The direct physiological effects that promote nicotine dependence (ND) are mediated by nicotinic acetylcholine receptors (nAChRs). In line with the genetic and pharmacological basis of addiction, many previous studies have revealed significant associations between variants in the nAChR subunit genes and various measures of ND in different ethnic samples. In this study, we first examined the association of variants in nAChR subunits α2 (CHRNA2) and α6 (CHRNA6) genes on chromosome 8 with ND using a family sample consisting of 1,730 European Americans (EAs) from 495 families and 1,892 African Americans (AAs) from 424 families (defined as the discovery family sample). ND was assessed by two standard quantitative measures: smoking quantity (SQ) and the Fagerström Test for ND (FTND). We found nominal associations for all seven tested SNPs of the genes with at least one ND measure in the EA sample and for two SNPs in CHRNA2 in the AA sample. Of these, associations of SNPs rs3735757 with FTND (P = 0.0068) and rs2472553 with both ND measures (with a P value of 0.0043 and 0.00086 for SQ and FTND, respectively) continued to be significant in the EA sample even after correction for multiple tests. Further, we found several haplotypes that were significantly associated with ND in the EA sample in CHRNA6 and in the both EA and AA samples in CHRNA2. To confirm the associations of the two genes with ND, we conducted a replication study with an independent case-control sample from the SAGE study, which showed a significant association of the two genes with ND, although the significantly associated SNPs were not always the same in the two samples. Together, these findings indicate that both CHRNA2 and CHRNA6 play a significant role in the etiology of ND in AA and EA smokers. Further replication in additional independent samples is warranted.