High-Resolution Calcium Imaging Method for Local Calcium Signaling.

All eukaryotic cells respond to extracellular signals in a physiologically meaningful way. For multicellular organisms, physiologically relevant cooperation is only possible, if cell-cell communication works properly. Here, the extracellular signals must be translated into intracellular signals that ultimately result in cellular responses. This process is termed signal transduction or signaling. Ca signaling has been developed in almost all eukaryotic cells. The cellular components used for this highly versatile signaling system are often termed "Ca toolbox". Besides Ca pumps and Ca-binding proteins, the Ca channels that are located in the plasma membrane and intracellular membranes and the Ca-mobilizing second messengers are major players in shaping the four-dimensional nature of Ca signals.Here, we report on methodological developments to acquire and analyze cellular Ca signals with high temporal and spatial resolution with specific focus on (1) photobleaching of Ca indicators at high acquisition rate, (2) determination of system noise and spatiotemporal detection limits, and (3) image processing.