Pharmacological characterization of muscarinic receptors in mouse isolated urinary bladder smooth muscle

Abstract

The pharmacological characteristics of muscarinic receptors in the male mice urinary bladder smooth muscle were studied. (+)-Cis-dioxolane, oxotremorine-M, acetylcholine, carbachol and pilocarpine induced concentration-dependent contractions of the urinary bladder smooth muscle (pEC(50)=6.6+/-0.1, 6.9+/-0.1, 6.7+/-0.1, 5.8+/-0.1 and 5.8+/-0.1, E(Max)=3.2+/-0.8 g, 2.7+/-0.4 g, 1.0+/-0.1 g, 2.7+/-0.3 and 0.9+/-0.2 g, respectively, n=4). These contractions were competitively antagonized by a range of muscarinic receptor antagonists (pK(B) values): atropine (9.22+/-0.09), pirenzepine (6.85+/-0.08), 4-DAMP (8.42+/-0.14), methoctramine (5.96+/-0.05), p-F-HHSiD (7.48+/-0.09), tolterodine (8.89+/-0.13), AQ-RA 741 (7.04+/-0.12), s-secoverine (8.21+/-0.09), zamifenacin (8.30+/-0.17) and darifenacin (8.70+/-0.09). In this tissue, the pK(B) values correlated most favourably with pK(i) values for these compounds at human recombinant muscarinic M(3) receptors. A significant correlation was also noted at human recombinant muscarinic m5 receptors given the poor discriminative ability of ligands between M(3) and m5 receptors. In recontraction studies, in which the muscarinic M(3) receptor population was decreased, and conditions optimized to study M(2) receptor activation, methoctramine exhibited an affinity estimate consistent with muscarinic M(3) receptors (pK(B)=6.23+/-0.14; pA(2)=6.16+/-0.03). Overall, these data study suggest that muscarinic M(3) receptors are the predominant, if not the exclusive, subtype mediating contractile responses to muscarinic agonists in male mouse urinary bladder smooth muscle.

Figure 1

Effects of muscarinic agonists on mouse urinary bladder smooth muscle. Contractile effects were expressed as percentages of the maximum response of the control curve. The values shown are means±s.e.mean, n⩾4 animals.

Figure 2

Correlation between the functional affinities (pK_i values) of muscarinic antagonists at muscarinic receptor in mouse isolated urinary bladder smooth muscle and binding affinities (pK_i values) at human recombinant muscarinic receptors (m1-m5; a–e respectively). The binding data were taken from Dörje et al., 1991; Eglen et al., 1997; Hegde et al., 1997; Nilvebrant et al., 1996. The broken line is the line of identity (x=y) while the solid line is the correlation plot (the inserts give the correlation factors (r) and the sum of squares values (ssq)).

Figure 3

Recontraction experiments in mouse urinary bladder smooth muscle: effect of methoctramine on the recontractile concentration-effect to (+)-cis-dioxolane obtained after elevation of adenylyl cyclase activity following preferential alkylation of muscarinic M₃ receptors (n=4). (a) Time control; (b) +0.3 μM methoctramine; (c) +1.0 μM methoctramine; (d) +3.0 μM methoctramine.