The brain efflux index (BEI), a measurement of blood–brain barrier (BBB) efflux transport, was estimated at 15 s, 30 s, 1 min, 3 min and 10 min after intracerebral injection of [14C]pyrimidines. An initial steep increase of the BEI values over time was observed for uracil and [14C]thymine, followed by a more moderate increase after 1 min. For the corresponding nucleosides, [14C]uridine and [14C]thymidine, the increase of BEI values over time was less steep and linear between 30 s and 3 min. The apparent BBB efflux clearances for [14C]uridine, [14C]thymidine, [14C]uracil and [14C]thymine were (μl/min/g): 95.2 ± 12.1, 125.3 ± 18.4, 290.4 ± 28 and 358.5 ± 32.5, respectively, which is at least several folds higher than the predicted BBB influx clearances of uridine, uracil and thymidine. Quick depletion of brain parenchyma from brain microvasculature has revealed that [14C] radioactivity accumulated in brain microvessels after injection of nucleosides [14C]thymidine and [14C]uridine, but that was not observed when nucleobases, [14C]thymine and [14C]uracil, were injected. Reverse transcriptase-PCR revealed that the rat brain and liver (positive control) express dihydropyrimidine dehydrogenase, a key enzyme in pyrimidine nucleobase catabolism. Two bands representing spliced variants have been detected with the relative density of the bands (expressed relative to the density of glyceraldehyde3-phosphate dehydrogenase bands, mean ± SEM from 3 separate samples) 0.16 ± 0.06 and 0.04 ± 0.01 (brain) and 0.49 ± 0.1 and 0.07 ± 0.01 (liver). Overall, these results indicate that the net direction of pyrimidine BBB transport is the efflux transport; rapid BBB efflux transport and metabolic breakdown of pyrimidine nucleobases appear to be important for brain homeostasis.
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We acknowledge financial support of the Department of Physiology, Kuwait University and of the ICN Pharmaceuticals (their ex branch ICN Yugoslavia) for their kind support with radiolabeled nucleosides and nucleobases. We also acknowledge Mrs Nada Selakovic Bojovic for her technical assistance in the BEI experiments.
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