The effects of H-89 (N-[2-(p-bromocinnamylamino)ethyl]-5-isoquinolinesulfonamide), a potent and selective inhibitor of protein kinase A (PKA), were examined on Kv1.3 channels stably expressed in Chinese hamster ovary (CHO) cells using the patch clamp technique. In whole-cell recordings, H-89 decreased Kv1.3 currents and accelerated the decay rate of current inactivation in a concentration-dependent manner with an IC(50) value of 1.70 microM. These effects were completely reversible after washout. Intracellular infusion with PKA inhibitors, adenosine 3', 5'-cyclic phosphorothioate-Rp (Rp-cAMPS) or protein kinase A inhibitor 5-24 (PKI 5-24) had no effect on Kv1.3 currents and did not prevent the inhibitory action of H-89 on the current. H-89 applied to the cytoplasmic surface also inhibited Kv1.3 currents in excised inside-out patches. These findings suggest that H-89 inhibits Kv1.3 currents independently of PKA.