Hydrogen sulfide (H2S) is a neuromodulator in the brain and a relaxant for smooth muscle. H2S protects primary cortical neurons from oxidative stress by increasing the intracellular concentrations of glutathione, the major antioxidant in cells. However, changes in glutathione alone are not sufficient to account for full protection in all types of nerve cells. H2S is here shown to protect an immortalized mouse hippocampal cell line from oxidative glutamate toxicity by activating ATP-dependent K+ (KATP) and Cl- channels, in addition to increasing the levels of glutathione. The present study therefore identifies a novel pathway for H2S protection from oxidative stress.