Regional differences in repolarisation have been shown to contribute to ventricular arrhythmias in animal models of heart failure, however little is known about their presence and role in the human heart. In this study we investigated regional electrophysiological properties in the failing human ventricle using cardiac slices prepared from the left ventricular free wall (LV) and from the left side of the ventricular septum (LVS). Cardiac slices (300 µm thick) were prepared from ventricular biopsies obtained from end-stage heart failure patients undergoing heart transplant (Camelliti et al, 2011). Slices were point-stimulated at cycle lengths (CL) ranging from 500 ms to 4000 ms, and analysed using a multi-electrode array system. Field potential duration (FPD), an index of action potential duration, was measured. Values are mean ± S.E.M., compared by t-test or ANOVA. FPD was significantly longer in slices from the LVS versus slices from the LV at all pacing cycle lengths (CL 500 ms: 368 ± 13 ms in LVS vs 330 ± 7 ms in LV, n=7 slices/2 hearts, p < 0.05; CL 1000 ms: 498 ± 20 ms in LVS vs 426 ± 19 ms in LV, n=7/2, p < 0.05; CL 2000 ms: 580 ± 27 ms in LVS vs 472 ± 27 ms in LV, n=7/2, p < 0.05; CL 4000 ms: 667 ± 40 ms in LVS vs 520 ± 33 ms in LV, n=7/2, p < 0.05 ). All slices displayed restitution of FPD, with FPD prolongation at longer pacing cycle lengths in both LV and LVS slices (ANOVA, p < 0.0001). FPD/pacing cycle length relationship was steeper in LVS than LV (mean lengthening: 299 ± 26 ms in LVS, 191 ± 26 ms in LV, n=7/2, p < 0.05). Our results indicate that significant electrophysiological differences exists within the failing human ventricle between free wall and septum. These regional heterogeneities may have pro-arrhythmic implications in the failing human heart.