Anwendung der Fluoreszenz-Korrelations-Spektroskopie zur Untersuchung dynamischer Prozesse in lebenden Zellen

Abstract

In this thesis the Fluorescence Correlation Spectroscopy (FCS) technique is applied to cell-physiologically relevant reaction and diffusion problems in living cells. In the nucleus as well as in the cytoplasm of bovine adrenal chromaffin cells the FCS method is used to determine diffusion coefficients of calcium chelators that are often used as calcium indicators. At the interface of hippocampal cell membrane and extracellular solution the specific molecular interaction of a dye-labelled benzodiazepine with its receptor is examined. The affinity of the ligand-receptor-complex as well as the reaction rates of the complex are determined. The applicability of different dyes for ligand-labelling in FCS experiments is investigated. The FCS technique is suited to resolve the dynamics of single synaptic vesicles in hippocampal nerve endings. The effect of kinases, phosphatases and the cytoskeleton on the vesicle mobility are investigated. The mobility of these vesicles in unstimulated boutons is mainly due to active transport and under the control of the myosin light chain kinase. Inhibition of the phosphatases 1 and 2A leads to an increased vesicle mobility which can be further increased by destruction of the actin filaments in the synapse.