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Shigeki Watanabe, Ph.D.
Shigeki Watanabe, Ph.D.
BCMB Graduate Program
Research topic: Cellular and molecular characterizations of rapid changes during synaptic plasticity
For information processing and integration, neurons undergo rapid cellular and molecular reorganization. At the level of a synapse, the entire structure of the postsynaptic compartment, dendritic spines, can alter rapidly to mediate synaptic plasticity. At the molecular level, the addition of receptors to the surface of spines is associated with strengthening of synapses while their removal is associated with weakening and neurodegeneration. Many of these changes take place on a millisecond time scale. Despite the importance of these changes to the organism, remarkably little is known about either how the morphology of spines is regulated or how the surface occupancy of receptors is regulated. What are the morphological changes that trigger synaptic plasticity? How are receptors redistributed during this process? What are the molecular pathways that mediate the redistribution? We aim to answer these questions using cutting-edge electron microscopy techniques in combination with molecular and biochemical approaches.
Membrane dynamics. We pioneered the “flash-and-freeze” approach that adds temporal information to electron micrographs. Electron microscopy traditionally only captures a static image of cells. To visualize membrane dynamics, we combined optogenetic stimulation of neurons with high-pressure freezing. By varying the intervals between stimulation and freezing, we can essentially make a “flip-book” of cellular dynamics at a millisecond temporal resolution. Using this approach, we are studying the cellular and molecular mechanisms underlying rapid membrane trafficking at synapses.
Molecular topology and dynamics. We developed a correlative super-resolution fluorescence microscopy and electron microscopy approach that visualizes proteins in electron micrographs. We found a method that preserves fluorescence through harsh fixation and plastic embedding. We perform super-resolution and electron microscopy imaging on the same ultrathin sections of tissues and map the molecular topology onto the subcellular structures. This technique can be used to pinpoint the locations of proteins within their subcellular context. In combination with the “flash-and-freeze” approach, we are studying the molecular dynamics at synapses.