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J Biol Chem, Vol. 274, Issue 45, 31891-31895, November 5, 1999
,,
From the Na+-dependent
transporters for glutamate exist on astrocytes (EAAT1 and EAAT2) and
neurons (EAAT3). These transporters presumably assist in keeping the
glutamate concentration low in the extracellular fluid of brain.
Recently, Na+-dependent glutamate transport was
described on the abluminal membrane of the blood-brain barrier. To
determine whether the above-mentioned transporters participate in
glutamate transport of the blood-brain barrier, total RNA was extracted
from bovine cerebral capillaries. cDNA for EAAT1, EAAT2, and EAAT3
was observed, indicating that mRNA was present. Western blot
analysis demonstrated all three transporters were expressed on
abluminal membranes, but none was detectable on luminal membranes of
the blood-brain barrier. Measurement of transport kinetics demonstrated
voltage dependence, K+-dependence, and an apparent
Km of 14 µM (aggregate of the three
transporters) at a transmembrane potential of Department of Physiology and Biophysics,
Finch University of Health Science, The Chicago Medical School,
North Chicago, Illinois 60064-3095 and § Departamento de
Bioquímica and Biología Molecular Facultades de
Medicina y Farmacia, Universitat de Valencia,
Valencia 46010 Spain
61 mV. Inhibition of
glutamate transport was observed using inhibitors specific for EAAT2
(kainic acid and dihydrokainic acid) and EAAT3 (cysteine). The relative
activity of the three transporters was found to be approximately 1:3:6
for EAAT1, EAAT2, and EAAT3, respectively. These transporters may
assist in maintaining low glutamate concentrations in the extracellular fluid.
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