title: Protein kinase C activation by serotonin potentiates agonist-induced stimulation of cAMP production in cultured rat retinal pigment epithelial cells.
abstract: Serotonin stimulates inositol phosphate production and intracellular calcium mobilization in cultured rat retinal pigment epithelial (RPE) cells through interaction with 5-HT2A receptors, but decreases cAMP production in cultured human RPE cells via 5-HT1A receptors. Studies were therefore undertaken to investigate the effect of serotonin on the cAMP system in rat RPE cells. Exposure of cultured rat RPE cells to serotonin (100 microM) for 10 minutes had no effect on the basal levels of cAMP. However, a 5 minute preincubation with serotonin potentiated the production of cAMP induced by a 5 minute exposure to forskolin (5 microM), isoproterenol (1 microM) and 5'-[N-ethylcarboxamido]-adenosine (10 microM) by 133.0%, 296.8% and 651.9%, respectively. This effect of serotonin was dose-dependent on forskolin and 5'-[N-ethylcarboxamido]-adenosine with half-maximal effects close to those reported for its action on inositol phosphates production. The antagonists ketanserin, methysergide and spiperone attenuated the action of serotonin, while yohimbine and spiroxatrine were ineffectual, thus indicating that the potentiating effect was through the 5-HT1A receptor. Incubation of cultured rat RPE cells with bradykinin stimulates inositol phosphates production with half-maximal effect observed at 1 nM. Bradykinin also potentiates the action of forskolin, isoproterenol and 5'-[N-ethylcarboxamido]-adenosine on cAMP production in a dose-dependent manner with little effect on basal levels. RPE cells exposed to serotonin (500 microM) or phorbol 12-13 dibutyrate (1 microM) for 30 minutes showed translocation of protein kinase C to the membrane from the cytosol, with 53.3% and 29.4% increases in membrane activity, respectively. Forskolin- and 5'-[N-ethylcarboxamido]-adenosine-induced cAMP production was potentiated by phorbol 12-13 dibutyrate (1 microM) treatment. The effect of both serotonin and phorbol 12-13 dibutyrate on forskolin-induced cAMP production was attenuated by pretreatment of cell cultures with the protein kinase C antagonists staurosporin and calphostin C at 1 microM. Thus, the production of cAMP in cultured rat RPE cells is potentiated by 5-HT2A receptors through activation of protein kinase C. This effect is, however, not specific since bradykinin, which stimulates inositol phosphates turnover, also potentiates stimulated cAMP production.
authors.affiliation: Nuffield Laboratory of Ophthalmology, Oxford University, U.K.
authors.forename: M S
authors.initials: MS
authors.lastname: Nash
authors.forename: J P
authors.initials: JP
authors.lastname: Wood
authors.forename: N N
authors.initials: NN
authors.lastname: Osborne
mesh_terms: D000241:Adenosine / Q000031:analogs & derivatives / Q000494:pharmacology; D019830:Adenosine-5'-(N-ethylcarboxamide); D000818:Animals; D001920:Bradykinin / Q000494:pharmacology; D002478:Cells, Cultured; D005576:Colforsin / Q000494:pharmacology; D000242:Cyclic AMP / Q000096:biosynthesis*; D004357:Drug Synergism; D004789:Enzyme Activation; D004791:Enzyme Inhibitors / Q000494:pharmacology; D007545:Isoproterenol / Q000494:pharmacology; D009281:Naphthalenes / Q000494:pharmacology; D015240:Phorbol 12,13-Dibutyrate / Q000494:pharmacology; D010857:Pigment Epithelium of Eye / Q000187:drug effects / Q000378:metabolism*; D011493:Protein Kinase C / Q000187:drug effects / Q000378:metabolism*; D051381:Rats; D011985:Receptors, Serotonin; D012701:Serotonin / Q000494:pharmacology*; D012702:Serotonin Antagonists / Q000494:pharmacology; D019311:Staurosporine / Q000494:pharmacology
chemical_list: D004791:Enzyme Inhibitors; D009281:Naphthalenes; D011985:Receptors, Serotonin; D012702:Serotonin Antagonists; D005576:Colforsin; D012701:Serotonin; D019830:Adenosine-5'-(N-ethylcarboxamide); D015240:Phorbol 12,13-Dibutyrate; D000242:Cyclic AMP; D011493:Protein Kinase C; D019311:Staurosporine; C058819:calphostin C; D000241:Adenosine; D007545:Isoproterenol; D001920:Bradykinin