Pharmacological evidence for the concept of spare glutamate transporters

• Published in: Neurochemistry international 2021-10, Vol.149, p.105142-105142, Article 105142
• Main Authors: Belo do Nascimento, Inês, Damblon, Jonathan, Ingelbrecht, Caroline, Goursaud, Stéphanie, Massart, Marion, Dumont, Amélie, Desmet, Nathalie, Hermans, Emmanuel
• Format: Article
• Language: English
• Subjects: Animals; Astrocytes; Brain - drug effects; Brain - metabolism; Cells, Cultured; Dose-Response Relationship, Drug; Doxycycline - pharmacology; Excitatory Amino Acid Antagonists - pharmacology; Excitatory Amino Acid Transporter 2 - antagonists & inhibitors; Excitatory Amino Acid Transporter 2 - metabolism; Excitotoxicity; Glial cells; GLT-1; Glutamate uptake; Glutamine - metabolism; HEK293 Cells; Humans; Neurons - drug effects; Neurons - metabolism; Rats; Rats, Sprague-Dawley; Transporter reserve
• ISSN: 0197-0186; 1872-9754
• DOI: 10.1016/j.neuint.2021.105142

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. [Display omitted] •An inducible promoter system was used to express GLT-1 in transfected HEK293 cells.•Glutamate uptake capacity is not strictly dependent on GLT-1 density.•The potency of the specific blocker WAY-213613 depends on the density of GLT-1.•WAY-213613 potency is higher in brain synaptosomes compared to cultured astrocytes.•Functional studies suggest the existence of glutamate transporter reserve.