Abstract
The monoamine, 5-hydroxytryptamine (5-HT), stimulates contraction of human uterine smooth muscle (myometrium), but the receptor subtypes involved have not been characterized. We studied the effects of a range of 5-HT receptor subtype-selective agonists and antagonists in isolated strips of myometrium obtained at the time of caesarean section. The 5-HT1A receptor agonist, 8-hydroxy-2-dipropylaminotetralin, produced an increase in contractions that was highly variable, of low potency, and was not significantly inhibited by the 5-HT1A antagonist WAY100635 [[O-methyl-3H]-N-(2-(4-(2-methoxyphenyl)-1-piperazinyl)ethyl)-N-(2-pyridinyl)cyclohexanecarboxamide]. The 5-HT2 receptor agonist, α-methyl-5-hydroxytryptamine (α-Me-5-HT), produced a strong, consistent, and concentration-dependent stimulation of contractions (pEC50 = 7.60 ± 0.10, n = 5). The 5-HT2A receptor antagonist, ketanserin [3-[2-[4-(4-fluoro benzoyl)-piperidin-1-yl]ethyl]-1H-quinazoline-2,4-dione], caused a parallel shift in the response to α-Me-5-HT, with a pKB value consistent with its known affinity for the 5-HT2A receptor (pKB = 8.47 ± 0.16, n = 5), but it had no effect on the response to oxytocin. The 5-HT2B and 5-HT2C receptor agonists, BW723C86 [(α-methyl-5-(2-thienylmethoxy)-1H-indole-3-ethanamine)] and Ro-60-01-75 [(S)-2-(6-chloro-5-fluoro-indol-1-yl)-1-methyl-ethylamine fumarate], produced inconsistent responses at potencies that were lower than expected for activation of their cognate receptors. The response to α-Me-5-HT was unaffected by the 5-HT2B and 5-HT2C receptor antagonists, SB204741 [(N-(1-methyl-1H-indolyl-5-yl)-N-(3-methyl-5-isothiazolyl)urea)] and RS102221 [8-[5-(2,4-dimethoxy-5-(4-trifluoromethyl phenylsulphonamido)phenyl-5-oxopentyl]-1,3,8-triazaspiro[4.5]decane-2,4-dione]. The 5-HT1B/1D receptor agonist, sumatriptan [1-[3-(2-dimethylaminoethyl)-1H-indol-5-yl]-N-methyl-methanesulfonamide], the 5-HT4 agonist, cisapride [4-amino-5-chloro-N-[1-[3-(4-fluorophenoxy)propyl]-3-methoxy-4-piperidyl]-2-methoxy-benzamide], and the 5-HT7 agonist, AS19 [(2S)-(+)-5-(1,3,5-trimethylpyrazol-4-yl)-2-(dimethylamino)tetralin], all had no effect on myometrial contractility. 5-HT2A receptor mRNA and immunoreactivity were identified using reverse transcriptase-polymerase chain reaction, Western blotting, and immunohistochemistry. Specific binding of [3H]ketanserin was demonstrated. This study provides strong evidence for the expression of contractile 5-HT2A receptors in pregnant human myometrium, and this receptor is a potential target for novel uterotonic therapies.
Footnotes
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This work was supported by the Evelyn Trust and by the National Institute for Health Research Cambridge Comprehensive Biomedical Research Centre.
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doi:10.1124/jpet.108.143040.
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ABBREVIATIONS: 5-HT, 5-hydroxytryptamine; RT, reverse transcriptase; PCR, polymerase chain reaction; AUC, area(s) under the curve; ANOVA, analysis of variance; RIPA, radioimmunoprecipitation assay; TBST, Tris-buffered saline/Tween 20; HRP, horseradish peroxidase; ketanserin, 3-[2-[4-(4-fluoro benzoyl) piperidin-1-yl]ethyl]-1H-quinazoline-2,4-dione; WAY100635, [O-methyl-3H]-N-(2-(4-(2-methoxyphenyl)-1-piperazinyl)-ethyl)-N-(2-pyridinyl)cyclohexanecarboxamide; 8-OH-DPAT, 8-hydroxy-2-dipropylaminotetralin; AS19, (2S)-(+)-5-(1,3,5-trimethylpyrazol-4-yl)-2-(dimethylamino)tetralin; BW723C86, (α-methyl-5-(2-thienylmethoxy)-1H-indole-3-ethanamine); cisapride, 4-amino-5-chloro-N-[1-[3-(4-fluorophenoxy)-propyl]-3-methoxy-4-piperidyl]-2-methoxy-benzamide; Ro 60-0175, (S)-2-(6-chloro-5-fluoro-indol-1-yl)-1-methyl-ethylamine fumarate; RS102221, 8-[5-(2,4-dimethoxy-5-(4-trifluoromethyl phenylsulphonamido)phenyl-5-oxopentyl]-1,3,8-triazaspiro[4.5]decane-2,4-dione; SB269970, (R)-3-(2-(2-(4-methyl-piperidin-1-yl)ethyl)-pyrrolidine-1-sulfonyl)phenol hydrochloride; SB204741, (N-(1-methyl-1H-indolyl-5-yl)-N-(3-methyl-5-isothiazolyl)urea); sumatriptan, 1-[3-(2-dimethylaminoethyl)-1H-indol-5-yl]-N-methyl-methanesulfonamide; DMSO, dimethyl sulfoxide; α-Me-5-HT, α-methyl-5-hydroxytryptamine; Ro-60-01-75, (S)-2-(6-chloro-5-fluoro-indol-1-yl)-1-methyl-ethylamine fumarate.
- Received July 3, 2008.
- Accepted December 12, 2008.
- The American Society for Pharmacology and Experimental Therapeutics
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