exposure were analyzed for their interaction with CRHR1 haplotypes and single-nucleotide polymorphisms (SNPs) in predicting the risk for alcoholism. Phenotypic data on 2533 non-related Caucasian individuals (1167 alcoholics and 1366 controls) were culled from the publically available Study of Addiction: Genetics and Environment genome-wide association study. Genotypes were available for 19 tag SNPs. Logistic regression models examined the interaction between CRHR1 haplotypes/SNPs and adulthood traumatic stress exposure in predicting alcoholism risk. Two haplotype blocks spanned CRHR1. Haplotype analyses identified one haplotype in the proximal block 1 (P = 0.029) and two haplotypes in the distal block 2 (P = 0.026, 0.042) that showed nominally significant (corrected P < 0.025) genotype × traumatic stress interactive effects on the likelihood of developing alcoholism. The block 1 haplotype effect was driven by SNPs rs110402 (P = 0.019) and rs242924 (P = 0.019). In block 2, rs17689966 (P = 0.018) showed significant and rs173365 (P = 0.026) showed nominally significant, gene × environment (G × E) effects on alcoholism status. This study extends the literature on the interplay between CRHR1 variation and alcoholism, in the context of exposure to traumatic stress. These findings are consistent with the hypothesized role of the extra hypothalamic corticotropin-releasing factor system dysregulation in the initiation and maintenance of alcoholism. Molecular and experimental studies are needed to more fully understand the mechanisms of risk and protection conferred by genetic variation at the identified loci.
Project description:Chronic alcoholism leads to neurotoxic effects in the central nervous system. Neuroadaptive changes in the brain may lead to tolerance to, and dependence on, alcohol as a result of alterations in synaptic complexity. G-proteins are negatively regulated by RGS proteins, which are integral to many neural pathways that include neurotransmission, hormonal responses, and chemotactic signals. These considerations, together with findings from microarray analyses of human autopsy brain, suggest that proteins involved in G-protein signalling, specifically the RGS protein family, may play an important role in the functioning of neural systems that are affected by chronic alcohol abuse. We used Real Time PCR to measure the expression of two members of the RGS family, RGS4 and RGS7, in the superior frontal gyrus and primary motor cortex from alcoholic and non-alcoholic cases. Overall, cirrhotic alcoholics had lower expression levels of RGS4 mRNA than controls and non-cirrhotic alcoholics. We also report that the four RGS4 SNPs (SNP1, 4, 7 and 18) may be associated with alcoholism in European Caucasians at the haplotype level. The haplotype T-C-G (SNP1-4-18) may exert a protective effect against alcoholism.
Project description:Alcohol use disorders (AUD) with co-morbid antisocial personality disorder (ASPD) have been associated with serotonin (5-HT) dysfunction. 5-HT3 receptors are potentiated by ethanol and appear to modulate reward. 5-HT3 receptor antagonists may be useful in the treatment of early-onset alcoholics with co-morbid ASPD. Low-voltage alpha electroencephalogram (EEG) power, a highly heritable trait, has been associated with both AUD and ASPD. A recent whole genome linkage scan in one of our samples, Plains American Indians (PI), has shown a suggestive linkage peak for alpha power at the 5-HT3R locus. We tested whether genetic variation within the HTR3A and HTR3B genes influences vulnerability to AUD with comorbid ASPD (AUD+ASPD) and moderates alpha power. Our study included three samples: 284 criminal alcoholic Finnish Caucasians and 234 controls; two independent community-ascertained samples with resting EEG recordings: a predominantly Caucasian sample of 191 individuals (Bethesda) and 306 PI. In the Finns, an intronic HTR3B SNP rs3782025 was associated with AUD+ASPD (P=.004). In the Bethesda sample, the same allele predicted lower alpha power (P=7.37e(-5)). Associations between alpha power and two other HTR3B SNPs were also observed among PI (P=.03). One haplotype in the haplotype block at the 3' region of the gene that included rs3782025 was associated with AUD+ASPD in the Finns (P=.02) and with reduced alpha power in the Bethesda population (P=.00009). Another haplotype in this block was associated with alpha power among PI (P=.03). No associations were found for HTR3A. Genetic variation within HTR3B may influence vulnerability to develop AUD with comorbid ASPD. 5-HT3R might contribute to the imbalance between excitation and inhibition that characterize the brain of alcoholics.
Project description:The expression of the galanin gene (GAL) in the paraventricular nucleus (PVN) and in the amygdala of higher vertebrates suggests the requirement for highly conserved, but unidentified, regulatory sequences that are critical to allow the galanin gene to control alcohol and fat intake and modulate mood. We used comparative genomics to identify a highly conserved sequence that lay 42?kb 5' of the human GAL transcriptional start site that we called GAL5.1. GAL5.1 activated promoter activity in neurones of the PVN, arcuate nucleus and amygdala that also expressed the galanin peptide. Analysis in neuroblastoma cells demonstrated that GAL5.1 acted as an enhancer of promoter activity after PKC activation. GAL5.1 contained two polymorphisms; rs2513280(C/G) and rs2513281(A/G), that occurred in two allelic combinations (GG or CA) where the dominant GG alelle occurred in 70-83?% of the human population. Intriguingly, both SNPs were found to be in LD (R(2) of 0.687) with another SNP (rs2156464) previously associated with major depressive disorder (MDD). Recreation of these alleles in reporter constructs and subsequent magnetofection into primary rat hypothalamic neurones showed that the CA allele was 40?% less active than the GG allele. This is consistent with the hypothesis that the weaker allele may affect food and alcohol preference. The linkage of the SNPs analysed in this study with a SNP previously associated with MDD together with the functioning of GAL5.1 as a PVN and amygdala specific enhancer represent a significant advance in our ability to understand alcoholism, obesity and major depressive disorder.
Project description:There is increasing physiological evidence in rodents connecting the neuropeptide galanin to triglyceride (TG) levels. We hypothesized that variation in the galanin preproprotein (GAL) gene may contribute to hypertriglyceridemia (HTG) in humans.We investigated GAL as a TG candidate gene by genotyping 4 tagSNPs in Dutch, Finnish, and Mexican familial combined hyperlipidemia (FCHL) families as well as in white combined hyperlipidemia cases/controls (n=2471). The common allele of rs2187331, residing in the promoter region of GAL, was significantly associated with HTG (probability value=0.00038). In an unascertained population sample of 4463 Finnish males, the rare allele of rs2187331 was associated with higher TGs (probability value=0.0028 to 0.00016). We also observed an allele specific difference with rs2187331 in reporter gene expression and nuclear factor binding in vitro. Furthermore, we detected differential expression of many key lipid genes in adipose tissue based on rs2187331 genotypes.The SNP rs2187331 is associated with HTG in FCHL and white combined hyperlipidemia cases/controls and influences TG levels in the population. Further studies are warranted to elucidate the allelic difference observed between FCHL and the general population. Functional evidence shows that rs2187331 has an allele specific cis-regulatory function and influences the expression of lipid related genes in adipose.
Project description:Alcoholism is a complex disease with both genetic and environmental risk factors. To identify genes that affect the risk for alcoholism, we systematically ascertained and carefully assessed individuals in families with multiple alcoholics. Linkage and association analyses suggested that a region of chromosome 4p contained genes affecting a quantitative endophenotype, brain oscillations in the beta frequency range (13-28 Hz), and the risk for alcoholism. To identify the individual genes that affect these phenotypes, we performed linkage disequilibrium analyses of 69 single-nucleotide polymorphism (SNPs) within a cluster of four GABA(A) receptor genes, GABRG1, GABRA2, GABRA4, and GABRB1, at the center of the linked region. GABA(A) receptors mediate important effects of alcohol and also modulate beta frequencies. Thirty-one SNPs in GABRA2, but only 1 of the 20 SNPs in the flanking genes, showed significant association with alcoholism. Twenty-five of the GABRA2 SNPs, but only one of the SNPs in the flanking genes, were associated with the brain oscillations in the beta frequency. The region of strongest association with alcohol dependence extended from intron 3 past the 3' end of GABRA2; all 43 of the consecutive three-SNP haplotypes in this region of GABRA2 were highly significant. A three-SNP haplotype was associated with alcoholism, with P=.000000022. No coding differences were found between the high-risk and low-risk haplotypes, suggesting that the effect is mediated through gene regulation. The very strong association of GABRA2 with both alcohol dependence and the beta frequency of the electroencephalogram, combined with biological evidence for a role of this gene in both phenotypes, suggest that GABRA2 might influence susceptibility to alcohol dependence by modulating the level of neural excitation.