Dronedarone is a noniodinated benzofuran derivative that has been synthesized to overcome the limiting iodine-associated adverse effects of the potent antiarrhythmic drug amiodarone. In this study, the acute electrophysiological effects of dronedarone on repolarizing potassium channels were investigated to determine the class III antiarrhythmic action of this compound. HERG and KvLQT1/minK potassium channels conduct the delayed rectifier potassium current IK in human heart, being a primary target for class III antiarrhythmic therapy. HERG and KvLQT1/minK were expressed heterologously in Xenopus laevis oocytes, and the respective potassium currents were recorded using the two-microelectrode voltage-clamp technique. Dronedarone blocked HERG channels with an IC50 value of 9.2 microM and a maximum tail current reduction of 85.2%. HERG channels were blocked in the closed, open, and inactivated states. The half-maximal activation voltage was shifted by -6.1 mV, and HERG current block by dronedarone was voltage-dependent, but not use-dependent. Dronedarone exhibited a weaker block of KvLQT1/minK currents (33.2% at 100 microM drug concentration), without causing significant changes in the corresponding current-voltage relationships. In conclusion, these data demonstrate that dronedarone is an antagonist of cloned HERG potassium channels, with additional inhibitory effects on KvLQT1/minK currents at higher drug concentrations, providing a molecular mechanism for the class III antiarrhythmic action of the drug.