Hamling, K. | bioRxiv Science

bioRxiv2021

Phenotypic analysis of catastrophic childhood epilepsy genes: The Epilepsy Zebrafish Project

Genetic engineering techniques have contributed to the now widespread use of zebrafish to investigate gene function, but zebrafish-based human disease studies, and particularly for neurological disorders, are limited. Here we used CRISPR-Cas9 to generate 40 single-gene mutant zebrafish lines representing catastrophic childhood epilepsies. We evaluated larval phenotypes using electrophysiological, behavioral, neuro-anatomical, survival and pharmacological assays. Phenotypes with unprovoked electrographic seizure activity (i.e., epilepsy) were identified in zebrafish lines for 8 genes; ARX, EEF1A, GABRB3, GRIN1, PNPO, SCN1A, STRADA and STXBP1. A unifying epilepsy classification scheme was developed based on local field potential recordings and blinded scoring from ~3300 larvae. We also created an open-source database containing sequencing information, survival curves, behavioral profiles and representative electrophysiology data. We offer all zebrafish lines as a resource to the neuroscience community and envision them as a starting point for further functional analysis and/or identification of new therapies.

neuroscience↗

bioRxiv2019

Phenotype-based screening of synthetic cannabinoids in a Dravet Syndrome zebrafish model

Dravet syndrome (DS) is a catastrophic epilepsy of childhood, characterized by cognitive impairment, severe seizures and increased risk for sudden unexplained death in epilepsy (SUDEP). Although refractory to conventional antiepileptic drugs, emerging preclinical and clinical evidence suggests that modulation of the endocanniboid system could be therapeutic in these patients. Here we used a validated zebrafish model of DS, scn1lab homozygous mutants, to screen a commercially available library containing 370 synthetic cannabinoid (SC) compounds for compounds effective in reducing spontaneous seizures. Primary phenotype-based screening was performed using a locomotion-based assay in 96-well plates, and a secondary local field potential recording assay was then used to confirm suppression of electrographic epileptiform events. Identified SCs with anti-seizure activity, in both assays, included five SCs structurally classified as indole-based cannabinoids: JWH 018 N-(5-chloropentyl) analog, JWH 018 N-(2-methylbutyl) isomer, 5-fluoro PB-22 5-hydroxyisoquinoline isomer, 5-fluoro ADBICA, and AB-FUBINACA 3-fluorobenzyl isomer. Our approach demonstrates that two-stage phenotype-based screening in a zebrafish model of DS successfully identifies synthetic cannabinoids with anti-seizure activity, and supports further investigation of SCs for refractory epilepsies.

neuroscience↗