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Inscribing Optical Excitability to Non-Excitable Cardiac Cells: Viral Delivery of Optogenetic Tools in Primary Cardiac Fibroblasts

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Optogenetics

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1408))

Abstract

We describe in detail a method to introduce optogenetic actuation tools, a mutant version of channelrhodopsin-2, ChR2(H134R), and archaerhodopsin (ArchT), into primary cardiac fibroblasts (cFB) in vitro by adenoviral infection to yield quick, robust, and consistent expression. Instructions on adjusting infection parameters such as the multiplicity of infection and virus incubation duration are provided to generalize the method for different lab settings or cell types. Specific conditions are discussed to create hybrid co-cultures of the optogenetically modified cFB and non-transformed cardiomyocytes to obtain light-sensitive excitable cardiac syncytium, including stencil-patterned cell growth. We also describe an all-optical framework for the functional testing of responsiveness of these opsins in cFB. The presented methodology provides cell-specific tools for the mechanistic investigation of the functional bioelectric contribution of different non-excitable cells in the heart and their electrical coupling to cardiomyocytes under different conditions.

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Acknowledgements

This work was supported by NIH-NHLBI grant R01-HL-111649 and NSF-Biophotonics grant 1511353 (to E.E.), and partially by a NYSTEM grant C026716 to the Stony Brook Stem Cell Center. We thank Christina Ambrosi and Aleks Klimas for helpful discussions.

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Authors and Affiliations

  1. Department of Biomedical Engineering, Stony Brook University, Stony Brook, NY, USA

    Jinzhu Yu

  2. Department of Biomedical Engineering, Institute for Molecular Cardiology, Stony Brook University, ST-6, Room 120B, Stony Brook, NY, 11794-8661, USA

    Emilia Entcheva Ph.D.

Authors
  1. Jinzhu Yu
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  2. Emilia Entcheva Ph.D.
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Corresponding author

Correspondence to Emilia Entcheva Ph.D. .

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Editors and Affiliations

  1. University of Bielefeld, Dept of Cellular & Dev Biology of Plants, Bielefeld, Germany

    Arash Kianianmomeni

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Yu, J., Entcheva, E. (2016). Inscribing Optical Excitability to Non-Excitable Cardiac Cells: Viral Delivery of Optogenetic Tools in Primary Cardiac Fibroblasts. In: Kianianmomeni, A. (eds) Optogenetics. Methods in Molecular Biology, vol 1408. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-3512-3_21

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  • DOI: https://doi.org/10.1007/978-1-4939-3512-3_21

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  • Publisher Name: Humana Press, New York, NY

  • Print ISBN: 978-1-4939-3510-9

  • Online ISBN: 978-1-4939-3512-3

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