We investigated the effects of the selective inhibitor of Na(+)/Ca(2+) exchanger (NCX), 2',4'- and 3',4'-dichlorobenzamil (DCB), on large-conductance Ca(2+)-activated K(+) (BK(Ca)) channels in cultured human umbilical vein endothelial cells (HUVECs) and fresh isolated mouse aortic smooth muscle cells (MASMCs) using the patch clamp techniques. Both kinds of DCB reversibly activated BK(Ca) currents in whole-cell clamped HUVECs or MASMCs. The EC(50) of 2',4'-DCB for BK(Ca) current activation in HUVECs was 2.64 +/- 0.10 muM. In inside-out and outside-out patches, 2',4'-DCB remarkably increased BK(Ca) channels activity. 2',4'-DCB increased open frequency, but had no significant effect on mean open time. In inside-out patches, 2',4'-DCB shifted the relationship curve between [Ca(2+)](i) and open probability (NP(o)) to the left; the [Ca(2+)](i) required to evoke half-maximal activation changed from 1087.45 +/- 142.91 nM to 500.24 +/- 66.83 nM by 10 muM 2',4'-DCB. In addition, 2',4'-DCB shifted the relationship curve between membrane potential and NP(o) to the left; the membrane potential to evoke half-maximal activation changed from 81.1 +/- 2.4 to 64.7 +/- 3.1 mV by 10 muM 2',4'-DCB. 3',4'-DCB also increased BK(Ca) channels activity. There was no significant difference in the effect of DCB on BK(Ca) channels between both excised patches. These results suggested that 2',4'- and 3',4'-DCB activate BK(Ca) channels activity in HUVECs and MASMCs by increasing the sensitivity of BK(Ca) channels to cytosolic free Ca(2+) and membrane potential. Our report would provide a consideration if they are used as NCX blocker in vascular endothelial cells or smooth muscle cells.