Lead-induced neurotoxicity is modulated by phosphorylation of neuronal nitric oxide synthase by PKA and CaM-KII

초록

This study aimed to identify the neuroprotective mechanism by the phosphorylation of neuronal nitric oxide synthase (nNOS) against Pb-caused cytotoxicity. We have used a neuroblastoma dopaminergic cell line, SH-SY5Y that has constitutive nNOS activity and thus produces NO spontaneously. Treatment of these cells with high concentration (100uM) of lead (Pb) acetate led to the phosphorylation of nNOS at Ser847, as determined by Western-blot analysis with a selective anti-phospho-nNOS antibody. This phosphorylation was accompanied by the decreased reductase activity and the activation of protein kinase A (PKA) and C (PKC) and calcium-calmodulin-dependent kinase II (CaM-KII). Pretreatment of these cells with selective inhibitors of PKA (H-89) or CaM-KII (KN-93) restored NOS activity, while PKC inhibitor (Ro-31-8220) did not affect the phosphorylation of nNOS. The results suggested that CaM-KII or PKA might negatively regulate nNOS to modulate NO production as neuroprotective mechansism against the overproduced NO and/or oxidative stress in brain