To investigate the roles of CuZn superoxide dismutase (CuZnSOD) and Mn superoxide dismutase (MnSOD) in oxygen radical-mediated cytotoxicity and to distinguish the actions of these two enzymes, fetal fibroblasts were derived from mouse fetuses that are either deficient in CuZnSOD (Sod1-/+ and -/-) or MnSOD (Sod2-1+ and -/-) for in vitro studies. Whereas the phenotype of the Sod1 mutant animals did not differ from that of their normal littermates, the growth of Sod1-/- fetal fibroblasts was only 25% of that of the -/+ and +/+ cells. On the other hand, although almost all homozygous Sod2 mutant animals (-/-) died within 10 days after birth, cultivation of Sod2-/- fetal fibroblasts was possible and their growth was about 60% that of -/+ and +/+ cells. When cultured cells were subjected to treatment with paraquat to assess their ability to grow in the presence of high levels of superoxide radicals, Sod1-/- cells were 80 times more sensitive and Sod2-/- cells were 12 times more sensitive to paraquat than wild-type cells. In addition, whereas the loss of 50% CuZnSOD rendered Sod1-/+ cells almost twice more sensitive to paraquat than +/+ cells, loss of 50% MnSOD had no effect on paraquat sensitivity. Our results suggest that CuZnSOD-deficient cells are more sensitive to oxygen toxicity than are MnSOD-deficient cells, that paraquat causes free radical-induced damage in both the mitochondria and cytoplasm, and that SOD compartmentalized in the cytosol cannot compensate for the loss of SOD in the mitochondria and vice versa.