SUBCELLULAR-LOCALIZATION AND COMPARTMENTATION OF THIAMINE DERIVATIVESIN RAT-BRAIN

The subcellular distribution of thiamine derivatives in rat brain was studied. Thiamine diphosphate content was highest in the mitochondrial and synaptosomal fractions, and lowest in microsomal, myelin and cyto solic fractions. Only 3-5% of total thiamine diphosphate was bound to transketolase, a cytosolic enzyme. Thiamine triphosphate was barely de tectable in the microsomal and cytosolic fraction, but synaptosomes we re slightly enriched in this compound compared to the crude homogenate . Both myelin and mitochondrial fractions contained significant amount s of thiamine triphosphate. In order to estimate the relative turnover rates of these compounds, the animals received an intraperitoneal inj ection of either [C-14]thiamine or [C-14]sulbutiamine (isobutyrylthiam ine disulfide) 1 h before decapitation. The specific radioactivities o f thiamine compounds found in the brain decreased in the order: thiami ne > thiamine triphosphate > thiamine monophosphate > thiamine diphosp hate. Incorporation of radioactivity into thiamine triphosphate was mo re marked with [C-14]sulbutiamine than with [C-14]thiamine. The highes t specific radioactivity of thiamine diphosphate was found in the cyto solic fraction of the brain, though this pool represents less than 10% of total thiamine diphosphate. Cytosolic thiamine diphosphate had a t wice higher specific radioactivity when [C-14]sulbutiamine was used as precursor compared with thiamine though no significant differences we re found in the other cellular compartments. Our results suggest the e xistence of two thiamine diphosphate pools: the bound cofactor pool is essentially mitochondrial and has a low turnover; a much smaller cyto solic pool (6-7% of total TDP) of high turnover is the likely precurso r of thiamine triphosphate.