Michel_1989_Naunyn.Schmiedebergs.Arch.Pharmacol_340_62

Kinetic, saturation and competition binding studies were conducted on the muscarinic receptor binding sites labeled by [3H]N-methylscopolamine ([3H]NMS) in membranes prepared from NG108-15 cells. The pharmacology of the NG108-15 cell muscarinic receptors was compared to that of the M1 receptors of rat cortex labeled using [3H]pirenzepine, the M2 and M3 receptors of rat heart and submaxillary gland, respectively, labeled using [3H]NMS and the muscarinic receptors of the PC12 cell line also labeled using [3H]NMS. The rate of dissociation of [3H]NMS from the NG108-15 cell muscarinic receptor was similar to that obtained at the M3 receptor and at the muscarinic receptor of the P12 cells but was slower that the dissociation rate obtained at the M2 cardiac muscarinic receptor. The Kd of [3H]NMS in the NG108-15 cells was significantly lower than that obtained at the M2 and M3 receptor but was similar to the Kd obtained in PC12 cells. In competition studies the affinity estimates for AF-DX 116, 4-DAMP, methoctramine and pirenzepine were not consistent with the presence of either an M1, M2 or M3 receptor but were identical to the affinity estimates obtained at the muscarinic receptor of the PC12 cell line. On the basis of these data we conclude that the muscarinic receptor present in the NG108-15 cells is different to the M1, M2 or M3 subtypes already described but is similar to the muscarinic receptor present in the PC12 cell line. Since NG108-15 cells expresses mRNA for the m4 muscarinic receptor gene described by Bonner et al. (1987) we propose that the muscarinic receptors present in this cell line be denoted as M4 receptors.