Abrupt changes in cycle length (CL) occur frequently in the clinical setting of atrial fibrillation. However, the effects of such changes on the His-Purkinje system (HPS) have not previously been considered during aberrant ventricular conduction (VAb). In 12 patients who manifested VAb with atrial premature stimulation (A2) during sinus rhythm, the relative refractory period (RRP) of the HPS was evaluated during a constant atrial CL (method I) and during abrupt changes in the CL (method II), wherein the A2 was coupled to an atrial CL (last A1A1) comparable to method I. This last A1A1 during method II was preceded by a series of constant atrial CLs 100-200 msec longer (method IIA) in 11 of 12 patients, or 109-300 msec shorter (method IIB) in all 12 patients. Although abrupt changes in the atrial CL using method IIA resulted in a longer HH interval (by 0-30 msec; mean 13.2 +/- 9.2 msec) preceding the A2, the RRP-HPS was 5-20 msec shorter (mean 9.3 +/- 5.3 msec) compared with method I in eight patients. The effect of abrupt changes was further evaluated in nine patients using method III, with a constant atrial CL, with a duration equal to the last H1H1 interval of method IIA. The VAb that occurred with method III was not manifested with method IIA in seven of nine patients, and in two patients the RRP-HPS was the same or less. Conversely, method IIB resulted in a shorter HH interval (by 0.110 msec; mean 28.9 +/- 21.1 msec) preceding A2, but in 10 patients, RRP-HPS was 5-40 msec longer (mean 20.7 +/- 10.5 msec) than that of method I and in two, VAb was only manifested using method IIB. Further scanning with method III, derived from the HH interval immediately preceding A2 of method IIB, was performed in seven patients and compared with method IIB. Prolongation in the RRP-HPS was shown using the latter method. The results indicate that abrupt changes in CL influence the functional behavior of the HPS in a manner not anticipated. Such changes may have important implications in determining the occurrence of VAb during atrial fibrillation.