Desipramine is a tricyclic antidepressant for psychiatric disorders that can induce QT prolongation, which may lead to torsades de pointes. Since blockade of cardiac human ether-a-go-go-related gene (hERG) channels is an important cause of acquired long QT syndrome, we investigated the acute effects of desipramine on hERG channels to determine the electrophysiological basis for its pro-arrhythmic potential. We examined the effects of desipramine on the hERG channels expressed in Xenopus oocytes using two-microelectrode voltage-clamp techniques. Desipramine-induced concentration-dependent decreases in the current amplitude at the end of the voltage steps and hERG tail currents. The IC(50) for desipramine needed to block the hERG current in Xenopus oocytes decreased progressively relative to the degree of depolarization. Desipramine affected the channels in the activated and inactivated states but not in the closed states. The S6 domain mutations, Tyr-652 located in the S6 domain of the hERG channel reduced the potency of the channel block by desipramine more than a mutation of Phe-656 in the same region. These results suggest that desipramine is a blocker of the hERG channels, providing a molecular mechanism for the arrhythmogenic side effects during the clinical administration of desipramine.