Laser Capture Microdissection System to Study Neuromuscular Biology

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Gene and protein expression profiling and the delineation of specific signaling pathways activated in response to a variety of stimuli within skeletal muscle have for the most part focused on the analysis of whole tissue extracts obtained from human biopsies or animal models. While these studies have yielded important insights into the aggregate response of skeletal muscle to various physiological stimuli, such approaches do not allow elucidation of the specific molecular responses of individual myofiber types, satellite cells, or capillaries within the muscle. Similarly, in animal models of neuromuscular adaptation, gene expression in motoneurons specifically is convoluted when whole spinal cords are analyzed. To address these knowledge gaps, we require a methodology to study cell-specific gene expression in muscles and spinal cord during conditions of phenotypic plasticity. Therefore, the purpose of this proposal is to obtain a laser capture microdissection (LCM) system. LCM is an approach that permits the rapid and reliable dissection and capture of homogeneous cell populations (e.g., myofibers, endothelium, motoneurons) or defined subcellular regions (e.g., neuromuscular junctions, myonuclear domains) without contamination from surrounding tissues, which enables a truly mechanistic approach for assaying the quantitative transcriptomic and proteomic status of these components in various phenotypes, as well as during transitions between these states. Few studies have investigated the cell-specific biology or subcellular biology of skeletal muscle or spinal cord in vivo. Furthermore, none have focused on mechanisms of gene expression during conditions of neuromuscular remodelling in basic rodent experiments, as well as in applied and basic human studies with particular emphasis on the effects of acute and chronic exercise in young and older males and females. Thus, acquisition of a LCM system represents a timely opportunity to innovate, to substantially enhance the research capabilities of our laboratories, as well as to expand HQP training opportunities using state-of-the-art technologies.