Daidzein, a phytoestrogen, has been reported to produce vasodilation via inhibition of Ca 2+ inflow. However, the involvement of large-conductance Ca 2+ -activated K + (BK Ca ) channels in the effect of daidzein is debated. Therefore, the present study was designed to investigate the effect of daidzein on the rat cerebral basilar artery and the underlying molecular mechanisms. Isolated cerebral basilar artery rings and single vascular smooth muscle cells (VSMCs) were used for vascular reactivity and electrophysiology measurements, to investigate the effect of daidzein on BK Ca channels in cerebral basilar artery smooth muscle. In addition, the human BK Ca channel α-subunit gene (hslo) was transfected into HEK293 cells, to directly assess whether daidzein activates BK Ca channels. The results showed that daidzein produced a concentration-dependent but endothelium-independent relaxation in rat cerebral basilar arteries. Paxilline, a selective BK Ca channel blocker, significantly inhibited the daidzein-induced vasodilation, whereas NS1619, a selective BK Ca channel opener, enhanced the vasodilation. In the whole-cell configuration, daidzein increased noisy oscillation currents in cerebral basilar artery VSMCs in a concentration-dependent manner, and washout of daidzein or blockade of BK Ca channels with paxilline fully reversed the increase. However, daidzein did not substantially affect hSlo currents in HEK293 cells when applied to the outside of the cell membrane. In conclusion, these results indicate that the activation of BK Ca channels in VSMCs at least partly contributes to the daidzein-induced vasodilation of the rat cerebral basilar artery. The β1-subunit of BK Ca channels plays a critical role in the activation of BK Ca currents by daidzein.