Fast isotopic exchange between mitochondria and cytosol in brain revealed by relayed C-13 magnetization transfer spectroscopy

In vivo C-13 magnetic resonance spectroscopy has been applied to studying brain metabolic processes by measuring C-13 label incorporation into cytosolic pools such as glutamate and aspartate. However, the rate of exchange between mitochondrial alpha-ketoglutarate/oxaloacetate and cytosolic glutamate/aspartate (V-x) extracted from metabolic modeling has been controversial. Because brain fumarase is exclusively located in the mitochondria, and mitochondrial fumarate is connected to cytosolic aspartate through a chain of fast exchange reactions, it is possible to directly measure V-x from the four-carbon side of the tricarboxylic acid cycle by magnetization transfer. In isoflurane-anesthetized adult rat brain, a relayed C-13 magnetization transfer effect on cytosolic aspartate C2 at 53.2 ppm was detected after extensive signal averaging with fumarate C2 at 136.1 ppm irradiated using selective radiofrequency pulses. Quantitative analysis using Bloch-McConnell equations and a four-site exchange model found that V-x approximate to 13-19 mu mol per g per min (>> V-TCA, the tricarboxylic acid cycle rate) when the longitudinal relaxation time of malate C2 was assumed to be within +/- 33% of that of aspartate C2. If V-x approximate to V-TCA, the isotopic exchange between mitochondria and cytosol would be too slow on the time scale of C-13 longitudinal relaxation to cause a detectable magnetization transfer effect.