Rapid reconstruction of neural circuits using tissue expansion and light sheet microscopy

  1. Joshua L Lillvis  Is a corresponding author
  2. Hideo Otsuna
  3. Xiaoyu Ding
  4. Igor Pisarev
  5. Takashi Kawase
  6. Jennifer Colonell
  7. Konrad Rokicki
  8. Cristian Goina
  9. Ruixuan Gao
  10. Amy Hu
  11. Kaiyu Wang
  12. John Bogovic
  13. Daniel E Milkie
  14. Linus Meienberg
  15. Brett D Mensh
  16. Edward S Boyden
  17. Stephan Saalfeld
  18. Paul W Tillberg
  19. Barry J Dickson  Is a corresponding author
  1. Janelia Research Campus, United States
  2. University of Illinois at Chicago, United States
  3. ETH Zurich, Switzerland
  4. Massachusetts Institute of Technology, United States
  5. University of Queensland, Australia

Abstract

Brain function is mediated by the physiological coordination of a vast, intricately connected network of molecular and cellular components. The physiological properties of neural network components can be quantified with high throughput. The ability to assess many animals per study has been critical in relating physiological properties to behavior. By contrast, the synaptic structure of neural circuits is presently quantifiable only with low throughput. This low throughput hampers efforts to understand how variations in network structure relate to variations in behavior. For neuroanatomical reconstruction there is a methodological gulf between electron-microscopic (EM) methods, which yield dense connectomes at considerable expense and low throughput, and light-microscopic (LM) methods, which provide molecular and cell-type specificity at high throughput but without synaptic resolution. To bridge this gulf, we developed a high-throughput analysis pipeline and imaging protocol using tissue expansion and light sheet microscopy (ExLLSM) to rapidly reconstruct selected circuits across many animals with single-synapse resolution and molecular contrast. Using Drosophila to validate this approach, we demonstrate that it yields synaptic counts similar to those obtained by EM, enables synaptic connectivity to be compared across sex and experience, and can be used to correlate structural connectivity, functional connectivity, and behavior. This approach fills a critical methodological gap in studying variability in the structure and function of neural circuits across individuals within and between species.

Data availability

All software and code used for data analysis is available at Github (https://github.com/JaneliaSciComp/exllsm-circuit-reconstruction). Ground truth data used to train the synapse classifier is available at Dryad (https://doi.org/10.5061/dryad.5hqbzkh8b). All genetic reagents are available upon request. The data used to generate the figures and videos in this manuscript exceeds 100TB. Therefore, it is not practical to upload the data to a public repository. However, all data used in this paper will be made freely available to those who request and provide a mechanism for feasible data transfers (physical hard drives, cloud storage, etc.). Documentation for construction of a lattice light-sheet microscope can be obtained by execution of a research license agreement with HHMI.

The following data sets were generated

Article and author information

Author details

  1. Joshua L Lillvis

    Janelia Research Campus, Ashburn, United States
    For correspondence
    lillvisj@janelia.hhmi.org
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-6235-8759
  2. Hideo Otsuna

    Janelia Research Campus, Ashburn, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2107-8881
  3. Xiaoyu Ding

    Janelia Research Campus, Ashburn, United States
    Competing interests
    No competing interests declared.
  4. Igor Pisarev

    Janelia Research Campus, Ashburn, United States
    Competing interests
    No competing interests declared.
  5. Takashi Kawase

    Janelia Research Campus, Ashburn, United States
    Competing interests
    No competing interests declared.
  6. Jennifer Colonell

    Janelia Research Campus, Ashburn, United States
    Competing interests
    No competing interests declared.
  7. Konrad Rokicki

    Janelia Research Campus, Ashburn, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2799-9833
  8. Cristian Goina

    Janelia Research Campus, Ashburn, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0003-2835-7602
  9. Ruixuan Gao

    University of Illinois at Chicago, Chicago, United States
    Competing interests
    Ruixuan Gao, is a co-inventor on multiple patents related to expansion microscopy..
  10. Amy Hu

    Janelia Research Campus, Ashburn, United States
    Competing interests
    No competing interests declared.
  11. Kaiyu Wang

    Janelia Research Campus, Ashburn, United States
    Competing interests
    No competing interests declared.
  12. John Bogovic

    Janelia Research Campus, Ashburn, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4829-9457
  13. Daniel E Milkie

    Janelia Research Campus, Ashburn, United States
    Competing interests
    No competing interests declared.
  14. Linus Meienberg

    ETH Zurich, Zurich, Switzerland
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6793-1422
  15. Brett D Mensh

    Janelia Research Campus, Ashburn, United States
    Competing interests
    No competing interests declared.
  16. Edward S Boyden

    MIT McGovern Institute for Brain Research, Massachusetts Institute of Technology, Cambridge, United States
    Competing interests
    Edward S Boyden, is a co-inventor on multiple patents related to expansion microscopy and is also a co-founder of a company that aims to pursue commercial deployment of expansion microscopy-related technology..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0419-3351
  17. Stephan Saalfeld

    Janelia Research Campus, Ashburn, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-4106-1761
  18. Paul W Tillberg

    Janelia Research Campus, Ashburn, United States
    Competing interests
    Paul W Tillberg, is a co-inventor on multiple patents related to expansion microscopy..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2568-2365
  19. Barry J Dickson

    Queensland Brain Institute, University of Queensland, Queensland, Australia
    For correspondence
    b.dickson@uq.edu.au
    Competing interests
    No competing interests declared.

Funding

Howard Hughes Medical Institute

  • Barry J Dickson

The funders had no role in study design, data collection and interpretation, or the decision to submit the work for publication.

Reviewing Editor

  1. Matthieu Louis, University of California, Santa Barbara, United States

Version history

  1. Preprint posted: November 15, 2021 (view preprint)
  2. Received: June 20, 2022
  3. Accepted: October 25, 2022
  4. Accepted Manuscript published: October 26, 2022 (version 1)
  5. Version of Record published: November 11, 2022 (version 2)

Copyright

© 2022, Lillvis 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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  1. Joshua L Lillvis
  2. Hideo Otsuna
  3. Xiaoyu Ding
  4. Igor Pisarev
  5. Takashi Kawase
  6. Jennifer Colonell
  7. Konrad Rokicki
  8. Cristian Goina
  9. Ruixuan Gao
  10. Amy Hu
  11. Kaiyu Wang
  12. John Bogovic
  13. Daniel E Milkie
  14. Linus Meienberg
  15. Brett D Mensh
  16. Edward S Boyden
  17. Stephan Saalfeld
  18. Paul W Tillberg
  19. Barry J Dickson
(2022)
Rapid reconstruction of neural circuits using tissue expansion and light sheet microscopy
eLife 11:e81248.
https://doi.org/10.7554/eLife.81248

Share this article

https://doi.org/10.7554/eLife.81248

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