title: Prevention of ginsenoside-induced desensitization of Ca2+-activated Cl- current by microinjection of inositol hexakisphosphate in Xenopus laevis oocytes: involvement of GRK2 and beta-arrestin I.
abstract: We demonstrated that ginsenosides, the active ingredient of Panax ginseng, enhance endogenous Ca(2+)-activated Cl(-) currents via Galpha(q/11)-phospholipase C-beta3 pathway in Xenopus laevis oocytes. Moreover, prolonged treatment of ginsenosides induced Cl(-) channel desensitization. However, the molecular mechanisms involved in ginsenoside-induced Cl(-) channel desensitization have not yet been determined precisely. To provide answers to these questions, we investigated the changes in ginsenoside-induced Cl(-) channel desensitization after intraoocyte injection of inositol hexakisphosphate (InsP(6)), which is known to bind beta-arrestins and interfere with beta-arrestin-induced receptor down-regulation, and cRNAs coding beta-arrestin I/II and G-protein receptor kinase 2 (GRK2), which is known to phosphorylate G protein-coupled receptors and attenuate agonist stimulations. When control oocytes were stimulated with ginsenosides, the second, third, and fourth responses to ginsenosides were 69.6 +/- 4.1, 9.2 +/- 2.3, and 2.6 +/- 2.2% of the first responses, respectively. Preintraoocyte injection of InsP(6) before ginsenoside treatment restored ginsenoside effect to initial response levels in a concentration-, time-, and structurally specific manner, in that inositol hexasulfate had no effect. The EC(50) was 13.9 +/- 8.7 microM. Injection of cRNA coding beta-arrestin I but not beta-arrestin II blocked InsP(6) effect on prevention of ginsenoside-induced Cl(-) channel desensitization. Injection of cRNA coding GRK2 abolished ginsenoside effect enhancing Cl(-) current. However, the GRK2-caused loss of ginsenoside effect on Cl(-) current was prevented by coinjection of GRK2 with GRK2-K220R, a dominant-negative mutant of GRK. These results indicate that ginsenoside-induced Cl(-) channel desensitization is mediated via activation of GRK2 and beta-arrestin I.
authors.affiliation: Research Laboratory for the Study of Ginseng Signal Transduction and Department of Physiology, College of Veterinary Medicine, College of Medicine, Konkuk University, Seoul 143-701 Korea.
authors.forename: Jun-Ho
authors.initials: JH
authors.lastname: Lee
authors.forename: Sang Min
authors.initials: SM
authors.lastname: Jeong
authors.forename: Byung-Hwan
authors.initials: BH
authors.lastname: Lee
authors.forename: Hye-Sung
authors.initials: HS
authors.lastname: Noh
authors.forename: Bo-Kyung
authors.initials: BK
authors.lastname: Kim
authors.forename: Jai-Il
authors.initials: JI
authors.lastname: Kim
authors.forename: Hyewhon
authors.initials: H
authors.lastname: Rhim
authors.forename: Hyoung-Chun
authors.initials: HC
authors.lastname: Kim
authors.forename: Kyeong-Man
authors.initials: KM
authors.lastname: Kim
authors.forename: Seung-Yeol
authors.initials: SY
authors.lastname: Nah
mesh_terms: D000818:Animals; D019390:Arrestins / Q000737:chemistry / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism*; D002712:Chlorides / Q000378:metabolism*; D002713:Chlorine / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D015536:Down-Regulation; D004594:Electrophysiology; D036145:Ginsenosides / Q000494:pharmacology*; D009865:Oocytes / Q000378:metabolism*; D010766:Phosphorylation; D010833:Phytic Acid / Q000378:metabolism*; D018075:RNA, Complementary / Q000378:metabolism; D015398:Signal Transduction; D013997:Time Factors; D014982:Xenopus laevis; D051552:beta-Adrenergic Receptor Kinases; D000071557:beta-Arrestins
chemical_list: D019390:Arrestins; D002712:Chlorides; D036145:Ginsenosides; D018075:RNA, Complementary; D000071557:beta-Arrestins; D002713:Chlorine; D010833:Phytic Acid; D017868:Cyclic AMP-Dependent Protein Kinases; D051552:beta-Adrenergic Receptor Kinases; D002118:Calcium