The location of conduction abnormality including activation slowing and wavefront discontinuity, which often corresponds to the critical zones of reentrant circuit in scar-mediated ventricular tachycardia (VT), is affected by the wavefront direction during substrate mapping. We hypothesized that an abrupt change in paced-QRS morphologies may occur at the region with conduction abnormality. This study assessed the conduction velocity in substrate mapping during at least two rhythms at the region where an abrupt change in paced-QRS morphologies was obtained. In a retrospective review of 57 patients who underwent functional substrate mapping for scar-mediated VT, 21 patients underwent it during multiple rhythms, that is intrinsic rhythm and paced rhythm with high-density pace-mapping in the area of interest. In 10 of these patients, those critical sites, as determined by direct termination during ablation (4 VTs, 33%) or pace-mapping findings (8 VTs, 67%) were identified in a total of 12 VTs. Conduction velocities during an intrinsic rhythm or paced rhythm at the isthmus area were assessed based on the difference of local activation time between two sites along the site where pace-mapping scores changed abruptly. Pace-mapping scores, which were obtained at two adjacent sites along the presumed isthmus, showed 92% [interquartile range [IQR] 89-96%] at the exit side and 48% [IQR 41-56%] at the adjacent entrance side. Ten patients (71±9 years, 9 (90%) male, 6 (60%) nonischemic) underwent high-density substrate mapping during multiple rhythms, that is intrinsic rhythm including sinus rhythm (n=7) or right ventricular pacing (n=5) and paced rhythm including right (n=5) or left ventricular pacing (n=7). Conduction velocity during intrinsic rhythm at the location of abrupt pacemap change was 0.37m/sec [IQR 0.19-0.43 m/sec], which was no slower than 0.25 m/sec, defined as activation slowing in previous studies. However, conduction velocity at abrupt pacemap change site that was faster in one of the multiple rhythms, was significantly slowed down when changing to the other rhythm (0.41 m/sec [IQR 0.25-1.05 m/sec] vs. 0.09 m/sec [IQR0.07-0.22 m/sec], P<0.01) (Fig.1). The abrupt change in paced-QRS morphology in a novel high-density pace-mapping strategy, which is shown independent of wavefront direction during substrate mapping, can identify the high-yield regions with conduction abnormality being critical for reentry.