Characterization of recombinant human cardiac KCNQ1/KCNE1 channels (I (Ks)) stably expressed in HEK 293 cells.
The present study was designed to characterize pharmacological, biophysical and electrophysiological properties of the recombinant human cardiac I (Ks) (KCNQ1/KCNE1) channels at physiological temperature. Human cardiac KCNQ1 and KCNE1 genes were cotransfected into HEK 293 cells, and a cell clone stably expressing both genes was selected. Membrane currents were recorded using a perforated patch-clamp technique. The typical I (Ks) was slowly activated upon depolarization voltages in HEK 293 cells stably expressing human cardiac KCNQ1 and KCNE1 genes, and the current was inhibited by I (Ks) blockers HMR 1556 and chromanol 293B, with 50% inhibitory concentrations (IC(50)s) of 83.8 nM: and 9.2 muM: , respectively. I (Ks) showed a significant temperature-dependent increase in its magnitude upon elevating bath temperature to 36 degrees C from room temperature (21 degrees C). The current was upregulated by the beta-adrenoceptor agonist isoproterenol, and the effect was reversed by H89. In addition, I (Ks) was inhibited by Ba(2+) in a concentration-dependent manner (IC(50) = 1.4 mM). Action potential clamp revealed a "bell-shaped" time course of I (Ks) during the action potential, and maximal peak current was seen at the plateau of the action potential. A significant use- and frequency-dependent increase of I (Ks) was observed during a train of action potential clamp. These results indicate that the recombinant human cardiac I (Ks) stably expressed in HEK 293 cells is similar to native I (Ks) in drug sensitivity and regulated by Ba(2+) and beta-adrenoceptor via the cyclic adenosine monophosphate/protein kinase A pathway. Importantly, the current exhibits significant temperature dependence, a bell-shaped time course during action potential and prominent use- or frequency-dependent accumulation during a train of action potentials.
Dong
Ming-Qing
MQ
Department of Medicine and Research Centre of Heart, Brain, Hormone, and Healthy Aging, Faculty of Medicine, University of Hong Kong, Pokfulam, Hong Kong, People's Republic of China.
Lau
Chu-Pak
CP
Gao
Zhan
Z
Tseng
Gea-Ny
GN
Li
Gui-Rong
GR
D000200:Action Potentials / Q000187:drug effects / Q000502:physiology*; D000318:Adrenergic beta-Agonists / Q000494:pharmacology; D001464:Barium / Q000494:pharmacology; D002478:Cells, Cultured; D002839:Chromans / Q000494:pharmacology; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D005786:Gene Expression Regulation / Q000502:physiology*; D006801:Humans; D007545:Isoproterenol / Q000494:pharmacology; D007546:Isoquinolines / Q000494:pharmacology; D051657:KCNQ1 Potassium Channel / Q000037:antagonists & inhibitors / Q000502:physiology*; D007668:Kidney / Q000196:embryology / Q000378:metabolism*; D008564:Membrane Potentials / Q000187:drug effects; D018408:Patch-Clamp Techniques; D026902:Potassium Channel Blockers / Q000494:pharmacology; D024642:Potassium Channels, Voltage-Gated / Q000037:antagonists & inhibitors / Q000502:physiology*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011994:Recombinant Proteins / Q000378:metabolism; D013449:Sulfonamides / Q000494:pharmacology; D014162:Transfection
D000318:Adrenergic beta-Agonists; D002839:Chromans; C420023:HMR 1556; D007546:Isoquinolines; C489622:KCNE1 protein, human; D051657:KCNQ1 Potassium Channel; D026902:Potassium Channel Blockers; D024642:Potassium Channels, Voltage-Gated; D047428:Protein Kinase Inhibitors; D011994:Recombinant Proteins; D013449:Sulfonamides; C103474:6-cyano-4-(N-ethylsulfonyl-N-methylamino)-3-hydroxy-2,2-dimethylchromane; D001464:Barium; D017868:Cyclic AMP-Dependent Protein Kinases; D007545:Isoproterenol; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide