Kn-62, A Specific Ca++ Calmodulin-Dependent Protein-Kinase Inhibitor, Reversibly Depresses The Rate Of Beating Of Cultured Fetal Mouse Cardiac Myocytes

Effects of KN-62 {1-[N,O-bis(5-isoquinolinesulfonyl)-N-methyl-L- tyrosyl]-4-phenylpiperazine}, a specific Ca++/calmodulin (CaM)-dependent protein kinase inhibitor, were examined on the rate of spontaneous beating and the intracellular Ca++ transient of cultured myocytes from fetal mouse ventricle. KN-62 depressed the rate of beating in a dose-dependent fashion. Spontaneous beating ceased 10 min after the administration of 1 mu M KN-62 and recovered gradually after washing with cultured medium. Addition of KN-04 [N-{1-1[P-(5-isoquinolinsulfonyl)benzyl]-2-(4-phenylpiperazinyl)ethyl}-5-isoquinolinsulfonamide; 1 mu M], an analog of KN-62, did not change the rate of beating. In the experiment using an intracellular Ca++ fluorescence indicator, fluo-3, KN-62 depressed the fluo-3 intensity at a systolic phase. The kinase activity to syntide-2 of Ca++/CaM kinase II purified from the rabbit heart was inhibited by KN-62, but not by KN-04. Addition of KN-62 inhibited the phosphorylation of phospholamban by Ca++/CaM kinase II in a dose-dependent manner. KN-62 depressed the Ca++-pumping ATPase activity in the presence of Ca++ and CaM by 32%. These findings indicate that Ca++/CaM kinase II changes an intracellular Ca++ transient and modulates the rate of beating at least in part.