Effect of sublethal 6‐hydroxydopamine on the response to subsequent oxidative stress in dopaminergic cells: evidence for preconditioning

Exposure to sublethal stress can trigger endogenous protection against subsequent, higher levels of stress. We tested for this preconditioning phenomenon in a model of Parkinson's disease by applying 6‐hydroxydopamine to the dopaminergic MN9D cell line. Exposure to sublethal concentrations of 6‐hydroxydopamine (5–10 µ m ) protected against the toxic effects of a subsequent exposure to a higher concentration (50 µ m ), as measured by the Hoechst assay for nuclear viability. This was accompanied by little or no protection against 6‐hydroxydopamine‐induced lactate dehydrogenase release, decline in ATP, or reduction in 3 H‐dopamine uptake. The antioxidant, N ‐acetyl cysteine (20 m m ), when applied during preconditioning, abolished protection, as did the protein synthesis inhibitor, cycloheximide (0.2 µ m ). Preconditioning did not affect superoxide dismutase or glutathione peroxidase enzymes, or levels of heat shock protein‐72. However, Bcl‐2 protein levels rose with preconditioning. Preconditioning rapidly increased phosphorylation of kinases ERK1/2, Akt and JNK, and was abolished by pharmacological inhibitors of their activity. Finally, sublethal 6‐hydroxydopamine preconditioned against the toxicity of proteasome inhibitor, MG‐132 (1 µ m ). Thus, exposure of a dopaminergic cell line to sublethal oxidative stress can protect against additional oxidative stress due to translational and post‐translational modifications, as well as confer ‘cross‐tolerance’ against a different insult, proteasome inhibition.