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Page 240 Mohammadi et al. J Transl Genet Genom 2020;4:238-50 I https://doi.org/10.20517/jtgg.2020.29

and back to the patients, can be often realized by repurposed drugs (i.e., those that are already licensed
for a different indication), for specific disease-related targets or pathways, thereby providing an immediate
benefit for affected patients [7,8,13] . However, knowing the functional effect (LoF or GoF) of a variant is crucial
and can assist in avoiding ineffective or even disease-aggravating treatments. For instance, individuals with
GoF variants in SCN2A/3A/8A respond well to treatment with sodium channel blockers such as phenytoin,
carbamazepine and oxcarbazepine, whereas the clinical picture of the individuals with LoF variants in
[7,8]
SCN1A/2A/8A worsens with the same drugs .
To apply stratified treatment options for individual patients, it is of great advantage to determine the
functional effect of the variant and potentially test drug efficacy in patient-specific systems.

Pathophysiology of epilepsy
The pathophysiological mechanisms underlying epileptiform activity and seizures are hyperexcitability of
cortical neurons and hypersynchronous simultaneous firing of numerous neurons, which is considered to be
caused by an imbalance between inhibitory (GABAergic) and excitatory (glutamatergic) neurotransmission.
Hyperexcitability of individual neurons can result from different mechanisms including decreased inhibitory
or increased excitatory neurotransmission, alterations in voltage-gated ion channels, or alterations of intra-
or extracellular ion concentrations favoring membrane depolarization and leading to epileptiform activity
[17]
and seizures . However, epileptogenesis is far more complex, and generation of epileptiform activity and
seizures may include many other processes such as cellular diversity, synaptic spatiotemporal dynamics of
interneuronal connectivity, synaptic reorganization of neuronal microcircuits, modifying network synchrony
[18]
and brain oscillations . Any brain region can potentially generate a seizure if the net excitation in a cortical
area exceeds the net inhibition, and each step in seizure initiation, propagation, and termination is ultimately
[17]
governed by the balance between excitation and inhibition . Most currently available AEDs are aimed at
[19]
restoring the excitatory/inhibitory balance either by decreasing excitation or by increasing inhibition .

Experimental models of epilepsy
Recent studies have identified > 500 epilepsy-associated genes, among which ion channel genes are
predominant [6,20,21] . To investigate disease pathology and assess optimal treatment options for monogenic
epilepsies, a wide range of cellular and animal models including fly, worm, zebrafish and rodent models
[22]
have been developed . In this review, we mainly focus on the cellular models [non-neuronal and neuronal
models, and human stem cell-derived two-dimensional (2D) and three-dimensional (3D) models], which
have clearly increased our knowledge on the functional consequences of genetic variants in, for example,
ion channel genes, the pathophysiology of epilepsy, and predicting drug efficacies. However, the knowledge
gained has also raised many new questions to be answered. One of the crucial questions is how both GoF
and LoF variants of the same channel gene can cause epilepsy, as exemplified by variants detected in SCN2A,
SCN8A, KCNQ2, KCNQ3, KCNA2, CACNA1A and GRIN2A.

Cellular expression systems utilizing human embryonic kidney cells (HEK293), Xenopus laevis oocytes
or Chinese hamster cells, are widely used as high throughput platforms for determining the biophysical
consequences of genetic variants compared to the wild-type. The observed functional effects of the variants
on ion channels often include shifts in activation and inactivation curves, changes in overall current
[23]
amplitudes and altered persistent currents and kinetics of inactivation . Currently, the gold standard for
investigating the functional consequence of a certain variant is to express the mutant channel in Xenopus
[24]
oocytes and use two-electrode voltage clamp methods to assess the function . Voltage clamping allows the
monitoring of minute changes in the electric currents across membranes. It is a fast and low-cost screening
system where the biophysical consequences can be studied using heterologous expression systems that are
[22]
not contaminated by endogenous channels . Furthermore, Xenopus oocytes have the advantages of small
size, durability and effectiveness in exogenous protein expression. These expression systems have not only