Comparative pharmacology of recombinant human M 3 and M 5 muscarinic receptors expressed in CHO‐K1 cells

Affinity estimates were obtained for several muscarinic antagonists against carbachol‐stimulated [ 3 H]‐inositol phosphates accumulation in Chinese hamster ovary (CHO‐K1) cells stably expressing either human muscarinic M 3 or M 5 receptor subtypes. The rationale for these studies was to generate a functional antagonist affinity profile for the M 5 receptor subtype and compare this with that of the M 3 receptor, in order to identify compounds which discriminate between these two subtypes. The rank order of antagonist apparent affinities (p K B ) at the muscarinic M 5 receptor was atropine (8.7)tolterodine (8.6)=4‐diphenylacetoxy‐N‐methylpiperidine (4‐DAMP, 8.6)>darifenacin (7.7)zamifenacin (7.6)>oxybutynin (6.6)=para‐fluorohexahydrosiladifenidol (p‐F‐HHSiD, 6.6)>pirenzepine (6.4)methoctramine (6.3)=himbacine (6.3)>AQ‐RA 741 (6.1). Antagonist apparent affinities for both receptor subtypes compare well with published binding affinity estimates. No antagonist displayed greater selectivity for the muscarinic M 5 subtype over the M 3 subtype, but himbacine, AQ‐RA 741, p‐F‐HHSiD, darifenacin and oxybutynin displayed between 9‐ and 60 fold greater selectivity for the muscarinic M 3 over the M 5 subtype. This study highlights the similarity in pharmacological profiles of M 3 and M 5 receptor subtypes and identifies five antagonists that may represent useful tools for discriminating between these two subtypes. Collectively, these data show that in the absence of a high affinity M 5 selective antagonist, affinity data for a large range of antagonists is critical to define operationally the M 5 receptor subtype. British Journal of Pharmacology (1999) 127 , 590–596; doi: 10.1038/sj.bjp.0702551