The astroglial ASCT2 amino acid transporter as a mediator of glutamine efflux

Glutamine release from astrocytes is an essential part of the glutamate-glu tamine cycle in the brain. Uptake of glutamine into cultured rat astrocytes occurs by at least four different routes. In agreement with earlier studie s, a significant contribution of amino acid transport systems ASC, A, L, an d N was detected. It has not been determined whether these systems are also involved in glutamine efflux or whether specific efflux transporters exist . We show here that ASCT2, a variant of transport system ASC, is strongly e xpressed in rat astroglia-rich primary cultures but not in neuron-rich prim ary cultures. The amino acid sequence of rat astroglial ASCT2 is 83% identi cal to that of mouse ASCT2. In Xenopus laevis oocytes expressing rat ASCT2, we observed high-affinity uptake of [U-C-14]glutamine (K-m =70 mu M) that was Na+-dependent, concentrative, and unaffected by membrane depolarization . When oocytes were preloaded with [U-C-14]glutamine, no glutamine efflux w as detected in the absence of extracellular amino acids. Neither lowering i ntracellular pH nor raising the temperature elicited efflux. However, addit ion of 0.1 mM unlabeled alanine, serine, cysteine, threonine, glutamine, or leucine to the extracellular solution resulted in a rapid release of gluta mine from the ASCT2-expressing oocytes. Amino acids that are not recognized as substrates by ASCT2 were ineffective in this role. Extracellular glutam ate stimulated glutamine release weakly at pH 7.5 but was more effective on lowering pH to 5.5, consistent with the pH dependence of ASCT2 affinity fo r glutamate. Our findings suggest a significant role of ASCT2 in glutamine efflux from astrocytes by obligatory exchange with extracellular amino acid s. However, the relative contribution of this pathway to glutamine release from cells in vivo or in vitro remains to be determined.