Acetylcholinesterase histochemistry and functional characterization of the muscarinic receptor mediating the contraction of the bovine oesophageal groove

1 By using acetylcholinesterase (AChE) histochemistry and invitro isometric techniques, we have studied the presence and distribution of AChE-positive nerves, as well as the effects of muscarinic cholinoceptor agonists and selective antagonists, in the bovine oesophageal groove. 2 AChE-positive nerves and cells were distributed widely on the oesophageal groove floor. These fibres originated from adventitial ganglia containing bodies with high AChE activity and were shown grouped as large adventitial nerve bundles. 3 Both in the presence and absence of physostigmine, acetylcholine (ACh) induced concentration dependent contractions of bovine oesophageal groove strips. The rank order of the pD₂ values for muscarinic agonists was: oxotremorine-M (7.37) = carbachol (7.14) > acetylcholine plus physostigmine (6.46) > bethanechol (5.42) > McN-A-343 (4.45) > acetylcholine (4.06). 4 Hexamethonium (10^?6–10^?4M ), a nicotinic receptor blocker, did not affect the carbachol concentration–response curve, which was significantly inhibited by the muscarinic antagonist, atropine (10^?9–10^?8M ). 5 The preferential muscarinic antagonists pirenzepine (M₁), 11-(2(-(diethyl-amino)methyl)-1-piperidinylacetyl)-5,-11-dihydro-6H-pyrido(2,3-b)-(1,4)-benzodiazepine-6-one (AF-DX 116) and methoctramine (M₂), 4-diphenyl-acetoxy-N-methyl-piperidine methiodide (4-DAMP) and p-fluoro-hexahydrosiladiphenidol (p-F-HHSiD) (M₃) and tropicamide (M₄) evoked rightwards displacements in a parallel manner of the carbachol control curve, and there was no decrease of the maximum response with the highest concentration of antagonist utilized. The muscarinic antagonist affinities, expressed in terms of pA₂ values, were: atropine (9.51) = 4-DAMP (9.32) > p-F-HHSiD (7.78) > tropicamide (7.40) > pirenzepine (6.91) = AF-DX 116 (6.88) = methoctramine (6.71). This muscarinic antagonist profile suggests that an M₃ receptor is involved in the carbachol induced contraction. 6 The present results suggest that a rich network of AChE- positive fibres is present in the oesophageal groove floor, where they form a nerve trunk and thinner branches accompanying blood vessels and sometimes around ganglia. The muscarinic cholinergic contraction of the bovine oesohageal groove seems to be mediated via activation of an M₃ postsynaptic muscarinic receptor.