Enaminone Modulators of Extrasynaptic α4β3δ γ-Aminobutyric AcidA Receptors Reverse Electrographic Status Epilepticus in the Rat After Acute Organophosphorus Poisoning

Abstract

Seizures induced by organophosphorus nerve agent exposure become refractory to treatment with benzodiazepines because these drugs engage synaptic γ-aminobutyric acid-A receptors (GABA A Rs) that rapidly internalize during status epilepticus (SE). Extrasynaptic GABA A Rs, such as those containing α 4 β 3 δ subunits, are a putative pharmacological target to comprehensively manage nerve agent-induced seizures since they do not internalize during SE and are continuously available for activation. Neurosteroids related to allopregnanolone have been tested as a possible replacement for benzodiazepines because they target both synaptic and extrasynaptic GABA A Rs receptors. A longer effective treatment window, extended treatment efficacy, and enhanced neuroprotection represent significant advantages of neurosteroids over benzodiazepines. However, neurosteroid use is limited by poor physicochemical properties arising from the intrinsic requirement of the pregnane steroid core structure for efficacy rendering drug formulation problematic. We tested a non-steroidal enaminone GABA A R modulator that interacts with both synaptic and extrasynaptic GABA A Rs on a binding site distinct from neurosteroids or benzodiazepines for efficacy to control electrographic SE induced by diisopropyl fluorophosphate or soman intoxication in rats. Animals were treated with standard antidotes, and experimental therapeutic treatment was given following 1 h (diisopropyl fluorophosphate model) or 20 min (soman model) after SE onset. We found that the enaminone 2-261 had an extended duration of seizure termination (>10 h) in the diisopropyl fluorophosphate intoxication model in the presence or absence of midazolam (MDZ). 2-261 also moderately potentiated MDZ in the soman-induced seizure model but had limited efficacy as a stand-alone anticonvulsant treatment due to slow onset of action. 2-261 significantly reduced neuronal death in brain areas associated with either diisopropyl fluorophosphate- or soman-induced SE. 2-261 represents an alternate chemical template from neurosteroids for enhancing extrasynaptic α 4 β 3 δ GABA A R activity to reverse SE from organophosphorous intoxication.

Refractory and Super-Refractory Status Epilepticus in Nerve Agent-Poisoned Rats Following Application of Standard Clinical Treatment Guidelines

“…Cohorts of eight rats were placed into individual Plexiglas R chambers and connected to an EEG monitoring system (MP160, BIOPAC Systems Inc., Goleta, CA, United States) to enable realtime display of brain activity using AcqKnowledge R 5.0 software. After approximately 30 min of baseline, NA exposure proceeded as previously described and shown in Figure 1 (Althaus et al, 2017;McCarren et al, 2018;Jackson et al, 2019;Johnstone et al, 2019;Barker et al, 2020). The NA used was soman (150 µg/kg s.c.).…”

New efficient synthesis, spectroscopic characterization, and X-ray analysis of novel β-enaminocarboxamide derivatives

“…In the literature, the introduction of the amide group into β-enaminones structure has been described and have interesting biological properties [33,34]. In Fig.…”

Delayed Adjunctive Treatment of Organophosphate-Induced Status Epilepticus in Rats with Phenobarbital, Memantine, or Dexmedetomidine

“…As part of the CounterACT Neurotherapeutics Screening (CNS) Program, we have previously developed a rat model of delayed treatment of OP-induced SE, using the OP insecticide diisopropyl fluorophosphates (DFP), to screen compounds for efficacy in the termination of seizures and reduction of neuronal death (Pouliot et al, 2016;Johnstone et al, 2019;Spampanato et al, 2019;Barker et al, 2020). These previous studies have demonstrated that delayed treatment with MDZ (including standardof-care antidotes) results in a transient reduction in seizure activity that is also associated with a reduction in neuronal loss; however, the duration and magnitude of seizure suppression and the reduction of neuronal loss were minimal (Spampanato et al, 2019) and could be enhanced by adjunct treatment (Johnstone et al, 2019;Barker et al, 2020). To further validate our model for screening of test compounds, we have tested three adjunct therapies -each with unique mechanisms.…”

“…To further validate our model for screening of test compounds, we have tested three adjunct therapies -each with unique mechanisms. Three drugs were selected by the NIH CNS Program because (1) they are currently approved for use in humans, (2) they have demonstrated some efficacy in reduction of seizure activity and/or neuronal death in humans All experimental procedures were conducted as similarly described (Johnstone et al, 2019;Spampanato et al, 2019;Barker et al, 2020).…”