1 The objective of the study was to determine the role of muscarinic receptor subtypes in mediating contraction of the porcine detrusor smooth muscle in vitro. 2 Strips of pig detrusor muscle were set up in physiological salt solution and the tensions developed by the tissues were recorded. Responses to carbachol were obtained in the absence and presence of a range of muscarinic antagonists (4-DAMP, methoctramine, darifenacin, oxybutynin, tolterodine and pirenzepine). Antagonist affinity values (pKB values) were calculated and compared with those quoted in the literature for these antagonists at each of the muscarinic receptor subtypes. 3 The M3-selective antagonists, 4-DAMP and darifenacin had high affinities (pKB values of 9.4 and 8.6, respectively). Oxybutynin, tolterodine and pirenzepine had affinities of 8.2, 8.1 and 6.8, respectively, whilst the M2-selective agent methoctramine had a relatively low affinity (pKB = 6.1). The rank order of affinities was, therefore, 4-DAMP > darifenacin > oxybutynin > tolterodine > pirenzepine > methoctramine for the pig detrusor. Correlation of the antagonist affinities obtained on the bladder with those published for these antagonists at the five muscarinic receptor subtypes identified the M3(m3)-receptor as the muscarinic subtype mediating detrusor contractile responses in vitro. 4 These data suggest that a small population of M3-muscarinic receptors must mediate direct contractile responses of the pig detrusor muscle to muscarinic receptor stimulation in vitro.