title: The inwardly rectifying potassium channel Kir1.1: development of functional assays to identify and characterize channel inhibitors.
abstract: The renal outer medullary potassium (ROMK) channel is a member of the inwardly rectifying family of potassium (Kir) channels. ROMK (Kir1.1) is predominantly expressed in kidney where it plays a major role in the salt reabsorption process. Loss-of-function mutations in the human Kir1.1 channel are associated with antenatal Bartter's syndrome type II, a life-threatening salt and water balance disorder. Heterozygous carriers of Kir1.1 mutations associated with antenatal Bartter's syndrome have reduced blood pressure and a decreased risk of developing hypertension by age 60. These data suggest that Kir1.1 inhibitors could represent novel diuretics for the treatment of hypertension. Because little is known about the molecular pharmacology of Kir1.1 channels, assays that provide a robust, reliable readout of channel activity-while operating in high-capacity mode-are needed. In the present study, we describe high-capacity, 384- and 1,536-well plate, functional thallium flux, and IonWorks electrophysiology assays for the Kir1.1 channel that fulfill these criteria. In addition, 96-well (86)Rb(+) flux assays were established that can operate in the presence of 100% serum, and can provide an indication of the effect of a serum shift on compound potencies. The ability to grow Madin-Darby canine kidney cells expressing Kir1.1 in Transwell supports provides a polarized cell system that can be used to study the mechanism of Kir1.1 inhibition by different agents. All these functional Kir1.1 assays together can play an important role in supporting different aspects of drug development efforts during lead identification and/or optimization.
authors.affiliation: Department of Ion Channels, Merck Research Laboratories, Rahway, New Jersey, USA.
authors.forename: John P
authors.initials: JP
authors.lastname: Felix
authors.forename: Birgit T
authors.initials: BT
authors.lastname: Priest
authors.forename: Kelli
authors.initials: K
authors.lastname: Solly
authors.forename: Timothy
authors.initials: T
authors.lastname: Bailey
authors.forename: Richard M
authors.initials: RM
authors.lastname: Brochu
authors.forename: Chou J
authors.initials: CJ
authors.lastname: Liu
authors.forename: Martin G
authors.initials: MG
authors.lastname: Kohler
authors.forename: Laszlo
authors.initials: L
authors.lastname: Kiss
authors.forename: Magdalena
authors.initials: M
authors.lastname: Alonso-Galicia
authors.forename: Haifeng
authors.initials: H
authors.lastname: Tang
authors.forename: Alexander
authors.initials: A
authors.lastname: Pasternak
authors.forename: Gregory J
authors.initials: GJ
authors.lastname: Kaczorowski
authors.forename: Maria L
authors.initials: ML
authors.lastname: Garcia
mesh_terms: D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D003412:Cricetulus; D004285:Dogs; D055808:Drug Discovery / Q000379:methods*; D006801:Humans; D061985:Madin Darby Canine Kidney Cells; D026902:Potassium Channel Blockers / Q000097:blood / Q000737:chemistry / Q000378:metabolism* / Q000494:pharmacology*; D024661:Potassium Channels, Inwardly Rectifying / Q000037:antagonists & inhibitors* / Q000097:blood / Q000378:metabolism*; D051381:Rats; D013793:Thallium / Q000378:metabolism
chemical_list: C086989:KCNJ1 protein, human; C510692:Kcnj1 protein, rat; D026902:Potassium Channel Blockers; D024661:Potassium Channels, Inwardly Rectifying; D013793:Thallium