The Cell Biology/Microscopy Core facility of the Massachusetts General Hospital Program in Membrane Biology provides intellectual and technical support for a large number of NIH-funded investigators within the MGH and Harvard community. Major funding for this facility is derived through a Membrane Biology Program Project (PI: Dennis Brown, Ph.D.) that comprises four individual grants and a Microscopy Core. These projects are brought together by a common interest in vesicle/protein trafficking and signal transduction events. Many other MGH investigators also share an interest in membrane trafficking, and are associate members of the cross-departmental Program in Membrane Biology. A major thrust of the entire Program is to dissect intracellular trafficking events. To allow these events to be examined and followed in real-time with high temporal and spatial resolution, a spinning disk confocal microscope system is requested in this application. The spinning disk system, coupled to an inverted microscope, is an ideal technology for examining real-time cellular events. It is marketed by Perkin Elmer as the """"""""Ultraview Live Cell Imager'. It allows rapid acquisition of confocal images, has low photobleaching and phototoxicity for lengthy observation periods on live material, and captures images with a sensitive cooled CCD camera with a high signal to noise ration and high resolution. At the last competing renewal of our Program Project (January 2002), the NIH committee found the Microscopy Core to be an """"""""Outstanding"""""""" facility, but strongly suggested the acquisition of state-of-the-art live cell imaging methodology to allow us to take advantage of many of the novel tools, cell lines and GFP-coupled constructs that were developed during the previous funding period. Four individual components of the Program Project, all of which deal with protein and vesicle trafficking events (aquaporin trafficking, V-ATPase recycling, endosomal coat protein recruitment, phospholipase A2 translocation), will be supported by this equipment. In addition, five individual grants with a major emphasis on protein and receptor recycling (e. g., vasopressin receptor, PTH receptor, vacuolar H-ATPase), as well as renal development in a Zebrafish model will be supported. Several minor users will benefit from the requested equipment, including two projects examining cytosol to nucleus translocation of regulatory proteins. Finally, the equipment requested will support two Center grants related to diabetes and inflammatory bowel disease research will be supported by the equipment requested. Thus, a wide-range of projects and investigators will benefit from this equipment, which will be integrated into a Core facility that has been operational for over 15 years.
