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CELLULAR AND MOLECULAR

Pharmacological Evidence for a Functional Serotonin-2B Receptor in a Human Uterine Smooth Muscle Cell Line

Molecular Pharmacology Unit, Alcon Research, Ltd., Fort Worth, Texas

The present study investigated the serotonin-induced increase in phosphoinositide hydrolysis and mobilization of intracellular Ca²⁺ ([Ca²⁺]_i) in human uterine smooth muscle cells (HUSMCs) to identify the serotonergic receptor positively coupled to phospholipase C in these cells. In phosphoinositide (PI) assays, serotonin (5-HT) and -methyl-5-HT were potent, full agonists (EC₅₀ = 20 and 4.1 nM, respectively), whereas the phenylethylamine, R-(–)-1-(4-iodo-2,5-dimethoxyphenyl)-2-aminopropane hydrochloride, was less active (EC₅₀ = 63 nM). Proposed 5-HT_2B-selective agonists, BW-723C86 [-methyl-5-(2-thienylmethoxy)-1H-indole-3-ethanamine hydrochloride] and (+)-norfenfluramine, exhibited strong agonist potency and efficacy comparable with 5-HT (EC₅₀ = 18 and 33 nM, respectively) and 15-fold more potency than (–)-norfenfluramine (EC₅₀ = 500 nM). 5-HT_2C receptor agonists m-chlorophenylpiperazine and MK-212 [6-chloro-2-(1-piperaxinyl)pyrazine] were weak agonists in these cells, with potencies of 110 and 880 nM, respectively. A similar rank order of potency was observed in [Ca²⁺]_i mobilization assays (r = 0.9, p < 0.005) in the HUSMC and with contraction of rat stomach fundus strips that contain a 5-HT_2B receptor (r = 0.9, p < 0.001). Antagonist studies revealed that a 5-HT_2B-selective antagonist, RS-127445 [2-amino-4-(4-fluoronaphth-1-yl)-6-isopropylpyrimidine] (K_i = 0.13 nM), was significantly more effective at inhibiting 5-HT-induced activity than a 5-HT_2A antagonist, M-100907 (R-(+)--(2,3-dimethoxyphenyl)-1-[2-(4-fluorophenyl)ethyl]-4-piperidinemethanol]) (K_i = 914 nM) and the 5-HT_2C antagonists RS-102221 (8-[5-(2,4-dimethoxy-5-(4-trifluoromethylsulfo-amido)phenyl-5-oxopentyl]-1,3,8-triazaspiro[4.5]decane-2,4-dione hydrochloride) (K_i = 2.5 µM) and SB-242084 (6-chloro-5-methyl-1-[6-92-methylpyridin-3-yloxy) pyridine-3-ylcarbamoyl] indoline) (K_i = 42.4 nM) in the HUSMC PI turnover assays. Taken together, these studies strongly suggest the presence of a functionally active 5-HT_2B receptor subtype in HUSMCs. The physiological role of this receptor in these cells remains to be defined.

Address correspondence to: Dr. Curtis R. Kelly, Molecular Pharmacology (R2-43), Alcon Research, Ltd., 6201 South Freeway, Fort Worth, TX 76134. E-mail: curtis.kelly{at}alconlabs.com

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