It is well documented that manganese neurotoxicity induces clinical symptoms similar to those of idiopathic Parkinson's disease. Although microglial cytotoxic mediator-induced neurotoxicity is suggested, the mechanism by which manganese up-regulates cytotoxic mediator, such as nitric oxide (NO), remains poorly understood. Therefore, in this study, we investigated the mechanism of manganese on induction of iNOS in microglial cells. iNOS promoter/luciferase assay revealed that manganese (500 (M) regulated the iNOS expression at the transcriptional level. Immunoblot analysis also revealed that phosphorylation levels of ERK, JNK MAPKs and Akt (PKB, PI 3-kinase downstream effector), were increased. Both protein and mRNA levels of iNOS expression were abrogated by specific inhibitors, SP600125 (JNK inhibitor, 20 microM), PD98059 (ERKs inhibitor, 50 microM), or LY294002 (PI 3-kinase inhibitor, 20 microM), but not by SB203580 (20 microM), a p38 specific inhibitor. These data lead to the conclusion that manganese regulates the iNOS expression at the transcriptional level in BV2 microglial cells and the increased iNOS protein expression is mediated via both JNK-ERK MAPK and PI3K/Akt signaling pathways, but not via p38 MAPK pathway. Increased iNOS protein level was also found in RAW264.7 murine macrophage cells.