Objective To silence the expression of α-synuclein in MN9D dopaminergic cells using vector mediated RNA interference (RNAi) and examined its effects on cell proliferation and viability. Methods We identified two 19-nucleotide stretches within the coding region of the α-synuclein gene and designed three sets of oligonuc leotides to generate double stranded (ds) oligos. The ds ofigos were inserted into the pENTRTM/Hl/TO vector and transfected into MN9D dopaminergic cells. α-synuclein expression was detected by RT-PCR, real-time PCR, immunocytochemistry staining and Western blot. Inaddition, we measured cell proliferation using growth curves and cell viability by 3-(4,5)-dimethylthiahiazo (-z-yl)-3,5-diphenytetrazoliumromide (MTT). Results The mRNA and protein levels of α-synuclein gene were significantly down-regulated in pSH2/α-SYN transfected cells compared with control MN9D and pSH/CON-transfected MN9D cells, while pSH1/α-SYN-transfected cells showed no significant difference. Silencing α-synuclein expression does not affect cell proliferation but may decrease cell viability. Conclusion Our results demonstrated pSH2/a-SYN is an effective small interfering RNA (siRNA) sequence and potent silencing of mouse α-synuclein expression in MN9D cells by vector-based RNAi, which provides the tools for studying the normal function of α-synuclein and examining its role in Parkinson's disease (PD) pathogenesis. α-synuclein may be important for the viability of MN9D cells. and loss of α-synuclein may induce cell injury directly or indirectl