Status epilepticus: Role for etiology in determining response to benzodiazepines.

OBJECTIVE: Clinical factors contributing to benzodiazepine failure in treating status epilepticus (SE) include suboptimal dosing and seizure duration. As many benzodiazepine-refractory episodes of SE arise from acute etiologies, we sought to determine whether etiology impacts SE treatment. METHODS: The potency of diazepam to terminate SE induced by lithium-pilocarpine (LiPilo-SE) or kainic acid (KA-SE) in 3-week-old rats was studied by video-electroencephalography. Synaptic γ-aminobutyric acid type A receptor (GABAR)-mediated currents were recorded from dentate granule cells using voltage-clamp electrophysiology. Surface expression of γ2 subunit-containing GABARs and Kv4.2 potassium channels in hippocampal slices was determined using a biotinylation assay. Expression of phosphorylated forms of ß2/3 and γ2 subunits was determined using phosphospecific antibodies and Western blotting. RESULTS: Diazepam failed to terminate late SE in LiPilo-SE animals but was successful in terminating KA-SE of 1- and 3-hour duration. One hour after SE onset, GABAR-mediated synaptic inhibition and γ2 subunit-containing GABAR surface expression were reduced in LiPilo-SE animals. These were unchanged in KA-SE animals at 1 and 3 hours. Phosphorylation of γ2 subunit residue S327 was unchanged in both models, although GABAR ß3 subunit S408/409 residues were dephosphorylated in the LiPilo-SE animals. Kv4.2 potassium channel surface expression was increased in LiPilo-SE animals but reduced in KA-SE animals. INTERPRETATION: SE-model-dependent differences support a novel hypothesis that the development of benzodiazepine pharmacoresistance may be etiologically predetermined. Further studies are required to investigate the mechanisms that underlie such etiological differences during SE and whether etiology-dependent protocols for the treatment of SE need to be developed. Ann Neurol 2018;83:830-841.

Phosphatase inhibition prevents the activity-dependent trafficking of GABAA receptors during status epilepticus in the young animal.

OBJECTIVES: To determine if the activity-dependent trafficking of γ2 subunit-containing γ-aminobutyric acid type A receptors (GABAA Rs) that has been observed in older animals and posited to contribute to benzodiazepine pharmacoresistance during status epilepticus (SE) is age-dependent, and to evaluate whether blockade of protein phosphatases can inhibit or reverse the activity-dependent plasticity of these receptors. METHODS: The efficacy and potency of diazepam 0.2-10 mg/kg administered 3 or 60 min after the onset of a lithium/pilocarpine-induced seizure in postnatal day 15-16 rats was evaluated using video-electroencephalography (EEG) recordings. The surface expression of γ2 subunit-containing GABAA Rs was assessed using a biotinylation assay, and GABAA R-mediated miniature inhibitory postsynaptic currents (mIPSCs) were recorded using whole-cell patch-clamp recording techniques from dentate granule cells in hippocampal slices acutely obtained 60 min after seizure onset (SE-treated). The effect of the protein phosphatase inhibitors FK506 and okadaic acid (OA) on the surface expression of these receptors was determined in organotypic slice cultures exposed to high potassium and N-methyl-d-aspartate (NMDA) or in SE-treated slices. RESULTS: Diazepam terminated seizures of 3 min but not 60 min duration, even at the highest dose. In the SE-treated slices, the surface expression of γ2 subunit-containing GABAA Rs was reduced and the amplitude of the mIPSCs was diminished. Inhibition of protein phosphatases prevented the activity-induced reduction of the γ2 subunit-containing GABAA Rs in organotypic slice cultures. Furthermore, treatment of SE-treated slices with FK506 or OA restored the surface expression of the γ2 subunit-containing GABAA Rs and the mIPSC amplitude. SIGNIFICANCE: This study demonstrates that the plasticity of γ2 subunit-containing GABAA Rs associated with the development of benzodiazepine resistance in young and adult animals is similar. The findings of this study suggest that the mechanisms regulating the activity-dependent trafficking of GABAA Rs during SE can be targeted to develop novel adjunctive therapy for the treatment of benzodiazepine-refractory SE.