ON THE MECHANISM OF NUCLEOTIDE DIPHOSPHATE ACTIVATION OF THE ATP-SENSITIVE K+ CHANNEL IN VENTRICULAR CELL OF GUINEA-PIG

1. Effects of intracellular nucleotide diphosphates (NDPs) on the ATP-sensitive K+ channel (K(ATP)+ channel) were examined in ventricular cells of guinea-pig heart, using the inside-out patch clamp technique. On formation of inside-out patches in the ATP-free internal solution, the K(ATP)+ channel appeared and then ran down spontaneously. This run-down of the K(ATP)+ channel activity was probably due to dephosphorylation. 2. Millimolar concentrations of various NDPs, e.g. UDP (uridine diphosphate), IDP (inosine diphosphate), CDP (cytidine diphosphate) and GDP (guanosine diphosphate), applied to the internal side of the patch membrane, induced openings of the K(ATP)+ channel after run-down, i.e. in the dephosphorylated state. ADP opened the channel weakly at low concentrations (100-mu-M) but inhibited it at higher concentrations (1-10 mM). 3. NDP-induced openings of the channel were Mg2+ dependent and inhibited by ATP (100-mu-M) and glibenclamide (1-mu-M). None of nucleosides, nucleotide monophosphates nor nucleotide triphosphates induced openings of the channel. Thus, the K(ATP)+ channel may have a Mg2+-dependent NDP-binding site, which induces openings of the dephosphorylated channel in ATP-free solution, in addition to the Mg2+-independent ATP-binding inactivation site and phosphorylation site. 4. In inside-out patches, pinacidil (a K(ATP)+ channel opener) activated the K(ATP)+ channel in the phosphorylated state but not in the dephosphorylated state. In the presence of NDPs (UDP, IDP, CDP, GDP), however, pinacidil (30-mu-M) enhanced openings of the dephosphorylated K(ATP)+ channel prominently. 5. From the above results, we concluded that NDP-binding to the specific site has similar effects to channel phosphorylation, i.e. it keeps the K(ATP)+ channel in an operative state in ATP-free solution and enhances the pinacidil-induced channel openings.