2-photon excitation fluorescence microscopy enables deeper high-resolution imaging of voltage and Ca2+ in intact mice, rat, and rabbit hearts

Authors

  • Iffath A. Ghouri,

    1. Institute of Cardiovascular and Medical Sciences; College of Medical, Veterinary, and Life Sciences, University of Glasgow, Glasgow, UK
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  • Allen Kelly,

    1. Institute of Cardiovascular and Medical Sciences; College of Medical, Veterinary, and Life Sciences, University of Glasgow, Glasgow, UK
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  • Francis L. Burton,

    1. Institute of Cardiovascular and Medical Sciences; College of Medical, Veterinary, and Life Sciences, University of Glasgow, Glasgow, UK
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  • Godfrey L. Smith,

    1. Institute of Cardiovascular and Medical Sciences; College of Medical, Veterinary, and Life Sciences, University of Glasgow, Glasgow, UK
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  • Ole Johan Kemi

    Corresponding author
    1. Institute of Cardiovascular and Medical Sciences; College of Medical, Veterinary, and Life Sciences, University of Glasgow, Glasgow, UK
    • Phone: +44(0)1413305962, Fax: +44(0)1413305481

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Abstract

We describe a novel two-photon (2P) laser scanning microscopy (2PLSM) protocol that provides ratiometric transmural measurements of membrane voltage (Vm) via Di-4-ANEPPS in intact mouse, rat and rabbit hearts with subcellular resolution. The same cells were then imaged with Fura-2/AM for intracellular Ca2+ recordings. Action potentials (APs) were accurately characterized by 2PLSM vs. microelectrodes, albeit fast events (<1 ms) were sub-optimally acquired by 2PLSM due to limited sampling frequencies (2.6 kHz). The slower Ca2+ transient (CaT) time course (>1ms) could be accurately described by 2PLSM. In conclusion, Vm - and Ca2+-sensitive dyes can be 2P excited within the cardiac muscle wall to provide AP and Ca2+ signals to ∼400 µm. (© 2013 WILEY-VCH Verlag GmbH &Co. KGaA, Weinheim)

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