Abstracts

Rationale: The organophosphate, DFP (diisopropyl fluorophosphate) is considered a surrogate for nerve agents soman and sarin as acute exposure inhibits acetylcholinesterase and induces status epilepticus (SE), triggering epileptogenesis. Although agents such as diazepam are used to suppress seizures following intoxication, further intervention is needed to suppress the long-term brain injury that could lead to epileptogenesis. We have previously shown that SE upregulates the inducible nitric oxide synthase (iNOS) in glial cells which mediates the production of free radicals to cause nitro-oxidative stress in the kainic acid (KA) rat model. Treating with 1400W suppressed nitro-oxidative stress, neuroinflammation, neurodegeneration, and significantly reduced spontaneous seizures (>90%) in a six months study. We therefore hypothesized that 1400W could be a disease modifier in the rat DFP model of toxicity. Methods: Adult male Sprague Dawley rats were randomized, grouped and coded, and measured for baseline behavior before experimentation. Some rats were implanted with radio-telemetry devices 10 days prior exposure to DFP (3-4 mg/kg, s.c) followed immediately by atropine sulfate (2 mg/kg, i.m.) and pralidoxime (25 mg/kg. i.m). After two hours, behavioral SE was halted by diazepam administration (5 mg/kg, i.p.). Following a further two hours, rats were administered 6 doses 1400W (20 mg/kg, i.m.) or vehicle every 12 hours. Rats were video EEG monitored continuously for three months; spontaneous seizures and epileptiform spikes were quantified using Neuroscore software. Additional rats were given a similar treatment, without telemetry devices, and various behavioral measures such as forced swim test, rotarod, horizontal bar test, elevated T-maze, and Morris water maze (MWM) were undertaken.  All rats were euthanized with pentobarbital (100 mg/kg, i.p) for histological analysis as well as multiplex assay. Results: DFP treated rats developed SE in <10 min. Two hours SE was quantified and there was no statistical significance between the 2 groups. Three months EEG quantification revealed a significant reduction in epileptiform spikes, spontaneous seizures, and spike trains in 1400W group (p<0.05). The DFP+ vehicle group compared to the DFP+1400W group had an increase in microglia (IBA1) with reactive-type morphology (p<0.05) and Flurojade-B +NeuN positive cells in the CA1 and CA3 regions of the hippocampus, as well as certain pro-inflammatory/chemokines such as IL-13, TNF-a, LIX, fractalkine, and MCP-1  (p<0.05). Learning was unaffected, however, 1400W appears to mitigate DFP-induced progressive loss of memory. Although overall motor deficits and depression-like behavior were marginal, 1400W appears to mitigate the deficit during acceleration. Conclusions: 1400W treatment during the first three days following the acute exposure to DFP suppressed long-term hyperexcitability, neuroinflammation and neurodegeneration. Although behavioral deficits were marginal, 1400W prevented the progressive loss of memory. Funding: This project is funded by the NIH (NS099007).