ATP-dependent regulation of inwardly rectifying K+ current in bovine retinal pigment epithelial cells

Inwardly rectifying K+ current (IKir) in freshly isolated bovine retinal pigment epithelial (RPE) cells was studied in the whole cell recording configuration of the patch-clamp technique. When cells were dialyzed with pipette solution containing no ATP, IKir ran down completely in <10 min [half time (t1/2) = 1.9 min]. In contrast, dialysis with 2 mM ATP sustained IKir for 10 min or more. Rundown was also prevented with 4 mM GTP or ADP. When 0.5 mM ATP was used, IKir ran down by approximately 71%. Mg2+ was a critical cofactor because rundown occurred when the pipette solution contained 4 mM ATP but no Mg2+ (t1/2 = 1.8 min). IKir also ran down when the pipette solution contained 4 mM Mg2+ + 4 mM 5'-adenylylimidodiphosphate (t1/2 = 2.7 min) or 4 mM adenosine 5'-O-(3-thiotriphosphate) (t1/2 = 1.9 min), nonhydrolyzable and poorly hydrolyzable ATP analogs, respectively. We conclude that the sustained activity of IKir in bovine RPE requires intracellular MgATP and that the underlying mechanism may involve ATP hydrolysis.