A medium-throughput functional assay of KCNQ2 potassium channels using rubidium efflux and atomic absorption spectrometry

Anal Biochem. 2003 Aug 15;319(2):251-7. doi: 10.1016/s0003-2697(03)00328-2.


Heterologous expression of KCNQ2 (Kv7.2) results in the formation of a slowly activating, noninactivating, voltage-gated potassium channel. Using a cell line that stably expresses KCNQ2, we developed a rubidium flux assay to measure the functional activity and pharmacological modulation of this ion channel. Rubidium flux was performed in a 96-well microtiter plate format; rubidium was quantified using an automated atomic absorption spectrometer to enable screening of 1000 data points/day. Cells accumulated rubidium at 37 degrees C in a monoexponential manner with t(1/2)=40min. Treating cells with elevated extracellular potassium caused membrane depolarization and stimulation of rubidium efflux through KCNQ2. The rate of rubidium efflux increased with increasing extracellular potassium: the t(1/2) at 50mM potassium was 5.1 min. Potassium-stimulated efflux was potentiated by the anticonvulsant drug retigabine (EC(50)=0.5 microM). Both potassium-induced and retigabine-facilitated efflux were blocked by TEA (IC(50)s=0.4 and 0.3mM, respectively) and the neurotransmitter release enhancers and putative cognition enhancers linopirdine (IC(50)s=2.3 and 7.1 microM, respectively) and XE991 (IC(50)s=0.3 and 0.9 microM, respectively). Screening a collection of ion channel modulators revealed additional inhibitors including clofilium (IC(50) = 27 microM). These studies extend the pharmacological profile of KCNQ2 and demonstrate the feasibility of using this assay system to rapidly screen for compounds that modulate the function of KCNQ2.

MeSH terms

  • Anthracenes / pharmacology
  • Carbamates / pharmacology
  • Cell Line
  • Clone Cells
  • Dose-Response Relationship, Drug
  • Humans
  • Indoles / pharmacology
  • KCNQ2 Potassium Channel
  • Kinetics
  • Phenylenediamines / pharmacology
  • Potassium Channel Blockers / pharmacology
  • Potassium Channels / drug effects
  • Potassium Channels / genetics
  • Potassium Channels / metabolism*
  • Potassium Channels, Voltage-Gated
  • Potassium Chloride / pharmacology
  • Pyridines / pharmacology
  • Quaternary Ammonium Compounds / pharmacology
  • Rubidium / analysis
  • Rubidium / metabolism*
  • Spectrophotometry, Atomic
  • Tetraethylammonium / pharmacology
  • Transfection


  • 10,10-bis(4-pyridinylmethyl)-9(10H)-anthracenone
  • Anthracenes
  • Carbamates
  • Indoles
  • KCNQ2 Potassium Channel
  • KCNQ2 protein, human
  • Phenylenediamines
  • Potassium Channel Blockers
  • Potassium Channels
  • Potassium Channels, Voltage-Gated
  • Pyridines
  • Quaternary Ammonium Compounds
  • ezogabine
  • Tetraethylammonium
  • Potassium Chloride
  • clofilium
  • linopirdine
  • Rubidium