Component of nicotine-induced intracellular calcium elevation mediated through a3- And a5-containing nicotinic acetylcholine receptors are regulated by cyclic AMP in SH-SY 5Y cells

Abstract

The pathway from the medial habenular nucleus to the interpeduncular nucleus, in which nicotinic acetylcholine receptor (nAChR) including the a3 and a5 subunits (a3 * and a5 * nAChRs) are expressed, is implicated in nicotine dependence. We investigated whether a3 * and a5 * nAChRs are regulated by cAMP using SH-SY5Y cells to clarify the significance of these receptors in nicotine dependence. We analyzed the nicotine-induced elevation of intracellular Ca2+ ([Ca2+]i). Nicotine induces a concentration-dependent increase in [Ca2+]i. The elimination of Ca2+ from extracellular fluid or intracellular stores demonstrated that the nicotine-induced [Ca2+]i elevation was due to extracellular influx and intracellular mobilization. The effects of tubocurarine on nicotine-induced [Ca2+]i elevation and current suggest that intracellular mobilization is caused by plasma membrane-permeating nicotine. The inhibition of a3 *, a5 *, a7 nAChR and voltage-gated Ca2+ channels by using siRNAs and selective antagonists revealed the involvement of these nAChR subunits and channels in nicotine-induced [Ca2+]i elevation. To distinguish and characterize the a3 * and a5 * nAChR-mediated Ca2+ influx, we measured the [Ca2+]i elevation induced by nonmembranepermeating acetylcholine when muscarinic receptors, a7nAChR and Ca2+ channels were blocked. Under this condition, the [Ca2+]i elevation was significantly inhibited with a 48-h treatment of dibutyryl cAMP, which was accompanied by the downregulation of a3 and ß4 mRNA. These findings suggest that a3 * and a5 * nAChR-mediated Ca2+ influx is possibly regulated by cAMP at the transcriptional level.