22 February 2013 Probing the spatiotemporal relationship between intracellular Ca2+ release and action potential propagation in cardiomyocytes by ultrafast multi-photon random access microscopy
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Abstract
Action potential, via the transverse axial tubular system (TATS), synchronously triggers uniform Ca2+ release throughout the cardiomyocyte. Cardiac diseases associated with TATS structural remodeling preclude a uniform Ca2+ release across the myocyte, contributing to contractile dysfunction. A simultaneous recording of intracellular local Ca2+ release and action potential in tubular network can be useful to unravel the link between TATS abnormality and dysfunctional EC coupling. Here we combine the advantage of an ultrafast random access multi-photon (RAMP) microscope with a double staining approach to optically record AP in several TATS elements and, simultaneously, the corresponding local Ca2+ transient. Isolated rat cardiomyocytes were labeled with a novel voltage sensitive dye (VSD) and a calcium indicator. RAMP microscope rapidly scans between lines drawn across the TATS of the cardiomyocyte to perform a multiplexed measurement of the two fluorescence signals. Although the calcium and voltage indicators can be excited at the same wavelength, the large Stokes shift of the VSD emission allows us to use spectral unmixing to resolve the voltage and calcium responses. In healthy cardiomyocytes, we found uniform AP propagation within the TATS and homogeneous Ca2+ release throughout the whole cell. The capability of our technique in probing spatiotemporal relationship between Ca2+ and electrical activity was then explored in a model of acute detubulation in which failure to conduct AP in disconnected TATS may cause local delay of Ca2+ transient rise leading to non-homogenous Ca2+ release.
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L. Sacconi, L. Sacconi, C. Crocini, C. Crocini, R. Coppini, R. Coppini, C. Ferrantini, C. Ferrantini, C. Tesi, C. Tesi, P. Yan, P. Yan, L. Loew, L. Loew, E. Cerbai, E. Cerbai, C. Poggesi, C. Poggesi, F. S. Pavone, F. S. Pavone, } "Probing the spatiotemporal relationship between intracellular Ca2+ release and action potential propagation in cardiomyocytes by ultrafast multi-photon random access microscopy", Proc. SPIE 8588, Multiphoton Microscopy in the Biomedical Sciences XIII, 85881B (22 February 2013); doi: 10.1117/12.2006109; https://doi.org/10.1117/12.2006109
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