Abstract
The ability of prostanoid receptors to regulate the volume-dependent efflux of the organic osmolyte taurine from murine fibroblasts (L cells) via a cAMP-dependent mechanism has been examined. Incubation of L cells under hypoosmotic conditions resulted in a time-dependent efflux of taurine, the threshold of release occurring at 250 mOsM. Addition of prostaglandin E1 (PGE1) potently (EC50 = 2.5 nM) enhanced the volume-dependent efflux of taurine at all time points examined and increased the threshold for osmolyte release to 290 mOsM. Maximal PGE1 stimulation (250–300% of basal) of taurine release was observed at 250 mOsM. Of the PGE analogs tested, only the EP2-selective agonist butaprost (9-oxo-11α,16S-dihydroxy-17-cyclobutyl-prost-13E-en-1-oic acid) was able to enhance taurine efflux. Inclusion of 1,9-dideoxyfoskolin, 5-nitro-2-(3-phenylpropylamino) benzoic acid, or 4-[(2-butyl-6,7-dicloro-2-cyclopentyl-2,3-dihydro-1-oxo-1H-inden-5-yl)oxy]-butanoic acid blocked the ability of PGE1 to enhance taurine release, indicating the mediation of a volume-sensitive organic osmolyte and anion channel. The ability of PGE1 to increase osmolyte release from L cells was mimicked by the addition of agents that inhibit cAMP breakdown, directly activate adenylyl cyclase, or are cell-permeant analogs of cAMP. Taurine release elicited by either PGE1 or 8-(4-chlorophenylthio)-cAMP was attenuated by >70% in L cells that had been stably transfected with a mutant regulatory subunit of cAMP-dependent protein kinase (PKA). PGE1 stimulation of taurine efflux was not attenuated by either depletion of intracellular calcium or inhibition of protein kinase C. The results indicate that activation of prostanoid receptors on murine fibroblasts enhances osmolyte release via a cAMP and PKA-dependent mechanism.
Footnotes
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This work was supported by National Institutes of Health Grant NS23831 (to S.K.F.).
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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doi:10.1124/jpet.106.109496.
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ABBREVIATIONS: VSOAC, volume-sensitive organic osmolyte and anion channel; ICl,swell, outwardly rectifying Cl– current activated by hypotonicity; DDF, 1,9-dideoxyforskolin; NPPB, 5-nitro-2-(3-phenylpropylamino) benzoic acid; DCPIB, 4-[(2-butyl-6,7-dichloro-2-cyclopentyl-2,3-dihydro-1-oxo-1H-inden-5-yl)oxy]butanoic acid; GPCR, G protein-coupled receptor; EP, prostaglandin receptor; mAChR, muscarinic cholinergic receptor; PKC, protein kinase C; PKA, cAMP-dependent protein kinase; 8-CPT-cAMP, 8-(4-chlorophenylthio)-cAMP; Gö6983, 2-[1-(3-dimethyl-aminopropyl)-5-methoxyindol-3-yl]-3-(1H-indol-3-yl)maleimide; PG, prostaglandin; IBMX, 3-isobutyl-1-methylxanthine; AM, acetoxymethyl ester; DMEM, Dulbecco's modified Eagle's medium; ANOVA, analysis of variance.
- Received June 19, 2006.
- Accepted August 24, 2006.
- The American Society for Pharmacology and Experimental Therapeutics
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