A variety of neurons in gastrointestinal and genitourinary smooth muscle express muscarinic auto- as well as heteroreceptors. These receptors are found on the soma and dendrites of many cholinergic, sympathetic and NANC neurons and on axon terminals. A given neuron may contain both excitatory and inhibitory presynaptic muscarinic receptors. The subtypes involved are species- and tissue-dependent, and neuronal m 1 to M 4 receptors have been shown to be expressed in smooth muscle tissues. In this study, the ability of several selective muscarinic receptor antagonists to inhibit the effect of arecaidine propargyl ester (APE) on prejunctional muscarinic receptors on sympathetic nerve endings in the rabbit anococcygeus muscle (RAM) was investigated to characterise the receptor subtype involved. Electrical field stimulation (EFS) resulted in a release of noradrenaline (NA) eliciting monophasic contractions due to stimulation of postjunctional α 1-adrenoceptors. The selective muscarinic agonist APE did not reduce contractions to exogenous NA, but caused a concentration-related and N-methylatropine-sensitive inhibition of neurogenic responses. All muscarinic antagonists investigated failed to affect the EFS-induced contractions, but shifted the concentration-response curve of APE to the right in a parallel and surmountable fashion. Schild analysis yielded regression lines of unit slope, indicating competitive antagonism. The following rank order of antagonist potencies (pA 2 values) was found: tripitramine (9.10) > AQ-RA 741 (8.26) ≥ himbacine (8.04) ≥ (S)-dimethindene (7.69) ≥ pirenzepine (6.46) ≥ p-F-HHSiD (6.27). A comparison of the pA 2 values determined in the present study with literature binding and functional affinities obtained at native or recombinant m 1 to M 5 receptors strongly suggests that NA release from sympathetic nerve endings in RAM is inhibited by activation of prejunctional muscarinic M 2 receptors.