Mitochondrial Glutathione Transport Is a Key Determinant of Neuronal Susceptibility to Oxidative and Nitrosative Stress

• Published in: The Journal of biological chemistry 2013-02, Vol.288 (7), p.5091-5101
• Main Authors: Wilkins, Heather M., Kirchhof, Danielle, Manning, Evan, Joseph, Jamie W., Linseman, Daniel A.
• Format: Article
• Language: English
• Subjects: 2-Oxoglutarate Carrier (OGC, Slc25a11); Animals; Astrocytes - cytology; Astrocytes - metabolism; Biological Transport; Central Nervous System - metabolism; Cerebellum - cytology; Cytosol - metabolism; Dicarboxylate Carrier (DIC, Slc25a10); Free Radicals; Glutathione; Glutathione - metabolism; Malonates - pharmacology; Mitochondria; Mitochondria - metabolism; Mitochondrial Glutathione Transport; Mitochondrial Oxidative Stress; Mitochondrial Transport; Models, Biological; Molecular Bases of Disease; Neurons - metabolism; Nitrogen - metabolism; Nitrosative Stress; Oxidative Stress; Rats; Rats, Sprague-Dawley; Succinates - pharmacology
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.M112.405738

Mitochondrial oxidative stress significantly contributes to the underlying pathology of several devastating neurodegenerative disorders. Mitochondria are highly sensitive to the damaging effects of reactive oxygen and nitrogen species; therefore, these organelles are equipped with a number of free radical scavenging systems. In particular, the mitochondrial glutathione (GSH) pool is a critical antioxidant reserve that is derived entirely from the larger cytosolic pool via facilitated transport. The mechanism of mitochondrial GSH transport has not been extensively studied in the brain. However, the dicarboxylate (DIC) and 2-oxoglutarate (OGC) carriers localized to the inner mitochondrial membrane have been established as GSH transporters in liver and kidney. Here, we investigated the role of these carriers in protecting neurons from oxidative and nitrosative stress. Immunoblot analysis of DIC and OGC in primary cultures of rat cerebellar granule neurons (CGNs) and cerebellar astrocytes showed differential expression of these carriers, with CGNs expressing only DIC and astrocytes expressing both DIC and OGC. Consistent with these findings, butylmalonate specifically reduced mitochondrial GSH in CGNs, whereas both butylmalonate and phenylsuccinate diminished mitochondrial GSH in astrocytes. Moreover, preincubation with butylmalonate but not phenylsuccinate significantly enhanced susceptibility of CGNs to oxidative and nitrosative stressors. This increased vulnerability was largely prevented by incubation with cell-permeable GSH monoethylester but not malate. Finally, knockdown of DIC with adenoviral siRNA also rendered CGNs more susceptible to oxidative stress. These findings demonstrate that maintenance of the mitochondrial GSH pool via sustained mitochondrial GSH transport is essential to protect neurons from oxidative and nitrosative stress. Background: Mitochondrial glutathione transport has not been extensively studied within the CNS. Results: Cerebellar neurons and astrocytes use distinct mechanisms of mitochondrial glutathione transport. Conclusion: Inhibition of a single mitochondrial glutathione transporter renders neurons more susceptible to oxidative and nitrosative stress. Significance: Mitochondrial glutathione transport is essential to protect neurons from oxidative and nitrosative stress conditions common to neurodegenerative diseases.