α-Synuclein, a gene whose mutations, duplication, and triplication has been linked to autosomal dominant familial Parkinson's disease (fPD), appears to play a central role in the pathogenesis of sporadic PD (sPD) as well. Enhancement of neurodegeneration induced by mutant α-synuclein has been attributed to date largely to faster formation of α-synuclein aggregates in neurons. Recently, we reported that microglial activation enhances wild type (WT) α-synuclein-elicited dopaminergic neurodegeneration. In the present study, using a primary mesencephalic culture system, we tested whether mutated α-synuclein could activate microglia more powerfully than WT α-synuclein, thereby contributing to the accelerated neurodegeneration observed in fPD. The results showed that α-synuclein with the A30P or A53T mutations caused greater microglial activation than WT α-synuclein. Furthermore, the extent of microglial activation paralleled the degree of dopaminergic neurotoxicity induced by WT and mutant α-synuclein. Mutant α-synuclein also induced greater production of reactive oxygen species than WT α-synuclein by NADPH oxidase (PHOX), and PHOX activation was linked to direct activation of macrophage antigen-1 (Mac-1) receptor, rather than α-synuclein internalization via scavenger receptors. These results have, for the first time, demonstrated that microglia are also critical in enhanced neurotoxicity induced by mutant α-synuclein. © 2007 Wiley-Liss, Inc.