Abstracts

Rationale: Despite epilepsy being one of the most common neurological disorders, one-third of patients remain without effective anticonvulsant treatments. Unfortunately, anticonvulsant discovery remains static. The lack of drug-discovery can be partly attributed to species-specific effects and bottle-necks during in-vivo rodent testing. By utilising the high-throughput models C. elegans (nematode worm) and D. rerio (Zebrafish) as front-line screening tools, larger scale compound testing is feasible, and species-specific effects would be reduced; increasing the translational potential of positive compounds. Methods: Both D. rerio and C. elegans were exposed to the pro-convulsant pentylenetetrazol (PTZ) via their bathing medium before analysis and behavioural phenotyping. Validation of acute anti-seizure effects was then established using PTZ, maximal electroshock seizure, 6-Hz psychomotor seizure and corneal kindling models in mice. Results: This approach was validated through the compound LIV001 identified in a D. rerio screen of 2000 compounds utilising c-fos expression in response to PTZ. LIV001 pre-treatment significantly reduced PTZ induced convulsive phenotypes in both D. rerio and C. elegans (p>0.05). A molecular target was identified using a C. elegans genetic screen against a LIV001induced paralysis phenotype. Here, worms with mutant lgc-37 (a GABAA receptor subunit orthologue) were significantly resistant to the compound p>0.05. Paralysis resistance in these mutants was reversed through reintroducing the native lgc-37 gene and, in wild type worms, pharmacological antagonism of the GABAA channel protected against LIV001-induced paralysis. This GABAA mechanism was conserved in mouse thalamocortical relay neurons, as demonstrated by LIV001’s stimulatory effect on both tonic (LIV001 tonic current 589±157pA vs -324±111pA control) and phasic inhibition (LIV001 mIPSC peak amplitude -51.1± 3.1 vs -43.3± 2.2). LIV001 was shown to be anticonvulsant in a battery of mouse acute seizure tests. Notably, efficacy was demonstrated in the 6Hz 44mA assay of pharmacoresistant seizures, protective index (PI) 2.3 (PI=median toxic dose/median effective dose), distinguishing LIV001 from several FDA approved anti-convulsants (ED50=66.2mg/kg, CI=45.8-102). Conclusions: We describe here conserved anti-convulsive effects in all three models, rapid characterisation of molecular targets and identification of a compound effective in a mammalian pharmacoresistant seizure model using our pipeline, highlighting the benefits of this approach. Funding: Medical Research Council UK