SUMMARYThe availability of animal models of epileptic seizures provides opportunities to identify novel anticonvulsants for the treatment of people with epilepsy. We found that exposure of 2-day-old zebrafish embryos to the convulsant agent pentylenetetrazole (PTZ) rapidly induces the expression of synaptic-activity-regulated genes in the CNS, and elicited vigorous episodes of calcium (Ca2+) flux in muscle cells as well as intense locomotor activity. We then screened a library of ∼2000 known bioactive small molecules and identified 46 compounds that suppressed PTZ-inducedtranscription of the synaptic-activity-regulated gene fos in 2-day-old (2 dpf) zebrafish embryos. Further analysis of a subset of these compounds, which included compounds with known and newly identified anticonvulsant properties, revealed that they exhibited concentration-dependent inhibition of both locomotor activity and PTZ-induced fos transcription, confirming their anticonvulsant characteristics. We conclude that this in situ hybridisation assay for fos transcription in the zebrafish embryonic CNS is a robust, high-throughput in vivo indicator of the neural response to convulsant treatment and lends itself well to chemical screening applications. Moreover, our results demonstrate that suppression of PTZ-induced fos expression provides a sensitive means of identifying compounds with anticonvulsant activities.

Gene-regulatory network analysis is a powerful approach to elucidate the molecular processes and pathways underlying complex disease. Here we employ systems genetics approaches to characterize the genetic regulation of pathophysiological pathways in human temporal lobe epilepsy (TLE). Using surgically acquired hippocampi from 129 TLE patients, we identify a gene-regulatory network genetically associated with epilepsy that contains a specialized, highly expressed transcriptional module encoding proconvulsive cytokines and Toll-like receptor signalling genes. RNA sequencing analysis in a mouse model of TLE using 100 epileptic and 100 control hippocampi shows the proconvulsive module is preserved across-species, specific to the epileptic hippocampus and upregulated in chronic epilepsy. In the TLE patients, we map the trans-acting genetic control of this proconvulsive module to Sestrin 3 (SESN3), and demonstrate that SESN3 positively regulates the module in macrophages, microglia and neurons. Morpholino-mediated Sesn3 knockdown in zebrafish confirms the regulation of the transcriptional module, and attenuates chemically induced behavioural seizures in vivo.

Drug discovery in epilepsy has historically been biased towards transmembrane channels and receptors. Ibhazehiebo et al. describe a novel, metabolism-based drug discovery platform that assays for bioenergetic changes in zebrafish models, and demonstrate that histone deacetylases 1 and 3 decrease mitochondrial hyperexcitability and represent a combined target for anti-seizure drug development.

Neurovascular coupling (through which local cerebral blood flow changes in response to neural activation are mediated) is impaired in many diseases including diabetes. Current preclinical rodent models of neurovascular coupling rely on invasive surgery and instrumentation, but transgenic zebrafish coupled with advances in imaging techniques allow non-invasive quantification of cerebrovascular anatomy, neural activation, and cerebral vessel haemodynamics. We therefore established a novel non-invasive, non-anaesthetised zebrafish larval model of neurovascular coupling, in which visual stimulus evokes neuronal activation in the optic tectum that is associated with a specific increase in red blood cell speed in tectal blood vessels. We applied this model to the examination of the effect of glucose exposure on cerebrovascular patterning and neurovascular coupling. We found that chronic exposure of zebrafish to glucose impaired tectal blood vessel patterning and neurovascular coupling. The nitric oxide donor sodium nitroprusside rescued all these adverse effects of glucose exposure on cerebrovascular patterning and function. Our results establish the first non-mammalian model of neurovascular coupling, offering the potential to perform more rapid genetic modifications and high-throughput screening than is currently possible using rodents. Furthermore, using this zebrafish model, we reveal a potential strategy to ameliorate the effects of hyperglycemia on cerebrovascular function.