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Identification of oxygen-sensitive neuroepithelial cells through an endogenous reporter gene in larval and adult transgenic zebrafish

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Abstract

In teleost fish, specialized oxygen (O2) chemoreceptors, called neuroepithelial cells (NECs), are found in the gill epithelium in adults. During development, NECs are present in the skin before the formation of functional gills. NECs are known for retaining the monoamine neurotransmitter, serotonin (5-HT) and are conventionally identified through immunoreactivity with antibodies against 5-HT or synaptic vesicle protein (SV2). However, identification of NECs in live tissue and isolated cell preparations has been challenging due to the lack of a specific marker. The present study explored the use of the transgenic zebrafish, ETvmat2:GFP, which expresses green fluorescent protein (GFP) under the control of the vesicular monoamine transporter 2 (vmat2) regulatory element, to identify NECs. Using immunohistochemistry and confocal microscopy, we confirmed that the endogenous GFP in ETvmat2:GFP labelled serotonergic NECs in the skin of larvae and in the gills of adults. NECs of the gill filaments expressed a higher level of endogenous GFP compared with other cells. The endogenous GFP also labelled intrabranchial neurons of the gill filaments. Flow cytometric analysis demonstrated that filamental NECs could be distinguished from other dissociated gill cells based on high GFP expression alone. Acclimation to 2 weeks of severe hypoxia (PO2 = 35 mmHg) induced an increase in filamental NEC frequency, size and GFP gene expression. Here we present for the first time a transgenic tool that labels O2 chemoreceptors in an aquatic vertebrate and its use in high-throughput experimentation.

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Acknowledgements

The authors thank the Becker Laboratory at the University of Edinburgh for graciously providing the ETvmat2:GFP transgenic line.

Funding

This study was funded by the Natural Sciences and Engineering Research Council of Canada (grant nos. 342303 and 05571), the Canadian Foundation for Innovation, and the Ontario Research Fund (grant no. 16589).

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Authors and Affiliations

  1. Department of Biology, University of Ottawa, 30 Marie Curie Pvt., Ottawa, ON, K1N 6N5, Canada

    Wen Pan & Michael G. Jonz

  2. Department of Pathology and Molecular Medicine, McMaster University, Hamilton, ON, L8S 4K1, Canada

    Angela L. Scott

  3. Department of Biology, McMaster University, Hamilton, ON, L8S 4K1, Canada

    Colin A. Nurse

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  1. Wen Pan
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  2. Angela L. Scott
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  3. Colin A. Nurse
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  4. Michael G. Jonz
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Correspondence to Michael G. Jonz.

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The authors declare that there is no conflict of interest.

Ethical approval

All applicable international, national, and/or institutional guidelines for the care and use of animals were followed. All procedures for animal use were carried out in accordance with institutional guidelines according to protocol BL-1760, along with guidelines provided by the Canadian Council on Animal Care (CCAC). This article does not contain any studies with human participants performed by any of the authors.

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Pan, W., Scott, A.L., Nurse, C.A. et al. Identification of oxygen-sensitive neuroepithelial cells through an endogenous reporter gene in larval and adult transgenic zebrafish. Cell Tissue Res 384, 35–47 (2021). https://doi.org/10.1007/s00441-020-03307-5

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  • DOI: https://doi.org/10.1007/s00441-020-03307-5

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