This subproject is one of many research subprojects utilizing the resources provided by a Center grant funded by NIH/NCRR. The subproject and investigator (PI) may have received primary funding from another NIH source, and thus could be represented in other CRISP entries. The institution listed is for the Center, which is not necessarily the institution for the investigator. This activity has two components to be selected by the user. These activities are in addition to weekly for teaching and discussion in various aspects of 13C NMR and metabolism in research group meetings. First, a hands-on course conducted over 3-4 days is made available on request. We offer this course for experienced investigators already familiar with conducting NMR experiments who would like an introduction to experiments based on the use of deuterium and carbon-13 in their own laboratories. The material covered includes the use of D2O and 13C propionate for measuring fluxes in vivo. The provision of D2O to subjects and measurement of deuterium exchange via 2H NMR of glucose provides the relative contributions of gluconeogenesis (from PEP or glycerol) and glycogenolysis. Giving [U-13C]proprionate in capsules and analysis of the C2 carbon of glucose via 13C NMR gives the relative rate of gluconeogenesis from PEP and flux through the citric acid cycle. When measurements from both approaches are combined, glucose production can be expressed in terms of citric acid cycle turnover. Combining this with measurement of glucose turnover (via the dilution in blood of glucose infused at a constant rate), the various relative measurements mentioned above (glucose production from various sources, citric acid cycle activity) can be expressed as absolute fluxes. The student has the opportunity to be involved in all aspects these measurements, from designing protocols to data analysis and interpretation. The student will observe the procedures for handling the preparation, processing samples, collecting NMR data, performing spectral analysis and finishing with applying metabolic models. Instruction in the use of software required at various stages is provided. The course is tailored to individual needs as closely as possible. Second, users are taught, one-on-one, how to interpret 1H NMR spectra from human subjects. This involves showing them how to open spectral data files, convert to format readable by ACD, transformation, fitting and conversion of raw data to physiological variables.
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