Brain capillaries isolated from 2-month-old male and female Sprague-Dawley rats were used to study the transport of neutral amino acids. The uptake of alanine, leucine, and alpha-methylaminoisobutyric acid (MeAIB) was a linear function of time for the first minute of incubation. Based on these observations, an incubation time of 1 min was used to measure transport activities. The intracellular water volume of the isolated capillaries was 2.2 μl/mg protein. This value was significantly lower (1.8 μl/mg protein) when measured in the absence of sodium. L-Alanine, L-serine, and L-cysteine were taken up from the abluminal surface of brain capillaries by a sodium- and energy-dependent, carrier-mediated system. This uptake, for the most part, was not inhibited by MeAIB. System ASC (alanine-serine-cysteine) appeared to be of primary importance for the transport of these amino acids in isolated brain capillaries. The apparent Km and Vmax values for L-alanine uptake by ASC transport, based on the Hofstee plot presentation, were 1.3 mM and 0.975 nmol/μl water content/min, respectively. The results also indicate that the transport of MeAIB and 2-aminobicyclo(2,2,1)heptane-2-carboxylic acid (BCH) was limited to the sodium-dependent system A (alanine) and the sodium-independent system L (leucine), respectively.
AudusKLBorchardtRT, (1986) Characteristics of the large neutral amino acid transport system of bovine brain microvessel endothelial cell monolayers. J Neurochem40:484–488.
2.
BetzALGilboeDDDrewesLR, (1975) Kinetics of unidirectional leucine transport into brain: Effects of isoleucine, valine, and anoxia. Am J Physiol228:895–900.
3.
BetzALGoldsteinGW, (1978) Polarity of the blood–brain barrier: Neutral amino acid transport into isolated brain capillaries. Science202:225–227.
4.
BetzALCsejteyJGoldsteinGW, (1979) Hexose transport and phosphorylation by capillaries isolated from rat brain. Am J Physiol236:C96–C102.
5.
BetzALFirthJGoldsteinGW, (1980) Polarity of the blood–brain barrier: Distribution of enzymes between the luminal and antiluminal membranes of brain capillary endothelial cells. Brain Res192:17–28.
6.
CancillaPADeBaultLE, (1983) Neutral amino acid transport properties of cerebral endothelial cells in vitro. J Neuropath Exp Neurol42:191–199.
7.
Cardelli-CangianoPCangianoCJamesJHJeppssonBBrennerWFisherJE, (1981) Uptake of amino acids by brain microvessels isolated from rats after portal caval anastomosis. J Neurochem36:627–632.
8.
ChristensenHNLiangMArcherEG, (1967) A distinct Na-requiring transport system for alanine, serine, cysteine, and similar amino acids. J Biol Chem242:5237–5246.
9.
ChristensenHN, (1975) Biological Transport, ed 2. Reading, MA, W. A. Benjamin
10.
ChristensenHN, (1979) Exploiting amino acid structure to learn about membrane transport. Adv Enzymol49:41–101.
11.
CohenSR, (1985) Rate equations and kinetics of uptake of alpha-aminoisobutyric acid and gamma-aminobutyric acid by mouse cerebrum slices incubated in media containing L(+)-lactate or a mixture of succinate, L-malate, and pyruvate as the energy source. J Neurochem44:455–464.
12.
Gazzola-FranchiRGazzolaGCDall'astVGuidottiGG, (1982) The transport of alanine, serine, and cysteine in cultured human fibroblasts. J Biol Chem257:9582–9587.
13.
GoldsteinGWWolinskyJSCsejteyJDiamondI, (1975) Isolation of metabolically active capillaries from rat brain. J Neurochem25:715–717.
14.
GuidottiGGBorghettiAFGazzolaGC, (1978) The regulation of amino acid transport in animal cells. Biochim Biophys Acta515:329–366.
15.
HawkinsRA, (1986) Transport of essential nutrients across the blood–brain barrier of individual structures. Fed Proc45:2055–2059.
16.
HjelleJTBaird-LambertJCardinaleGSpectorSUdenfriendS, (1978) Isolated microvessels: The blood–brain barrier in vitro. Proc Natl Acad Sci USA75:4544–4548.
17.
HwangSMWeissSSegalS, (1980) Uptake of L-(35S)cysteine by isolated rat brain capillaries. J Neurochem35:417–424.
18.
HwangSMMillerMSegalS, (1983) Uptake of L-(14C)proline by isolated rat brain capillaries. J Neurochem40:317–323.
19.
KletzienRFParizaMWBeckerJEPotterVR, (1975) A method using 3-o-methyl-D-glucose and phloretin for the determination of intracellular water space of cells in monolayer culture. Analyt Biochem68:537–544.
20.
LefauconnierJMTrouvéR, (1983) Developmental changes in the pattern of amino acid transport at the blood–brain barrier in rats. Dev Brain Res6:175–182.
21.
LefauconnierJMTayaraniIRouxF, (1985a) System ASC for amino acid transport in isolated rat brain capillaries. J Neurochem44:S66.
22.
LefauconnierJMTayaraniIRouxF, (1985b) Delineation of transport systems for some neutral amino acids in isolated rat brain microvessels. J Cereb Blood Flow Metabol5(suppl 1):S91–S92.
23.
LowryOHRosebroughNJFarrACRandallRJ, (1951) Protein measurement with the Folin phenol reagent. J Biol Chem193:265–275.
24.
MannGETradattiCEYudilevichDL, (1982) Unidirectional uptake of L-alanine and L-serine by system ASC in the baso-lateral membrane of cat salivary epithelium. Selective inhibition of rapid alanine metabolism by aminooxyacetate. Biochim Biophys Acta692:157–160.
25.
NewtonRBSullivanJLDebuskAG, (1984) Neutral amino acid transport and in vitro aging. Mech Aging Dev27:63–72.
26.
OldendorfWH, (1971) Brain uptake of radiolabeled amino acids, amines and hexoses after arterial injection. Am J Physiol221:1629–1639.
27.
OxenderDLChristensenHN, (1963) Distinct mediating systems for the transport of neutral amino acids by the Ehrlich cell. J Biol Chem238:3686–3699.
28.
PardridgeWM, (1977) Kinetics of competitive inhibition of neutral amino acid transport across the blood–brain barrier. J Neurochem28:103–108.
29.
PardridgeWMOldendorfWH, (1977) Transport of metabolic substrates through the blood–brain barrier. J Neurochem28:5–12.
30.
SershenHLajthaA, (1976) Capillary transport of amino acids in the developing brain. Exp Neurol53:465–474.
31.
SershenHLajthaA, (1979) Inhibition pattern by analogs indicates the presence of ten or more transport systems for amino acids in brain cells. J Neurochem32:719–726.
32.
ShotwellMAJaymeDWKilbergMSOxenderDL, (1981) Neutral amino acid transport systems in Chinese Hamster ovary cells. J Biol Chem256:5422–5427.
33.
WadeLABradyHM, (1981) Cysteine and cystine transport at the blood–brain barrier. J Neurochem37:730–734.
34.
WondergemRHarderDR, (1980) Transmembrane potential and amino acid transport in rat hepatocytes in primary monolayer culture. J Cell Physiol104:53–60.
35.
YudilevichDLDeRoseNSepulvedaFV, (1972) Facilitated transport of amino acids through the blood–brain carrier of the dog studied in a single capillary circulation. Brain Res44:569–578.
