title: PKC-dependent activation of human K(2P) 18.1 K(+)  channels.
abstract: OBJECTIVE
  Two-pore-domain K(+) channels (K(2P) ) mediate K(+) background currents that modulate the membrane potential of excitable cells. K(2P) 18.1 (TWIK-related spinal cord K(+) channel) provides hyperpolarizing background currents in neurons. Recently, a dominant-negative loss-of-function mutation in K(2P) 18.1 has been implicated in migraine, and activation of K(2P) 18.1 channels was proposed as a therapeutic strategy. Here we elucidated the molecular mechanisms underlying PKC-dependent activation of K(2P) 18.1 currents.
  
  
  METHODS
  Human K(2P) 18.1 channels were heterologously expressed in Xenopus laevis oocytes, and currents were recorded with the two-electrode voltage clamp technique.
  
  
  RESULTS
  Stimulation of PKC using phorbol 12-myristate-13-acetate (PMA) activated the hK(2P) 18.1 current by 3.1-fold in a concentration-dependent fashion. The inactive analogue 4α-PMA had no effect on channel activity. The specific PKC inhibitors bisindolylmaleimide I, Ro-32-0432 and chelerythrine reduced PMA-induced channel activation indicating that PKC is involved in this effect of PMA. Selective activation of conventional PKC isoforms with thymeleatoxin (100 nM) did not reproduce K(2P) 18.1 channel activation. Current activation by PMA was not affected by pretreatment with CsA (calcineurin inhibitor) or KT 5720 (PKA inhibitor), ruling out a significant contribution of calcineurin or cross-talk with PKA to the PKC-dependent hK(2P) 18.1 activation. Finally, mutation of putative PKC phosphorylation sites did not prevent PMA-induced K(2P) 18.1 channel activation.
  
  
  CONCLUSIONS
  We demonstrated that activation of hK(2P) 18.1 (TRESK) by PMA is mediated by PKC stimulation. Hence, PKC-mediated activation of K(2P) 18.1 background currents may serve as a novel molecular target for migraine treatment.
authors.affiliation: Department of Cardiology, Medical University Hospital Heidelberg, Heidelberg, Germany.
authors.forename: Ann-Kathrin
authors.initials: AK
authors.lastname: Rahm
authors.forename: Jakob
authors.initials: J
authors.lastname: Gierten
authors.forename: Jana
authors.initials: J
authors.lastname: Kisselbach
authors.forename: Ingo
authors.initials: I
authors.lastname: Staudacher
authors.forename: Kathrin
authors.initials: K
authors.lastname: Staudacher
authors.forename: Patrick A
authors.initials: PA
authors.lastname: Schweizer
authors.forename: Rüdiger
authors.initials: R
authors.lastname: Becker
authors.forename: Hugo A
authors.initials: HA
authors.lastname: Katus
authors.forename: Dierk
authors.initials: D
authors.lastname: Thomas
mesh_terms: D000818:Animals; D006801:Humans; D009865:Oocytes / Q000187:drug effects / Q000502:physiology; D024683:Potassium Channels, Tandem Pore Domain / Q000502:physiology*; D011493:Protein Kinase C / Q000502:physiology*; D015398:Signal Transduction; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014982:Xenopus laevis
chemical_list: D024683:Potassium Channels, Tandem Pore Domain; D011493:Protein Kinase C; D013755:Tetradecanoylphorbol Acetate