Quantitative Magic Angle Spinning Detection of Deuteration in Small Biopsies of Rat Brain

The replacement of the 12C present in cerebral metabolites by 13C derived from appropriate 13C enriched precursors, as detected by 13C NMR, has allowed the determination of the cerebral tricarboxylic acid and glutamine cycle fluxes. However, the 13C NMR method is relatively insensitive and requires significantly long acquisitions, a circumstance limiting the time resolution of the method. Up to now, only fluxes slower than the tricarboxylic acid cycle flux have been determined after fitting the 13C turnover curves to appropriate models of cerebral metabolism. To overcome this limitation we proposed earlier the investigation of the 1H-2H exchange of specific metabolite protons from 13C isotopomers, a process depicting faster kinetics and thus potentially able to investigate faster reaction rates. However, the High Resolution 13C NMR approach we proposed earlier, requires the use of relatively large amounts of brain samples for the preparation of extracts. More recently, it has become possible to obtain high resolution 13C and 1H NMR spectra directly from the tissue biopsies avoiding extract preparation, by using High Resolution-Magic Angle Spinning (HR-MAS) Spectroscopy. Here, we report on the use of 1D and 2D 2H, 1H-2H and 1H-13C HR MAS methodologies to investigate quantitatively 1H-2H turnover in small (< 10 mg), intact and unprocessed, biopsy samples prepared from the brain of deuterated rats.