Manganese induces sustained Ser40 phosphorylation and activation of tyrosine hydroxylase in PC12 cells

Manganese (Mn2+) is an essential metal involved in normal functioning of a range of physiological processes. However, occupational overexposure to Mn2+ causes neurotoxicity. The dopaminergic system is a particular target for Mn2+ neurotoxicity. Tyrosine hydroxylase (TH) is the rate limiting enzyme for dopamine synthesis and is regulated acutely by phosphorylation at Ser40 and chronically by protein synthesis. In this study we used pheochromocytoma 12 cells to investigate the effects of Mn2+ exposure on the phosphorylation and activity of TH. Mn2+ treatment for 24 h caused a sustained increase in Ser40 phosphorylation and TH activity at a concentration of 100 mu M, without altering the level of TH protein or PC12 cell viability. Inhibition of protein kinase A and protein kinase C and protein kinases known to be involved in sustained phosphorylation of TH in response to other stimuli did not block the effects of Mn2+ on Ser40 phosphorylation. A substantial increase in H2O2 production occurred in response to 100 mu M Mn2+. The antioxidant Trolox(TM) completely inhibited H2O2 production but did not block TH phosphorylation at Ser40, indicating that oxidative stress was not involved. Sustained TH phosphorylation at Ser40 and the consequent activation of TH both occurred at low concentrations of Mn2+ and this provides a potential new mechanism for Mn2+-induced neuronal action that does not involve H2O2-mediated cell death.