New Screening System for Selective Blockers of Voltage-Gated K⁺ Channels Using Recombinant Cell Lines Dying Upon Single Action Potential

  • Fujii Masato
    Department of Molecular and Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, Japan
  • Hayashi Keisuke
    Department of Molecular and Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, Japan
  • Ohya Susumu
    Department of Pharmacology, Division of Pathological Sciences, Kyoto Pharmaceutical University, Japan
  • Yamamura Hisao
    Department of Molecular and Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, Japan
  • Imaizumi Yuji
    Department of Molecular and Cellular Pharmacology, Graduate School of Pharmaceutical Sciences, Nagoya City University, Japan

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タイトル別名
  • New Screening System for Selective Blockers of Voltage-Gated K<sup>+</sup> Channels Using Recombinant Cell Lines Dying Upon Single Action Potential

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To develop a simple screening system for blockers of voltage-gated Kv1.3 and Kv1.5 channels, new cell lines co-expressing mutated Nav1.5 (IFM/Q3), Kir2.1 (Kir), and Kv1.3 or Kv1.5 were introduced as IFM/Q3+Kir+Kv1.3 and IFM/Q3+Kir+Kv1.5, respectively. Electrical stimulation (ES) of a cell line, IFM/Q3+Kir, induced prolonged action potentials due to the slow inactivation of IFM/Q3 and subsequent cell death. Additional co-expression of Kv1.3 or Kv1.5 to IFM/Q3+Kir shortened the evoked action potentials and prevented cell death. In the presence of margatoxin, a selective Kv1.3-blocker, ES induced cell death in IFM/Q3+Kir+Kv1.3, but not in IFM/Q3+Kir+Kv1.5. In the presence of 4-aminopyridine, a non-selective Kv-channel blocker, ES application elicited cell death in both cell lines. The IC50s of acacetin, a Kv1.5-blocker, was 10.2 μM in IFM/Q3+Kir+Kv1.3 and almost identical to that in IFM/Q3+Kir+Kv1.5 (7.6 μM). The IC50s of citalopram, a 5-HT uptake-inhibitor, were 1.8 μM in IFM/Q3+Kir+Kv1.3 and 1.5 μM in IFM/Q3+Kir+Kv1.5, respectively. These IC50s were comparable to those determined electrophysiologically. In conclusion, acacetin and citalopram block both Kv1.3 and Kv1.5 without selectivity. The Kv1.3 or Kv1.5 channel inhibition assay using these new cell lines may be applicable to high–throughput screening because of its simplicity, accuracy, and high cost-performance.

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