Fast two-photon imaging of subcellular voltage dynamics in neuronal tissue with genetically encoded indicators

Abstract

Monitoring voltage dynamics in defined neurons deep in the brain is critical for unraveling the function of neuronal circuits, but is challenging due to the limited performance of existing tools. In particular, while genetically encoded voltage indicators have shown promise for optical detection of voltage transients, many indicators exhibit low sensitivity when imaged under two-photon illumination. Previous studies thus fell short of visualizing voltage dynamics in individual neurons in single trials. Here, we report ASAP2s, a novel voltage indicator with improved sensitivity. By imaging ASAP2s using random-access multi-photon microscopy, we demonstrate robust single-trial detection of action potentials in organotypic slice cultures. We also show that ASAP2s enables two-photon imaging of graded potentials with subcellular resolution in organotypic slice cultures and in Drosophila. These results demonstrate that the combination of ASAP2s and fast two-photon imaging methods enables detection of neural electrical activity with subcellular spatial resolution and millisecond-timescale precision.

Article and author information

Author details

  1. Simon Chamberland

    Department of Psychiatry and Neuroscience, Université Laval, Québec, Canada
    Competing interests
    No competing interests declared.
  2. Helen H Yang

    Department of Neurobiology, Stanford University, Stanford, United States
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-5140-9664
  3. Michael M Pan

    Department of Bioengineering, Stanford University, Stanford, United States
    Competing interests
    No competing interests declared.
  4. Stephen W Evans

    Department of Neurobiology, Stanford University, Stanford, United States
    Competing interests
    No competing interests declared.
  5. Sihui Guan

    Department of Neuroscience, Baylor College of Medicine, Houston, United States
    Competing interests
    No competing interests declared.
  6. Mariya Chavarha

    Department of Neurobiology, Stanford University, Stanford, United States
    Competing interests
    No competing interests declared.
  7. Ying Yang

    Department of Neurobiology, Stanford University, Stanford, United States
    Competing interests
    No competing interests declared.
  8. Charleen Salesse

    Department of Psychiatry and Neuroscience, Université Laval, Québec, Canada
    Competing interests
    No competing interests declared.
  9. Haodi Wu

    Stanford Cardiovascular Institute, Stanford University, Stanford, United States
    Competing interests
    No competing interests declared.
  10. Joseph C Wu

    Stanford Cardiovascular Institute, Stanford University, Stanford, United States
    Competing interests
    No competing interests declared.
  11. Thomas R Clandinin

    Department of Neurobiology, Stanford University, Stanford, United States
    For correspondence
    trc@stanford.edu
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-6277-6849
  12. Katalin Toth

    Department of Psychiatry and Neuroscience, Université Laval, Québec, Canada
    For correspondence
    katalin.toth@fmed.ulaval.ca
    Competing interests
    No competing interests declared.
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-2300-4536
  13. Michael Z Lin

    Department of Neurobiology, Stanford University, Stanford, United States
    Competing interests
    Michael Z Lin, Holds a US patent for a voltage sensor design based on ASAP-family indicators (patent number US9606100 B2)..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0002-0492-1961
  14. François St-Pierre

    Department of Neuroscience, Baylor College of Medicine, Houston, United States
    For correspondence
    francois.st-pierre@bcm.edu
    Competing interests
    François St-Pierre, Holds a US patent for a voltage sensor design based on ASAP-family indicators (patent number US9606100 B2)..
    ORCID icon "This ORCID iD identifies the author of this article:" 0000-0001-8618-4135

Funding

Burroughs Wellcome Fund

  • Michael Z Lin

Rita Allen Foundation

  • Michael Z Lin

Stanford University (Graduate and Interdisciplinary Graduate Fellowships)

  • Helen H Yang

Stanford University (Stanford Neuroscience Microscopy Service pilot grant)

  • Michael Z Lin
  • François St-Pierre

Canadian Institutes of Health Research (MOP-81142)

  • Katalin Toth

Natural Sciences and Engineering Research Council of Canada (RGPIN-2015-06266)

  • Katalin Toth

Natural Sciences and Engineering Research Council of Canada (Graduate fellowship)

  • Simon Chamberland

National Institutes of Health (1U01NS090600)

  • Joseph C Wu

National Institutes of Health (HL12652701)

  • Joseph C Wu

National Institutes of Health (R01 EY022638)

  • Thomas R Clandinin

National Institutes of Health (R21 NS081507)

  • Thomas R Clandinin

National Science Foundation (1707359)

  • François St-Pierre

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

Reviewing Editor

  1. Kristin Scott, University of California, Berkeley, Berkeley, United States

Ethics

Animal experimentation: Animal experiments were performed in accordance with either (1) the recommendations in the Guide for the Care and Use of Laboratory Animals of the National Institutes of Health and the guidelines of the Stanford Institutional Animal Care and Use Committee under Protocol APLAC-23407, or (2) the guidelines for animal welfare of the Canadian Council on Animal Care and protocols approved by the Université Laval Animal Protection Committee (protocol number 2014-149-3). All surgery was performed under sodium pentobarbital anesthesia, and every effort was made to minimize suffering.

Version history

  1. Received: February 3, 2017
  2. Accepted: July 21, 2017
  3. Accepted Manuscript published: July 27, 2017 (version 1)
  4. Version of Record published: September 5, 2017 (version 2)
  5. Version of Record updated: September 14, 2017 (version 3)

Copyright

© 2017, Chamberland 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. Simon Chamberland
  2. Helen H Yang
  3. Michael M Pan
  4. Stephen W Evans
  5. Sihui Guan
  6. Mariya Chavarha
  7. Ying Yang
  8. Charleen Salesse
  9. Haodi Wu
  10. Joseph C Wu
  11. Thomas R Clandinin
  12. Katalin Toth
  13. Michael Z Lin
  14. François St-Pierre
(2017)
Fast two-photon imaging of subcellular voltage dynamics in neuronal tissue with genetically encoded indicators
eLife 6:e25690.
https://doi.org/10.7554/eLife.25690

Share this article

https://doi.org/10.7554/eLife.25690

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