Lead-induced alterations of dopaminergic signaling systems in pheochromocytoma cells

Lead, perhaps the oldest known toxicant to man, remains of serious environmental concern because of its low dose toxicity, the variety of adverse health effects it produces on biological systems, and the population at risk of the adverse effects, i.e. children. Lead neurotoxicity in children, caused by chronic, low level lead exposure, manifests as learning and memory deficits, cognitive impairments, and behavioral abnormalities (Needleman, 1996). These findings have been supported by animal studies (Jadhav and Areola, 1997). Alterations in the dopaminergic neurotransmitter system in specific regions of the brain appear to be closely associated with manifestations of behavioral changes (Cory-Slechta, 1998). Low level lead exposure has been associated with alterations of Tyrosine Hydroxylase (TH), the rate limiting enzyme of the dopaminergic biosynthetic pathway. However, the underlying biochemical and molecular events leading to these changes are obscure. Therefore, the objective of this study was to determine the effect of lead on the dopaminergic signaling system (i.e. Calcium Calmodulin dependent TH phosporylation and activity) in Pheochromocytoma Cells. The PC12 cell line is a particularly useful in vitro tool because it synthesizes, stores and releases dopamine in a manner similar to that of neuronal cells (Reams, 1997). Hence, 2 × 106 cells/ml were exposed to 0, .01, .05, .1, .5, 1, 5, 10 μM Pb2+ for 60 min. Total dopamine levels were analyzed via High Performance liquid chromatography. CaMK II α and β, TH and D1, D2 receptor protein expressions were analyzed via Western blot analysis; and CaMK II activity, TH activity and TH phosphorylation were analyzed via immunoradioassays.; This comprehensive study of the dopaminergic pathway revealed that 1μM lead decreased CaMK II β expression and inhibited CaMK II activity. The inhibition of CaMK II activity attenuated TH phosphorylation and TH activity, all of which subsequently lead to decreased dopamine levels. Hence, these findings suggest that lead-induced decrease in dopamine levels in PC12 cells are due to its effects on the CaMK II dependent TH phosphorylation and TH activity. However, close scrutiny of the three significant toxicological levels (NOEL, LOEL, MOEL) of lead's effects on each parameter further indicates that the dynamics of interrelations between these mechanisms may be more complex resulting in the broad spectrum of toxicity of pathways may explain the toxic consequences independent of this neurotransmitter (Supported by ATSDR Cooperative Agreement #U50/ATU 398948).