ORIGINAL ARTICLE Haplotype-Based Localization of an Alcohol Dependence Gene to the 5q34 ␥-Aminobutyric Acid Type A Gene Cluster Marta Radel, MD, PhD; Roger L. Vallejo, PhD; Nakao Iwata, MD, PhD; Richard Aragon, PhD; Jeffrey C. Long, PhD; Matti Virkkunen, MD; David Goldman, MD Context: Pharmacobehavioral and pharmacogenetic evi- Results: Sib-pair linkage of GABRG2 to alcohol depen- ␥ dence links -aminobutyric acid type A (GABAA) recep- dence was observed in Finns (P=.008). Association of tors and chromosomal regions containing GABAA recep- the GABRB2 1412T allele with alcohol dependence was tor genes to ethanol-related responses. The GABAA gene detected in both populations (Finns, P=.01; Southwest- cluster on chromosome 5q34 is of particular interest in ern Native Americans, P=.008), and the GABRA6 1519T the genetics of alcohol dependence because of the ␥2 sub- allele was associated in both Finns (P=.01) and South- unit requirement for ethanol’s modulatory action on western Native Americans (P=.03). Linkage disequilib- rium mapping with 3-locus haplotypes yielded evi- GABAA receptors, previous linkage findings in mice and humans implicating both GABRA6 and GABRG2, and re- dence for an alcohol-dependence locus at the GABAA gene ported associations of GABRA6, GABRB2, and GABRG2 cluster region in both populations. The most highly sig- nificant signals were at 3-locus haplotypes that in- alleles with alcohol dependence. cluded 1 or more GABRA6 polymorphisms, with the peak signal at a GABRA6 3-locus haplotype (Finns, empirical Objective: To determine whether variation at the 5q34 P=.004; Southwestern Native Americans, empirical GABAA gene cluster is implicated in differential suscep- P=.02). tibility to alcohol dependence. Conclusions: We detected sib-pair linkage of 5q34 Methods: Two large psychiatrically interviewed samples, GABAA receptor genes to alcohol dependence in Finns a Southwestern Native American population sample and found association both in Finns and in Southwest- (N=433) and a Finnish sample (N=511) with alcohol- ern Native Americans. In both populations, the haplo- dependent subjects and unaffected individuals, were geno- type localization implicates the region containing the typed for 6 single nucleotide polymorphisms at the 5q34 Pro385Ser GABRA6 polymorphism and 2 other polymor- GABAA gene cluster. In addition to sib-pair linkage and phisms at GABRA6. case-control association analyses, linkage disequilib- rium mapping with haplotypes was used. Arch Gen Psychiatry. 2005;62:47-55 LCOHOL DEPENDENCE (“AL- lated in the pharmacokinetic genetic risk coholism”) is a common domain (ie, in genes coding for aldehyde (10% in men, 4% in wom- dehydrogenase type 26 and for alcohol de- en),1 genetically influ- hydrogenase type 27), they are common enced disorder with heri- in East Asians but not in most other popu- tability estimates ranging from 40% to lations. Pharmacodynamic-specific gene A2-4 Author Affiliations: Laboratory 60%. The vulnerability/protective alle- targets for predisposition to alcohol de- of Neurogenetics, National les for alcoholism represent both the phar- pendence include the genes that encode Institute on Alcohol Abuse and macokinetic (detected and widely repli- receptors at which ethanol (alcohol) first Alcoholism, National Institutes cated) and the pharmacodynamic acts in the brain and that may act as gate- of Health, Rockville, Md (detected but still putative) domains. A keepers in alcohol response. (Drs Radel, Vallejo, Aragon, substantial part of the genetic compo- ␥-Aminobutyric acid (GABA) is the ma- Long, and Goldman); nent of variance in alcoholism vulnerabil- jor inhibitory neurotransmitter in the Department of Psychiatry, Fujita ity is substance specific, that is, not cross- mammalian central nervous system8 and Health University School of 5 Medicine, Toyoake, Japan inherited with vulnerability to other drugs. exerts most of its actions at GABA type A (Dr Iwata); and Department of Such genetic epidemiological data impli- (GABAA) receptors, which are ligand- Psychiatry, University of cate genes that are relatively substance spe- gated chloride-channel complexes. ␥-Ami- Helsinki, Helsinki, Finland cific. Although a few substance-specific nobutyric acid type A receptor com- (Dr Virkkunen). polymorphisms have already been iso- plexes are generally heteropentamers (REPRINTED) ARCH GEN PSYCHIATRY/ VOL 62, JAN 2005 WWW.ARCHGENPSYCHIATRY.COM 47 ©2005 American Medical Association. All rights reserved. Downloaded From: https://jamanetwork.com/ on 09/27/2021 composed of genetically distinct subunits; 16 related mam- unit gene clusters appear to have ethanol response– 9,10 23 malian subunits have been reported. The human GABAA associated quantitative trait loci nearby. In the mouse, receptor subunit genes are grouped into 7 classes (␣, ␤, the Ala11Thr variant of the ␥2 subunit correlates with ␥, ␦, ε, ␲, and ␪).10 Within these classes, protein se- acute ethanol withdrawal severity,24 and in the rat, an ␣6 quence identity is approximately 70% (ie, ␣1-␣6, ␤1- Arg100Gln amino acid substitution that differentiates al- ␤3, or ␥1-␥3) and protein sequence identity is approxi- cohol tolerant and nontolerant has been reported to al- 25 mately 30% between classes. Most of the human GABAA ter response to benzodiazepines. receptor subunit genes have been assigned to chromo- In the human, preliminary linkage findings have also some regions, and subunit gene clusters have been iden- indicated that variation in GABAA receptor subunit genes tified on chromosomes 4, 5, 15, and X.10 Because of the plays a role in differential vulnerability to alcohol de- ␣ number of subunits, the diversity of expressed GABAA pendence, and a specific polymorphism of the 6 sub- receptor pentamers is large and would be larger except unit gene (ie, GABRA6) encoding an amino acid substi- that these receptors are usually assembled from defined tution (Pro385Ser) was implicated.26 As measured by proportions of ␣, ␤, and ␥ subunits and except that there slowing of saccadic eye movement velocity, sons of al- is considerable variation in the localization and devel- coholics appear to have diminished sensitivity to ben- opmental timing of expression of the different sub- zodiazepines as compared with young men who are at units.11-14 The most abundant subunit combinations ob- lower risk for alcohol dependence, leading to the pro- served so far are ␣1␤2␥2, ␣2␤3␥2, and ␣3␤3␥2, which posal that differences in the expression or function of make up about 80% of all GABAA receptors. The recep- GABAA receptors alter vulnerability to alcohol depen- ␣ ␤ ␥ 27 ␣ tors containing a 1 2 2 combination, believed to as- dence. In these same subjects, the GABAA 6 subunit semble following the coordinated expression of GABRA1, Pro385Ser polymorphism was shown to predict benzo- GABRB2, and GABRG2 genes on 5q34, constitute the ma- diazepine sensitivity.28 jor GABAA receptor subtype in the adult central nervous When ethanol itself is administered as a drug chal- system (about 50%), have been identified in neurons at lenge, decreased intoxication, motor effects, and hor- all levels of the neuraxis, and are believed to mediate the monal responses are observed in sons of alcoholics,29 basic pharmacological spectrum of the classical, high- which has been extensively replicated. In a 15-year fol- affinity BZ site ligands, except CL 218872.12,15 A variety low-up study of 450 men, Schuckit30,31 showed that low of central nervous system–depressant drugs that show level of response to alcohol in young adulthood is a pre- cross-tolerance, including ethanol, benzodiazepines, and dictor of later alcoholism. The risk attributable to the level barbiturates, as well as inhalant anesthetics and some en- of response is substantial (approximately 40% of the vari- dogenous neuroactive steroids, positively modulate the ance in vulnerability), largely independent of family his- 8,16-18 31 GABAA receptors. Many behavioral effects of etha- tory, and largely unshared with other drugs of abuse. nol (eg, anxiolytic, ataxic, and sedative/hypnotic) may By selective genotyping of a subsample of this same co- 26 be explained by allosteric enhancement of GABAA recep- hort, Schuckit et al found preliminary evidence that the ␣ tor–mediated ionic influx and consequent hyperpolar- GABAA 6 Pro385Ser amino acid substitution is associ- ization of the neuronal membrane.8 Agents that in- ated both with alcohol sensitivity and with increased risk crease GABAA receptor activity in the central nervous for alcohol dependence. Associations of markers at the system by acting as GABA positive modulators (ie, ben- 5q34 GABAA gene cluster have also been reported in Scot- zodiazepines, barbiturates, and depressant steroids) en- tish,32 German,33 and Japanese34 samples. hance acute sensitivity to ethanol and maintain ethanol Here we report both locus-based linkage analyses in preference, whereas drugs that act as GABA antagonists sib pairs and allele-based localization by association us- at GABAA receptors, such as picrotoxin, decrease many ing GABAA haplotypes in this Finnish population and in acute actions of ethanol and reduce ethanol prefer- a second semi-isolated population of Southwestern Na- ence.19 In addition, signs of ethanol withdrawal are di- tive Americans. minished following treatments with those GABA ago- nists that increase GABAA receptor function, whereas METHODS GABA antagonists at GABAA receptors increase such 19 signs. Effects of ethanol on GABAA receptor function and expression make the GABAA receptor