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Practical Methods for In Vivo Cortical Physiology with 2-Photon Microscopy and Bulk Loading of Fluorescent Calcium Indicator Dyes

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Neural Tracing Methods

Part of the book series: Neuromethods ((NM,volume 92))

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Abstract

In vivo 2-photon imaging of neurons that have been bulk-loaded with fluorescent calcium indicator dyes is permitting many fundamental principles of neural circuit organization and development to be uncovered. In this article, we describe the materials and procedures that we have used in our investigations of ferrets, tree shrews, and mice. Special attention is given to the design and construction of custom stereotaxic devices and the prevention of stray light from entering the 2-photon microscope during vision experiments.

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Acknowledgments

We thank Prakash Kara, Tom Mrsic-Flogel, Aaron Kerlin, and Clay Reid for their valuable advice as we were learning to perform 2-photon imaging. We also thank David Fitzpatrick and Leonard E. White for support and mentoring. We thank Frank Mello, machinist at Brandeis University, Don Pearce of the Medical Instrument Shop at Duke University Medical Center, and Janet Patterson of the Physics Machine Shop at Duke University for their creative input and expertise in building the stereotaxic devices we describe here. This work was supported by the National Institutes of Health, National Science Foundation, and the John Merck Foundation.

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Authors and Affiliations

  1. Department of Biology, Brandeis University, 415 South Street, Waltham, MA, 02453, USA

    Stephen D. Van Hooser, Arani Roy & Kelly Flavahan

  2. Department of Neurobiology, Duke University Medical Center, Durham, NC, USA

    Stephen D. Van Hooser, Elizabeth N. Johnson, Ye Li, Mark Mazurek, Julie H. Culp & Rishabh Kasliwal

  3. Duke Institute for Brain Science, Duke University, Durham, NC, USA

    Elizabeth N. Johnson

  4. Howard Hughes Medical Institute, University of California, Berkeley, CA, USA

    Ye Li

  5. Department of Biology, Metropolitan State University of Denver, Denver, CO, USA

    Mark Mazurek

  6. Center for Neuroscience, University of California, Davis, CA, USA

    Julie H. Culp

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  1. Stephen D. Van Hooser
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  2. Elizabeth N. Johnson
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  3. Ye Li
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  5. Julie H. Culp
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Corresponding author

Correspondence to Stephen D. Van Hooser .

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Editors and Affiliations

  1. Baylor College of Medicine, Houston, Texas, USA

    Benjamin R. Arenkiel

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Van Hooser, S.D. et al. (2015). Practical Methods for In Vivo Cortical Physiology with 2-Photon Microscopy and Bulk Loading of Fluorescent Calcium Indicator Dyes. In: Arenkiel, B. (eds) Neural Tracing Methods. Neuromethods, vol 92. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1963-5_6

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  • DOI: https://doi.org/10.1007/978-1-4939-1963-5_6

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-1962-8

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