EFFECTS OF MUSCARINIC M(2) AND M(3) RECEPTOR STIMULATION AND ANTAGONISM ON RESPONSES TO ISOPRENALINE OF GUINEA-PIG TRACHEA IN-VITRO

1 In guinea-pig and canine airway smooth muscle, there is reduced beta-adrenoceptor agonist sensitivity in tissues pre-contracted with muscarinic agonists when compared to tissues pre-contracted with other spasmogens, such as histamine or leukotriene D-4. This reduced sensitivity may be the result,of an interaction between muscarinic receptors and beta-adrenoceptors. In this study the effects of M(2) receptor antagonism and stimulation have been investigated on the relaxant potency of isoprenaline in guinea-pig isolated tracheal smooth muscle. 2 (+)-cis-Dioxolane contracted isolated tracheal strips in a concentration-dependent manner (EC(50) = 11.5 +/- 0.9 nM). The rank order of antagonist apparent affinities (with pA(2) values in parentheses) was atropine (9.4 +/- 0.1) > zamifenacin (8.2 +/- 0.1) > para-fluoro-hexahydro-siladiphenidol (p-F-HHSiD, 7.2 +/- 0.1) > pirenzepine (6.5 +/- 0.1) > methoctramine (5.5 +/- 0.1). Schild slopes were not significantly different from unity. This was consistent with a role of muscarinic M(3) receptors in mediating contraction. 3 In tissues pre-contracted to 3 g isometric tension using (+)-cis-dioxolane (0.2 mu M, approximately EC(50)), the relaxant potency of isoprenaline was significantly (P < 0.05) increased by 0.3 mu M methoctramine (control EC(50) = 32.2 +/- 4.3 nM, plus methoctramine EC(50) = 19.1 +/- 4.5 nM). This concentration of methoctramine had no effect on contractile responses to (+)-cis-dioxolane (control, EC(50) = 17.6 +/- 3.2 nM, plus methoctramine, EC(50) = 21.0 +/- 4.4 nM). 4 When acetylcholine (non-selective), (+)-cis-dioxolane (non-selective), L-660,863 ((+/-)-3-(3-amino-1,2,4-oxadiazole-5-yl)-quinuclidine, M(2)-selective) or SDZ ENS 163 (thiopilocarpine, mixed M(2) antagonist, partial M(3) agonist) were used to achieve isometric tensions of 3 g, the relaxant potency of isoprenaline ranged from 3.7 +/- 0.3 nM (SDZ ENS 163) to 49.4 +/- 3.2 nM ((+)-cis-dioxolane). Reducing the concentration of these agonists (and therefore the level of developed tension to 2 g), significantly (P < 0.05) increased the relaxant potency of isoprenaline. In contrast, when histamine was used to pre-contract tissues to either 2 or 3 g (EC(50) = 4.2 +/- 0.6 and 3.8 +/- 1.1 nM, respectively), there was no significant effect on the relaxant potency of isoprenaline. 5 There was a slight but significant (P < 0.05) reduction in the relaxant potency of isoprenaline, in tissues pre-contracted to 3 g using histamine in combination with (+)-cis-dioxolane (30 nM). This effect was reversed by M(2) receptor antagonism, using methoctramine (1 mu M). 6 These data suggest that in guinea-pig isolated trachea, the relaxant potency of isoprenaline may depend not only on the level of developed tension but also, on the level of muscarinic M(2) receptor stimulation/blockade of the spasmogen inducing the tension. However, the lack of selective M(2) agonist and the low M(2)/M(3) selectivity of antagonists in this tissue do not permit definitive conclusions to be made about the role of these receptors in modulating isoprenaline potency.