The effects of imipramine on the Na+ current of guinea-pig ventricular myocytes were examined by the whole-cell clamp method. Imipramine inhibited the Na+ current with a dissociation constant value of 25 microM at a -130 mV holding potential. At 1 microM, imipramine caused a negative shift of the channel availability curve by 4.0 +/- 1.03 mV with its steepness unaffected. The inactivation time constants were not changed by 30 microM imipramine. Paired pulse experiments revealed that imipramine binds to the inactivated Na+ channels with time constants of 3.7 +/- 0.27 sec at -65 mV and 2.4 +/- 0.58 sec at -20 mV, and that it dissociates from the channels with time constants of 5.9 +/- 1.05 sec at -90 mV and 2.0 +/- 0.87 sec at -130 mV. From these paired pulse experiments, the dissociation constant for the interactions between imipramine and inactivated channels was calculated to be 0.67 microM, a value within its therapeutic plasma concentration. These slow interactions of imipramine with inactivated Na+ channels resulted in a slow onset of the frequency-dependent extrablock in the effects of imipramine on the Na+ current. Consequently, the imipramine-induced extrablock sufficient to terminate re-entrant tachyarrhythmias would not develop shortly after their initiation. Short depolarizations of 1- to 3-msec duration sustained appreciable extra blockage when a high concentration of 10 microM imipramine was used, or they were repeatedly applied at a high frequency. However, access of imipramine to the open channels seems to play a minor role in the drug-channel interactions.