title: Regulation of the human organic cation transporter hOCT1.
abstract: The human organic cation transporter type 1 (hOCT1) is an important transport system for small organic cations in the liver. Organic cation transporters are regulated by different signaling pathways, but the regulation of hOCT1 has not yet been studied. In this work, we have for the first time investigated the regulation of hOCT1. hOCT1 was expressed in Chinese hamster ovary cells (CHO-hOCT1) and in human embryonic kidney cells (HEK293-hOCT1). Its activity was monitored using microfluorimetry with the fluorescent organic cation 4-(4-(dimethylamino)styryl)-N-methylpyridinium (ASP(+)) as substrate. hOCT1 expressed in CHO-cells was inhibited by protein kinase A (PKA) activation (1 microM forskolin, -58 +/- 6%, n = 12), calmodulin inhibition (0.1 microM calmidazolium, -68 +/- 3%, n = 6; 10 microM ophiobolin A, -48 +/- 10%, n = 7), calmodulin-dependent kinase II inhibition (1 microM KN62, -78 +/- 4%, n = 12), and inhibition of p56(lck) tyrosine kinase (10 microM aminogenistein, -35 +/- 7%, n = 12). The apparent affinities for TEA(+) were lower in CHO-hOCT1 than in HEK293-hOCT1, while those for TPA(+) and quinine were almost identical; the rank order of EC(50) values (TPA(+) > quinine > TEA(+)) was independent of the expression system. EC(50) values for TEA(+) in CHO-hOCT1 or HEK293-hOCT1 were increased under calmidazolium incubation (6.3 and 1.4 mM, respectively). hOCT1 was inhibited by PKA and endogenously activated by calmodulin, calmodulin-dependent kinase II, and p56(lck) tyrosine kinase. Regulation pathways were the same in the two expression systems. Since apparent substrate affinities depend on activity of regulatory pathways, the expression system plays a role in determining the substrate affinities.
authors.affiliation: Medizinische Klinik und Poliklinik D, Experimentelle Nephrologie, Universitätsklinikum Münster, D-48149 Münster, Germany.
authors.forename: Giuliano
authors.initials: G
authors.lastname: Ciarimboli
authors.forename: Katja
authors.initials: K
authors.lastname: Struwe
authors.forename: Petra
authors.initials: P
authors.lastname: Arndt
authors.forename: Valentin
authors.initials: V
authors.lastname: Gorboulev
authors.forename: Hermann
authors.initials: H
authors.lastname: Koepsell
authors.forename: Eberhard
authors.initials: E
authors.lastname: Schlatter
authors.forename: Jochen R
authors.initials: JR
authors.lastname: Hirsch
mesh_terms: D000818:Animals; D002147:Calmodulin / Q000037:antagonists & inhibitors / Q000378:metabolism; D002460:Cell Line; D006224:Cricetinae; D006152:Cyclic GMP / Q000378:metabolism; D004789:Enzyme Activation; D005453:Fluorescence; D005786:Gene Expression Regulation; D006801:Humans; D027702:Organic Cation Transporter 1 / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D011494:Protein Kinases / Q000378:metabolism; D011726:Pyridinium Compounds / Q000494:pharmacology; D043562:Receptors, G-Protein-Coupled / Q000378:metabolism; D013050:Spectrometry, Fluorescence; D013379:Substrate Specificity
chemical_list: C032315:4-(4-dimethylaminostyryl)-1-methylpyridinium; D002147:Calmodulin; D027702:Organic Cation Transporter 1; D011726:Pyridinium Compounds; D043562:Receptors, G-Protein-Coupled; D011494:Protein Kinases; D006152:Cyclic GMP