Computational 3D histological phenotyping of whole zebrafish by X-ray histotomography
- Penn State College of Medicine, United States
- Duke University, United States
- The University of Chicago, United States
- Motorola Mobility, United States
- Argonne National Laboratory, United States
Abstract
Organismal phenotypes frequently involve multiple organ systems. Histology is a powerful way to detect cellular and tissue phenotypes, but is largely descriptive and subjective. To determine how synchrotron-based X-ray micro-tomography (micro-CT) can yield 3-dimensional whole-organism images suitable for quantitative histological phenotyping, we scanned whole zebrafish, a small vertebrate model with diverse tissues, at ~1-micron voxel resolutions. Using micro-CT optimized for cellular characterization (histotomography), brain nuclei were computationally segmented and assigned to brain regions. Shape and volume were computed for populations of nuclei such as those of motor neurons and red blood cells. Striking individual phenotypic variation was apparent from color maps of computed cell density. Unlike histology, histotomography allows the detection of phenotypes that require millimeter scale context in multiple planes. We expect the computational and visual insights into 3D tissue architecture provided by histotomography to be useful for reference atlases, hypothesis generation, comprehensive organismal screens, and diagnostics.
Data availability
ViewTool is publically available (http://3D.fish). Digital histology is publicly available from our Zebrafish Lifespan Atlas (http://bio-atlas.psu.edu) (Cheng, 2004). Registered and unregistered 8-bit reconstructions of the heads of five zebrafish larvae involved in analysis are available on Dryad (https://datadryad.org/) along with scripts written for cell nuclei detection, analysis, and sample registration. Full resolution scans, including raw projection data, are available from researchers upon request as a download or by transfer to physical media.
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Data from: Computational 3D histological phenotyping of whole zebrafish by X-ray histotomographyDryad Digital Repository, doi:10.5061/dryad.4nb12g2.
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ViBE-Z: A Framework for 3D Virtual Colocalization Analysis in Zebrafish Larval Brainshttp://vibez.informatik.uni-freiburg.de.
Article and author information
Author details
Patrick La Riviere
Funding
NIH Office of the Director (R24-OD018559)
- Patrick La Riviere
- Keith Cheng
National Institutes of Health (R24-RR017441)
- Patrick La Riviere
- Keith Cheng
The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.
Reviewing Editor
- Richard M White, Memorial Sloan Kettering Cancer Center, United States
Ethics
Animal experimentation: All procedures on live animals were approved by the Institutional Animal Care and Use Committee (IACUC) at the Pennsylvania State University, ID: PRAMS201445659, Groundwork for a Synchrotron MicroCT Imaging Resource for Biology (SMIRB).
Version history
- Received: January 5, 2019
- Accepted: May 4, 2019
- Accepted Manuscript published: May 7, 2019 (version 1)
- Version of Record published: June 11, 2019 (version 2)
Copyright
© 2019, Ding et al.
This article is distributed under the terms of the Creative Commons Attribution License permitting unrestricted use and redistribution provided that the original author and source are credited.
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Computational 3D histological phenotyping of whole zebrafish by X-ray histotomography
eLife 8:e44898.
https://doi.org/10.7554/eLife.44898
Further reading
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- Developmental Biology
By enabling researchers to image whole zebrafish with cellular resolution, X-ray histotomography will improve our understanding of the biological differences between individuals of the same species.
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- Developmental Biology
Inhibitory G alpha (GNAI or Gαi) proteins are critical for the polarized morphogenesis of sensory hair cells and for hearing. The extent and nature of their actual contributions remains unclear, however, as previous studies did not investigate all GNAI proteins and included non-physiological approaches. Pertussis toxin can downregulate functionally redundant GNAI1, GNAI2, GNAI3, and GNAO proteins, but may also induce unrelated defects. Here, we directly and systematically determine the role(s) of each individual GNAI protein in mouse auditory hair cells. GNAI2 and GNAI3 are similarly polarized at the hair cell apex with their binding partner G protein signaling modulator 2 (GPSM2), whereas GNAI1 and GNAO are not detected. In Gnai3 mutants, GNAI2 progressively fails to fully occupy the sub-cellular compartments where GNAI3 is missing. In contrast, GNAI3 can fully compensate for the loss of GNAI2 and is essential for hair bundle morphogenesis and auditory function. Simultaneous inactivation of Gnai2 and Gnai3 recapitulates for the first time two distinct types of defects only observed so far with pertussis toxin: (1) a delay or failure of the basal body to migrate off-center in prospective hair cells, and (2) a reversal in the orientation of some hair cell types. We conclude that GNAI proteins are critical for hair cells to break planar symmetry and to orient properly before GNAI2/3 regulate hair bundle morphogenesis with GPSM2.
