Roles of Autophagy in MPP(+)-Induced Neurotoxicity In Vivo: The Involvement of Mitochondria and α-Synuclein Aggregation

Macroautophagy (also known as autophagy) is an intracellular self-eating mechanism and has been proposed as both neuroprotective and neurodestructive in the central nervous system (CNS) neurodegenerative diseases. In the present study, the role of autophagy involving mitochondria and α-synuclein was investigated in MPP(+) (1-methyl-4-phenylpyridinium)-induced oxidative injury in chloral hydrate-anesthetized rats in vivo. The oxidative mechanism underlying MPP(+)-induced neurotoxicity was identified by elevated lipid peroxidation and heme oxygenase-1 levels, a redox-regulated protein in MPP(+)-infused substantia nigra (SN). At the same time, MPP(+) significantly increased LC3-II levels, a hallmark protein of autophagy. To block MPP(+)-induced autophagy in rat brain, Atg7siRNA was intranigrally infused 4 d prior to MPP(+) infusion. Western blot assay showed that in vivo Atg7siRNA transfection not only reduced Atg7 levels in the MPP(+)-infused SN but attenuated MPP(+)-induced elevation in LC3-II levels, activation of caspase 9 and reduction in tyrosine hydroxylase levels, indicating that autophagy is pro-death. The immunostaining study demonstrated co-localization of LC3 and succinate dehydrogenase (a mitochondrial complex II) as well as LC3 and α-synuclein, suggesting that autophagy may engulf mitochondria and α-synuclein. Indeed, in vivo Atg7siRNA transfection mitigated MPP(+)-induced reduction in cytochrome c oxidase. In addition, MPP(+)-induced autophagy differentially altered the α-synuclein aggregates in the infused SN. In conclusion, autophagy plays a prodeath role in the MPP(+)-induced oxidative injury by sequestering mitochondria in the rat brain. Moreover, our data suggest that the benefits of autophagy depend on the levels of α-synuclein aggregates in the nigrostriatal dopaminergic system of the rat brain.