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Cationic amino acid (CAA) transport is brought about by two families of proteins that are found in various tissues: Cat (CAA transporter), referred to as system [y.sup.+], and Bat [broad-scope amino acid (AA) transporter], which comprises systems [b.sup.0,+], [B.sup.0,+], and [y.sup.+]L. CAA traverse the blood-brain barrier (BBB), but experiments done in vivo have only been able to examine the BBB from the luminal (blood-facing) side. In the present study, plasma membranes isolated from bovine brain microvessels were used to identify and characterize the CAA transporter(s) on both sides of the BBB. From these studies, it was concluded that system [y.sup.+] was the only transporter present, with a prevalence of activity on the abluminal membrane. System [y.sup.+] was voltage dependent and had a [K.sub.m] of 470 [+ or -] 106 [micro]M (SE) for lysine, a [K.sub.i] of 34 [micro]M for arginine, and a [K.sub.i] of 290 [micro]M for omithine. In the presence of [Na.sup.+], system y + was inhibited by several essential neutral AAs. The [K.sub.i] values were 3-10 times the plasma concentrations, suggesting that system [y.sup.+] was not as important a point of access for these AAs as system L1. Several small nonessential AAs (serine, glutamine, alanine,and glycine) inhibited system [y.sup.+] with [K.sub.i] values similar to their plasma concentrations, suggesting that system [y.sup.+] may account for the permeability of the BBB to these AAs. System [y.sup.+] may be important in the provision of arginine for NO synthesis. Real-time PCR and Western blotting techniques established the presence of the three known nitric oxide synthases in cerebral endothelial cells: NOS-1 (neuronal), NOS-2 (inducible), and NOS-3 (endothelial). These results confirm that system [y.sup.+] is the only CAA transporter in the BBB and suggest that NO can be produced in brain endothelial cells. amino acid active transport; brain capillaries; endothelial cells; essential amino acids; nonessential amino acids; polarity doi:10.1152/ajpendo.00007.2006

Brain capillary endothelial cells form the blood-brain barrier (BBB). They are connected by extensive tight junctions, and are polarized into luminal (blood-facing) and abluminal (brain-facing) plasma membrane domains. The polar distribution of transport proteins mediates amino acid (AA) homeostasis in the brain. The existence of two facilitative transporters for neutral amino acids (NAAs) on both membranes provides the brain access to essential AAs. Four [Na.sup.+]-dependent transporters of NAA exist in the abluminal membranes of the BBB. Together these systems have the capability to actively transfer every naturally occurring NAA from the extracellular fluid (ECF) to endothelial cells and from there into circulation. The presence of [Na.sup.+]-dependent carriers on the abluminal membrane provides a mechanism by which NAA concentrations in the ECF of brain are maintained at ~10% those of the plasma. Also present on the abluminal membrane are at least three [Na.sup.+]-dependent systems transporting acidic AAs (EAAT) and a [Na.sup.+]-dependent system transporting glutamine (N). Facilitative carriers for glutamine and glutamate are found only in the luminal membrane of the BBB. This organization promotes the net removal of acidic- and nitrogen-rich AAs from the brain and accounts for the low level of glutamate penetration into the central nervous system. The presence of a [gamma]-glutamyl cycle at the luminal membrane and [Na.sup.+]-dependent AA transporters at the abluminal membrane may serve to modulate movement of AAs from blood to the brain. The [gamma]-glutamyl cycle is expected to generate pyroglutamate (synonymous with oxyproline) within the endothelial cells. Pyroglutamate stimulates secondary active AA transporters at the abluminal membrane, thereby reducing the net influx of AAs to the brain. It is now clear that BBB participates in the active regulation of the AA content of the brain. KEY WORDS: * brain capillaries * endothelial cells * polarity * sodium dependent transport * facilitative transport

[Na.sup.+] -dependent neutral amino acid transporters A, ASC, and N of the blood-brain barrier: mechanisms for neutral amino acid removal. Am J Physiol Endocrinol Metab :287: E622-E629, 2004. First published May 27, 2004; 10.1152/ajpendo.00187.2004.--Four [Na.sup.+]-dependent transporters of neutral amino acids (NAA) are known to exist in the abluminal membranes (brain side) of the blood-brain barrier (BBB). This article describes the kinetic characteristics of systems A, ASC, and N that, together with the recently described [Na.sup.+]-dependent system for large NAA ([Na.sup.+]-LNAA), provide a basis for understanding the functional organization of the BBB. The data demonstrate that system A is voltage dependent (3 positive charges accompany each molecule of substrate). Systems ASC and N are not voltage dependent. Each NAA is a putative substrate for at least one system, and several NAA are transported by as many as three. System A transports Pro, Ala, His, Ash, Ser, and Gin; system ASC transports Ser, Gly, Met, Val, Leu, Ile, Cys, and Thr; system N transports Gin, His, Set, and Asn; [Na.sup.+]-LNAA transports Leu, Ile, Val, Trp, Tyr, Phe, Met, Ala, His, Thr, and Gly. Together, these four systems have the capability to actively transfer every naturally occurring NAA from the extracellular fluid (ECF) to endothelial cells and thence to the circulation. The existence of facilitative transport for NAA (L1) on both membranes provides the brain access to essential NAA. The presence of [Na.sup.+]-dependent carriers on the abluminal membrane provides a mechanism by which NAA concentrations in the ECF of brain are maintained at ~10% of those of the plasma. active transport; brain extracellular fluid; capillaries; endothelial cells; essential amino acids