Analysis of Mechanotransducers in Endothelial Cells with Atomic Force Microscopy

Abstract

Several types of physiological stimuli increase cytosolic free Ca^<2+> concentration ([Ca^<2+>]i) in endothelial cells (ECs), and the spatiotemporal pattern of the [Ca^<2+>]i mediates the signal transduction from cell surfaces to internal cell organelles. We therefore investigated how mechanical stimuli were transduced to the spatiotemporal pattern of [Ca^<2+>]i increase, with an Atomic Force Microscope (AFM) for stimulating the subcellular domain. Mechanical stimulation resulted in an immediate and transient increase in [Ca^<2+>]i, as depending on not force but deflection of cells. In the subcellular level, fast Ca^<2+> response domains were different between mechanical stimulation and ATP-induced intracellular Ca^<2+> release. The mechanical stimulus-induced fast Ca^<2+> response started at the cell peripheral domains, and its response reduced when extracellular Ca^<2+> was removed. These results indicate that the earliest Ca^<2+> mobilization depends on extracellular Ca^<2+> through stretch-activated (SA) channels and domains of primary Ca^<2+> response reflect the distribution of SA channels.