The effects of cromakalim on ATP-sensitive potassium channels in insulin-secreting cells

1. The single-channel current recording technique has been used to investigate the effects of cromakalim, diazoxide and ATP, separately and combined, on the opening of ATP-sensitive potassium channels in the insulin-secreting cell-line RINm5F. The actions of these drugs have been studied using the permeabilized open-cell variation of the patch-clamp technique. 2. In the absence of internal ATP, cromakalim (80-200 microM) was unable to open ATP-sensitive K+ channels but when ATP was present both cromakalim and diazoxide caused channel openings. 3. Interactions between ATP and cromakalim seemed competitive. Concentrations of cromakalim in the range 80-200 microM readily activated channels inhibited by 0.1 mM ATP, but had no effects when the concentration of ATP was increased to 0.5-2 mM. Only when the concentration of cromakalim was increased to 400-800 microM could opening of 0.5-2 mM ATP-inhibited channels be regularly observed. In the continued presence of cromakalim (400-800 microM), an increase in the internal concentration of ATP from either 0.25 to 0.5 mM or 1 to 2 mM, inhibited cromakalim-activated K+ channels. 4. Activation of ATP-inhibited K+ channels was abolished by replacing ATP with ATP gamma S and cromakalin had no effects on ATP gamma S-inhibited channels. This suggests that cromakalim may open KATP channels in insulin-secreting cells by a mechanism which involves protein phosphorylation.