Relaxation of rat distal colon by activation of muscarinic, neuronal receptors: possible involvement of P2y-purinoceptors (original) (raw)
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Contractile roles of the M2 and M3 muscarinic receptors in the guinea pig colon
The Journal of pharmacology and experimental therapeutics, 1998
The contractile roles of the M2 and M3 muscarinic receptors were investigated in guinea pig longitudinal colonic smooth muscle. Prior treatment of the colon with N-(2-chloroethyl)-4-piperidinyl diphenylacetate (4-DAMP mustard) (40 nM) in combination with [[2-[(diethylamino)methyl]-1-piperidinyl]acetyl]-5,11- dihydro-6H-pyrido[2,3b][1,4]benzodiazepine-6-one (AF-DX 116) (1.0 microM) caused a subsequent, irreversible inhibition of oxotremorine-M-induced contractions when measured after extensive washing. The estimate of the degree of receptor inactivation after 2 hr (97%) was not much greater than that measured after 1 hr (95%), which suggests that both 4-DAMP mustard-sensitive and -insensitive muscarinic subtypes contribute to the contractile response. Pertussis toxin treatment had no significant inhibitory effect on the control contractile response to oxotremorine-M, but caused an 8.8-fold increase in the EC50 value measured after a 2-hr treatment with 4-DAMP mustard. These results s...
Acute desensitization of muscarinic receptors in the isolated guinea-pig ileal longitudinal muscle
Journal of Autonomic Pharmacology, 1992
The effects of acute desensitization of muscarinic receptors mediating contractile responses of the guinea-pig ileal longitudinal muscle were studied in vitro, using similar conditions for both functional and radioligand binding studies. 2 The pA, values for a number of muscarinic antagonists (pirenzepine, methoctramine, (k)para-fluoro-hexahydro-siladifenidol and 4-diphenylacetoxy-N-methyl piperidine-methiodide) indicated that the contractile response to carbachol was mediated through an M3 muscarinic receptor. In binding experiments the muscarinic receptor subtype population in ileal longitudinal muscle was found to be heterogeneous, consisting of approximately 77% M2 and 23% M3 receptors. 4 It is concluded that desensitization of the contractile responses of the guineapig ileal longitudinal muscle is a result of M3 but not M2 muscarinic receptor desensitization. Acute desensitization, therefore, is not accompanied by meaningful changes in the total number of both M2 and M3 receptors or by alterations in the affinity of the receptor to ligands.
Journal of Autonomic Pharmacology, 1997
1 The present study examined the role of muscarinic receptors in the modulation of noradrenaline (NA) release in the guinea-pig isolated distal colon. The spontaneous endogenous NA over¯ow assayed by HPLC-ED was taken as an index of NA release from enteric noradrenergic nerve terminals. 2 Physostigmine (10 mM) signi®cantly enhanced spontaneous endogenous NA over¯ow. Hyoscine (muscarinic antagonist), (R)-(-)-trihexyphenidyl and telenzepine (M 1 -selective antagonists), and 11[[2-[(diethylamino)methyl]-1-piperydil]acetyl]-5,11-dihydro-6H-pyrido[2,3-b][1,4]benzodiazepine-6-one (AF-DX 116, M 2 -selective antagonist) inhibited NA over¯ow in a concentration dependent manner, with the following EC 50 values: 131.74 (18.19±953.96), 101.62 (58.83±175.60), 150 (60±330), 30 (5±170) nM, respectively. 4-diphenylacetoxy-N-methylpiperidine methiodide (4-DAMP, M 1 -and M 3 -selective antagonist) had no signi®cant eect up to 100 mM. 3 The muscarinic agonist oxotremorine inhibited NA over¯ow in a concentration dependent manner, with an EC 50 value of 0.67 (0.30±1.51) mM. The response to oxotremorine was inhibited by muscarinic antagonists with the following order of potency: hyoscine = (R)-(-)-trihexyphenidyl = telenzepine 4 4-DAMP 44 AF-DX 116. 4 In the presence of 3 mM tetrodotoxin (TTX), the eect of oxotremorine and 4-DAMP was unchanged, while hyoscine, (R)-(-)-trihexyphenidyl, telenzepine and AF-DX 116, instead of inhibiting, signi®cantly enhanced NA over¯ow. 5
European Journal of Pharmacology, 1984
The myenteric plexus-longitudinal muscle preparation of the guinea-pig ileum was preincubated with [3 H]choline, then superfused and stimulated electrically (1 Hz 120 pulses). Oxotremorine reduced the evoked outflow of [3H]acetylcholine in a concentration-dependent manner. Each of the six antagonists (scopolamine, methylatropine, trihexyphenidyl, 4-DAMP, clozapine, pirenzepine) produced parallel shifts of the concentration-response curves for the prejunctional effects of oxotremorine. Similarly, in contraction experiments, the antagonists competitively antagonized the postjunctional responses to oxotremorine. The pre-and postjunctional pA 2 values did not differ significantly for any of the antagonists. It is concluded that pre-and postjunctional muscarinic receptors in the guinea-pig ileum are pharmacologically similar. Guinea-pig ileum Muscarinic antagonists Pre-and postjunctional receptors pA 2 values
British Journal of Pharmacology, 2000
1 The muscarinic acetylcholine receptors mediating the contractile response elicited to endogenous acetylcholine released by the selective P2X receptor agonist a,b-methylene ATP (mATP) were investigated in guinea-pig ileum. 2 mATP (0.1 ± 30 mM) elicited a concentration-dependent neurogenic contractile response inhibited by tetrodotoxin (TTX) and antagonized by the non-selective muscarinic receptor antagonist Nmethylscopolamine (NMS). 3 The contractile response to mATP was pertussis toxin-insensitive, irreversibly antagonized by N-(2-chloroethyl)-4-piperidinyl diphenylacetate (4-DAMP mustard), and unaected by the muscarinic M 2 /M 4 receptor selective antagonist AF-DX 116 (1 mM). 4 When measured in the presence of histamine and isoproterenol after treatment with 4-DAMP mustard, mATP elicited a pertussis toxin-sensitive contractile response potently antagonized by AF-DX 116. 5 Collectively, our data suggest that endogenous acetylcholine released by mATP can elicit a direct contractile response through the muscarinic M 3 receptor and an indirect contractile response through the muscarinic M 2 receptor by antagonizing the relaxant eects of isoproterenol on histamine induced contraction.
The Journal of pharmacology and experimental therapeutics, 1997
The purpose of this study was to characterize the role of M2 muscarinic receptors in inhibiting relaxant effects of drugs that stimulate cyclic AMP (cAMP) accumulation in the guinea pig ileum. We investigated the ability of oxotremorine-M (oxo-M) to inhibit cAMP accumulation in the presence of agonists that stimulate adenylyl cyclase in other cells and tissues. Appreciable stimulation of cAMP (> 50% over basal levels) was achieved with forskolin and maximally effective concentrations of isoproterenol, cicaprost, prostaglandin E1, prostaglandin E2 and prostaglandin I2, with the stimulation over basal levels of cAMP being 14.9-, 2.51-, 2.45-, 2.27-, 2.28- and 1.52-fold, respectively. Moderate or no cAMP stimulation was observed with dopamine, 5-hydroxytryptamine, 5-methoxytryptamine, dimaprit, vasoactive intestinal peptide, SKF-38393, 2-chloroadenosine, CGS-21680, prostaglandin D2, secretin and vasopressin. Oxo-M (1 microM) inhibited cAMP accumulation by 35% under basal conditions....
Canadian Journal of Physiology and Pharmacology, 2001
Muscarinic receptor mediated membrane currents and contractions were studied in isolated canine colon circular smooth muscle cells. Carbachol (10 -5 M) evoked a slow transient inward current that was superimposed by a transient outward current at holding potentials greater than -50 mV. Carbachol contracted the cells by 70 ± 2%. The effects of carbachol were blocked by atropine (10 -6 M), tetraethyl ammonium (20 mM), and BAPTA-AM (25 mM applied for 20 min). The inward current and contraction were not sensitive to diltiazem (10 -5 M), nitrendipine (3 × 10 -7 M), niflumic acid (10 -5 M), or N-phenylanthranilic acid (10 -4 M), but were gradually inhibited after repetitive stimulations in Ca 2+ free solution. Ni 2+ (2 mM) inhibited the inward current by 67 ± 4%. The inward current reversed at +15 mV. The outward component could be selectively inhibited by iberiotoxin (20 nM) or by intracellular Cs + . Repeated stimulation in the presence of cyclopiazonic acid (CPA, 3 µM) inhibited the carbachol-induced outward current and partially inhibited contraction. CPA did not inhibit the inward current. In conclusion, muscarinic receptor stimulation evoked a CPA-sensitive calcium release that caused contraction and a CPA-insensitive transient inward current was activated that is primarily carried by Ca 2+ ions and is sensitive to Ni 2+ .
British Journal of Pharmacology, 1993
The characterization of muscarinic receptors on single cells of the guinea-pig ileum longitudinal smooth muscle, devoid of neuronal elements, was functionally studied by estimating the affinities of muscarinic antagonists on acetylcholine-induced contractions. 2 Atropine (5 x 10-" to 5 x 10-6 M), 4-diphenylacetoxy-N-methyl-piperidine methiodide (4-DAMP, 5 x 10-8 to 5 x 10-6 M), cyclohexyl(4-fluoro-phenyl) (3-piperidinopropyl) silanol (pFHHSiD, 5 x 10-7 to 5 x 10-' M) as well as pirenzepine (5 x 10' to 5 x 10-M) competitively antagonized the acetylcholinedependent contractions with different affinities (atropine > 4-DAMP > pFHHSiD > pirenzepine). 3 Methoctramine (5 x 10-7 to 5 x 10-5 M), and AF-DX 116 (5 x 10-6 and 5 x 10-5M) also showed antagonist properties but these deviated from simple competition. These compounds, which discriminate between M2 and M3 receptors, showed a potency lower than that of pirenzepine, the rank order of potencies being pirenzepine > methoctramine > AF-DX 116. When concentrations of AF-DX 116, methoctramine and pirenzepine were increased an unspecific contractile effect occurred. 4 McN-A-343, a partial agonist on intact guinea-pig longitudinal smooth muscle strips, on this preparation induced a weak contraction (about 7% in comparison to control) that was not reversed by antimuscarinic agents. 5 These data indicate that M3 rather than M2 receptor sites are present on this tissue.
Cholinergic-mediated secretion in the rat colon: neuronal and epithelial muscarinic responses
European Journal of Pharmacology, 1989
10~4 M), carbachol (5 x 10e6-5 x lOA M), bethanechol (5 x lo-'-5 x 10m4 M) and dimethylphenylpiperazinium (DMPP, lop5 M) mcreased the short-circuit current (1s~) in the rat colon descendens by a tetrodotoxin (TTX)-sensitive mechanism. Blockade by TTX was still observed after removal of the submucosa, indicating the involvement of neurons of the mucosal plexus. Hexamethonium (10e5 M) and atropine (10W6 M) were used to distinguish between nicotinic and muscarinic neuronally mediated effects. The inhibitor of choline uptake, hemicholinium-3 (1 mM), reversibly inhibited the effect of repeated electric field stimulation (EFS). The EFS response was only inhibited by high concentrations of atropine (2 lop5 M). In mucosa-submucosa preparations 4-diphenylacetoxy-N-methylpiperidine methiodide (CDAMP) was more effective than telenzepine whereas pirenzepine was ineffective. Pirenzepine inhibited the EFS response in mucosa preparations as did telenzepine and 4-DAMP. It was not possible to differentiate between the muscarinic receptors involved in the different parts of the enteric nervous system on the basis of our results.
Functional role of M2 muscarinic receptors in the guinea pig ileum
Life Sciences, 1995
Muscarinic agonists elicit contraction in the standard guinea pig ileum bioassay through activation of M3 muscarinic receptors that are also linked to phosphoinositide hydrolysis. Surprisingly, the most abundant muscarinic receptor in the ileum is the M2 which causes a specific inhibition of cyclic AMP accumulation elicited by the 13-adrenergic receptor. After most of the M3 receptors are inactivated, the ileum still retains high sensitivity to muscarinic agonists provided that the contractile responses are measured in the presence of histamine and forskolin, which together, have no effect on contraction.