Exposure of hepatoma 1c1c7 cells to 2,3-dimethoxy-1,4-naphthoquinone (DMNQ) resulted in a sustained elevation of cytosolic Ca2+, DNA single strand breaks and cell killing. DNA single strand break formation was prevented when cells were preloaded with either of the intracellular Ca2+ chelators, Quin 2 or BAPTA, to buffer the increase in cytosolic Ca2+ concentration induced by the quinone. DMNQ caused marked NAD+ depletion which was prevented when cells were preincubated with 3-aminobenzamide, an inhibitor of nuclear poly-(ADP-ribose)-synthetase activity, or with either of the two Ca2+ chelators. However, 3-aminobenzamide did not protect the hepatoma cells from loss of viability. Our results indicate that quinone-induced DNA damage, NAD+ depletion and cell killing are mediated by a sustained elevation of cytosolic Ca2+.