Background:  Nuclear receptors play an important role in bone metabolism. In bone cells, the vitamin D receptor (VDR) and the steroid and xenobiotic receptor (SXR) are activated by vitamin D and vitamin K2, respectively. VDR and SXR are the NR1I subfamily members of nuclear receptors. We speculated that the constitutive androstane receptor (CAR), the third member of the NR1I subfamily, also could be implicated in the regulation of bone metabolism. Therefore, we analyzed expression of CAR mRNA in osteoblasts and then examined association of a single nucleotide polymorphism (SNP) in the human CAR gene at intron 2 (IVS2–99C>T, rs2502815) with bone mineral density (BMD).Methods:  Expression levels of CAR mRNA were analyzed during the culture course of rat primary osteoblasts. Association of an SNP in the CAR gene with BMD was examined in 548 healthy Japanese postmenopausal women.Results:  CAR mRNA increased at day 16 and then increased during culture of rat primary osteoblasts. The increase of CAR mRNA was parallel with the increase of alkaline phosphatase expression, a differentiation marker of osteoblasts. As a result of association study of an SNP in the CAR gene at intron 2, subjects with the CC genotype (n = 208) had significantly higher BMD than subjects with the TT or CT genotype (n = 340) (lumbar spine BMD, P = 0.0185; total body BMD, P = 0.0416).Conclusion:  CAR mRNA was expressed and regulated in primary osteoblasts. A genetic variation at the CAR gene locus is associated with BMD, suggesting an involvement of the CAR gene in bone metabolism.