title: Competitive antagonism of the mouse 5-hydroxytryptamine3 receptor by bisindolylmaleimide I, a "selective" protein kinase C inhibitor.
abstract: We examined the effects of several protein kinase C (PKC) inhibitors on the murine 5-hydroxytryptamine3 (5-HT3) receptor to determine whether they acted directly on the receptor. The 5-HT-evoked currents in Xenopus laevis oocytes expressing the recombinant 5-HT3 receptor were measured with the two-electrode voltage-clamp technique. The PKC inhibitors bisindolylmaleimide I (BIM, GF109203x) and staurosporine, but not calphostin C or chelerythrine, decreased the 5-HT3 receptor-mediated currents when coapplied with 5-HT. BIM blocked 0.5 microM 5-HT-elicited currents with an IC50 value of 7 nM, whereas in the presence of 5 microM staurosporine, 42% inhibition of 0.5 microM 5-HT-mediated currents was observed. Increasing concentrations of BIM resulted in a rightward shift of the 5-HT concentration-response curve, without altering efficacy. A Schild plot was generated, which had a slope of -1.01, suggesting competitive antagonism. The Ki value of BIM was determined to be 29 nM. To confirm competitive antagonism, a competitive binding assay was performed on Sf21 insect cells infected with the mouse 5-HT3 receptor cDNA in a baculovirus expression vector. BIM completely displaced binding of the selective 5-HT3 receptor antagonist [3H]GR65630. BIM bound to the 5-HT3 receptor with a Ki value of 61 nM, which was slightly less potent than that of the selective 5-HT3 receptor antagonist MDL72222 (27 nM). The PKC inhibitor BIM is a potent competitive antagonist at the 5-HT3 receptor.
authors.affiliation: Department of Pharmacology, Texas Tech University Health Sciences Center, Lubbock, Texas, USA.
authors.forename: S J
authors.initials: SJ
authors.lastname: Coultrap
authors.forename: H
authors.initials: H
authors.lastname: Sun
authors.forename: T E
authors.initials: TE
authors.lastname: Tenner
authors.forename: T K
authors.initials: TK
authors.lastname: Machu
mesh_terms: D000470:Alkaloids; D000818:Animals; D053119:Benzophenanthridines; D001667:Binding, Competitive; D002462:Cell Membrane / Q000378:metabolism; D004247:DNA / Q000096:biosynthesis; D004791:Enzyme Inhibitors / Q000494:pharmacology*; D066298:In Vitro Techniques; D007211:Indoles / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D008301:Maleimides / Q000037:antagonists & inhibitors / Q000494:pharmacology*; D051379:Mice; D009281:Naphthalenes / Q000494:pharmacology; D009865:Oocytes / Q000378:metabolism; D018408:Patch-Clamp Techniques; D010617:Phenanthridines / Q000494:pharmacology; D011493:Protein Kinase C / Q000037:antagonists & inhibitors*; D012313:RNA / Q000096:biosynthesis; D011985:Receptors, Serotonin / Q000096:biosynthesis / Q000187:drug effects* / Q000235:genetics; D044406:Receptors, Serotonin, 5-HT3; D012702:Serotonin Antagonists / Q000494:pharmacology*; D019311:Staurosporine / Q000494:pharmacology; D014982:Xenopus laevis
chemical_list: D000470:Alkaloids; D053119:Benzophenanthridines; D004791:Enzyme Inhibitors; D007211:Indoles; D008301:Maleimides; D009281:Naphthalenes; D010617:Phenanthridines; D011985:Receptors, Serotonin; D044406:Receptors, Serotonin, 5-HT3; D012702:Serotonin Antagonists; D012313:RNA; D004247:DNA; C016299:chelerythrine; D011493:Protein Kinase C; D019311:Staurosporine; C058819:calphostin C; C070515:bisindolylmaleimide I