Direct inhibition of the pancreatic β-cell ATP-regulated potassium channel by α-ketoisocaproate

The ATP-regulated potassium (K-ATP) channel plays an essential role in the control of insulin release from the pancreatic beta-cell. In the present study we have used the patch-clamp technique to study the direct effects of alpha-ketoisocaproate on the K-ATP channel in isolated patches and intact pancreatic beta-cells. In excised inside-out patches, the activity of the K-ATP channel was dose dependently inhibited by alpha-ketoisocaproate, half-maximal concentration being approximately 8 mM. The blocking effect of alpha-ketoisocaproate was fully reversible. Stimulation of channel activity by the addition of ATP/ADP (ratio 1) did not counteract the inhibitory effect of alpha-ketoisocaproate. In the presence of the metabolic inhibitor sodium azide, alpha-ketoisocaproate was still able to inhibit single channel activity in excised patches and to block whole cell K-ATP, currents in intact cells. No effect of alpha-ketoisocaproate could be obtained on either the large or the small conductance Ca2+-regulated K+ channel. Enzymatic treatment of the patches with trypsin prevented the inhibitory effect of alpha-ketoisocaproate. Based on these observations, it is unlikely that the blocking effect of alpha-ketoisocaproate is due to an unspecific effect on K+ channel pores. Leucine, the precursor of alpha-ketoisocaproate, did not affect K-ATP, channel activity in excised patches. Our findings are compatible with the view that alpha-ketoisocaproate not only affects the beta-cell stimulus secretion coupling by generation of ATP but also by direct inhibition of the K-ATP, channel.