A novel type of ATP block on a Ca2+-activated K+ channel from bullfrog erythrocytes

Using the patch-clamp technique, we have identified an intermediate conductance Ca2+-activated K+ channel from bullfrog (Rana catesbeiana) erythrocytes and have investigated the regulation of channel activity by cytosolic ATP, The channel was highly selective for K+ over Na+, gave a linear I-V relationship with symmetrical 117.5 mM K+ solutions and had a single-channel conductance of 60 pS, Channel activity was dependent on Ca2+ concentration (K-1/2 = 600 nM) but voltage-independent, These basic characteristics are similar to those of human and frog erythrocyte Ca2+-activated K+ (Gardos) channels previously reported. However, cytoplasmic application of ATP reduced channel activity with block exhibiting a novel bell-shaped concentration dependence. The channel was inhibited most by similar to 10 mu M ATP (P-O reduced to 5% of control) but less blocked by lower and higher concentrations of ATP, Moreover, the novel type of ATP block did not require Mg2+, was independent of PKA or PKC, and was mimicked by a nonhydrolyzable ATP analog, AMP-PNP. This suggests that ATP exerts its effect by direct binding to sites on the channel or associated regulatory proteins, but not by phosphorylation of either of these components.