title: Pregabalin activates ROMK1 channels via cAMP-dependent protein kinase and protein kinase C.
abstract: Pregabalin (PGB) displays analgesic and anticonvulsant activities. Regulation of the resting membrane potential (RMP) by renal outer medullary potassium (ROMK1) channels may provide a mechanism for these activities. We examined the effects of PGB on ROMK1 channel activity. To investigate the regulatory effect of PGB on the activity of ROMK1 channel, we used inside-out excised membrane patches to measure the K+ current in Xenopus oocytes expressed either the wild-type (WT) or mutant ROMK1 channels. PGB concentration-dependently enhanced the activity of ROMK1 channels. PGB increases the WT channels, pHi gating residue mutant channels (K80M) and the mutant channels at phosphatidylinositol bisphosphate (PIP2)-binding sites (R188Q, R217A, and K218A). Our study suggests that PGB in the regulating of ROMK1 channel function are neither by pHi- nor PIP2-dependent mechanism. We found PGB failed to prompt the activity of consensus phosphorylation sites for protein kinase C (PKC) mutated channels (S183A, T191A, T193A, S201A and T234A). Furthermore, PGB did not stimulate the activity of channels in the presence of cAMP-dependent protein kinase (PKA) inhibitors, the mutants of the C-terminal PKA-phosphorylation sites (S219A and S313A), and the mutants constructed (S219D and S313D) which mimic the addition of negative charged associated with phosphorylation bound to a serine. These results demonstrated that PKA- and PKC-mediated phosphorylation represents a novel mechanism for PGB-activated ROMK1 channels. The enhancement of ROMK1 currents proves to an important molecular mechanism underlying the analgesic/anticonvulsant property of PGB for the restoration of RMP.
authors.affiliation: Department of Pharmacology, College of Medicine, National Taiwan University, Taipei, Taiwan; Department of Nursing, Min-Hwei Junior College of Health Care Management, Tainan, Taiwan.
authors.forename: Chien-Hsing
authors.initials: CH
authors.lastname: Lee
authors.affiliation: Department of Pharmacology, College of Medicine, National Taiwan University, Taipei, Taiwan; Department of Neurology, National Taiwan University Hospital, Taipei, Taiwan. Electronic address: hhl@ntu.edu.tw.
authors.forename: Horng-Huei
authors.initials: HH
authors.lastname: Liou
mesh_terms: D000700:Analgesics / Q000494:pharmacology*; D000818:Animals; D000927:Anticonvulsants / Q000494:pharmacology*; D002227:Carbazoles / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D005260:Female; D007546:Isoquinolines / Q000494:pharmacology; D009865:Oocytes; D024661:Potassium Channels, Inwardly Rectifying / Q000235:genetics / Q000378:metabolism*; D000069583:Pregabalin; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011758:Pyrroles / Q000494:pharmacology; D013449:Sulfonamides / Q000494:pharmacology; D014982:Xenopus laevis; D005680:gamma-Aminobutyric Acid / Q000031:analogs & derivatives* / Q000494:pharmacology
chemical_list: D000700:Analgesics; D000927:Anticonvulsants; D002227:Carbazoles; D007546:Isoquinolines; D024661:Potassium Channels, Inwardly Rectifying; D047428:Protein Kinase Inhibitors; D011758:Pyrroles; D013449:Sulfonamides; D000069583:Pregabalin; D005680:gamma-Aminobutyric Acid; C057416:KT 5720; D017868:Cyclic AMP-Dependent Protein Kinases; D011493:Protein Kinase C; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide
keywords: PKA; PKC; Pregabalin; ROMK1 channels