Human Pluripotent Stem Cell-Derived Neurons to Model Epilepsy

Epilepsy is a complex neurological disorder that is associated with unpredictable reoccurring seizures. The seizures are triggered by abnormal neuronal activity within the brain. The cause of the seizures is largely unknown, however, a range of insults, such as structural, metabolic, immunological or genetic alterations can lead to the onset of seizures. The disorder is known to affect over 70 million people of all ages globally. To date there is no cure for epilepsy. Though the majority of the patients obtain symptom relief from the anti-seizure medication, about 30% of patients do not benefit from them. Most of the pharmaceutical research is conducted in various animal models, however the translatability to humans has proven challenging. Therefore, the need for human predictive pre-clinical models stands as a necessity.

The focus of this thesis was to establish relevant and human-specific seizure models in vitro by using human pluripotent stem cell (hPSC)-derived neuronal networks. In the developed models, key pathological aspects were studied including genetic background and inflammation associated with epilepsy, and mimicry of human brain connectivity.

For this reason, hPSCs were firstly differentiated into the appropriate neuronal cell-types which are excitatory- (glutamate producing) and inhibitory (GABA producing) neurons known to be affected in epilepsy. To mimic inter-connectivity of the brain, a microphysiological platform, presented as brain-on-a-chip, was developed to create physiologically relevant interconnected networks and study seizure like events. The seizure-like events were induced chemically on hPSC-derived neurons using kainic acid (KA). Here, seizure-like events were captured at network level. Furthermore, the role of key inflammatory mediator interleukin-6 (IL-6) cytokine, which known to have an active role following onset of seizures, was assessed in the developed model. Finally, focus was set on a genetic form of epilepsy, Dravet syndrome (DS), to assess and characterize the genotype-to-phenotype correlation at a functional level.

The overall results presented in this thesis, showed that hPSC-derived neuronal networks display a prominent seizure-like phenotype following response to the seizure inducing agent, KA. This suggests the efficacy of the model as a foundation to investigate further aspects of the disease. The established microphysiological platform incorporated with the disease relevant-cell types, demonstrated support of neuronal structural and functional development. Upon KA induction, hPSC-derived neuronal networks in the platform mimicked a more focal seizure-type. Assessment of IL-6 on KA induced cultures, showed that IL-6 specific receptor, IL-6R, was increased, however at the functional level, changes in activity behavior were not detected. Finally, DS patient-derived hiPSC-neurons when cultured as either GABAergic or mixed neurons, displayed patient specific differences in functional behavior which unveiled distinct seizure-like phenotypes that corresponded to their clinical state.

In conclusion, the work of this thesis describes a significant human-based platform that holds promise in advancing future studies related to epilepsy research.