Regional Variation of Calcium Transients within Neonatal Cardiomyocyte

Abstract

Cardiomyocyte contraction is preceded by a calcium transient, an influx of calcium into the cytosol causing a contraction to occur followed by relaxation which occurs as the intracellular calcium returns to resting level.We are investigating how calcium transient signals from the nuclear, perinuclear and more distant cytosolic regions of neonatal cardiomyocytes differ. Calcium transients exhibit significant differences in the rise and decay time courses between regions close to and relatively distant from the nucleus. Nuclear calcium transients are slower to occur and have broader peaks than cytosolic transients more distant from the nucleus. The distribution of SR around nuclear and perinuclear regions influences the behavior of calcium transient signals. We targeted four regions of interest (ROI) in the cell: (1) ROI 1 is the region with the least amount of nuclear calcium contribution; (2) ROI 2 refers to regions adjacent the nucleus where there is significant nuclear calcium contribution; (3) ROI 3 is within the nucleus; (4) ROI 4 refers to the signal averaged over the entire cell. In order to accurately define these four regions of interest the cells were stained with ER‐TrackerTM, a dye that binds to the SR, and with Fluo‐3. Calcium transients were recorded using Leica SP2 Fluorescent Laser Scanning Confocal Microscope. XT Scans were used to record all three regions of interest simultaneously for 30 sec and to compare these transients from an averaged signal recorded from the entire cell when cells were paced at 0.3 Hz. Neonatal ventricular myocyte measurements were obtained in an in‐line scan mode at 200 lines per second. Data files were exported in text format and graphed as bar charts and scatter plots. An averaged time to peak and averaged time to 50 percent signal decay were used to compare signal behavior between regions.