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Research ArticleMethods/New Tools, Novel Tools and Methods

3D Imaging of Axons in Transparent Spinal Cords from Rodents and Nonhuman Primates

Cynthia Soderblom, Do-Hun Lee, Abdul Dawood, Melissa Carballosa, Andrea Jimena Santamaria, Francisco D. Benavides, Stanislava Jergova, Robert M. Grumbles, Christine K. Thomas, Kevin K. Park, James David Guest, Vance P. Lemmon, Jae K. Lee and Pantelis Tsoulfas
eNeuro 26 March 2015, 2 (2) ENEURO.0001-15.2015; DOI: https://doi.org/10.1523/ENEURO.0001-15.2015
Cynthia Soderblom
Miami Project to Cure Paralysis, Department of Neurological Surgery, University of Miami School of Medicine, Miami, Florida 33136
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Do-Hun Lee
Miami Project to Cure Paralysis, Department of Neurological Surgery, University of Miami School of Medicine, Miami, Florida 33136
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Abdul Dawood
Miami Project to Cure Paralysis, Department of Neurological Surgery, University of Miami School of Medicine, Miami, Florida 33136
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Melissa Carballosa
Miami Project to Cure Paralysis, Department of Neurological Surgery, University of Miami School of Medicine, Miami, Florida 33136
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Andrea Jimena Santamaria
Miami Project to Cure Paralysis, Department of Neurological Surgery, University of Miami School of Medicine, Miami, Florida 33136
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  • ORCID record for Andrea Jimena Santamaria
Francisco D. Benavides
Miami Project to Cure Paralysis, Department of Neurological Surgery, University of Miami School of Medicine, Miami, Florida 33136
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Stanislava Jergova
Miami Project to Cure Paralysis, Department of Neurological Surgery, University of Miami School of Medicine, Miami, Florida 33136
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Robert M. Grumbles
Miami Project to Cure Paralysis, Department of Neurological Surgery, University of Miami School of Medicine, Miami, Florida 33136
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Christine K. Thomas
Miami Project to Cure Paralysis, Department of Neurological Surgery, University of Miami School of Medicine, Miami, Florida 33136
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Kevin K. Park
Miami Project to Cure Paralysis, Department of Neurological Surgery, University of Miami School of Medicine, Miami, Florida 33136
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James David Guest
Miami Project to Cure Paralysis, Department of Neurological Surgery, University of Miami School of Medicine, Miami, Florida 33136
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Vance P. Lemmon
Miami Project to Cure Paralysis, Department of Neurological Surgery, University of Miami School of Medicine, Miami, Florida 33136
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Jae K. Lee
Miami Project to Cure Paralysis, Department of Neurological Surgery, University of Miami School of Medicine, Miami, Florida 33136
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Pantelis Tsoulfas
Miami Project to Cure Paralysis, Department of Neurological Surgery, University of Miami School of Medicine, Miami, Florida 33136
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Abstract

The histological assessment of spinal cord tissue in three dimensions has previously been very time consuming and prone to errors of interpretation. Advances in tissue clearing have significantly improved visualization of fluorescently labelled axons. While recent proof-of-concept studies have been performed with transgenic mice in which axons were prelabeled with GFP, investigating axonal regeneration requires stringent axonal tracing methods as well as the use of animal models in which transgenic axonal labeling is not available. Using rodent models of spinal cord injury, we labeled axon tracts of interest using both adeno-associated virus and chemical tracers and performed tetrahydrofuran-based tissue clearing to image multiple axon types in spinal cords using light sheet and confocal microscopy. Using this approach, we investigated the relationships between axons and scar-forming cells at the injury site as well as connections between sensory axons and motor pools in the spinal cord. In addition, we used these methods to trace axons in nonhuman primates. This reproducible and adaptable virus-based approach can be combined with transgenic mice or with chemical-based tract-tracing methods, providing scientists with flexibility in obtaining axonal trajectory information from transparent tissue.

  • 3DISCO
  • axon regeneration
  • spinal cord injury
  • tissue clearing

Footnotes

  • ↵1 The authors declare no competing financial interests.

  • ↵3 This study was funded by NINDS Grants R01NS081040 and R21NS082835, NEI Grant R01EY022961, NICHD Grant RO1HD057632, Craig Neilsen Foundation, Department of Defense, International Spinal Research Trust, State of Florida Specific Appropriation 538, The Miami Project to Cure Paralysis, and the Buoniconti Fund.

This is an open-access article distributed under the terms of the Creative Commons Attribution 4.0 International, which permits unrestricted use, distribution and reproduction in any medium provided that the original work is properly attributed.

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3D Imaging of Axons in Transparent Spinal Cords from Rodents and Nonhuman Primates
Cynthia Soderblom, Do-Hun Lee, Abdul Dawood, Melissa Carballosa, Andrea Jimena Santamaria, Francisco D. Benavides, Stanislava Jergova, Robert M. Grumbles, Christine K. Thomas, Kevin K. Park, James David Guest, Vance P. Lemmon, Jae K. Lee, Pantelis Tsoulfas
eNeuro 26 March 2015, 2 (2) ENEURO.0001-15.2015; DOI: 10.1523/ENEURO.0001-15.2015

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3D Imaging of Axons in Transparent Spinal Cords from Rodents and Nonhuman Primates
Cynthia Soderblom, Do-Hun Lee, Abdul Dawood, Melissa Carballosa, Andrea Jimena Santamaria, Francisco D. Benavides, Stanislava Jergova, Robert M. Grumbles, Christine K. Thomas, Kevin K. Park, James David Guest, Vance P. Lemmon, Jae K. Lee, Pantelis Tsoulfas
eNeuro 26 March 2015, 2 (2) ENEURO.0001-15.2015; DOI: 10.1523/ENEURO.0001-15.2015
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Keywords

  • 3DISCO
  • axon regeneration
  • spinal cord injury
  • tissue clearing

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