1. By using acetylcholinesterase (AChE) histochemistry and in vitro 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(-4) M), a nicotine receptor blocker, did not affect the carbachol concentration-response curve, which was significantly inhibited by the muscarinic antagonist, atropine (10(-9)-10(-8) M). 5. The preferential muscarinic antagonists pirenzepine (M1), 11-(2(diethyl-amino)methyl)-1- piperidinylacetyl)-5,-11-dihydro-6H-pyrido(2,3-b)-(1,4)-benzodiaze pine-6-one (AF-DX 116) and methoctramine (M2), 4-diphenyl-acetoxy-N-methyl-piperidine methiodide (4-DAMP) and p-fluorohexahydrosiladiphenidol (p-F-HHSiD) (M3) 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 oesophageal groove seems to be mediated via activation of an M3 postsynaptic muscarinic receptor.