Development of a high-throughput zebrafish model of blood-brain barrier disruption

Authors

• AQ Wang
• C Ryu
• NR Hagen
• X Xu

Abstract

The blood-brain barrier (BBB) excludes most drugs, including chemotherapeutics, from the brain. Transient disruption of the BBB is an effective strategy to increase drug delivery to the brain. Assessment of the BBB in vitro is technically challenging, and mammalian models are low throughput. Zebrafish, which have a remarkably conserved BBB, are an ideal model for high-throughput BBB disruption studies.
We have created a zebrafish model of BBB disruption, by expressing the luciferase NanoLuc under the control of the glial-specific promoter GFAP (glial fibrillary acidic protein) in the brain parenchyma, behind the BBB. The NanoLuc substrate furimazine is BBB impermeable as it is a substrate of the multi-drug efflux transporter ABCG2, which is expressed on the lumenal surface of BBB endothelial cells, and effluxes drugs back into the blood, preventing brain entry.
Zebrafish have 4 homologs of ABCG2 (abcg2a, -b, -c, -d). We identified abcg2a as the only paralog expressed at the BBB in larvae and adults, and it also shares the most substrates with human ABCG2, including furimazine. Treatment of zebrafish larvae with the ABCG2 inhibitor Ko 143 blocks Abcg2a from effluxing furimazine, allowing it to cross the BBB, producing a dose-dependent increase in bioluminescent light. Younger larvae (3 vs 7 days old) have higher baseline bioluminescence, indicating a less restrictive developing BBB.
Our future goal is to adapt this assay for high-throughput screening of BBB disruptors, and eventually combine these methods with brain tumor xenografts to assess the delivery of chemotherapeutics to brain tumors.

Scientific Focus Area: Cancer Biology

This page was last updated on Monday, September 25, 2023