A zebrafish model of benzodiazepine-refractory status epilepticus for whole organism discovery of novel anti-seizure treatments
Abstract number :
1.196
Submission category :
2. Translational Research / 2D. Models
Year :
2025
Submission ID :
1198
Source :
www.aesnet.org
Presentation date :
12/6/2025 12:00:00 AM
Published date :
Authors :
Presenting Author: Christopher McGraw, MD, PhD – Northwestern
Zoe Gardner, BA – Boston Children's Hospital
Rationale: Status epilepticus (SE) is a severe neurologic condition associated with significant morbidity and mortality. Over 40% of individuals with SE are refractory to 1st and 2nd line anti-seizure medications, such as benzodiazepines. Although mammalian models of SE have proven instrumental to understand mechanisms of SE and to test interventions, these models have limited potential for chemical and genetic screening.
Methods: We characterized a novel zebrafish model of status epilepticus in response to paraoxon-ethyl (PXN), an organophosphate associated with treatment refractory SE in rodents and humans. We monitored the response of larval zebrafish (dpf 5) expressing neuronal GCaMP6s to PXN exposure by a combined movement and fluorescence assay, light microscopy, and time-lapse fluorescence microscopy. Morphological effects of PXN exposure on zebrafish were scored visually at multiple timepoints after exposure (0.5 - 24 hrs). The pharmacology of PXN-related SE was probed using inhibitors of NMDAR (MK801) and brain-specific alpha7 nicotinic AChRs (MLA); a positive allosteric modulator of GABAR (diazepam, DZP); and the antidote atropine + 2-pralidoxime (A/2PAM).
Results: We show that PXN has LD50(6 hours)= 125uM and at earlier timepoints results in immobility, L-bend contracture, and dramatic brain region-specific increases in tonic calcium fluorescence in still living fish, consistent with status epilepticus. MK801 substantially delayed progression of PXN-related SE, with contracture and tonic calcium fluorescence completely blocked at 0.5 and 1 hour post-exposure, and still low at 3 hrs post-exposure. Similar although less potent effects were observed for MLA. Meanwhile, although A/2PAM significantly reduced progression of PXN-related SE, the addition of either early (10min) or late (40min) DZP post-exposure had no effect.
Conclusions: Similar to rodent models of PXN-related SE, our study confirms the key roles of NMDAR and central effects of cholinergic excess to PXN-related SE in zebrafish, as well as its poor response to benzodiazepine treatment. To the best of our knowledge, no existing zebrafish models of seizures demonstrate benzodiazepine resistance. These findings will now enable high-throughput screening to identify novel counter-measures that may control refractory SE related to organophosphate exposure and other causes more broadly.
Funding: This work was funded by the NIH CounterACT program (NINDS 5R21NS127345).
Translational Research