Characterization of an atypical muscarinic cholinoceptor mediating contraction of the guinea-pig isolated uterus

In many smooth muscle tissues a minor M3-muscarinic acetylcholine (mACh) receptor population mediates contraction, despite the presence of a larger M2-mACh receptor population. However, this is not the case for guinea-pig uterus where radioligand binding and functional studies exclude a dominant role for M3-mACh receptors. Using tissue from animals pre-treated with diethylstilboestrol, estimates of antagonist affinity were made before and after selective alkylation procedures, together with estimates of agonist affinity to characterise the mACh receptor population mediating carbachol-induced contraction of guinea-pig isolated uterus. Antagonist affinity estimates made at 'protected' receptors were not significantly different from those made in untreated tissues. However all estimations were significantly different from those reported in guinea-pig ileum and atria. The rank order of affinities were atropine>zamifenacin=tripitramine> methoctramine. Carbachol-induced contractions were insensitive to the M4-selective muscarinic toxin MTx-3, or PD102807 (0.1 microM) ruling out a role for M4-mACh receptors. The agonist affinity value for L-660,863, a putative 'M2-selective' agonist of 5.44+/-0.30 (n=6) was significantly different from that reported in guinea-pig atria. In contrast, the pKA value for carbachol (4.22+/-0.17 n = 8) agrees with that reported for guinea-pig ileum. Carbachol-induced contractions were insensitive to pertussis toxin although carbachol-induced inhibition of forskolin-stimulated cyclic AMP production was attenuated, ruling out the involvement of Gi-proteins in contraction. Radioligand binding studies revealed a KD for N-[3H]-methylscopolamine of 0.12+/-0.05 nM and a Bmax of 147+/-18 fmol mg protein(-1). Antagonist affinity estimates made using competition binding studies supported previous data suggesting the presence of a homogenous population of M2-mACh receptors. These data suggest a small population of mACh receptors with an atypical operational profile which can not be distinguished using radioligand binding studies may mediate carbachol-induced contraction of guinea-pig isolated uterus.