In the present study we confirmed our hypothesis that GABRA2
, childhood trauma and their interaction influence vulnerability to substance dependence, at least in African American men. Firstly, we found that the patients with heroin, alcohol and cocaine dependence had experienced significantly more childhood trauma than the controls. Furthermore, our results showed that the greater the severity of childhood trauma the greater the likelihood of polysubstance dependence. The latter finding is supported by previous studies (37
). Secondly, our results showed that GABRA2
variation predicted addiction vulnerability, particularly for heroin dependence. Thirdly, an interaction between childhood trauma and GABRA2
variation was found to influence addiction risk, particularly for cocaine dependence.
Earlier studies, together with HapMap, have identified the same two GABRA2
haplotype blocks within Caucasians, Asians, Native Americans and African Americans. The previously reported significant association signals with alcoholism have been within the haplotype block that extends downstream from intron 3 (called block 2 in our study). In Caucasians, Asians and Native Americans there are two major yin-yang haplotypes within this block that account for nearly all of the haplotype diversity (24
). Individuals of African origin also have these two yin-yang haplotypes however in addition they have two common haplotypes that are not present in other populations. In the present study these two unique haplotypes were associated with addiction vulnerability: one haplotype was associated with heroin dependence and the other haplotype was more common in controls and may be protective against addiction. In contrast, we found no association between the yin-yang haplotypes and alcohol dependence or drug dependence unlike the many earlier studies in Caucasians (12
). The two previously reported studies in African Americans showed no association between the yin-yang haplotypes and alcohol dependence (22
) or polysubstance abuse (23
). Moreover, a recent, dense genomewide linkage scan for alcohol dependence in African Americans did not find a linkage peak at the GABAA
receptor gene complex on chromosome 4 (45
), unlike earlier studies in Caucasians (46
) and Native Americans (47
In this study we also found that an intron 2 SNP, rs11503014 that is not in LD with any SNPs in haplotype blocks 1 or 2, was associated with heroin dependence. Rs11503014 is not located within a haplotype block in the three HapMap populations or within Finnish Caucasian and Plains Indian samples for which we have genotyped the same SNPs as in the present study (data not shown). Rs11503014 is located nearby to an alternatively spliced exon 2 and within an exon (5′ UTR) of the alternative GABRA2 transcript NM_001114175. The DNA sequence within which rs11503014 is located is similar to the exonic splicing enhancers: srp55, srp40, sf2 and sc35. Thus it is theoretically possible that rs11503014 may be implicated in exon splicing. Our results therefore suggest that GABRA2 may have at least two independent loci that are implicated in the vulnerability to heroin dependence.
It has been shown that there is a common genetic factor for addiction to illicit drugs, including cocaine and opiates (4
). However alcohol and drug dependence also have substantial disorder-specific genetic loading (6
). In the present study we detected both a general and specific effect for GABRA2
: one of the uniquely African haplotypes appeared to be protective against any addiction, however it was specifically protective for heroin dependence. In contrast, the other uniquely African haplotype predicted heroin dependence only. Likewise, rs11503014 variation was predictive for any addiction and specifically for heroin addiction.
Opiates and cocaine have different mechanisms of action in the CNS and may interact differently with GABRA2
variation and early life stress to influence the vulnerability to heroin or cocaine dependence. Our finding of a GABRA2
-heroin dependence association is backed up by preclinical studies that indicate that GABAA
receptors may influence the actions of opiates; for example, hyperpolarization of GABAergic neurons in the ventral tegmental area (where GABAA
receptors are highly expressed) by opiates results in increased firing of dopamine neurons within the dopamine reward pathway (8
In our study, childhood trauma had the least impact on heroin dependence. In contrast, we found a strong effect of childhood trauma on cocaine dependence. Although there was no main effect of GABRA2,
severe childhood trauma was associated with cocaine dependence only in individuals with the rs11503014 11/12 genotypes. Preclinical studies support our findings: rats subjected to early life stress are more sensitive to cocaine, demonstrate increased cocaine self-administration (27
) and, perhaps in line with this, have an altered pattern of distribution of GABAA
) that have been implicated in cocaine sensitization (56
). Therefore, bases on these preclinical findings, one speculative explanation for the gene × environment (G×E) interaction in our study might be that if rs11503014 (or a tightly linked SNP) is indeed implicated in exon splicing and thus might influence GABRA2
expression, carriers of the variant allele might be more sensitive to early life stress and subsequent vulnerability to cocaine addiction. However, one caveat should be discussed. Due to the limited sample size and loss of power stemming from categorization of variables we did not expect strong effects of G×E interactions and indeed the RSquare values (reflecting the proportion of the total uncertainty that is attributed to the model fit) of the whole model tests for rs111503014 (0.13 for cocaine dependence only, 0.06 for all cocaine dependence) are modest indicating that, if there were no a priori
biological hypothesis, the likelihood of expected cross validation in other samples on a statistical basis alone might be low.
Strengths of the present study include the large sample size of African-American subjects, a group that has been under-represented in genetic studies of addiction. Moreover, since the dataset included individuals with polysubstance dependence as well as individuals addicted to a single substance we were able to parse out both specific and general influences of GABRA2 variation on addiction. Furthermore, it should be noted that we corrected for the effects of population stratification by using ethnic factor scores derived from 186 AIMS as covariates in our analyses. The European factor had a significant effect on heroin addiction in most analyses but no effect on cocaine addiction or alcoholism.
There are some limitations to the present study. Data on other Axis 1 diagnoses were not available for the patients. Since it is known that there is high comorbidity between substance dependence and other psychiatric disorders, particularly major depression, and the controls were free of all Axis 1 diagnoses, it is possible that the signals for association found in our study derived from hidden comorbidity. Nevertheless it should be noted that the extensive literature on association studies with GABRA2
) has largely focused on alcohol and drug dependence and no published study has yet shown a GABRA2
association with depression. Moreover, Covault et al’s 2004 (19
) study showed that the association with alcoholism became stronger when alcoholics with major depression were removed.
Measures of childhood trauma were retrospectively derived from the CTQ. Longitudinal measures are preferable; for example a recent study (57
)suggests that the GABRA2
× childhood trauma interaction might be moderated across development by other environmental factors. Nevertheless, the CTQ is widely used and has been shown to have high reliability and validity in both sexes, in different ethnic groups and in psychiatric patients (42
). Although the overall dataset was large some of the analytical subsets were smaller hence the G×E interactions that we have detected may be an underestimate. Moreover, because of these power issues we had to combine the rs11503014 11/12 genotypes in all analyses. The controls were derived from two sources and although their mean age was appreciably lower than that of patients they had largely passed through the peak age of risk for onset of addictive disorders.
In conclusion, the present study has shown that childhood trauma is a strong predictor for alcohol, cocaine and heroin dependence in African American men. Together with childhood trauma, GABRA2
variation influences risk and resilience for all addiction but most strongly for heroin dependence. Our results suggest that GABRA2
may have at least two independent loci that are implicated in the vulnerability to heroin addiction. The two GABRA2
risk – resilience haplotypes are unique to African Americans. There were no findings with the yin-yang haplotypes that confer addiction risk in Caucasians. Moreover, since there is evidence for sexual dimorphism in the influence of GABRA2
on addiction vulnerability with previous results being significant only in men (16
), the results of our study may not extend to African American women. Thus the findings of the present study may be unique to African-American men and await replication in other similar datasets.