Chloride channel activity is one of the critical factors responsible for cell apoptotic volume decrease (AVD). However, the roles of chloride channels in apoptosis have not been fully understood. In the current study, we assessed the role of chloride channels in hydrogen peroxide (H2O2)-induced apoptosis of pheochromocytoma cells (PC12). Extracellular application of H2O2 activated a chloride current and induced cell volume decrease in a few minutes. Incubation of cells with H2O2 elevated significantly the membrane permeability to the DNA dye Hoechst 33258 in 1 h and induced apoptosis of most PC12 cells tested in 24 h. The chloride channel blocker NPPB (5-nitro-2-(3-phenylpropylamino)-benzoate) prevented appearance of H2O2-induced high membrane permeability and cell shrinkage, suppressed H2O2-activated chloride currents and protected PC12 cells from apoptosis induced by H2O2. The results suggest that chloride channels may contribute to H2O2-induced apoptosis by ways of elevation of membrane permeability and AVD in PC12 cells. (C) 2009 Elsevier Inc. All rights reserved.