UNLABELLED: Since electrical stimulation (ES) can significantly accelerate bone healing, a conductive scaffold that can deliver ES locally at the defect site is desirable for bone defect therapy. Herein, an electrically conductive scaffold was prepared via incorporation of polypyrrole (PPY) in a polycaprolactone (PCL) template scaffold. In vitro tests with mouse osteoblasts indicate that the PPY/PCL scaffold has good biocompatibility, and is suitable for use as an ES substrate. When human adipose-derived mesenchymal stem cells (AD-MSCs) were cultured in the PPY/PCL scaffold and subjected to 200 μA of direct current for 4h per day for 21 days, the amount of calcium deposited was 100% higher than that without ES. When these cells were subjected to ES together with blockers of voltage-gated calcium (Ca(2+)v), sodium (Na(+)v), potassium (K(+)v), or chloride (Cl(-)v) channels, the ES-induced enhancement of AD-MSCs' functions was reduced with Na(+)v, K(+)v, or Cl(-)v blockers and completely nullified with Ca(2+)v blocker. These results indicate that ion fluxes through these channels activated by ES induce different cascades of reactions in the cells, which subsequently regulate AD-MSCs' functions, and Ca(2+)v plays a more critical role than the other three channels. Our results further the current understanding of the mechanisms by which ES regulates stem cells' behavior, and also showed that the conductive PPY/PCL scaffold with application of ES has good potential in bone defect therapy. STATEMENT OF SIGNIFICANCE: In this work, an electrically conductive and biocompatible scaffold was prepared by incorporating polypyrrole in a polycaprolactone template scaffold. Application of 200 μA direct current for 4h per day to human adipose derived-mesenchymal stem cells cultured on this scaffold promoted migration of these cells into the inner region of the scaffold and enhanced their osteogenic differentiation. The roles of voltage-gated ion channels (Ca(2+)v, Na(+)v, K(+)v and Cl(-)v) in osteogenic differentiation stimulated by the electric current were investigated. The results from these experiments further the current understanding of the mechanisms by which electrical stimulation regulates stem cells' behavior, and also show that the polypyrrole-polycaprolactone scaffold with application of electrical stimulation has good potential in bone defect therapy.