Transport Of Α-aminoisobutyric Acid Research Articles

Inhibition of oxidative insult in cultured cells by a novel 6-chromanol-containing antioxidant

N18-RE-105 neuronal hybridoma cells were used in a cell culture system to evaluate the protective effects of a novel 6-chromanol-containing antioxidant, U78517F. First, the incorporation of the compound into the cells was evaluated, using a serum albumin carrier. Then the cells were exposed to peroxide-generating compounds, and the cell injury was estimated from the loss of α-aminoisobutyric acid (AIB) transport. We found that U78517F only protected the cells significantly when the degree of oxidative insult was below a certain limit; the measurable protection of cells by U78517F against either cumene hydroperoxide or H 2O 2 was limited to a narrow range of concentrations of the reactive oxygen species generator. Additionally, the protection provided by U78517F was largely localized to the cell membrane and did not extend to protection of mitochondrial function. The action of U78517 was fully consistent with a direct radical scavenging in the cells. The results indicate that the following factors must be taken into account for evaluation of antioxidants in cell culture: (a) the delivery of a compound to cells, especially when the compound is lipophilic; (b) the nature and extent of the oxidative insult used to evaluate protection; and (c) the location of the protective agent in the cells.

Transport of α-aminoisobutyric acid across the blood-brain barrier studied with in situ perfusion of rat brain

Transport of α-aminoisobutyric acid (AIB) from blood to brain in pentobarbital-anesthetized rats was examined using in situ perfusion. In situ perfusion with washed sheep red blood cells allowed the precise control of the composition of the perfusate that was necessary for a detailed examination of the transport of AIB. Retrograde perfusion at 4 ml/min through the left external carotid artery with oxygenated, artificial blood (hematocrit= 0.3) maintained a normal electroencephalogram during a 10 min experiment. The perfusate cerebral blood flow, at a value of 1.2 ± 0.1ml/g/min, and the perfusate cerebral plasma volume, at a value of 5.4 ± 1.9 μl/g, in the left frontal cortex were within the range of reported in vivo values. The in situ PS product for AIB (3.8 ± 0.4 μl/g/min) was higher than the value observed in vivo. AIB uptake was reduced to the in vivo value by 2 mM phenylalanine (1.3 ± 0.3 μl/g/min) and equally well by a mixture of neutral amino acids at their normal plasma concentrations but was unaffected by 2 mM methyl-AIB or removal of sodium from the perfusate. A kinetic analysis showed that the apparent K i for phenylalanine inhibition of AIB transport was 19.8 ± 4.9 μM. Thus, although AIB has affinity for the large neutral amino acid carrier in the blood-brain barrier, brain uptake by this mechanism in vivo is negligible due to competition by other amino acids in the plasma.

Characteristics ofα–aminoisobutyric acid transport by lactating rat mammary gland

The transport of alpha-aminoisobutyric acid (AIB) by lactating rat mammary tissue has been examined. AIB uptake by mammary tissue was via both Na(+)-dependent and Na(+)-independent pathways. AIB uptake via the Na(+)-dependent pathway was inhibited by (methylamino)isobutyric acid (MeAIB) whereas AIB uptake via the Na(+)-independent pathway was blocked by 2-aminobicyclo[2,2,1]-heptane-2-carboxylic acid (BCH). A small fraction of AIB influx persisted in the presence of both MeAIB and BCH. The Na(+)-independent moiety of AIB uptake was strongly inhibited by phenylalanine, tryptophan, leucine, isoleucine and methionine. AIB efflux from mammary tissue slices was found to be both Na(+)-dependent and Na(+)-independent. Trans-stimulation of AIB efflux by other amino acids was not observed; in contrast, external phenylalanine, tryptophan and leucine inhibited AIB efflux. The results are largely consistent with the presence of systems A and L in lactating rat mammary tissue. However, the Na(+)-independent fraction of AIB transport may represent transport via a tissue specific form of system L.

Effect of Xylose on the Synthesis of Phosphorylcholine and Phosphorylethanolamine in Rat Lenses

Choline, an essential phospholipid precursor, enters the lens by a facilitated transport system and is phosphorylated to form phosphorylcholine (P-choline). Intact lenses incubated with [ 3H]choline accumulate both [ 3H]choline and P-[ 3H]choline. The rate and extent of this accumulation have been used to study the effects of osmotic or oxidative cataractogenic stress, and also to test the ability of compounds to protect lenses from stress-related damage. The initial effect of oxidative stress on choline metabolism is decreased choline transport, but the mechanism by which osmotic stress affects the accumulation of [ 3H]choline is not understood. The effects of osmotic and oxidative stress on choline influx and metabolism were compared in rat lenses incubated in TC-199 medium. After osmotic stress by incubation with 30 mM xylose for up to 24 hr, lenses accumulated the same amount of radiolabel as controls during a 30 min pulse with [ 3H]choline. However, if the lenses were exposed to [ 3H]choline for 6 hr so that accumulation of radiolabel in the lenses was limited by the rate of P-choline synthesis. xylose treated lenses accumulated less choline than controls. Separation of the lenticular radiolabel into [ 3H]choline and P-[ 3H]choline confirmed that xylose decreased synthesis of P-choline, although adequate unphosphorylated [ 3H]choline was available in the lenses. A decrease in P-[ 3H]ethanolamine synthesis was also seen in xylose-treated lenses incubated with [ 3H]ethanolamine. Ethanolamine can enter lenses by a non-saturable process which is not dependent upon a transporter. Although xylose decreased P-choline synthesis in intact lenses, neither xylose nor xylitol inhibited choline kinase in lens homogenates, Xylose also decreased transport of αaminoisobutyric acid, which enters lenses by a different transporter than choline. In contrast to osmotic stress from xylose, oxidative stress from singlet oxygen decreased choline influx. Thus, the superficially similar effects of osmotic and oxidative stresses on accumulation of radiolabeled choline by cultured rat lenses actually occur by different mechanisms and reflect different types of damage to the lens.

Hormonal influence on the permeability of the blood-brain barrier: Effect of an analog of adrenocorticotropic hormone, β 1–24 corticotrophin

Regional unidirectional transport of α-aminoisobutyric acid (AIB) (mol. wt.: 104) and sucrose (mol wt.: 342) which have a low permeability across the intact endothelium was investigated in brain of rats either treated with synacthène: an analog of ACTH, tetracosactide retard (β-1–24 corticotrophin) or in brain of placebo-treated controls. Three days treatment with synacthène, reduced the rate of influx of AIB and sucrose in most of the brain regions studied especially in thalamus, hypothalamus, cortex, and caudate nucleus without affecting the vascular compartment. The brainstem, cerebellum and white matter were less affected. These experimental findings may suggest that ACTH exhibits significant influence on hormonal regulation of blood-brain barrier permeability. Thereby such a regulation may involve the entry of polar compounds into the CNS and may influence the central effects of diffusion-limited drugs.

Functional receptors for insulin-like growth factors I and II in rat thymocytes and mouse thymoma cells

Functional receptors for insulin-like growth factors (IGF) I and II have been identified in rat thymocytes and mouse thymoma cell lines R1.1 and S49.1. IGF-I receptor α-subunit (MW130 000) bind IGF-I and IGF-II with equal affinity ( K d ∼ 4–7 NM ), and insulin with ∼ 100 times lower affinity. Tyrosine kinase activity and autophosphorylation of the IGF-I receptor β-subunit (MW 95 000) are stimulated by IGF-I and IGF-II with equal potency (ED 50 ∼ 0.5 nM). IGF-II receptors (MW 250 000) bind IGF-II with K d ∼ 0.3 nM and IGF-I with 30 times lower affinity, but not insulin. IGF-I and IGF-II do not cross-react with the insulin receptor to which insulin bonds with an apparent K d ∼ 1 nM , and stimulates its tyrosine kinase activity with ED 50 ∼ 3 nM. In thymocytes, α-aminoisobutyric acid transport is stimulated 2-fold by IGF-I and IGF-II with identical potency (ED 50 ∼ 2 nM), and by insulin with ED 50 ∼ 10 nM. Activation of thymocytes by concanavalin A increased the number of IGF-II receptors 2-fold, whereas IGF-I receptor binding and IGF-stimulated amino acid transport were unaltered. We conclude that the effect of IGF-I and IGF-II in thymocytes is mediated via binding to the IGF-I receptor and stimulation of its tyrosine kinase. The presence of functional IGF receptors on thymocytes and thymoma cells suggests that IGF-I and IGF-II play a role in the regulation of thymic functions.

Regulation of amino acid transport in rat and human thyroid cells.

To clarify the role of insulin, IGF-I and TSH in thyroid cell regulation, their effects on amino acid transport were studied separately. These effects were noted and compared using both the Wistar rat thyroid cell line and human thyroid cell cultures. Insulin, IGF-I and TSH were able independently to induce the radiolabelled alpha-aminoisobutyric acid transport within the rat thyroid cells: TSH stimulated the amino acid transport in rat thyroid cells in a dose-dependent way from 1 pmol/l to 10 nmol/l. Similarly, insulin increased amino acid transport significantly from 0.17 nmol/l up to 0.17 mumol/l and IGF-I from 0.13 pmol/l up to 0.13 mumol/l. The combined effects of insulin and TSH on amino acid transport were equal to the theoretical sum of the activities, whereas those of IGF-I and TSH were greater than the theoretical one. When human thyroid cell cultures were used, a significant increase of labelled amino acid transport was induced by TSH, i.e. from 0.1 pmol/l to 10 pmol/l; IGF-I stimulated amino acid transport in a range from 0.13 pmol/l to 13 pmol/l, under the same conditions. Conversely, only large doses of insulin, i.e. 1.7 nmol/l, were able weakly to stimulate amino acid transport. When submaximal TSH and IGF-I doses were co-incubated in human thyroid cells, an additive effect on amino acid transport was observed.

α-Aminoisobutyric acid transport in isolated rat submandibular salivary acinar cells

Na +-dependent α-aminoisobutyric acid (AIB) transport by isolated submandibular cell aggregates (pmol min −1 mg protein −1) was greater in the presence than in the absence of insulin, V max (5220 compared with 2900). K m (1.78 and 1.40 mM, respectively) was unaffected by insulin. Na +-dependent methyl-aminoisobutyric acid (MeAIB) transport was also greater in the presence of insulin ( V max, 3120 compared with 2010 pmol min −1 mg protein −1; K m, 1.03 and 0.93 mM). In the presence of 10 mM MeAIB, Na +-dependent AIB transport was reduced to 76 pmol min −1 mg protein −1 in both control and insulin-treated cells. The remaining Na +-dependent uptake of AIB was inhibited by 10 mM serine. Na +-independent AIB transport was unaffected by insulin, and in the presence of 5mM 2-aminobicyclo-[2,2,1]-heptane-carboxylic acid (BCH) AIB uptake was reduced to 10% of that observed under Na +-replete conditions. In the absence of insulin, the rate of Na +-dependent AIB uptake rapidly decayed; however, following the addition of hormone the rate of transport was maintained. Thus in the rat submandibular gland AIB uptake is mediated by at least three transport systems (A, L and ASC), and maintenance of normal system A activity requires insulin.

Transport of α-aminoisobutyric Acid into Rat Parotid after X-irradiation

Rat parotid gland exposed to 20 Gy X-irradiation exhibits functional alteration 3 days after exposure. The flow rate of saliva and the uptake of alpha-aminoisobutyric acid by the gland was reduced to 50 per cent of values for the control nonirradiated glands. When the same gland was studied in an in vitro system it functioned normally. K+ release and alpha-aminoisobutyric acid uptake by the irradiated dispersed acinar cells was comparable to the control. Transport alteration from the circulatory system into the parotid gland may cause the initial radiation-induced damage.

Characteristics of a neutral amino acid transport system (system A) in osteoblastic rat osteosarcoma cells

Transport of α-aminoisobutyric acid (AIB) in the clonal, osteoblastic-like cell line, ROS 17/2, was characterized. AIB transport was time-, temperature- and Na +-dependent. Both ouabain and monensin inhibited AIB transport in these cells. AIB uptake followed Michaelis-Menten kinetics with an apparent K m = 0.57 mM and a V max = 4.07 nmol/30 min/plate. These characteristics are consistent with the presence of system A neutral amino acid transport in ROS 17/2 cells. Exposure of ROS 17/2 cells to either parathyroid hormone or dibutyryl cyclic AMP (db-cAMP), but not to dibutyryl cyclic GMP (db-cGMP), markedly stimulated AIB transport. This suggests that extracellular stimuli which enhance osteogenic responses in this cell type, coordinately upregulate system A transport.

Transport of α-aminoisobutyric acid across the blood-brain barrier after different durations of ischaemia

Transport of α-aminoisobutyric acid across the blood-brain barrier after different durations of ischaemia

The Influence of Temperature on the Functions of Cultured Sertoli Cells*

Three functions of Sertoli cells were examined with cells from rats aged 13 and 25 days at 34 C, 38 C, and 40 C, namely protein synthesis (incorporation of amino acids into material precipitated by trichloroacetic acid), transport of alpha-aminoisobutyric acid, and production of lactate. A fourth function, namely production of cAMP was examined at two temperatures (34 C and 38 C) in cells from rats aged 25 days. In all cases, activity was greater at 38 C than at 34 C and greater at 40 C than at 38 C. Lysate of Sertoli cells showed similar differences in protein synthesis at the three temperatures, again at both ages. Protein synthesis was higher at 38 C than at 34 C in Sertoli cell-enriched tubules from rats aged 25 days and in Sertoli cells from normal rats of the same age cultured on rat tail collagen. The difference in protein synthesis at the two temperatures was seen whether the cells were cultured, before the experiment, for 7 days at 38 C or at 34 C. Production of cAMP by Sertoli cells was greater at 38 C than at 34 C. It is concluded that Sertoli cells do not show in vitro any inhibitory effect of body temperature on any of the functions tested, but on the contrary, are more active at body than at scrotal temperature. The possible significance of the differences seen is discussed.

Energetic basis of development of salt-tolerant transport in a moderately halophilic bacterium, Vibrio costicola

Active transport of α-aminoisobutyric acid (AIB) in Vibrio costicola utilizes a system with affinity for glycine, alanine and, to some extent, methionine. AIB transport was more tolerant of high salt concentrations (3–4 M NaCl) in cells grown in the presence of 1.0 M NaCl than in those grown in the presence of 0.5 M NaCl. The former cells could also maintain much higher ATP contents than the latter in high salt concentrations. Transport kinetic studies performed with bacteria grown in 1.0 M NaCl revealed three effects of the Na+ ion: the first effect is to increase the apparent affinity (K t) of the transport system for AIB at Na+ concentrations <0.2 M, the second to increase the maximum velocity (V max) of transport (Na+ concentrations between 0.2 and 1.0 M), and the third to decrease the V max without affectig K t (Na+ concentrations >1.0 M). Cells grown in the presence of 0.5 M or 1.0 M NaCl had similar affinity for AIV. Thus, the differences in salt response of transport in these cells do not seem due to differences in AIB binding. Large, transport-inhibitory concentrations of NaCl resulted in efflux of AIB from cells preloaded in 0.5 M or 1.0 M NaCl, with most dramatic efflux occurring from the cells whose AIB transport was more salt-sensitive. Our results suggest that the degree to which high salt concentrations affect the transmembrane electrochemical energy source used for transport and ATP synthesis is an important determinant of salt tolerance.

Energetics of sodium-dependent α-aminoisobutyric acid transport in the moderate halophile Vibrio costicola

The energetics of α-aminoisobutyric acid transport were examined in Vibrio costicola grown in a medium containing the NaCl content (1 M) optimal for growth. Respiration rate, the membrane potential (Δψ) and α-aminoisobutyric acid transport had similar pH profiles, with optima at 8.5–9.0. Cells specifically required Na + ions to transport α-aminoisobutyric acid and to maintain the highest Δψ (150–160 mV). Sodium was not required to sustain high rates of O 2-uptake. Δψ (and α-aminoisobutyric acid transport) recovered fully upon addition of Na + to Na +-deficient cells, showing that Na + is required in formation or maintenance of the transmembrane gradients of ions. Inhibitions by protonophores, monensin, nigericin and respiratory inhibitors revealed a close correlation between the magnitudes of Δψ and α-aminoisobutyric acid transport. Also, dissipation of Δψ with triphenylmethylphosphonium cation abolished α-aminoisobutyric acid transport without affecting respiration greatly. On the other hand, alcohols which stimulated respiration showed corresponding increases in α-aminoisobutyric acid transport, without affecting Δψ. Similarly, N, N′-dicyclohexylcarbodiimide (10 μM) stimulated respiration and α-aminoisobutyric acid transport and did not affect Δψ, but caused a dramatic decline in intracellular ATP content. From these, and results obtained with artificially established energy sources (Δψ and Na + chemical potential), we conclude that Δψ is obligatory for α-aminoisobutyric acid transport, and that for maximum rates of transport an Na + gradient is also required.

VIRALLY INDUCED MEMBRANE PERMEABILITY CHANGES AS A POSSIBLE LINK BETWEEN INFLUENZA B AND REYE'S SYNDROME

The mechanism by which viral infection can lead to the multiple metabolic derangements characteristic of Reye's Syndrome is not understood. The possibility that influenza virus could induce changes in membrane permeability to nutrients ordinarily concentrated within the cell was examined. Madin Darbin canine kidney (MDCK) cells were infected by egg grown influenza B virus by two routes: at physiological conditions, pH 7.4, 37°C; and adsorption at 0°C followed by brief exposure to pH 5.0, 37°C. Control cells were mock-infected with allantoic fluid. Transport (uptake and release) of phosphate (Pi) 2-deoxyglucose (dGlc) and α-aminoisobutyric acid (AIB) were measured at various intervals 0 to 10 hours after infection. At physiological pH, Pi uptake by infected cells was inhibited (p< 0.01) at 0 to 2 hours as compared to controls. Uptake of AIB was also inhibited (p< 0.01) at 0 to 2 hours. The uptake of all nutrients was higher at 6 to 10 hours in infected cells as compared to controls (p< 0.01). When release of Pi and dGlc was measured, there was no difference between infected cells and controls at 0-10 hours. At low pH, fusion occurred in infected cells, but not in control cells. Fused infected cells demonstrated both significantly increased release and diminished uptake of nutrients compared to controls. Thus influenza B virus can induce changes in host cell membrane permeability that may have important implications for cell metabolism.

α-Aminoisobutyric Acid Transport In The Midgut Of Two Lepidopteran Larvae

ABSTRACT A net absorption of a-aminoisobutyric acid (AIB) takes place in vitro in the midgut of two lepidopteran larvae, Philosamia cynthia Drury and Bombyx mori L. In P. cynthia the midgut epithelium accumulates AIB from the lumen, while in the same conditions AIB accumulation is not observed in B. mori midgut cells. In P. cynthia, when the lumen is bathed with a low K solution, the net absorption of AIB is reduced and the intracellular accumulation from the lumen is abolished. Brush border membrane vesicles, prepared from the midgut of both species, show a transient K-dependent concentrative uptake of AIB. The relationship between AIB uptake and AIB concentration in the presence of a transmembrane K gradient was studied inB. mori vesicles and the kinetic constants were calculated. These results confirm that there is a K-amino acid co-transport system on the brush border of columnar cells in the midgut of both larvae.

Effects of NaCl on the uptake of alpha-[14C]aminoisobutyric acid by the halotolerant bacterium Halomonas elongata.

The alpha-aminoisobutyric acid (AIB) transport system of the halotolerant bacterium, Halomonas elongata, was examined. Cells were grown in L-alanine defined medium with 0.05, 0.375, 1.37, 2.5, or 3.4 M NaCl. Each group of cells was resuspended in buffered salts with different NaCl concentrations (0.05, 0.375, 1.37, 2.5, and 3.4 M) and the uptake of alpha-[14C]AIB was measured. Optimum AIB uptake occurred in the 0.375 M NaCl solution for the lower salt grown cells and the 1.37 M NaCl solution for the higher salt grown cells. When cells were grown in the higher salt media and suspended in hypoosmotic solutions, appreciable AIB uptake occurred; but for cells grown in lower salt media and suspended in hyperosmotic solutions, the uptake was dramatically reduced. This effect was mainly attributed to cell plasmolysis which in turn resulted in some cell death. The AIB uptake was Na+ specific and this analogue was not metabolized after being transported into the cells. An amino acid competition study gave a pattern similar to that of a marine bacterium.

Energy Parameters, Macromolecular Synthesis and Cell Cycle Progression of in vitro Grown Ehrlich Ascites Tumor Cells after Inhibition of Oxydative ATP Synthesis by Oligomycin

In order to elucidate the significance of oxidative ATP production for the proliferation of Ehrlich ascites tumor cells, cell cycle progression, energy metabolism and macromolecular synthesis in the presence of oligomycin were studied. In the presence of the inhibitor (20 micrograms/ml), lactate production and glucose uptake of the cells increased by about 30-35% as compared to controls; oxygen consumption was maximally inhibited by 30-45% and could not further be reduced by higher concentrations of the inhibitor. ATP/ADP ratios of the oligomycin treated cells and control cells were not significantly different. In the first passage in the presence of oligomycin proliferation of the cells is reduced to about 50% that of controls; without severely affecting viability (dye exclusion test). In the second passage with oligomycin cell proliferation completely arrests. As was shown by flow cytometric analysis and BrdU-H33258 technique of flow cytometry, cells accumulate in the early S phase; division of cells which are in the S- and G2M compartment at the beginning of oligomycin treatment accounts for the increase of cell number in the first passage in the presence of oligomycin. On recultivation in the third passage in the absence of the inhibitor cells take up proliferation again; an increase of cell number of about 60% of controls was observed within 24 h. In the presence of oligomycin incorporation of [2-14C]thymidine is reduced to about 20% of the controls within 8 h, incorporation of [U-14C]lysine begins to slow down immediately after treatment with the inhibitor, the same is true for the incorporation of [2-14C]uridine.(ABSTRACT TRUNCATED AT 250 WORDS)

Driving forces and current-voltage characteristics of amino acid transport in Riccia fluitans

The complete steady-state I–V relationship of α-aminoisobutyric acid transport across the plasmalemma of rhizoid cells from Riccia fluitans has been measured and analysed with special emphasis on α-aminoisobutyric acid equilibrium and saturation conditions. (A) The electrical data show that: (1) the amino acid-induced electrical current saturates after the addition of the amino acid, regardless of the concentration; (2) a steady state is reached 1–2 h after incubation in α-aminoisobutyric acid, but after less that 5 min in the presence of 1 mM CN −; (3) the steady-state I–V characteristic of α-aminoisobutyric acid transport is a sigmoid curve and fairly symmetric in current with respect to the voltage axis; and (4) the equilibrium potential is clearly a function of the amino acid accumulation ratio. It is suggested that the sigmoid curve represents the characteristic of carrier-mediated α-aminoisobutyric acid transport with a voltage-insensitive step, possibly the translocation of the unloaded carrier, rate-limiting. Since under normal conditions the voltage-sensitive rate constant k oi is much greater than k io , it is further suggested that the energy to drive this system is put into the transfer of positive charge from outside to the cytoplasm. (B) Accumulation ratios have been determined by inspection of current-voltage data, and additionally by compartmental analysis on green thalli from Riccia fluitans. Both methods give ratios far too low compared with the thermodynamically possible accumulation of about 10 4. It is suggested that substantial leakages via different non-electrical pathways prevent equilibrium at steady state, and it is concluded that in such leaky systems the thermodynamic equilibrium condition is not suitable for estimating stoichiometries.

Transport of alpha-aminoisobutyric acid across brain capillary and cellular membranes.

The transport of alpha-aminoisobutyric acid (AIB), N-methyl-AIB (MeAIB), and diethylenetriaminepentaacetic acid (DTPA) from blood to brain was measured over different experimental periods in eight regions of the rat brain. Unidirectional transfer rate constants were determined from multiple-time/graphical and single-time analysis of the experimental data; values of 0.0018, 0.00057, and 0.000021 ml g-1 min-1, respectively, were obtained for the thalamus by graphical analysis. The initial distribution volume of AIB and MeAIB in brain tissue was several-fold greater than that of DTPA and the tissue plasma volume, and this difference was not accounted for by red blood cell uptake. This discrepancy could be due to rapid transport of AIB and MeAIB into brain endothelial cells in addition to the relatively rapid uptake by choroidal, meningeal, and ependymal associated tissues that was demonstrated by autoradiography. Thus, it may be misleading and erroneous to consider the blood-brain barrier (BBB) to be a simple, single-membrane structure when analyzing the blood-brain transfer data of solutes such as amino acids. The data from the ventriculocisternal perfusion experiments and previously published AIB uptake data in mouse brain slices were used to estimate the transfer rate constants across brain cell membranes. These studies indicated that the transport of AIB into brain cells was approximately 110 to 265 times greater than that across normal brain capillaries per unit mass of brain tissue, and that the BBB limits blood-to-brain cell transport of this amino acid. These observations (low rate of transport across normal brain capillaries and rapid concentrative uptake by brain cells) indicate that AIB is a good marker for measuring moderate to large increases in BBB permeability by experiments that require unidirectional flux of the tracer.