We previously reported that hydrogen sulfide (H(2)S) produces protection in PC12 cells during 1-methy-4-phenylpyridinium ion (MPP(+)) challenge. The present study aims to clarify the mechanisms underlying the neuroprotective effects of H(2)S. We showed that both glybenclamide, an ATP-sensitive potassium (K(ATP)) channel blocker, and LY294002, a specific PI(3)K-AKT pathway inhibitor, reversed the neuroprotective effect of NaHS (a H(2)S donor) against MPP(+)-induced cytotoxicity to PC12 cells and that NaHS up-regulated the activity of AKT in PC12 cells, which was abolished by blockade of K(ATP) channels with glybenclamide. In addition, NaHS up-regulated the expression of Bcl-2 and blocked MPP(+)-induced down-regulation of Bcl-2, and this augmentation of Bcl-2 expression was prevented by both glybenclamide and LY294002. These data provided the evidence that the neuroprotective action of H(2)S against MPP(+) toxicity to PC12 cells is via the K(ATP)/PI(3)K/AKT/Bcl-2 pathway. We also demonstrated that NaHS attenuated the inhibitory effect of MPP(+) ERK1/2 activation in PC12 cells, whereas U0126, a specific MEK inhibitor, did not reverse the neuroprotective effect of NaHS, which indicated that attenuating MPP(+)-triggered down-regulation of ERK1/2 activation is involved in the protection of H(2)S against MPP(+) neurotoxicity, but ERK1/2 is not an essential effector mediating the neuroprotective effect of H(2)S. In conclusion, the present observations identify a crucial role of the K(ATP)/PI(3)K/AKT/Bcl-2 pathway in H(2)S-exerted neuroprotection against the toxicity of MPP(+). Findings from the present study will help shed light on the mechanisms of H(2)S-elicited neuroprotective effects on MPP(+) toxicity.