Coupling of m2 and m4 muscarinic acetylcholine receptor subtypes to Ca²⁺-dependent K⁺ channels in transformed NL308 neuroblastoma x fibroblast hybrid cells

Muscarinic acetylcholine receptor (mAChR) subtype (m1-m4)-specific cDNAs were transfected into NL308 neuroblastoma-fibroblast hybrid cells and clones expressing each of the individual mAChR subtypes m1, m2, m3 and m4 obtained. Acetylcholine increased phosphoinositide (PI) turnover in m1- and m3-transformed cells, but did not produce detectable changes in m2- and m4-transformed cells. In cells expressing m1 and m3 subtypes, ACh produced an initial outward K⁺ current, followed by a cationic current. In cells expressing m2 and m4 receptors, only the initial K⁺ current was detected. The outward currents were associated with a rise in intracellular Ca²⁺ as measured with Fura-2 or Indo-1, and were inhibited by chelating intracellular Ca²⁺ with external BAPTA-AM, or by external charybdotoxin or Ba²⁺: hence they were attributed to the activation of a Ca²⁺-dependent K⁺ current. However, the outward current produced in m2- and m4-transformed cells was blocked by pretreatment with 5 ng ml^-3 Pertussis toxin (PTX), whereas that in m1- and m3-transformed cells was not. These results suggest that m2- and m4-receptors in transformed NL308 cells coupled to PTX-sensitive G-protein which is capable of mobilizing intracellular Ca²⁺ and activate I(K(Ca)) whereas m1 and m3 receptors activate a similar process through a different, PTX-insensitive G-protein.