Oxidative stress on EAAC1 is involved in MPTP-induced glutathione depletion and motor dysfunction

Excitatory amino acid carrier 1 (EAAC1) is a glutamate transporter expressed on mature neurons in the CNS, and is the primary route for uptake of the neuronal cysteine needed to produce glutathione (GSH). Parkinson's disease (PD) is a neurodegenerative disorder pathogenically related to oxidative stress and shows GSH depletion in the substantia nigra (SN). Herein, we report that 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mice, an experimental model of PD, showed reduced motor activity, reduced GSH contents, EAAC1 translocation to the membrane and increased levels of nitrated EAAC1. These changes were reversed by pre-administration of n-acetylcysteine (NAC), a membrane-permeable cysteine precursor. Pretreatment with 7-nitroindazole, a specific neuronal nitric oxide synthase inhibitor, also prevented both GSH depletion and nitrotyrosine formation induced by MPTP. Pretreatment with hydrogen peroxide, L-aspartic acid beta-hydroxamate or 1-methyl-4-phenylpyridinium reduced the subsequent cysteine increase in midbrain slice cultures. Studies with chloromethylfluorescein diacetate, a GSH marker, demonstrated dopaminergic neurons in the SN to have increased GSH levels after NAC treatment. These findings suggest that oxidative stress induced by MPTP may reduce neuronal cysteine uptake, via EAAC1 dysfunction, leading to impaired GSH synthesis, and that NAC would exert a protective effect against MPTP neurotoxicity by maintaining GSH levels in dopaminergic neurons.