Distinct induction of c-fos mRNA in NG108-15 cells transfected with muscarinic m1 and m3 receptors

The differences of intracellular signalling mechanisms between muscarinic acetylcholine m1 and m3 receptors, which are coupled with polyphosphoinositide turnover, were examined by using m1- and m3-transfected NG108-15 cells. The c-fos mRNA was induced by 1 mM acetylcholine peak at 60 min in both m1 and m3 cells. The c-fos induction in m1 cells was inhibited by 1,2-bis(2-aminophenoxy)ethane-N,N,N′,N′-tetraacetic acid tetraacetoxymethyl ester (BAPTA-AM) and N-(6-aminohexyl)-5-chloro-1-naphthalenesulfonamide hydrochloride (W-7), but was not inhibited by prolonged treatment with 12-O-tetradecanoylphorbol 13-acetate (TPA), suggesting that intracellular Ca²⁺ and calmodulin are involved in the induction. The c-fos induction in m3 cells was inhibited by BAPTA-AM and prolonged treatment with TPA, but was not influenced by W-7, suggesting that protein kinase C is mainly involved in m3-induced c-fos expression. Acetylcholine induced an increase in inositol phosphates and a transient increase in the intracellular concentration of Ca²⁺ in both m1 and m3 cells. Sustained stimulation of acetylcholine strongly increased the inositol monophosphate content in m3 cells, but that of inositol triphosphate and inositol diphosphate in m1 cells. These results suggest that the difference between m1- and m3-induced c-fos mRNA induction mechanisms is due to the difference in respective properties in polyphosphoinositide turnover.