Activation of α7 nicotinic acetylcholine receptor augments nerve growth factor action on PCtrk cells.

Abstract

Although cigarette smoking is known to be a critical risk factor for various organ systems and cancers, accumulating evidence indicates that nicotine - a main constituent of cigarette smoking - can exert neuroprotective effects on neuronal cells through nicotinic acetylcholine receptors (nAChRs). However, the precise molecular mechanisms for nicotinic neuroprotective actions remain to be fully elucidated. In this study, we examine the effects of agonists, such as nicotine and PNU282987, on tropomyosin-related kinase (Trk)-dependent neuroprotective pathways in PC12 cells overexpressing a Trk neurotrophin receptor (PCtrk cells). We found that even considerably higher concentrations (mM range for nicotine and µM range for PN282987) of nAChR agonists exert favorable effects, such as the augmentation of nerve growth factor (NGF)-induced Trk neurotrophin receptor autophosphorylation of tyrosine residues and NGF-induced neurite extension. Moreover, nicotine upregulated reactive oxygen species (ROS) levels in the cells. ROS production was completely cancelled by pretreatment with Mito-Tempo, a mitochondria-targeted antioxidant, indicating that the main source of ROS production by nicotine was mitochondria. Furthermore, treatment with nAChR agonists appeared to induce autophagic flux, as evidenced by the upregulation of LC3-II expression in cells. Furthermore, sucrose density ultracentrifugation of nicotine-treated cells clearly disclosed the augmented recruitment of α7nAChR protein into the lipid rafts fraction of the membrane. Intriguingly, a pull-down assay of anti-Trk antibody immunoprecipitates clearly included α7nAChR protein, indicating that Trk and α7nAChR proteins form a complex. These results reveal a new molecular interaction between activated α7nAChR and Trk protein that may serve as a new molecular basis of nicotine-induced neuroprotective action.