1 The present study attempted to pharmacologically characterize the muscarinic receptor subtypes mediating contraction of human umbilical vein (HUV). 2 HUV rings were mounted in organ baths and concentration-response curves were constructed for acetylcholine (ACh) (pEC 50 : 6.1670.04; maximum response 80.0071.98% of the responses induced by serotonin 10 mM). The absence of endothelium did not modify the contractile responses of ACh in this tissue. 3 The role of cholinesterases was evaluated: neither neostigmine (acetylcholinesterase inhibitor) nor iso-OMPA (butyrylcholinesterase inhibitor) modified ACh responses. When both enzymes were simultaneously inhibited, a significantly but little potentiation was observed (control: pEC 50 6.3370.03; double inhibition: pEC 50 6.5770.05). 4 Atropine, nonselective muscarinic receptors antagonist, inhibited ACh-induced contraction (pK B 9.67). The muscarinic receptors antagonists pirenzepine (M 1 ), methoctramine (M 2 ) and pFHHSiD (M 3 ) also antagonized responses to ACh. The affinity values estimated for these antagonists against responses evoked by ACh were 7.58, 6.78 and 7.94, respectively. On the other hand, PD 102807 (M 4 selective muscarinic receptors antagonist) was ineffective against ACh-induced contraction. 5 In presence of a blocking concentration of pirenzepine, pFHHSiFD produced an additional antagonism activity on ACh-induced responses. 6 The M 1 muscarinic receptors agonist McN-A-343 produced similar maximum but less potent responses than ACh in HUV. The calculated pA 2 for pirenzepine against McN-A-343 induced responses was 8.54. 7 In conclusion, the data obtained in this study demonstrate the role of M 1 muscarinic receptor subtypes and suggest the involvement of M 3 muscarinic receptor subtypes in ACh-induced vasoconstriction in HUV rings. In addition, the vasomotor activity evoked by ACh does not seem to be modulated by endothelial factors, and their enzymatic degradation appears to have little functional relevance in this tissue.