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

1 In many smooth muscle tissues a minor M(3)-muscarinic acetylcholine (mACh) receptor population mediates contraction, despite the presence of a larger M(2)-mACh receptor population. However, this is not the case for guinea-pig uterus where radioligand binding and functional studies exclude a dominant role for M(3)-mACh receptors. 2 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. 3 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 M(4)-selective muscarinic toxin MTx-3, or PD102807 (0.1 mu M) ruling out a role for M(4)-mACh receptors. 4 The agonist affinity value for L-660,863, a putative 'M(2)-selective' agonist of 5.44+/-0.30 (n=6) was significantly different from that reported in guinea-pig atria. In contrast, the pK(A) value for carbachol (4.22+/-0.17; n=8) agrees with that reported for guinea-pig ileum. 5 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 G(i)-proteins in contraction. 6 Radioligand binding studies revealed a K(D) for N-[(3)H]-methylscopolamine of 0.12+/-0.05 nM and a B(max) 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 M(2)-mACh receptors. 7 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.