Involvement of the M(2) muscarinic receptor in contractions of the guinea pig trachea, guinea pig esophagus, and rat fundus

The involvement of the M(2) muscarinic receptor in contractile responses of the guinea pig trachea, guinea pig esophagus, and rat fundus was investigated. In the standard assay, oxotremorine-M elicited contractions of the trachea with an EC(50) value of approximately 73 nM. [[2-[(Diethylamino)methyl]-1-piperidinyl]acetyl]-5,11 -dihydro-6H-pyrido[2,3-b][1,4]benzodiazepine-6-one (AF-DX 116) at 1 and 10 mu M antagonized these contractions by 2.1- and 9.0-fold increases in the EC(50) value for oxotremorine-M. These effects are consistent with antagonism of an M(3)-mediated contractile response. In subsequent experiments, the M(3) receptors were first inactivated selectively by incubation with N-(2-chloroethyl)-4-piperidinyl diphenylacetate (4-DAMP mustard) (40 nM) for 1 hr in the presence of AF-DX 116 (1 mu M) followed by extensive washing. In 4-DAMP mustard treated trachea, oxotremorine-M elicited contractions with an EC(50) value of 0.31 mu M in the presence of histamine (10 mu M) and forskolin (4 mu M). Under these conditions, AF-DX 116 at 1 and 10 mu M antagonized contractions to oxotremorine-M by 8- and 59-fold increases in the EC(50), respectively, while parafluorohexahydrosiladiphenidol (p-F-HHSiD) (0.1 mu M) had no effect. These effects are consistent with a contraction being mediated by an M(2) receptor. In the guinea pig esophagus and rat fundus, AF-DX 116 and p-F-HHSiD blocked contractions measured under similar conditions with magnitudes intermediate between what would be expected from an M(2) and an M(3) receptor, suggesting that perhaps both subtypes contribute to the overall contractile response under these conditions. In addition, contractions of the guinea pig trachea measured in the presence of histamine and forskolin were pertussis toxin sensitive. These results indicate that, in the trachea, M(3) receptors can dominate the contractile response after a majority of the M(2) receptors have been inactivated, whereas in the guinea pig esophagus and rat fundus, M(2) receptors may contribute to, but do not play a dominant role in the overall response.