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

Abstract

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 a nity were made before and after selective alkylation procedures, together with estimates of agonist a nity to characterise the mACh receptor population mediating carbachol-induced contraction of guinea-pig isolated uterus. 3 Antagonist a nity estimates made at`protected' receptors were not signi®cantly di erent from those made in untreated tissues. However all estimations were signi®cantly di erent from those reported in guinea-pig ileum and atria. The rank order of a nities were atropine4zamifenacin=tripitramine4 methoctramine. Carbachol-induced contractions were insensitive to the M 4 -selective muscarinic toxin MTx-3, or PD102807 (0.1 mM) ruling out a role for M 4 -mACh receptors. 4 The agonist a nity value for L-660,863, a putative`M 2 -selective' agonist of 5.44+0.30 (n=6) was signi®cantly di erent 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 71 . Antagonist a nity 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 pro®le which can not be distinguished using radioligand binding studies may mediate carbachol-induced contraction of guinea-pig isolated uterus.

Characterization of Muscarinic Acetylcholine Receptors in the Rat Epididymis1

“…The muscarinic acetylcholine receptor antagonist affinities (pK i values) obtained in the caput and cauda epididymis were compared with those obtained by Doods et al [40]. In addition, the pK i values for methoctramine obtained in the present study in the caput (7.80 Ϯ 0.15) and cauda epididymis (7.26 Ϯ 0.11) were comparable to those reported in tissues that express muscarinic M 2 receptors, such as rat vas deferens (pK i ϭ 8.4 [46]), smooth muscle cell culture from human prostate (pK i ϭ 8.1 [47]), and guinea pig uterus (pK i ϭ 7.5 [40], and pK i ϭ 8.1 [48]). Both analyses indicated that muscarinic acetylcholine receptors detected in the epididymis have a highly significant correlation with the M 2 receptor subtype.…”

Influence of Tissue Integrity on Pharmacological Phenotypes of Muscarinic Acetylcholine Receptors in the Rat Cerebral Cortex

“…An early example is that several antidepressants such as chlorimipramine potently inhibited H 2 receptor-mediated stimulation of cAMP formation in the homogenates of guinea pig hippocampus but failed to affect the cAMP formation in the slice preparations (Dam Trung Tuong et al, 1980). The pharmacological profile of mAChRs mediating carbachol-induced contraction in the guinea pig and rat uterus was not in agreement with the profile obtained from radioligand binding studies with the homogenates (Boxall et al, 1998;Munns and Pennefather, 1998). In the rat stomach mucosa, M 1 mAChRs could be detected in the tissue segments but were undetectable in the homogenates (Anisuzzaman et al, 2008a).…”

“…However, there have been several reports of "atypical" pharmacological profiles in which anomalous estimates of the functional affinity of receptors for antagonists in native tissues play a significant role in defining these atypical phenotypes of mAChRs (Nelson and Chal-liss, 2007). For example, in the guinea pig and rat uterus the functional affinities for mAChR antagonists are not in agreement with the affinities of M 2 mAChRs identified in radioligand binding studies, suggesting a possible involvement of an atypical operational mAChR in cholinergic contraction (Boxall et al, 1998;Munns and Pennefather, 1998). In addition, certain mAChR antagonists, such as darifenacin and solifenacin that are used in the management of overactive bladder, display in vivo selectivity for inhibition of mAChRmediated urinary bladder contraction relative to salivary secretion, even though both responses are mediated by the same M 3 mAChR subtype (Abrams et al, 2006).…”

Characterization of the muscarinic receptor in isolated uterus of sham operated and ovariectomized rats

“…In guinea‐pig uterus, evidence for the presence of muscarinic M 2 ( Eglen et al ., 1989 ; 1992 ; Doods et al ., 1993 ), M 3 ( Leiber et al ., 1990 ), M 4 (Dörje et al ., 1991) and an atypical muscarinic receptor ( Boxall et al ., 1998 ) has been presented. The rabbit uterus has not been extensively studied but M 2 , M 3 and M 4 receptors have been identified using immunoprecipitation techniques ( Crankshaw, 1984 ; Dörje et al ., 1991 ).…”