Ion homeostasis in cells and cell organelles
Intracellular ion concentrations regulate various cellular processes. Changes in the cytoplasmic Ca2+ concentration represent a key signal for many cellular processes; Na+, K+, H+ and Cl- gradients provide the driving force for electrical processes, permit the accumulation of nutrients and neurotransmitters and allow cellular volume regulation. Sensors based on fluorescent dyes and fluorescent proteins allow quantification of ion concentrations in intact living cells in various preparations. We develop novel fluorescent protein-based sensors, and spectroscopically and microscopically characterize genetically encoded as well as organic fluorescent sensor molecules. We put particular emphasis on Fluorescence Lifetime Imaging (FLIM) based on time-correlated single photon counting (TCSPC) to quantify absolute ionic concentrations in cell culture, primary cultures as well as in acute slice preparations from normal or genetically altered animals. We combine these techniques with heterologous expression and electrophysiology to elucidate the molecular mechanisms underlying ion homeostasis in cytoplasm, lysosomes and synaptic vesicles in normal and pathologically altered cells.