The Interaction of Manganese Nanoparticles with PC-12 Cells Induces Dopamine Depletion

Abstract

This investigation was designed to determine whether nano-sized manganese oxide (Mn-40nm) particles would induce dopamine (DA) depletion in a cultured neuronal phenotype, PC-12 cells, similar to free ionic manganese (Mn²⁺). Cells were exposed to Mn-40nm, Mn²⁺ (acetate), or known cytotoxic silver nanoparticles (Ag-15nm) for 24 h. Phase-contrast microscopy studies show that Mn-40nm or Mn²⁺ exposure did not greatly change morphology of PC-12 cells. However, Ag-15nm and AgNO₃ produce cell shrinkage and irregular membrane borders compared to control cells. Further microscopic studies at higher resolution demonstrated that Mn-40nm nanoparticles and agglomerates were effectively internalized by PC-12 cells. Mitochondrial reduction activity, a sensitive measure of particle and metal cytotoxicity, showed only moderate toxicity for Mn-40nm compared to similar Ag-15nm and Mn²⁺ doses. Mn-40nm and Mn²⁺ dose dependently depleted DA and its metabolites, dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA), while Ag-15nm only significantly reduced DA and DOPAC at concentrations of 50 μg/ml. Therefore, the DA depletion of Mn-40nm was most similar to Mn²⁺, which is known to induce concentration-dependent DA depletion. There was a significant increase (> 10-fold) in reactive oxygen species (ROS) with Mn-40nm exposure, suggesting that increased ROS levels may participate in DA depletion. These results clearly demonstrate that nanoscale manganese can deplete DA, DOPAC, and HVA in a dose-dependent manner. Further study is required to evaluate the specific intracellular distribution of Mn-40nm nanoparticles, metal dissolution rates in cells and cellular matrices, if DA depletion is induced in vivo, and the propensity of Mn nanoparticles to cross the blood-brain barrier or be selectively uptaken by nasal epithelium.