The pulmonary vein is known as an important source of ectopic beats, initiating frequent paroxysms of atrial fibrillation. We compared effects of the class Ic antiarrhythmic drug pilsicainide on the electrophysiological parameters in the isolated pulmonary vein preparation from guinea pigs with those in the left atrium. Three pairs of bipolar electrodes were attached to the left atrium, pulmonary vein, and junctional region of the left atrium and pulmonary vein to measure intra-atrial and intra-pulmonary vein conduction velocity and effective refractory period. Pilsicainide (10 µM) decreased the conduction velocity in the pulmonary vein as well as the left atrium, whose effect on the pulmonary vein was relatively greater than that on the left atrium. The drug prolonged the effective refractory period in the pulmonary vein as well as the left atrium, and the effect of the drug on the pulmonary vein was less than that on the left atrium. The currently observed electrophysiological property of pilsicainide suggests that its effects on reentry within the pulmonary vein are estimated to be weaker than within the left atrium, which may be one of the key considerations for understanding its antiarrhythmic mechanisms in the atrium and pulmonary vein.