Phosphate transport by capillaries of the blood-brain barrier.

Abstract

3P8Capillaries were isolated from bovine brain cortex and used for phosphate transport studies.The influx of phosphate through capillary membranes was studied by incubation with [32Pi]phosphate followed by a rapid filtration technique.Phosphate uptake by brain capillaries was mediated by a saturable high-affinity system which is independent of the sodium concentration in the incubation medium.The apparent half-saturation constant (K,) and maximal influx (VmaX) were estimated to 160 g~ and 0.37 nmol/mg protein/30 s.Transport was inhibited by the phosphate analogues arsenate and phosphonoformic acid with apparent inhibition constants of 5 and 11 mM, respectively.The metabolic inhibitors cyanide and ouabain had no effect on the transport activity.Competition experiments showed that phosphate uptake was inhibited up to 41% by various anions (pyruvate, acetate, citrate, glutamate, and sulfate).In addition, phosphate uptake was significantly decreased by two selective inhibitors of anionic exchangers, 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid and 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid.Chloride was not a substrate of the phosphate carrier as the replacement of external chloride, by nitrate, thiocyanate, or gluconate, did not increase phosphate transport.Aminohippuric acid and N'-methylnicotinamide, two specific substrates of anionic and cationic drug exchangers, did not compete with the phosphate carrier of cerebral capillaries.However, trans-stimulation with bicarbonate increased phosphate transport by 28%, and this stimulation was inhibited by 1 m M 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid, suggesting that the carrier of the cerebral capillaries could exchange phosphate with bicarbonate.The plasma membrane of endothelial cells of cerebral capillaries forms a continuous barrier between blood and brain interstitium.These endothelial cells are sealed together by tight junctions and do not possess fenestrations or transendothelial channels (1,2).Brain capillaries also contain a large number of mitochondria and are able to metabolize a variety of substrates (3, 4).This specialized endothelium regulates the movement of solutes between blood and brain.This is accomplished through carrier-mediated transport systems for hexoses, monocarboxylic acids, amino acids, nucleosides, purines, and amines (5).Active transport pumps in the endothe-