title: A potassium leak channel silences hyperactive neurons and ameliorates status epilepticus.
abstract: OBJECTIVE
  To develop a constitutively active K(+) leak channel using TREK-1 (TWIK-related potassium channel 1; TREK-M) that is resistant to compensatory down-regulation by second messenger cascades, and to validate the ability of TREK-M to silence hyperactive neurons using cultured hippocampal neurons. To test if adenoassociated viral (AAV) delivery of TREK-M could reduce the duration of status epilepticus and reduce neuronal death induced by lithium-pilocarpine administration.
  
  
  METHODS
  Molecular cloning techniques were used to engineer novel vectors to deliver TREK-M via plasmids, lentivirus, and AAV using a cytomegalovirus (CMV)-enhanced GABRA4 promoter. Electrophysiology was used to characterize the activity and regulation of TREK-M in human embryonic kidney (HEK-293) cells, and the ability to reduce spontaneous activity in cultured hippocampal neurons. Adult male rats were injected bilaterally with self-complementary AAV particles composed of serotype 5 capsid into the hippocampus and entorhinal cortex. Lithium-pilocarpine was used to induce status epilepticus. Seizures were monitored using continuous video-electroencephalography (EEG) monitoring. Neuronal death was measured using Fluoro-Jade C staining of paraformaldehyde-fixed brain slices.
  
  
  RESULTS
  TREK-M inhibited neuronal firing by hyperpolarizing the resting membrane potential and decreasing input resistance. AAV delivery of TREK-M decreased the duration of status epilepticus by 50%. Concomitantly it reduced neuronal death in areas targeted by the AAV injection.
  
  
  CONCLUSIONS
  These findings demonstrate that TREK-M can silence hyperexcitable neurons in the brain of epileptic rats and treat acute seizures. This study paves the way for an alternative gene therapy treatment of status epilepticus, and provides the rationale for studies of AAV-TREK-M's effect on spontaneous seizures in chronic models of temporal lobe epilepsy.
authors.affiliation: Department of Pharmacology, Graduate Program University of Virginia School of Medicine, Charlottesville, Virginia, U.S.A; Department of Neuroscience, Graduate Program University of Virginia School of Medicine, Charlottesville, Virginia, U.S.A.
authors.forename: Deblina
authors.initials: D
authors.lastname: Dey
authors.forename: Veit-Simon
authors.initials: VS
authors.lastname: Eckle
authors.forename: Iuliia
authors.initials: I
authors.lastname: Vitko
authors.forename: Kyle A
authors.initials: KA
authors.lastname: Sullivan
authors.forename: Zofia M
authors.initials: ZM
authors.lastname: Lasiecka
authors.forename: Bettina
authors.initials: B
authors.lastname: Winckler
authors.forename: Ruth L
authors.initials: RL
authors.lastname: Stornetta
authors.forename: John M
authors.initials: JM
authors.lastname: Williamson
authors.forename: Jaideep
authors.initials: J
authors.lastname: Kapur
authors.forename: Edward
authors.initials: E
authors.lastname: Perez-Reyes
mesh_terms: D000818:Animals; D016923:Cell Death / Q000235:genetics; D016764:Cell Polarity / Q000235:genetics; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000008:administration & dosage / Q000235:genetics; D057809:HEK293 Cells; D006801:Humans; D008297:Male; D009433:Neural Inhibition / Q000235:genetics; D009474:Neurons / Q000473:pathology* / Q000502:physiology; D024683:Potassium Channels, Tandem Pore Domain / Q000008:administration & dosage / Q000235:genetics*; D051381:Rats; D017207:Rats, Sprague-Dawley; D013226:Status Epilepticus / Q000235:genetics* / Q000473:pathology / Q000517:prevention & control*
chemical_list: D024683:Potassium Channels, Tandem Pore Domain; C103742:potassium channel protein TREK-1
keywords: Gene therapy; Hippocampus; Potassium currents; Status epilepticus; T-type calcium channel; Temporal lobe epilepsy