A Caenorhabditis elegans assay of seizure-like activity optimised for identifying antiepileptic drugs and their mechanisms of action

Shi Quan Wong,Alistair Jones,Steven Dodd,Douglas Grimes,Jeff W Barclay,Anthony G Marson,Vincent T Cunliffe,Robert D Burgoyne,Graeme J Sills,Alan Morgan

doi:10.1016/j.jneumeth.2018.09.004

Shi Quan Wong, Alistair Jones + Show 8 more

Open Access

https://doi.org/10.1016/j.jneumeth.2018.09.004

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Abstract

BackgroundEpilepsy affects around 1% of people, but existing antiepileptic drugs (AEDs) only offer symptomatic relief and are ineffective in approximately 30% of patients. Hence, new AEDs are sorely needed. However, a major bottleneck is the low-throughput nature of early-stage AED screens in conventional rodent models. This process could potentially be expedited by using simpler invertebrate systems, such as the nematode Caenorhabditis elegans. New methodHead-bobbing convulsions were previously reported to be inducible by pentylenetetrazol (PTZ) in C. elegans with loss-of-function mutations in unc-49, which encodes a GABAA receptor. Given that epilepsy-linked mutations in human GABAA receptors are well documented, this could represent a clinically-relevant system for early-stage AED screens. However, the original agar plate-based assay is unsuited to large-scale screening and has not been validated for identifying AEDs. Therefore, we established an alternative streamlined, higher-throughput approach whereby mutants were treated with PTZ and AEDs via liquid-based incubation. ResultsConvulsions induced within minutes of PTZ exposure in unc-49 mutants were strongly inhibited by the established AED ethosuximide. This protective activity was independent of ethosuximide’s suggested target, the T-type calcium channel, as a null mutation in the worm cca-1 ortholog did not affect ethosuximide’s anticonvulsant action. Comparison with existing methodOur streamlined assay is AED-validated, feasible for higher throughput compound screens, and can facilitate insights into AED mechanisms of action. ConclusionsBased on an epilepsy-associated genetic background, this C. elegans unc-49 model of seizure-like activity presents an ethical, higher throughput alternative to conventional rodent seizure models for initial AED screens.

Highlights

Epilepsy is a brain disorder that results in the development of neuronal networks predisposed to the occurrence and recurrence of symptomatic epileptic seizures (Duncan et al, 2006)
Plate-based PTZ administration requires the preparation of PTZ-containing agar plates, which is labour-intensive and incompatible with modern high-throughput drug screens using compound libraries
We have demonstrated that PTZ responsiveness is unaffected by dimethyl sulfoxide (DMSO) concentrations up to 1%, thereby facilitating screening of DMSO-solubilised compound libraries

Summary

Introduction

Epilepsy is a brain disorder that results in the development of neuronal networks predisposed to the occurrence and recurrence of symptomatic epileptic seizures (Duncan et al, 2006). A major bottleneck is the low-throughput nature of early-stage AED screens in conventional rodent models. This process could potentially be expedited by using simpler invertebrate systems, such as the nematode Caenorhabditis elegans. Results: Convulsions induced within minutes of PTZ exposure in unc-49 mutants were strongly inhibited by the established AED ethosuximide. This protective activity was independent of ethosuximide’s suggested target, the T-type calcium channel, as a null mutation in the worm cca-1 ortholog did not affect ethosuximide’s anticonvulsant action. Conclusions: Based on an epilepsy-associated genetic background, this C. elegans unc-49 model of seizure-like activity presents an ethical, higher throughput alternative to conventional rodent seizure models for initial AED screens

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