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Updated: Jun 21, 2026
High-resolution Confocal Imaging of the Blood-brain Barrier: Imaging, 3D Reconstruction, and Quantification of Transcytosis
Published on: November 16, 2017
Identification of NanoLuciferase Substrates Transported by Human ABCB1 and ABCG2 and their Zebrafish Homologs at the
Collin T Inglut1, John A Quinlan1, Robert W Robey1
1Fischell Department of Bioengineering, University of Maryland, College Park, MD 20742 (C.T.I., J.A.Q., H.-C.H.); Laboratory of Cell Biology, Center for Cancer Research, National Cancer Institute, National Institutes of Health, Bethesda, MD, 20892 (C.T.I., J.A.Q., R.W.R, J.R.T, M.M.G.), Promega Corporation, San Luis Obispo, CA, 93401 (J.R.W., W.Z.).
Abstract:
ATP-binding cassette (ABC) transporters expressed at the blood-brain barrier (BBB) impede delivery of therapeutic agents to the brain, including agents to treat neurodegenerative diseases and primary and metastatic brain cancers. Two transporters, P-glycoprotein (P-gp, ABCB1) and ABCG2, are highly expressed at the BBB and are responsible for the efflux of numerous clinically useful chemotherapeutic agents, including irinotecan, paclitaxel, and doxorubicin. Based on a previous mouse model, we have generated transgenic zebrafish in which expression of NanoLuciferase (NanoLuc) is controlled by the promoter of glial fibrillary acidic protein, leading to expression in zebrafish glia. To identify agents that disrupt the BBB, including inhibitors of ABCB1 and ABCG2, we identified NanoLuc substrates that are also transported by P-gp, ABCG2, and their zebrafish homologs. These substrates will elevate the amount of bioluminescent light produced in the transgenic zebrafish with BBB disruption. We transfected HEK293 cells with NanoLuc and either human ABCB1, ABCG2, or their zebrafish homologs Abcb4 or Abcg2a, respectively, and expressed at the zebrafish BBB. We evaluated the luminescence of ten NanoLuc substrates, then screened the eight brightest to determine which are most efficiently effluxed by the ABC transporters. We identified one substrate efficiently pumped out by ABCB1, two by Abcb4, six by ABCG2, and four by Abcg2a. These data will aid in the development of a transgenic zebrafish model of the BBB to identify novel BBB disruptors and should prove useful in the development of other animal models that use NanoLuc as a reporter.
Insights
Researchers developed a transgenic zebrafish model using NanoLuciferase to identify blood-brain barrier (BBB) disruptors. This model helps find new drugs that can overcome efflux by P-glycoprotein (P-gp) and ABCG2 transporters.
Area of Science:
- Neuroscience
- Pharmacology
- Genetics
Background:
- ATP-binding cassette (ABC) transporters, including P-glycoprotein (P-gp, ABCB1) and ABCG2, are highly expressed at the blood-brain barrier (BBB).
- These transporters efflux therapeutic agents, hindering drug delivery for brain diseases and cancers.
- Existing models are limited in identifying novel BBB-disrupting agents or transporter inhibitors.
Conclusions:
- The developed transgenic zebrafish model provides a novel platform for identifying BBB disruptors and ABC transporter inhibitors.
- This model can accelerate the development of therapeutics for brain diseases by improving drug delivery across the BBB.
- The findings support the use of NanoLuciferase reporter systems in developing other animal models for drug discovery.
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00:49The Blood-brain Barrier
01:31ABC Transporters: Exporter
01:19Membrane Asymmetry Regulating Transporters
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