We investigated the effect of a specific protein kinase C (PKC) inhibitor, bisindolylmaleimide I [BIM (I)], on L-type Ca2+ channels in rat ventricular myocytes. BIM (I) alone inhibited the L-type Ca2+ current in a concentration-dependent manner, with a Kd value of 3.31±0.25μM, and a Hill coefficient of 2.34±0.23. Inhibition was immediate after applying BIM (I) in the bath solution and then it partially washed out. The steady-state activation curve was not altered by applying 3μM BIM (I), but the steady-state inactivation curve shifted to a more negative potential with a change in the slope factor. Other PKC inhibitors, PKC-IP and chelerythrine, showed no significant effects either on the L-type Ca2+ current or on the inhibitory effect of BIM (I) on the L-type Ca2+ current. The results suggest that the inhibitory effect of BIM (I) on the L-type Ca2+ current is independent of the PKC pathway. Thus, our results should be considered in studies using BIM (I) to inhibit PKC activity and ion channel modulation.