Quantitative fluorescence measurement of cardiac Na+/Ca2+ exchanger inhibition by kinetic analysis in stably transfected HEK293 cells.
We developed a method to quantitatively evaluate the potency of Na+/Ca2+ exchanger (NCX) inhibitors with fluorescence microscopy in NCX1-transfected HEK 293 cells. The reverse mode and forward mode NCX activities were measured as the ascending slope of the early phase increase in cytoplasmic Ca2+ concentration after change to low Na+ extracellular solution and the descending rate (inverse of the exponential time constant) on return to normal solution, respectively. Both modes of NCX were inhibited by SEA0400 (2-[4-[(2,5-difluorophenyl)methoxy]phenoxy]-5-ethoxyaniline) and KB-R7943 (2-[2-[4-(4-nitrobenzyloxy)phenyl]ethyl]isothiourea methanesulfonate), and the concentration-inhibition relationships for both inhibitors were in good agreement with those previously reported in voltage clamped cardiomyocytes.
Namekata
Iyuki
I
Department of Pharmacology, Toho University Faculty of Pharmaceutical Sciences, Funabashi, Chiba, Japan.
Kawanishi
Toru
T
Iida-Tanaka
Naoko
N
Tanaka
Hikaru
H
Shigenobu
Koki
K
D000814:Aniline Compounds / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism; D002417:Cattle; D002460:Cell Line; D005453:Fluorescence; D049452:Green Fluorescent Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D006801:Humans; D017136:Ion Transport / Q000187:drug effects; D007700:Kinetics; D010647:Phenyl Ethers / Q000494:pharmacology; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D012964:Sodium / Q000494:pharmacology; D019831:Sodium-Calcium Exchanger / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism*; D013890:Thiourea / Q000031:analogs & derivatives / Q000494:pharmacology; D014162:Transfection / Q000379:methods
C101670:2-(2-(4-(4-nitrobenzyloxy)phenyl)ethyl)isothiourea methanesulfonate; D000814:Aniline Compounds; D010647:Phenyl Ethers; D011993:Recombinant Fusion Proteins; C430918:SEA 0400; D019831:Sodium-Calcium Exchanger; D049452:Green Fluorescent Proteins; D012964:Sodium; D013890:Thiourea; D002118:Calcium