Inhibitory prostanoid EP receptors in human non-pregnant myometrium

Abstract

Prostanoid EP receptor agonists relaxed cloprostenol-stimulated contraction of human non-pregnant myometrium in vitro with pEC₅₀ values of (n=4): prostaglandin E₂, 7.8±0.2>1-OH prostaglandin E₁, 7.2±0.3>misoprostol, 6.6±0.1>16,16-dimethyl prostaglandin E₂, 6.3±0.7>butaprost, 5.7±0.3>11-deoxy prostaglandin E₁, 5.5±0.2=AH13205 ((±)-trans-2-[4-(1hydroxyhexyl)phenyl]-5-oxocyclopentaneheptanoic acid), 5.5±0.2. The EP₄ receptor antagonist AH23848B ([1α(z), 2β5α]-(±)-7-[5-[[(1,1′-biphenyl)-4-yl]methoxy]-2-(4-morpholinyl)-3-oxo-cyclopentyl]-4-heptenoic acid) (29 μM) had no effect on concentration–effect curves to the EP receptor agonists. The mixed prostanoid receptor antagonist AH6809 (6-isopropoxy-9-oxaxanthene-2-carboxylic acid) competitively antagonised prostaglandin E₂ with a pA₂ of 5.6±0.2. AH6809 (42 μM) antagonised misoprostol, 11-deoxy prostaglandin E₁, and the prostanoid DP receptor agonist BW245C (5-(6-carboxyhexyl)-1-(3-cyclohexyl-3-hydroxypropyl)hydantoin) with apparent pA₂ values of 5.6±0.3, 5.1±0.9 and 5.9±0.4 (n=4), respectively, but was ineffective against the IP receptor agonist cicaprost (n=4). The prostanoid DP receptor antagonist BW A868C (3-benzyl-5-(6-carboxyhexyl)-1-(2-cyclohexyl-2-hydroxyethylamino)hydantoin) (50 nM) had no effect on responses to prostaglandin E₂ or misoprostol. The presence of an AH6809-sensitive, AH23848B- and BW A868C-insensitive mechanism is consistent with the hypothesis that inhibitory EP receptor agonists cause relaxation of human non-pregnant myometrium by an EP₂ receptor-mediated process.

Introduction

Prostanoids inhibit the contractility of human non-pregnant myometrium by acting at DP receptors (Senior et al., 1992; Fernandes and Crankshaw, 1995), IP receptors (Senior et al., 1992) and inhibitory EP receptors (Senior et al., 1991; Brown and Crankshaw, 1995; Brown et al., 1997). The inhibitory EP receptors involved are thought to belong to the EP₂ subtype. This notion is based mainly upon the potency of the EP₂ receptor-selective agonist butaprost in a superfusion assay (Senior et al., 1991). Subsequent to the characterization of EP receptors in human myometrium (Senior et al., 1991), a novel prostanoid receptor, the EP₄ receptor, was identified pharmacologically (Coleman et al., 1994a). Cloning of the human EP₂ receptor by Regan et al. (1994)demonstrated that the human EP receptor previously cloned by Bastien et al. (1994)had been incorrectly identified. This earlier clone (Bastien et al., 1994) was subsequently identified as the EP₄ receptor (Toh et al., 1995). Both Northern blots of human uterus (Bastien et al., 1994) and reverse transcription-polymerase chain reaction (RT-PCR) of human myometrium (Senchyna and Crankshaw, 1995) revealed the presence of EP₄ receptor mRNA, but there is as yet no functional evidence for the presence of this receptor in human myometrium.

Although originally characterized as a TP receptor antagonist, the compound AH23848B is also an EP₄ receptor antagonist, with no antagonist activity at other EP receptors (Coleman et al., 1994a). On the other hand, AH6809 blocks DP and EP₁ receptors and these properties have been used extensively in prostanoid receptor classification (Coleman et al., 1994b), but it has recently been shown to antagonise the recombinant human EP₂ receptor (Woodward et al., 1995). Therefore, both AH23848B and AH6809 may be helpful in fully characterizing the inhibitory EP receptors mediating relaxation of human myometrium.

In the present study, we have determined the potencies of a number of inhibitory EP receptor agonists on human non-pregnant myometrium in vitro using equilibrium methods. Tissues were pre-stimulated with the prostanoid FP receptor agonist cloprostenol (Coleman et al., 1994b). We have investigated the effects of AH23848B and AH6809 on prostanoid-induced relaxation.