Gabrielle Mills
Gabrielle Mills
Gabrielle Mills
Helios Scholar
School: Arizona State University
Hometown: Phoenix, Arizona
Mentor: Vinodh Narayanan, M.D.
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Development of a Zebrafish Model to Study Childhood Epileptic Encephalopathy Caused by Dynamin 1 Mutations

Epileptic encephalopathies (EE) are genetic or environmentally-caused conditions that cause “catastrophic” damage or degradation of the sensory, cognitive, and behavioral centers of the brain. Next generation sequencing technology in C4RCD pediatric patients with EE has identified causal mutations in the DNM1 gene encoding dynamin-1. While recent studies have provided insights into dynamin 1’s structure and function, it is still unclear how the de novo missense mutation in DNM1—a core component of endocytosis machinery—leads to EE. In order to understand the phenotypes of EE caused by DNM1 mutations, the zebrafish model was utilized since it is an alternative model system with substantial benefits: cost-efficient breeding, 84% similarity to human disease genes, and the ability to engineer in vivo drug treatments. We hypothesized that the loss of DNM1 protein function (through the inhibition of DNM1 protein or microinjection of mutant DNM1 mRNA) in zebrafish could recapitulate the human EE phenotype. Our lab identified a novel DNM1 inhibitor, dynasore, through a seizure phenotype screening assay. To create the animal model, wildtype AB zebrafish between 6-24 months old were bred to produce embryos which were then randomly assigned to treatments of dynasore, the vehicle treatment, DMSO, or left untreated. Analysis of videography at 72 hours post fertilization identified significant increases in locomotion, seizure-like phenotype, and curvature phenotype in the dynasore-treated fish compared to both DMSO-treated and untreated fish. Microinjections of wild-type DNM1 mRNA into zebrafish embryos and treatment with dynasore verified this chemical inhibitor produced epileptic phenotypes via the dynamin-1 pathway. Biochemical characteristics of dynamin-1 epileptic zebrafish were validated through identification of differences in the localization versus quantity of dynamin-2, clathrin, and caveolin-1 endocytosis proteins via immunohistochemistry and western blotting, respectively. Our data indicates that the zebrafish model is a promising model system given that the disruption of dynamin-1 leads to seizure-like activity. This approach can potentially lead to the understanding of dynamin-1 biology in causing EE and the identification of novel treatments for epileptic encephalopathy caused by DNM1 mutations in humans.

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