In vivo und in vitro studies on second messenger homeostasis and synapse formation in dystrophic

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While molecular analyses in cell culture and other in vitro systems have previously revealed vast informations on the details of cell signaling cascades, our insights into their physiological relevance in the intact healthy or sick organism are still very limited. Green Fluorescent Protein (GFP)-technology and two-photon microscopy have now opened a new, fascinating window for the critical evaluation of numerous hypotheses in living tissue. In my previous work, I could establish a method which allows to express and subsequently monitor heterologous fluorescent proteins by means of two-photon microscopy in the live mouse skeletal muscle in situ. Using GFP-based cameleon Ca2+ sensors in this assay, I could show for the first time in situ that mitochondria take up Ca2+ both during tetanic stimulation and single twitches. In my future work I want to further broaden the range of applications with this methodology to gain insights into the cellular signaling of skeletal and cardiac muscle in health and disease in the intact organism. To this end, I will first develop and use new and existing probes, respectively, to study the handling of second messengers and the protein dynamics of the neuromuscular junction in normal and dystrophic skeletal muscle. In a second part, I want to establish a method to investigate the second messenger handling in cardiac muscle in normal and dystrophic mice in situ.