Abstract
Epilepsy is a complex neurological condition wherein patients have recurrent spontaneous seizures leading to numerous physical and mental challenges. De novo mutations cause genetic epilepsies like developmental and epileptic encephalopathy (DEE), which result in seizures and developmental delays in patients. Moreover, 30% of epilepsy patients develop drug resistance to antiepileptic drugs (AEDs). Patients with drug resistance must seek alternative treatments like ketogenic diets, lifestyle changes, and surgery if needed. Surgical procedures, however, are only appropriate for a small percentage of patients.
Current epilepsy research focuses on alternative therapy targets as a means of reducing seizure severity. One such target is neuroinflammation, as persistent elevated inflammation in brain seems to be associated with exacerbation of the epileptic condition. However, these research findings have not yet been translated into clinical practice. Therefore, functional studies of the numerous epilepsy genes and alternative therapy treatments are necessary to aid in the development of precision medicine for patients. Thus, a standardised, simple, and high-throughput model of epilepsy is urgently needed.
In order to bridge the gap between epilepsy research findings with clinical practice and to test alternative therapy targets through functional studies, this study developed a standardised Xenopus laevis tadpole model of epilepsy. Additionally, selected neuroinflammatory pathways were targeted in the model as it was essential to investigate whether targeting inflammation could reduce severe seizures in the tadpoles.
Xenopus laevis models were first used to create chemoconvulsant-induced models of acute seizures to determine if targeting specific neuroinflammatory components with anti-inflammatory drugs (AIDs) could reduce seizures in a Pentylenetetrazol (PTZ) induced epilepsy model. A thorough investigation of the induced seizure phenotype was also conducted with TopScan, an animal tracking software previously not utilized and calibrated for aquatic developmental models. Xenopus laevis tadpoles were then used to model specific DEEs associated with NEUROD2 and CHD2 genes via the CRISPR/Cas9 knockout (KO) method. Additionally, this genetic model was utilized to define the complex spontaneous chronic seizure phenotype in edited tadpoles, which has not been studied previously.
A key finding of the study was the importance of AIDs such as Losartan, which demonstrated a seizure-modifying effect in both acute and chronic seizure models. Additionally, it contributed to developing protocols for efficiently utilizing the tadpole model of epilepsy for future functional studies and drug testing. Furthermore, the CRISPR/Cas9 model proved to be accurate and sensitive enough for future screening for suspected DEE genes. The model highlighted the effect of different mutation types on the severity of the seizure phenotype. The results contribute to the overarching aim of the project, which was to set up functional tests for screening epilepsy genes, resulting in early intervention for patients and establishing how alternative therapy options like AIDs can help manage severe seizure conditions.