title: Differential sensitivity of TREK-1, TREK-2 and TRAAK background potassium channels to the polycationic dye ruthenium red.
abstract: OBJECTIVE
  Pharmacological separation of the background potassium currents of closely related K2P channels is a challenging problem. We previously demonstrated that ruthenium red (RR) inhibits TASK-3 (K2 P 9.1), but not TASK-1 (K2 P 3.1) channels. RR has been extensively used to distinguish between TASK currents in native cells. In the present study, we systematically investigate the RR sensitivity of a more comprehensive set of K2 P channels.
  
  
  METHODS
  K(+) currents were measured by two-electrode voltage clamp in Xenopus oocytes and by whole-cell patch clamp in mouse dorsal root ganglion (DRG) neurons.
  
  
  RESULTS
  RR differentiates between two closely related members of the TREK subfamily. TREK-2 (K2 P 10.1) proved to be highly sensitive to RR (IC50 = 0.2 μM), whereas TREK-1 (K2 P 2.1) was not affected by the compound. We identified aspartate 135 (D135) as the target of the inhibitor in mouse TREK-2c. D135 lines the wall of the extracellular ion pathway (EIP), a tunnel structure through the extracellular cap characteristic for K2 P channels. TREK-1 contains isoleucine in the corresponding position. The mutation of this isoleucine (I110D) rendered TREK-1 sensitive to RR. The third member of the TREK subfamily, TRAAK (K2 P 4.1) was more potently inhibited by ruthenium violet, a contaminant in some RR preparations, than by RR. DRG neurons predominantly express TREK-2 and RR-resistant TREK-1 and TRESK (K2 P 18.1) background K(+) channels. We detected the RR-sensitive leak K(+) current component in DRG neurons.
  
  
  CONCLUSIONS
  We propose that RR may be useful for distinguishing TREK-2 (K2P 10.1) from TREK-1 (K2P 2.1) and other RR-resistant K2 P channels in native cells.
authors.affiliation: Department of Physiology, Semmelweis University, Budapest, Hungary.
authors.forename: G
authors.initials: G
authors.lastname: Braun
authors.forename: M
authors.initials: M
authors.lastname: Lengyel
authors.forename: P
authors.initials: P
authors.lastname: Enyedi
authors.forename: G
authors.initials: G
authors.lastname: Czirják
mesh_terms: D000818:Animals; D004396:Coloring Agents / Q000494:pharmacology*; D005727:Ganglia, Spinal / Q000166:cytology; D051379:Mice; D009474:Neurons / Q000187:drug effects / Q000502:physiology; D009865:Oocytes / Q000502:physiology; D015221:Potassium Channels / Q000235:genetics / Q000502:physiology*; D012430:Ruthenium Red / Q000494:pharmacology*; D014981:Xenopus
chemical_list: D004396:Coloring Agents; D015221:Potassium Channels; D012430:Ruthenium Red