The role of M2-muscarinic receptors in mediating contraction of the pig urinary bladderin vitro

Abstract

In urinary bladder, M₂‐muscarinic receptors predominate, but it is the smaller population of M₃‐receptors which mediate detrusor contraction. This study examines the M₂ : M₃ ratio and the role of M₂‐receptors in contraction of pig urinary bladder. Competition experiments with [³H]‐QNB determined the ratio of M₂ : M₃. In functional studies, affinity values (pK_B) for 4‐DAMP, darifenacin and methoctramine were calculated. Similar experiments were performed on tissues following selective M₃‐inactivation (incubation with 40 nM 4‐DAMP mustard in the presence of 1 μM methoctramine to protect M₂‐receptors), precontraction with 50 mM KCl and relaxation with isoprenaline (30 μM) or forskolin (1 μM). In competition binding, displacement of [³H]‐QNB by 4‐DAMP, darifenacin and methoctramine best fitted a two‐site model suggesting a predominant (70–80%) population of M₂‐receptors. On normal detrusor in vitro, 4‐DAMP and methoctramine caused surmountable antagonism of responses to carbachol with pK_B values of 9.37±0.07 and 6.05±0.05 respectively. Darifenacin caused unsurmountable antagonism, the apparent pK_B value being 8.61±0.10. In tissues where the M₃‐receptors had been inactivated and cyclic AMP levels elevated, 4‐DAMP and darifenacin were less potent, with apparent pK_B values of 8.72±0.08 and 6.74±0.07. In contrast, methoctramine was more potent, the apparent pK_B value increasing significantly to 6.86±0.06. These data suggest that the pig bladder possesses a similar muscarinic receptor population to the human bladder and that the M₃‐receptor subtype mediates contraction of the normal detrusor muscle. However an involvement of M₂‐receptors in contraction can be observed following pharmacological manipulation of the receptor population. British Journal of Pharmacology (2000) 131, 1482–1488; doi:10.1038/sj.bjp.0703719