Induction of Ca2+-Dependent Exocytotic Processes by Laser Ablation of Endothelial Cells
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Ca2+ regulates a variety of cellular processes that are essential to maintain cell integrity and function. Different methods have been used to study these processes by increasing intracellular Ca2+ levels. Here, we describe a protocol to initiate Ca2+-dependent membrane-related events, using laser ablation by near-infrared irradiation. This creates a rupture in the plasma membrane that allows the extracellular Ca2+ to enter the cell and thereby induce a receptor-independent Ca2+ increase. We report laser ablation protocols to study two different Ca2+-induced processes in human endothelial cells—membrane resealing and exocytosis of secretory granules called Weibel-Palade bodies (WPBs). Thus, laser ablation represents a technique that permits the analysis of different Ca2+-regulated processes at high spatiotemporal resolution in a controlled manner.
Key wordsCa2+ Laser ablation Plasma membrane HUVEC FM4-64 WPB exocytosis
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