IMPROVING THE ELECTROPHYSIOLOGICAL FUNCTIONALITY OF HIPSC-DERIVED GLUTAMATERGIC NEURONAL MODELS TO EXAMINE <EM>DE NOVO</EM> POTASSIUM CHANNELOPATHIES

Developmental and Epileptic Encephalopathies (DEEs) are severe neurodevelopmental disorders typically caused by de novo variants in ion channels. Common clinical presentations include pharmacoresistant seizures, developmental delay/regression and intellectual disability. Genes encoding voltage-gated potassium channels (e.g. KCNQ2), responsible for setting the resting membrane potential and action potential firing patterns, account for a significant proportion of DEE cases.
Human induced pluripotent stem cells (hiPSCs) offer a valuable platform for assessing genetic epilepsy variants by providing the ability to interrogate specific neural cell lineages. Existing disease models use small molecule patterning or virally-mediated ectopic transgene expression (e.g. NGN2) to produce various neuronal types but struggle to generate neurons with sufficient functionality. Here, I have adapted a combined protocol that includes cortical patterning, lentiviral transduction and astrocyte co-culture.
This protocol generated glutamatergic neurons with robust action potential firing within 19 days. Importantly, spontaneous postsynaptic currents were observed by 35 days. Subsequently, I compared the electrophysiological properties of neurons from hiPSC lines carrying a KCNQ2 patient variant with a population control cell line. Notably, input resistance of both control and patient neurons significantly decreased compared to previous NGN2-based differentiations, indicating the combinatorial neurons were larger and expressed more ion channels leading to an accelerated maturation profile. A shift in frequency-current relationships, reduced accommodation and an increased rheobase were also seen in the KCNQ2 patient-derived neurons. Overall, I have developed a combined differentiation protocol that improves upon neuronal maturity for functional assessments of ion channel variants implicated in DEE.