Fast two-photon imaging of subcellular voltage dynamics in neuronal tissue with genetically encoded indicators
- Université Laval, Canada
- Stanford University, United States
- Baylor College of Medicine, United States
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
Thomas R Clandinin
François St-Pierre
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.
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.
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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