Abstract
Attention deficit hyperactivity disorder (ADHD) is a neurodevelopmental disorder of childhood with a strong genetic component. Despite the success of mapping ADHD risk loci, little work has been done to experimentally verify the contribution of these loci to ADHD phenotypes. Meta-analysis of four genome-wide association studies in ADHD suggested CHMP7 as a predisposing gene for ADHD. A DNA variant (rs2294123) mapped to CHMP7 has been shown (via bioinformatic analysis) to have a high likelihood for functionality and correlate with reduced transcript levels. We used CRISPR-Cas9 genome editing to generate a chmp7 zebrafish model for ADHD. chmp7+/− fish showed comparable reductions in mRNA levels to individuals homozygous for the CHMP7 ADHD risk allele. These fish displayed significant hyperactivity over a 24-h period at 6 days post-fertilisation compared to chmp7+/+, but this effect did not persist into juvenile and adulthood stages. In addition, chmp7+/− fish had significantly smaller total brain volumes than chmp7+/+ fish. Finally, the hyperactivity at 6 days post-fertilisation was significantly reduced through the application of methylphenidate, a mainstay pharmacological treatment for ADHD. Overall, this study highlights an important role for CHMP7 in the neurodevelopment of ADHD, and demonstrates the utility of zebrafish for modelling the functional effects of genes conferring risk to ADHD.
Highlights
Attention deficit hyperactivity disorder (ADHD) is a highly prevalent neurodevelopmental disorder that affects ~5% of school age children[1] and persists into adulthood in 30–60% of cases[2,3]
We demonstrate that chmp7 +/− fish are more active than wildtype and have decreased total brain volumes, similar to that reported in ADHD cases
We show that hyperactivity of chmp7 +/− fish can be significantly reduced through the application of methylphenidate, which is commonly used for the treatment of ADHD
Summary
Fish were maintained in the Monash University Fish Core facility under breeding colony license MARP/2015/ 004/BC. CDNA synthesis was performed on RNA extracted from wildtype embryos at the 8- and 16-somite stages, 1, 1.5, 2, 3, 4, and 5 dpf. RNA was extracted from chmp7 +/+, chmp7 +/−, and chmp7 −/− embryos at 6 dpf, and pooled with a constant number of fish (20–25) per genotype within each biological replicate. Confocal microscopy live imaging chmp7 +/− fish were crossed to a green fluorescent protein (GFP)-tagged HuC reporter (HuC:eGFP21) transgenic (Tg) line and raised to adulthood. In order to examine brain volume at a larval stage that was in line with the locomotion assays, at 6 dpf, embryos were again anesthetised and set in 1% low melting agarose in E3 containing tricaine in 0.8 mm fluorinated ethylene propylene (FEP) tubing (Bola, Grünsfeld, Germany).
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