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Zebrafish Brain Development Monitored by Long-Term In Vivo Microscopy: A Comparison Between Laser Scanning Confocal and 2-Photon Microscopy

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Laser Scanning Microscopy and Quantitative Image Analysis of Neuronal Tissue

Part of the book series: Neuromethods ((NM,volume 87))

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Abstract

Zebrafish is an attractive model organism to study vertebrate brain development. Its transparency makes it possible to follow development using live imaging. In a transgenic line where a subset of neurons is labeled by GFP expression, their migration, proliferation and the extension of axons can be observed by laser scanning confocal microscopy (LSCM) or 2-photon microscopy (2PM). However, when the whole brain is imaged, LSCM might result in phototoxicity. In contrast, 2PM allows for image acquisition over several days at intervals shorter than an hour. In this article, we describe a method to image a large region of the brain (500 × 500 μm) spanning 300 μm in depth by 2PM over 2 days or more. The results are compared with those obtained by the more widespread LSCM. Visualization and analysis of the resulting data is challenging, as they exceed the size that can be loaded into standard rendering software. We propose a routine to reduce the size by maximum projection while keeping and displaying three-dimensional information by a color code within ImageJ.

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Author information

Authors and Affiliations

  1. Nikon Imaging Center, University of Heidelberg, Heidelberg, Germany

    Nicolas Dross, Peter Bankhead & Ulrike Engel

  2. Center for Organismal Studies, Heidelberg University, Heidelberg, Germany

    Nicolas Dross, Peter Bankhead & Ulrike Engel

  3. Department of Cell and Molecular Biology, Medical Faculty Mannheim, Heidelberg University, Mannheim, Germany

    Carlo Antonio Beretta & Matthias Carl

Authors
  1. Nicolas Dross
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  2. Carlo Antonio Beretta
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  3. Peter Bankhead
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  4. Matthias Carl
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  5. Ulrike Engel
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Editor information

Editors and Affiliations

  1. Department of Neurobiology, University of Osnabrück, Osnabrück, Germany

    Lidia Bakota

  2. Department of Neurobiology, University of Osnabrück, Osnabrück, Germany

    Roland Brandt

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Dross, N., Beretta, C.A., Bankhead, P., Carl, M., Engel, U. (2014). Zebrafish Brain Development Monitored by Long-Term In Vivo Microscopy: A Comparison Between Laser Scanning Confocal and 2-Photon Microscopy. In: Bakota, L., Brandt, R. (eds) Laser Scanning Microscopy and Quantitative Image Analysis of Neuronal Tissue. Neuromethods, vol 87. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-0381-8_8

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  • DOI: https://doi.org/10.1007/978-1-4939-0381-8_8

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-0380-1

  • Online ISBN: 978-1-4939-0381-8

  • eBook Packages: Springer Protocols

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