Functional Selectivity of Muscarinic Receptor Antagonists for Inhibition of M3-Mediated Phosphoinositide Responses in Guinea Pig Urinary Bladder and Submandibular Salivary Gland

• Published in: The Journal of pharmacology and experimental therapeutics Vol. 310; no. 3; pp. 1255 - 1265
• Main Authors: Nelson, Carl P, Gupta, Paul, Napier, Carolyn M, Nahorski, Stefan R, Challiss, R A John
• Format: Journal Article
• Language: English
• Published: United States American Society for Pharmacology and Experimental Therapeutics 01.09.2004
• Subjects: Animals; CHO Cells; Cholinergic Antagonists - pharmacology; Cricetinae; Female; Guinea Pigs; In Vitro Techniques; Muscarinic Antagonists - pharmacology; Phosphatidylinositols - metabolism; Radioligand Assay; Receptor, Muscarinic M3 - metabolism; Receptors, Cholinergic - classification; Receptors, Cholinergic - drug effects; Receptors, Cholinergic - metabolism; Salivary Glands - metabolism; Submandibular Gland - metabolism; Urinary Bladder - metabolism
• ISSN: 0022-3565; 1521-0103
• DOI: 10.1124/jpet.104.067140

Binding and functional affinities of the muscarinic acetylcholine (mACh) receptor antagonists darifenacin, tolterodine, oxybutynin, and atropine were assessed in Chinese hamster ovary (CHO) cells expressing the human recombinant M 2 (CHO-m2) or M 3 (CHO-m3) receptors, and in guinea pig bladder and submandibular gland. In [ N -methyl- 3 H]scopolamine methyl chloride binding studies in CHO cells, darifenacin displayed selectivity (14.8-fold) for the M 3 versus M 2 mACh receptor subtype. Oxybutynin was nonselective, whereas atropine and tolterodine were weakly M 2 -selective (5.1- and 6.6-fold, respectively). Antagonist functional affinity estimates were determined by the inhibition of agonist-induced [ 3 H]inositol phosphate accumulation in CHO-m3 cells and antagonism of the agonist-induced inhibition of forskolin-stimulated cyclic AMP accumulation in CHO-m2 cells. Darifenacin was the most M 3 -selective antagonist (32.4-fold), whereas oxybutynin, atropine, and tolterodine exhibited lesser selectivity. Functional affinity estimates in guinea pig urinary bladder and submandibular salivary gland using indices of phosphoinositide turnover revealed that oxybutynin, darifenacin, and tolterodine each displayed selectivity for the response in the bladder, relative to that seen in the submandibular gland (9.3-, 7.9-, and 7.4-fold, respectively). In contrast, atropine displayed a similar affinity in both tissues. These data demonstrate that in bladder, compared with submandibular gland from a single species, the mACh receptor antagonists darifenacin, tolterodine, and oxybutynin display selectivity to inhibit agonist-mediated phosphoinositide responses. It is proposed that both responses are mediated via M 3 mACh receptor activation and that differential functional affinities displayed by some, but not all, antagonists are indicative of the influence of cell background upon the pharmacology of the M 3 mACh receptor.