Stimulation of large-conductance Ca2+-activated K+ channels by the Na+/Ca2+ exchanger inhibitor dichlorobenzamil in cultured human umbilical vein endothelial cells and mouse aortic smooth muscle cells

Abstract

We investigated the effects of the selective inhibitor of Na +/Ca2+ exchanger (NCX), 2′,4′- and 3′,4′-dichlorobenzamil (DCB), on large-conductance Ca 2+-activated K+ (BKCa) 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 BKCa currents in whole-cell clamped HUVECs or MASMCs. The EC50 of 2′,4′-DCB for BKCa current activation in HUVECs was 2.64 ± 0.10 μM. In inside-out and outside-out patches, 2′,4′-DCB remarkably increased BKCa 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 (NPo) to the left; the [Ca2+]i required to evoke half-maximal activation changed from 1087.45 ± 142.91 nM to 500.24 ± 66.83 nM by 10 μM 2′,4′-DCB. In addition, 2′,4′-DCB shifted the relationship curve between membrane potential and NPo 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 μM 2′,4′-DCB. 3′,4′-DCB also increased BKCa channels activity. There was no significant difference in the effect of DCB on BKCa channels between both excised patches. These results suggested that 2′,4′- and 3′,4′-DCB activate BKCa channels activity in HUVECs and MASMCs by increasing the sensitivity of BKCa channels to cytosolic free Ca2+ and membrane potential. Our report would provide a consideration if they are used as NCX blocker in vascular endothelial cells or smooth muscle cells.