Specific inhibition of cardiac and skeletal muscle sarcoplasmic reticulum Ca 2+ pumps by H-89

Abstract

The isoquinolinesulfonamide H-89, an inhibitor of cyclic AMP-dependent protein kinases (EC 2.7.1.37, cAPrK), inhibited the Ca 2+ -ATPase activity of cardiac and skeletal muscle sarcoplasmic reticulum (SR) with concentrations giving half-maximal inhibition of 8.1 ± 1.3 and 7.2 ± 0.9 μmol/L, respectively. The effect of H-89 on cardiac SR Ca 2+ -ATPase (EC 3.6.1.38) was the same irrespective of the presence or absence of inhibitors of cAPrK and furthermore, was not affected by a neutralising monoclonal antibody raised against phospholamban. Thus, the action of H-89 in inhibiting SR Ca 2+ -ATPase would not appear to be mediated by inhibition of cAPrK to reduce the phosphorylation state of phospholamban. In both cardiac and skeletal muscle SR, the inhibition by H-89 was noncompetitive with respect to ATP at a low concentration of ATP (< 1 mmol/L) and of a mixed pattern at high concentrations of ATP. H-89 produced a decrease in affinity of the SR Ca 2+ pump to Ca 2+ with an increase in the K m for Ca from 0.52 ± 0.01 to 0.94 ± 0.03 μmol/L ( P < 0.05) in cardiac SR and from 0.39 ± 0.01 to 0.79 ± 0.02 μmol/L ( P < 0.05) in skeletal muscle SR. These results suggest that H-89 inhibits SR Ca 2+ -ATPase by a direct action on the SR Ca 2+ pump to decrease its affinity to Ca 2+. Such an action may contribute to the pharmacological effect of H-89.