Role of nitric oxide in ethanol-induced up-regulation of muscarinic acetylcholine receptors in SH-SY5Y cells

Background: Ethanol abuse has been shown to up-regulate muscarinic acetylch oline receptors (mAChRs) in the central nervous system, both in vivo and in vitro, but the detailed mechanisms for this action are not known. Recent s tudies suggest that the actions of ethanol in several biological systems in volve nitric oxide (NO) pathways. The aim of this work was therefore to det ermine whether the effects of long-term ethanol treatment on mAChRs involve d actions upon NO pathways in a well defined cell culture system. Methods: Human neuroblastoma SH-SY5Y cells were used as an in vitro model s ystem. The time- and dose-dependent effects of ethanol on endogenous NO pro duction, as well as the effects of the NO donor 3-morpholino-sydnonimine-ch loride and of the neuronal NO synthase (nNOS) inhibitor N-omega-propyl-L-ar ginine on mAChR number and on ethanol effects upon these receptors, were st udied. Results: Ethanol time- and dose-dependently decreased the production of NO in the cells. Exogenous NO decreased the number of mAChRs and totally block ed the effects of ethanol upon these receptors. Inhibition of nNOS up-regul ated the number of mAChRs, but this effect was not additive to the effects of ethanol. Conclusions: The results of this study suggest that the number of cell-surf ace mAChRs in SH-SY5Y cells may be correlated with changes in NO levels. Th e number of cell surface mAChRs decreased with NO-elevating treatment and i ncreased with NO-lowering treatment. Because ethanol, which is known to up- regulate mAChRs in SH-SY5Y cells, also decreased NO levels and because nNOS inhibition and ethanol effects on mAChRs were not additive, it is conceiva ble that ethanol-induced up-regulation of mAChRs involves inhibition of nNO S.