Characterization of the interaction between muscarinic M2 receptors and β‐adrenoceptor subtypes in guinea‐pig isolated ileum
Open Access
- 1 January 1995
- journal article
- Published by Wiley in British Journal of Pharmacology
- Vol. 114 (1), 49-56
- https://doi.org/10.1111/j.1476-5381.1995.tb14904.x
Abstract
1 Contraction of guinea-pig ileum to muscarinic agonists is mediated by M3 receptors, even though they account for only 30% of the total muscarinic receptor population. The aim of this study was to characterize the biochemical and functional effects of stimulation of the predominant M2 muscarinic receptor (70%) and to investigate the hypothesis that M2 receptors specifically oppose β-adrenoceptor-mediated effects in the ileum. 2 In guinea-pig ileal longitudinal smooth muscle slices, isoprenaline, a non-selective β-adrenoceptor agonist, and BRL 37344 (sodium-4-[2-[2-hydroxy-2-(3-chlorophenyl)ethylamino]propyl]-phenoxyacetate sesquihydrate), a β3-adrenoceptor selective agonist, increased cyclic AMP accumulation with – log EC50 values of 6.6 ± 0.1 and 5.8 ± 0.1 respectively. Maximal stimulation by BRL 37344 (10 μm) was 26.4 ± 5.2% of that observed with isoprenaline (10 μm). Isoprenaline (10 μm)-stimulated cyclic AMP accumulation was significantly, but not completely, inhibited by propranolol (5 μm), with a propranolol-resistant component of 28.2 ± 6.8% of the maximal stimulation to isoprenaline. In contrast, basal and BRL 37344 responses were resistant to this antagonist. These data provide evidence that both β1- and β3-adrenoceptors activate adenylyl cyclase in guinea-pig ileum. 3 Isoprenaline (10 μm)-stimulated cyclic AMP accumulation was inhibited (67.4 ± 0.9%) by the muscarinic agonist (+)-cis-dioxolane (-log EC50 = 7.3 ± 0.1). The rank order of antagonist affinities against the (+)-cis-dioxolane response was (-log KB values in parentheses): atropine (9.0 ± 0.2) > methoctramine (7.1 ± 0.1) > p-fluoro-hexa-hydrosilaphenidol (p-F-HHSiD; 6.5 ± 0.2) ≥ pirenzepine (6.3 ± 0.2). (+)-cis-dioxolane also significantly inhibited BRL 37344 (10 μm; 56.5 ± 2.4%) stimulated cyclic AMP accumulation. These data suggest that M2 receptors mediate inhibition of cyclic AMP accumulation in response to both β1- and β3-adrenoceptor stimulation in guinea-pig ileum. 4 5-Hydroxytryptamine (5-HT), vasoactive intestinal peptide, prostaglandins E2 and E1, all at 10 μm, significantly increased cyclic AMP accumulation. (+)-cis-Dioxolane (10 μm) inhibited both basal and agonist-induced cyclic AMP accumulation. Thus the inhibitory effect of M2 receptor agonism does not appear to be restricted to β-adrenoceptor-stimulated cyclic AMP accumulation. 5 The potential for involvement of activation of M2 receptors on responses to β-adrenoceptor agonists was also studied functionally. Selective M3 receptor alkylation was achieved by pretreatment of tissues with 4-DAMP mustard (40 nm), in the presence of methoctramine (1 μm; to protect M2 receptors). After washing, tissues were pre-contracted with histamine (0.3 μm) and relaxed with isoprenaline (0.6 μm). Under these conditions, oxotremorine m caused concentration-dependent contractions (-log EC50 of 7.8 ± 0.1), that were surmountably antagonized by methoctramine (1 μm) with a -log KB estimate of 7.4 ± 0.1. Similar observations were seen versus relaxation produced by BRL 37344 (1 μm), where the -log KB value for methoctramine was 7.8 ± 0.2. These data suggest that M2 receptors mediate a functional inhibition of relaxant responses to isoprenaline and BRL 37344. 6 These findings are consistent with β1- and β3-adrenoceptors coupling to stimulation of adenylyl cyclase in guinea-pig ileum; a response that is inhibited by M2 receptor stimulation. Concordantly, M2 receptor stimulation also inhibits relaxation to both β1- and β3-adrenoceptor stimulation. These results implicate M2 receptors in the modulation of sympathetic control of ileal motility.Keywords
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