Background: A short-long-short sequence of cycle lengths predisposes to reentrant tachyarrhythmias. There is limited information about the effects of premature ventricular contractions (PVCs) on repolarization of postextrasystolic depolarizations (PEDs). Such information would contribute to understanding the mechanism for facilitating reentry with short-long short cycle lengths. Methods and Results: We introduced PVCs, over a range of coupling intervals and during a range of basic drive cycle lengths (BCLs), and determined PED repolarization. Our results from whole-animal experiments, isolated cell studies, and computer simulations are reported. In the whole- animal experiments, PED refractory periods (RPs) were longer than RP(BCT). The greatest difference between RP(PED) and RP(BCT) (ΔRP(max)) occurred after short coupling interval PVCs and was 4.3±0.8, 4.2±0.8, and 2.1±0.5 ms (mean±SEM) during drives with short, intermediate, and long BCLs, respectively. The diastolic interval preceding the PED (DI(PED)) was inversely related to the coupling interval between the basic drive beat and the PVC and directly related to RP(PED) PED action potential durations (APDs) of isolated canine myocytes were 9.8±4.9 ms (mean±SEM) longer than APD BCL (n = 19). The DiFrancesco-Noble membrane equations were used in simulations of action potential propagation in a one-dimensional cable, with stimulation protocols duplicating those in the animal experiments. PVCs prolonged APD(PED), and APD(PED) was prolonged more during short than during long BCL drives. There was a direct relation between DI(PED) and APD(PED). Analysis of the membrane currents over the time course of the PVCs and PEDs suggested that the ionic basis for PED repolarization prolongation was the interaction of I(to) and I(K). Hyperpolarizing constant-current injections introduced immediately after the spike of isolated myocyte action potentials caused APD prolongation. This observation is consistent with the I(to) and I(K) interaction causing PED repolarization prolongation. Conclusions: PED repolarization prolongation could provide sites for unidirectional block to propagation of PVCs after PEDs and could facilitate initiation of reentrant tachyarrhythmias after short-long-short sequences of cycle lengths.