Antiseizure and neuroprotective effects of delayed treatment with midazolam in a rodent model of organophosphate exposure

Abstract

Objective Exposure to organophosphates (OPs) and OP nerve agents (NAs) causes status epilepticus (SE) and irreversible brain damage. Rapid control of seizure activity is important to minimize neuronal injury and the resulting neurological and behavioral disorders; however, early treatment will not be possible after mass release of OPs or NAs. Methods We utilized a delayed‐treatment model of OP exposure in adult rats by administration of diisopropyl fluorophosphate (DFP) to study the relationship between the antiseizure and neuroprotective effects of the “standard‐of‐care” benzodiazepine, midazolam (MDZ), when given at 30, 60, and 120 minutes after SE onset. After electroencephalography (EEG) recordings, neural damage in serial brain sections was studied with Fluoro‐Jade B staining. Results MDZ‐induced seizure suppression was equivalent in magnitude regardless of treatment delay (ie, seizure duration). When assessed globally (ie, normalized across 10 different brain regions) for each treatment delay, MDZ administration resulted in only nonsignificant reductions in neuronal death. However, when data for MDZ treatment were combined from all three delay times, a small but significant reduction in global neuronal death was detected when compared to vehicle treatment, which indicated that the substantive MDZ‐induced seizure suppression led to only a small reduction in neuronal death. Significance In conclusion, MDZ significantly reduced DFP‐induced SE intensity when treatment was delayed 30, 60, and even up to 120 minutes; however, this reduction in seizure intensity had no detectable effect on neuronal death at each individual delay time. These data show that although MDZ suppressed seizures, additional neuroprotective therapies are needed to mitigate the effects of OP exposure.

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

“…The data presented in this report expand on our previously published studies concerning the development of a rodent model, where ASDs and neuroprotective agents can be assessed as adjunctive therapy with delayed administration of MDZ (Pouliot et al, 2016;Spampanato et al, 2019), which is currently the standard-of-care for a potential mass exposure of a nerve agent or other organophosphate chemical threat agent. Using the identical screening protocol, we found in our previous study (Spampanato et al, 2019), that treatment with MDZ alone at 1 h after DFP-induced SE resulted in little or no effect on neuronal loss (i.e., no statistically significant effect; the MDZ-treated rats had, on average, 72 + 47% of the neuronal death observed in untreated controls). Because this reduction in neuronal death was not significant, the effect of MDZ + vehicle controls in the present study would be equivalent in neuronal death to that of an untreated group, and was interpreted as such throughout this study.…”

“…Our previously published, delayed-treatment rodent model of OP exposure, whereby SE was induced with DFP, was used in these experiments (Pouliot et al, 2016;Spampanato et al, 2019). To decrease mortality due to the peripheral, lethal effects of the OP, and to model realistic antidote treatments, rats (150-240 g) were given pyridostigmine bromide (0.026 mg/kg, i.m.)…”

“…Exposure to DFP resulted in a rapid transition to electrographic SE (Pouliot et al, 2016) that was transiently reversed by treatment with MDZ (Spampanato et al, 2019), which can be seen in the compressed, raw EEG trace presented in Figure 1A for an MDZ + multisol (vehicle) treated rat (top trace). Addition of PHB (100 mg/kg) produced a more rapid offset of seizure activity that persisted for approximately 10 h ( Figure 1A, bottom trace).…”

“…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).…”