title: Persistent human cardiac Na+ currents in stably transfected mammalian cells: Robust expression and distinct open-channel selectivity among Class 1 antiarrhythmics.
abstract: Miniature persistent late Na(+) currents in cardiomyocytes have been linked to arrhythmias and sudden death. The goals of this study are to establish a stable cell line expressing robust persistent cardiac Na(+) currents and to test Class 1 antiarrhythmic drugs for selective action against resting and open states. After transient transfection of an inactivation-deficient human cardiac Na(+) channel clone (hNav1.5-CW with L409C/A410W double mutations), transfected mammalian HEK293 cells were treated with 1 mg/ml G-418. Individual G-418-resistant colonies were isolated using glass cylinders. One colony with high expression of persistent Na(+) currents was subjected to a second colony selection. Cells from this colony remained stable in expressing robust peak Na(+) currents of 996 ± 173 pA/pF at +50 mV (n = 20). Persistent late Na(+) currents in these cells were clearly visible during a 4-second depolarizing pulse albeit decayed slowly. This slow decay is likely due to slow inactivation of Na(+) channels and could be largely eliminated by 5 μM batrachotoxin. Peak cardiac hNav1.5-CW Na(+) currents were blocked by tetrodotoxin with an IC(50) value of 2.27 ± 0.08 μM (n = 6). At clinic relevant concentrations, Class 1 antiarrhythmics are much more selective in blocking persistent late Na(+) currents than their peak counterparts, with a selectivity ratio ranging from 80.6 (flecainide) to 3 (disopyramide). We conclude that (1) Class 1 antiarrhythmics differ widely in their resting- vs. open-channel selectivity, and (2) stably transfected HEK293 cells expressing large persistent hNav1.5-CW Na(+) currents are suitable for studying as well as screening potent open-channel blockers.
authors.forename: Ging Kuo
authors.initials: GK
authors.lastname: Wang
authors.forename: Gabriella
authors.initials: G
authors.lastname: Russell
authors.forename: Sho-Ya
authors.initials: SY
authors.lastname: Wang
mesh_terms: D000889:Anti-Arrhythmia Agents / Q000494:pharmacology*; D001496:Batrachotoxins / Q000494:pharmacology; D055724:Electrophysiological Phenomena / Q000187:drug effects*; D057809:HEK293 Cells; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects*; D009154:Mutation; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism*; D062554:NAV1.5 Voltage-Gated Sodium Channel / Q000235:genetics* / Q000378:metabolism*; D013997:Time Factors; D014162:Transfection
chemical_list: D000889:Anti-Arrhythmia Agents; D001496:Batrachotoxins; D062554:NAV1.5 Voltage-Gated Sodium Channel; C568320:SCN5A protein, human