Real time, confocal imaging of Ca2+ waves in arterially perfused rat hearts

Abstract

Objective: The aim of this study was to characterize the spatio-temporal dynamics of slow Ca2+ waves (SCW's) with cellular resolution in the arterially-perfused rat heart. Methods: Wister rat hearts were Langendorff-perfused with Tyrode solution containing bovine-albumine and Dextran. The heart was loaded with the Ca2+ sensitive dye Fluo-3 AM. Intracellular fluorescence changes reflecting changes in [Ca2+]i were recorded from subepicardial tissue layers using a slit hole confocal microscope with an image intensified video camera system at image rates of up to 50/s. Results: SCW's appeared spontaneously during cardiac rest or after trains of electrical stimuli. They were initiated preferentially in the center third of the cell and propagated to the cell borders, suggesting a relation between the cell nucleus and wave initiation. They were suppressed by Ca2+ transients and their probability of occurrence increased with the Ca2+ resting level. Propagation velocity within myocytes (40 to 180 μm/s) decreased with the resting Ca2+ level. Intercellular propagation was mostly confined to two or three cells and occurred bi-directionally. Intercellular unidirectional conduction block and facilitation of SCW's was occasionally observed. On average 10 to 20% of cells showed non-synchronized simultaneous SCW's within a given area in the myocardium. Conclusions: SCW's occurring at increased levels of [Ca2+]i in normoxic or ischemic conditions are mostly confined to two or three cells in the ventricular myocardium. Spatio-temporal summation of changes in membrane potential caused by individual SCW's may underlie the generation of triggered electrical ectopic impulse