The Fallopian tubes are sparsely innervated with cholinergic nerve fibers. Acetylcholine is released from these nerves and contracts the smooth muscles of the tubes. The aim of our study was to investigate the effect of acetylcholine on the isthmic segment of the Fallopian tubes using selective antagonists in different hormonal settings. We investigated the effects of acetylcholine on the isolated isthmus of Fallopian tubes taken from 83 patients during abdominal hysterectomy with adnexectomy. Twenty-eight patients were in the follicular phase, 36 were in the luteal phase of the menstrual cycle, and 19 patients were in menopause. Selective and non-selective muscarinic and nicotinic receptor antagonists were used. Acetylcholine (1.8-658.6 micro M) produced concentration-dependent tonic contraction of isthmus taken from the patients in the follicular phase, the luteal phase and menopause. The nicotinic receptor antagonist mecamylamine (6.5 micro M) and local anesthetic lidocaine (230.8 micro M) did not alter the effect of acetylcholine. While M(1) and M(2)-selective muscarinic receptor antagonists pirenzepine (1.6 micro M) and methoctramine (0.9 micro M) did not show specific effect, atropine (0.01 micro M) and the selective M(3)-receptor antagonist p-fluoro-hexahydro-sila-difenidol (pFHHSiD; 0.2 micro M) effectively blocked contractions caused by acetylcholine (maximal pA(2) values 9.74 and 7.54, respectively). The affinity of pFHHSiD for muscarinic receptors was highest in the follicular phase. The results of our study suggest the existence of functional M(3) muscarinic receptors in the isthmus of the Fallopian tubes, located on the smooth muscle cells.