CORE E. Molecular Mechanisms Core Plans: This core represents an expansion of the role of the previous Pharmacokinetics &Biomarker Core into what we now will call a Molecular Mechanisms Core in order to better meet the needs of the SCoBIRC faculty and other neuroscience investigators who use the SCoBIRC Core Facilities. In its former role, the core, managed by Core Director Dr. Edward D. Hall and Assistant Core Director and Supervisor Dr. Jeff Bosken, primarily employed GC-MS and HPLC technologies for bioanalytical measurements of drugs and potential injury-related biomarker compounds in biofluids (e.g blood, urine, cerebrospinal fluid). However, the core's scope was expanded very eariy during the current funding period to include equipment (LiCor Odyssey Infrared Image Analyzer) and expertise for highly sensitive analysis of immunoblots using infrared-emitting secondary anfibodies. This technology has been heavily employed by several SCoBIRC and other BBSRB neuroscience investigators. More recently, the core has added an Applied Biosystem StepOnePlus quantitative RT-PCR capability for gene expression analysis studies which is growing in use. However, going forward, we intend to add another functional component to the Core to support the large number of SCoBIRC and other neuroscience faculty who are studying changes in mitochondrial bioenergetics in in vitro and in vivo models of acute and chronic neurodegenerative disease models. The growing interest in mitochondrial functional studies across several funded and pilot projects has made it desirable to centralize this capability into the core to insure methodological consistency and quality control and comparability of data across laboratories. We plan to provide this centralized capability by first of all shifting existing equipment used for mitochondrial preparation and bioenergetic analyses of ex vivo mitochondrial isolates into the core and secondly to add new Seahorse technology for highly sensitive imaging of mitochondrial bioenergetics and dysfunction and whole cells in culture models. Requested Molecular Mechanisms Core Equipment and Supplies: We are specifically requesting funding for purchase of the following. Note: all equipment quotes are listed in Appendix I ? Seahorse Bioscience XF96 Dual Analyte Extracellular Analyzer- Justification: This state of art technology is capable of detecting real time changes in up to 3 analytes of cellular metabolism in a non-destructive manner in both energy producing pathways. $171,636 Molecular Mechanisms Core Usage: The following SCoBIRC and other neuroscience invesfigators who have and will continue to use the core include: Cambi, Geddes, Gerhardt, Gash, Guttman, Hall, Rabchevsky, Saatman, Smith, Snow, Springer, Sullivan, Zhang and Zhu. Molecular Mechanisms Core Staffing: The Center Core Grant PI Dr. Edward D. Hall (5% effort) will confinue to serve as Core Director for the Molecular Mechanisms Core. Dr. Jeff Bosken (100% effort) will also continue to serve as Core Supervisor. He will also be specifically responsible for conduct and training in regards to the bioanalytical instrumentation (GC-MS, HPLC) functional component. Dr. Indrapal Singh (25% effort), a Research Assistant Professor, who is highly expert in mitochondrial bioenergetic studies will be responsible for the mitochondrial bioenergetics component of the core in regards to equipment maintenance, training and pilot study conduct. . He will be responsible for maintenance, pilot studies and training of laboratory personnel in regards to the immunoblotting (western, slot, infra-red imaging) and quantitative RT-PCR gene expression functions of the core.
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