minoxidil-sulfate-ester and 1-oleoyl-2-acetylglycerol

We studied the mechanism of K+ channel activation by minoxidil-sulfate (MxSO4) in fused Madin-Darby canine kidney (MDCK) cells. Patch-clamp techniques were used to assess single channel activity, and fluorescent dye techniques to monitor cell calcium. A Ca(2+)-dependent inward-rectifying K+ channel with slope conductances of 53 +/- 3 (negative potential range) and 20 +/- 3 pS (positive potential range) was identified. Channel activity is minimal in cell-attached patches. MxSO4 initiated both transient channel activation and an increase of intracellular Ca2+ (from 94.2 +/- 9.1 to 475 +/- 12.6 nmol/liter). The observation that K+ channel activity of excised inside-out patches was detected only at Ca2+ concentrations in excess of 10 mumol/liter suggests the involvement of additional mechanisms during channel activation by MxSO4. Transient K+ channel activity was also induced in cell-attached patches by 10 mumol/liter of the protein kinase C activator 1-oleoyl-2-acetyl-glycerol (OAG). OAG (10 mumol/liter in the presence of 1.6 mmol/liter ATP) increased the Ca2+ sensitivity of the K+ channel in inside-out patches significantly by lowering the Km for Ca2+ from 100 mumol/liter to 100 nmol/liter. The channel activation by OAG was reversed by the protein kinase inhibitor H8. Staurosporine, a PKC inhibitor, blocked the effect of MxSO4 on K+ channel activation. We conclude that MxSO4-induced K+ channel activity is mediated by the synergistic effects of an increase in intracellular Ca2+ and a PKC-mediated enhancement of the K+ channel's sensitivity to Ca2+.

Topics: Alkaloids; Animals; Calcium; Cell Membrane; Cells, Cultured; Diglycerides; Dogs; Fluorescence; Kidney; Membrane Potentials; Minoxidil; Potassium Channels; Protein Kinase C; Staurosporine