Pharmacological evidence for the concept of spare glutamate transporters

Through the efficient clearance of extracellular glutamate, high affinity astrocytic glutamate transporters constantly shape excitatory neurotransmission in terms of duration and spreading. Even though the glutamate transporter GLT-1 (also known as EAAT2/SLC1A2) is amongst the most abundant proteins in the mammalian brain, its density and activity are tightly regulated. In order to study the influence of changes in the expression of GLT-1 on glutamate uptake capacity, we have developed a model in HEK cells where the density of the transporter can be manipulated thanks to a tetracycline-inducible promoter. Exposing the cells to doxycycline concentration-dependently increased GLT-1 expression and substrate uptake velocity. However, beyond a certain level of induction, increasing the density of transporters at the cell surface failed to increase the maximal uptake. This suggested the progressive generation of a pool of spare transporters, a hypothesis that was further validated using the selective GLT-1 blocker WAY-213613 of which potency was influenced by the density of the transporters. The curve showing inhibition of uptake by increasing concentrations of WAY-213613 was indeed progressively rightward shifted when tested in cells where the transporter density was robustly induced. As largely documented in the context of cell-surface receptors, the existence of ‘spare’ glutamate transporters in the nervous tissue and particularly in astrocytes could impact on the consequences of physiological or pathological regulation of these transporters.