Excitatory GABAergic signalling is associated with acquired benzodiazepine resistance in status epilepticus

Abstract

Status epilepticus (SE) is defined as a state of unrelenting seizure activity. It is associated with a rapidly rising mortality rate, and thus constitutes a medical emergency. Benzodiazepines, which act as positive modulators of chloride (Cl-) permeable GABAA receptors, are indicated as the first line of treatment, but this is ineffective in many cases. We found that 48% of children presenting with SE were unresponsive to benzodiazepine treatment, and critically, that the duration of SE at the time of treatment is an important predictor of non-responsiveness. We therefore investigated the cellular mechanisms that underlie acquired benzodiazepine resistance, using rodent organotypic brain slices. Removing Mg2+ ions leads to an evolving pattern of epileptiform activity, and eventually to a persistent state of repetitive discharges that strongly resembles clinical EEG recordings of SE. We show that the persistent SE-like activity is associated with a reduction in GABAA receptor conductance and Cl- extrusion capability. We explored the effect on intraneuronal Cl- using both gramicidin, perforated-patch clamp recordings and Cl- imaging. This showed that during SE-like activity, reduced Cl- extrusion capacity was further exacerbated by activity-dependent Cl- loading, resulting in a persistently high intraneuronal Cl-. Consistent with these results, we found that optogenetic stimulation of GABAergic interneurons in the SE-like state, actually enhanced epileptiform activity in a GABAAR dependent manner. Together our findings describe a novel potential mechanism underlying benzodiazepine-resistant SE, with relevance to how this life-threatening condition should be managed in the clinic.