Na+-dependent glutamate transporters (EAAT1, EAAT2, and EAAT3) of the blood-brain barrier - A mechanism for glutamate removal

Na+-dependent transporters for glutamate exist on astrocytes (EAAT1 and EAA T2) and neurons (EAAT3), These transporters presumably assist in keeping th e 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 trans porters participate in glutamate transport of the blood brain barrier, tota l RNA was extracted from bovine cerebral capillaries. cDNA for EAAT1, EAAT2 , and EAAT3 was observed, indicating that mRNA was present. Western blot an alysis demonstrated all three transporters were expressed on abluminal memb ranes, but none was detectable on luminal membranes of the blood brain barr ier. Measurement of transport kinetics demonstrated voltage dependence, K+- dependence, and an apparent K-m of 14 mu M (aggregate of the three transpor ters) at a transmembrane potential of -61 mV. Inhibition of glutamate trans port was observed using inhibitors specific for EAAT2 (kainic acid and dihy drokainic acid) and EAAT3 (cysteine). The relative activity of the three tr ansporters was found to be approximately 1:3:6 for EAAT1, EAAT2, and EAAT3, respectively. These transporters may assist in maintaining low glutamate c oncentrations in the extracellular fluid.