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1 In strips of human isolated detrusor muscle, the 5-hydroxytryptamine (5-HT) receptor (5-HT4) that mediates facilitation of neuromuscular cholinergic transmission was further characterized by using 5-HT and a series of ligands known for their 5-HT4 agonist (5-methoxytryptamine: 5-MeOT, cisapride, (R,S)-zacopride, BIMU 8) or antagonist (RS 23597, GR 125487, DAU 6285) properties. 4 Thus, in the human isolated detrusor muscle, the 5-HT4 receptors mediating facilitation of cholinergic neuromuscular transmission are activated by indoleamines (5-HT, 5-MeOT), substituted benzamide (cisapride, (R,S)-zacopride), benzoate (RS 23597) and benzimidazolone (BIMU 8) derivatives. The activities (in terms of both potency and efficacy) of most agonists, as well as the affinity estimates of the antagonists GR 125487 and DAU 6285, are comparable to those found in other peripheral tissues. Exceptions are RS 23597, which acted either as a partial agonist or as an antagonist of the response to 5-HT, and 5-MeOT that showed an unusually low potency. The latter findings may be ascribed to differences in the efficiency of receptor coupling mechanisms and/or in the molecular structure (i.e. splice variants) of the 5-HT4 receptor.

1 The effects of ten muscarinic antagonists on electrically evoked [3H]-acetylcholine release and muscle contraction were compared in an epithelium-free preparation of the guinea-pig trachea that had been preincubated with [3H]-choline. 2 The M3-selective antagonists UH-AH 37, 4-diphenyl-acetoxy-N-piperidine methobromide and parafluorohexahydrosiladiphenidol were more potent in reducing the contractile response than in facilitating the evoked [3H]-acetylcholine release. Hexahydrosiladiphenidol did not discriminate between pre-and postjunctional effects. The rank order of the postjunctional potencies of the ten antagonists as well as the postjunctional pA2 values obtained for hexahydrosiladiphenidol (7.95) and AQ-RA 741 (7.08) identified the muscular receptor as an M3 subtype. 3 The M2-selective antagonists methoctramine, AF-DX 116 and AQ-RA 741 were more potent in facilitating the evoked [3H]-acetylcholine release than in inhibiting the contractile response. The increase in release by low concentrations of methoctramine, AF-DX 116 and AQ-RA 741 was paralleled by an enhancement of the stimulation-evoked contractions.4 Comparison of the pre-and postjunctional potencies of the Ml-, M2-and M3-selective antagonists suggests that autoinhibition of acetylcholine release is mediated via an 'M2-like' receptor which differs from the cardiac type M2 receptor in its relatively high affinity for hexahydrosiladiphenidol.

1 Experiments were carried out in human detrusor strips to characterize muscarinic receptor subtypes involved in the prejunctional regulation of acetylcholine (ACh) release from cholinergic nerve terminals, and in the postjunctional smooth muscle contractile response. 2 In detrusor strips preincubated with [ 3 H]-choline, electrical ®eld stimulation (600 pulses) delivered in six trains at 10 Hz produced a tritium out¯ow and a contractile response. In the presence of 10 mM paraoxon (to prevent ACh degradation) the tritium out¯ow was characterized by HPLC analysis as [ 3 H]-ACh (76%) and [ 3 H]-choline (24%).3 Electrically-evoked [ 3 H]-ACh release was abolished by tetrodotoxin (TTX: 300 nM) and una ected by hexamethonium (10 mM), indicating a postganglionic event. It was reduced by physostigmine (100 nM) and the muscarinic receptor agonist, muscarone (10 nM ± 1 mM), and enhanced by atropine (0.1 ± 100 nM). These ®ndings indicate the presence of a muscarinic negative feedback mechanism controlling ACh release. 4 The e ects of various subtype-preferring muscarinic receptor antagonists were evaluated on [ 3 H]-ACh release and muscle contraction. The rank potency (7log EC 50 ) orders at pre-and postjunctional level were: atropine 54-diphenyl-acetoxy-N-piperidine (4-DAMP)4mamba toxin 3 (MT-3)4tripitramine4para-¯uorohexahydrosiladiphenidol (pF-HHSiD)5methoctramine5pirenze-pine4tripinamide, and atropine54-DAMP4pF-HHSiD> >pirenzepine=tripitramine4tripinami-de4methoctramine> >MT-3, respectively. 5 The comparison of pre-and post-junctional potencies and the relationship analysis with the a nity constants at human cloned muscarinic receptor subtypes indicates that the muscarinic autoreceptor inhibiting ACh release in human detrusor is an M 4 receptor, while the receptor involved in muscular contraction belongs to the M 3 subtype.

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Bladder overactivity (OAB) is a chronic and debilitating lower urinary tract (LUT) disorder that affects millions of individuals worldwide. LUT symptoms associated with OAB, such as urgency and urinary incontinence, cause a hygienic and social concern to patients, but their current pharmacological treatment is largely inadequate due to the lack of uroselectivity. Although OAB etiology remains multifactorial and poorly understood, increasing evidence indicates that serotonin [5-hydroxytryptamine (5-HT)] is an endogenous substance involved in the control of micturition at central and peripheral sites. In this study, we demonstrated the presence of three distinct 5-HT receptors localized at parasympathetic nerve terminals of the human bladder by measuring electrically evoked tritiated acetylcholine release in isolated detrusor strips. These prejunctional receptors, involved in both positive and negative feedback mechanisms regulating cholinergic transmission, have been characterized by means of three highly selective 5-HT antagonists for 5-HT 4 , 5-HT 7 , and 5-HT 1A receptors,. Under these conditions, we confirmed the facilitatory role of 5-HT 4 heteroreceptors on acetylcholine release and revealed for the first time the occurrence of 5-HT 7 and 5-HT 1A heteroreceptors with a facilitatory and an inhibitory action, respectively. Our findings strengthen the novel concept for the use of recently patented selective 5-HT agonists and antagonists for the control of OAB dysfunctions associated with inflammatory conditions, although their therapeutic efficacy needs to be explored in the clinical setting.