Excessive extracellular zinc may contribute to neuronal cell death following ischemia and seizures, although the mechanisms mediating zinc-induced cell death remain largely unknown. In this study, we examined potential cellular and molecular mechanisms associated with zinc neurotoxicity and determined the neuroprotective effects of the superoxide dismutase (SOD)/catalase mimetic, EUK-134. Cortical neuron cultures exposed to zinc for 24 h exhibited concentration-dependent increases in lactate dehydrogenase (LDH) release and number of apoptotic cell bodies. Both effects were prevented by treatment with EUK-134. Zinc exposure resulted in increased release of cytochrome c from the mitochondria into the cytosol. Treatment with EUK-134 blocked this parameter of mitochondrial dysfunction. Exposure of cultures to zinc for 4 h produced an elevation of reactive oxygen species (ROS) as determined by increased 2,7-dichlorofluorescein (DCF) fluorescence, which was followed by an increase in lipid peroxidation. EUK-134 completely attenuated ROS production and subsequent oxidative damage. Finally, zinc exposure activated NF-kappaB, an effect also prevented by EUK-134. These data indicate that multiple cellular and molecular mechanisms are involved in zinc neurotoxicity. As all these mechanisms appear to be sensitive to treatment with EUK-134, our data suggest that oxidative stress occurs early in the cascade of events triggered by zinc.