Abstracts

Rationale: Exposure to organophosphates (OP) or organophosphate nerve agents (NA), results in status epilepticus (SE) and neuronal damage in the central nervous system. Early control of seizure activity minimizes mortality and neuronal damage; however, in the event of a mass release (civilian or military), treatment is likely to be delayed, resulting in more severe long-term consequences. The CounterACT (Countermeasures Against Chemical Threats) Neurotherapeutic Screening Program was established to identify novel treatments for OP/NA-induced SE that can be administered at significant delay from exposure in a pre-hospital setting. In our recent year of screening, we have evaluated the antiseizure and neuroprotective effects of six novel compounds.  Methods: Male Sprague Dawley rats (150-200 g) were implanted with recording electrodes for collection of the electroencephalogram (EEG). Then, after a 1-week recovery period, SE was induced by administration of diisopropyl fluorophosphate (DFP). In addition to standard OP/NA anti-dotes, 1 h after the start of SE, rats were co-administered midazolam (MDZ) and test compound. EEG was recorded for 24 h, followed by perfusion, and then brains were collected for Fluoro-Jade B labeling. Fluoro-Jade B positive neurons were counted in 10 regions: dorsal CA1, dorsal CA3, hilus, ventral CA1, ventral CA3, amygdala, thalamus, and the parietal, entorhinal and piriform cortices. In conducting research using animals, the investigators adhered to the Animal Welfare Act Regulations and other Federal statutes relating to animals and experiments involving animals and the principles set forth in the current version of the Guide for Care and Use of Laboratory Animals, National Research Council. Results: Three compounds had no significant anti-seizure effects in the presence of MDZ. These compounds also had no effect on neuronal death in the presence of MDZ. In contrast, we observed that MDZ + dextromethorphan (n = 16, control MDZ+ vehicle n = 15) significantly reduced neuronal death (P < 0.05), although this compound did not alter the MDZ-induced reduction of SE. Finally, MDZ + retigabine (n = 19) significantly enhanced the anti-seizure effect of MDZ + vehicle (n = 18, P < 0.05), effectively eliminating seizure activity for the duration of the treatment period. MDZ + retigabine treatment also reduced neuronal death. Assessed on a global scale (i.e., 10 brain regions), we observed a reduction to approximately one-third of the neuronal death observed in the MDZ + vehicle controls (P < 0.05, Wilcoxon signed-rank test).  Conclusions: The data presented here demonstrate that compounds with dextromethorphan-like mechanisms may be useful in reducing neuronal death following OP/NA-induced SE, while compounds with retigabine-like mechanisms may be useful for reduction of both seizure activity and neuronal death, even when treatment is delayed.  Funding: This work is supported by the US Army Medical Research Institute of Chemical Defense under Contract No. W81XWH-18-C-0181. The views, opinions and/or findings contained in this report are those of the authors and should not be construed as an official Department of the Army position, policy or decision unless so designated by other documentation.