Induction of Ca²⁺-Dependent Exocytotic Processes by Laser Ablation of Endothelial Cells

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.