Targeted knockout of GABA receptor gamma 2 subunit provokes transient light-induced reflex seizures in zebrafish larvae

Abstract

Epilepsy is a common primary neurological disorder characterized by the chronic tendency of a patient to experience epileptic seizures, which are abnormal body movements or cognitive states that result from excessive, hypersynchronous brain activity. Epilepsy has been found to have numerous etiologies and, although about two-thirds of epilepsies were classically considered idiopathic, the majority of those are now believed to be of genetic origin. Mutations in genes involved in gamma-aminobutyric acid (GABA)-mediated inhibitory neurotransmission have been associated with a broad range of epilepsy syndromes. Mutations in the GABA-A receptor gamma 2 subunit gene (GABRG2), for example, have been associated with absence epilepsy and febrile seizures in humans. Several rodent models of GABRG2 loss of function depict clinical features of the disease; however, alternative genetic models more amenable for the study of ictogenesis and for high-throughput screening purposes are still needed. In this context, we generated a gabrg2 knockout (KO) zebrafish model (which we called R23X) that displayed light/dark-induced reflex seizures. Through high-resolution in vivo calcium imaging of the brain, we showed that this phenotype is associated with widespread increases in neuronal activity that can be effectively alleviated by the anti-epileptic drug valproic acid. Moreover, these seizures only occur at the larval stages but disappear after 1 week of age. Interestingly, our whole-transcriptome analysis showed that gabrg2 KO does not alter the expression of genes in the larval brain. As a result, the gabrg2−/− zebrafish is a novel in vivo genetic model of early epilepsies that opens new doors to investigate ictogenesis and for further drug-screening assays.

Functional Genomics of Epilepsy and Associated Neurodevelopmental Disorders Using Simple Animal Models: From Genes, Molecules to Brain Networks

“…One example is a recent study in which 132 schizophrenia risk variants were generated using CRISPR-Cas9 (Thyme et al, 2019). A wide array of zebrafish models have also been developed across epilepsy genes, and genes associated with broader neurodevelopmental phenotypes [e.g., scn1lab (Baraban et al, 2013), gabrg2 (Liao et al, 2019), GABRA1 (Samarut et al, 2018), mecp2 (Pietri et al, 2013), grin2a/b (Thyme et al, 2019), and others]. Importantly, despite different brain anatomy and physiology to mammalian counterparts, various zebrafish models exhibit phenotypes analogous to corresponding rodent models and clinical phenotypes.…”

“…Such recordings, obtained from field electrodes placed in the midbrain of agar immobilized larval zebrafish, appear as brief, small amplitude inter-ictal like events and prolonged, multi-spike ictal-like discharges which are qualitatively comparable to epileptiform discharges in patients and mammalian epilepsy models. Furthermore, GABRA1 −/− and gabrg2 −/− larvae, both modeling mutations reported in common epilepsy syndromes (Wallace et al, 2001;Johannesen et al, 2016), exhibit reflexive seizure-like events in response to light stimulation, reported as convulsive motor abnormalities and abnormal brain synchrony (Supplementary Video S2; Samarut et al, 2018;Liao et al, 2019).…”

“…It is, however, important to note that zebrafish lack a cortex and therefore qualitative homologies between zebrafish and human epilepsy (a putative cortical pathology) may be more useful for broad functional characterizations of genetic epilepsies, while specific seizure subtypes may be better modeled by more complex model organisms systems. Nonetheless, given that common anti-epileptic drugs correct electrographic, and motor abnormalities in these zebrafish models, the underlying neuropathology is likely to be conserved in genetic models (Baraban et al, 2013;Samarut et al, 2018;Liao et al, 2019). Finally, zebrafish larvae are also highly amenable to high-throughput behavioral drug screens, which have already identified novel drugs for the treatment of DS, thus closing the loop from fish tank to bedside (Griffin et al, 2018).…”

“…In this way the cellular mechanisms underlying observed network features in functional imaging data can be uncovered, to provide a conceptual bridge to explaining EEG phenomena during seizures, such as hypersynchrony and transitions between network states. As more genetic lines become available (Baraban et al, 2013;Samarut et al, 2018;Swaminathan et al, 2018;Liao et al, 2019), such imaging approaches may be harnessed to link gene mutation with network perturbation.…”

Comparison of three administration modes for establishing a zebrafish seizure model induced by N-Methyl-D-aspartic acid

“…The clinical manifestations are characterized by paroxysmal, transient, repetitive and stereotyped. The location of abnormal discharge neurons and the range of abnormal discharge spread are different, leading to different forms of seizure, manifested as sensory, motor, conscious, mental, behavioral, autonomic dysfunction or a combination of multiple dysfunctions[ 1 ]. According to the World Health Organization report, there are many causes of epilepsy, such as stroke, brain trauma, and central nervous system infection[ 2 ].…”