Pharmacological evidence for the concept of spare glutamate transporters

Nascimento, Inês Belo do; Damblon, Jonathan; Ingelbrecht, Caroline; Goursaud, Stéphanie; Massart, Marion; Dumont, Amélie; Desmet, Nathalie; Hermans, Emmanuel

doi:10.1016/j.neuint.2021.105142

Cited by 3 publications

(3 citation statements)

References 41 publications

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“…One notable feature of our iGluSnFR decay measures was that genotype differences were typically only detected after a longer train of neural activity was evoked with 100 pulses, and not with 5 pulses. A possible explanation for this finding is suggested by the concept of spare glutamate transporters [ 70 ], and that the glutamate released by 5 pulses is insufficient to overwhelm the glutamate uptake system. In other words, 3xTg mice still have sufficient GLT-1 protein to efficiently clear glutamate released by short bursts of activity.…”

Section: Discussionmentioning

confidence: 99%

Asymmetric dysregulation of glutamate dynamics across the synaptic cleft in a mouse model of Alzheimer’s disease

Brymer

Hurley

Barron

et al. 2023

acta neuropathol commun

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Most research on glutamate spillover focuses on the deleterious consequences of postsynaptic glutamate receptor overactivation. However, two decades ago, it was noted that the glial coverage of hippocampal synapses is asymmetric: astrocytic coverage of postsynaptic sites exceeds coverage of presynaptic sites by a factor of four. The fundamental relevance of this glial asymmetry remains poorly understood. Here, we used the glutamate biosensor iGluSnFR, and restricted its expression to either CA3 or CA1 neurons to visualize glutamate dynamics at pre- and postsynaptic microenvironments, respectively. We demonstrate that inhibition of the primarily astrocytic glutamate transporter-1 (GLT-1) slows glutamate clearance to a greater extent at presynaptic compared to postsynaptic membranes. GLT-1 expression was reduced early in a mouse model of AD, resulting in slower glutamate clearance rates at presynaptic but not postsynaptic membranes that opposed presynaptic short-term plasticity. Overall, our data demonstrate that the presynapse is particularly vulnerable to GLT-1 dysfunction and may have implications for presynaptic impairments in a variety of brain diseases.

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Section: Discussionmentioning

confidence: 99%

Asymmetric dysregulation of glutamate dynamics across the synaptic cleft in a mouse model of Alzheimer’s disease

Brymer

Hurley

Barron

et al. 2023

acta neuropathol commun

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show abstract

“…Although these assays use different readouts, one explanation for this discrepancy might be the high competing concentrations of substrate in the cell swelling assay, which are 100–10,000 times higher than in radioligand uptake assays ( Fontana, 2018 ). Alternatively, dox-induced overexpression of EAATs could create a pool of “spare” transporters on the plasma membrane that effectively buffers the extracellular concentration of inhibitor, resulting in a rightward shift of the concentration-inhibition curve and a lower apparent inhibitory potency ( Belo do Nascimento et al, 2021 ). Although the transporter expression levels and substrate concentrations should be thoroughly considered in further assay development, our impedance-based assay showed a robust window for the detection of transporter inhibitors.…”

Section: Discussionmentioning

confidence: 99%

Impedance-Based Phenotypic Readout of Transporter Function: A Case for Glutamate Transporters

et al. 2022

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Excitatory amino acid transporters (EAAT/SLC1) mediate Na+-dependent uptake of extracellular glutamate and are potential drug targets for neurological disorders. Conventional methods to assess glutamate transport in vitro are based on radiolabels, fluorescent dyes or electrophysiology, which potentially compromise the cell’s physiology and are generally less suited for primary drug screens. Here, we describe a novel label-free method to assess human EAAT function in living cells, i.e., without the use of chemical modifications to the substrate or cellular environment. In adherent HEK293 cells overexpressing EAAT1, stimulation with glutamate or aspartate induced cell spreading, which was detected in real-time using an impedance-based biosensor. This change in cell morphology was prevented in the presence of the Na+/K+-ATPase inhibitor ouabain and EAAT inhibitors, which suggests the substrate-induced response was ion-dependent and transporter-specific. A mechanistic explanation for the phenotypic response was substantiated by actin cytoskeleton remodeling and changes in the intracellular levels of the osmolyte taurine, which suggests that the response involves cell swelling. In addition, substrate-induced cellular responses were observed for cells expressing other EAAT subtypes, as well as in a breast cancer cell line (MDA-MB-468) with endogenous EAAT1 expression. These findings allowed the development of a label-free high-throughput screening assay, which could be beneficial in early drug discovery for EAATs and holds potential for the study of other transport proteins that modulate cell shape.

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“…One notable feature of our iGluSnFR decay measures was that genotype differences were typically only detected after a longer train of neural activity was evoked with 100 pulses, and not with 5 pulses. The most likely explanation for this finding is suggested by the concept of spare glutamate transporters (Belo do Nascimento et al, 2021), and that the glutamate released by 5 pulses is insufficient to overwhelm the glutamate uptake system. In other words, 3xTg still have sufficient GLT-1 protein to efficiently clear glutamate released by short bursts of activity.…”

Section: Activity-dependent Slowing Of Glutamate Clearancementioning

confidence: 99%

Asymmetric dysregulation of glutamate dynamics across the synaptic cleft in a mouse model of Alzheimer disease

Brymer

Hurley

Barron

et al. 2022

Preprint

View full text Add to dashboard Cite

SummaryMost research on glutamate spillover focuses on the deleterious consequences of postsynaptic glutamate receptor overactivation. However, two decades ago, it was noted that the glial coverage of hippocampal synapses is asymmetric: astrocytic coverage of postsynaptic sites exceeds coverage of presynaptic sites by a factor of four. The fundamental relevance of this glial asymmetry remains poorly understood. Here, we used the glutamate biosensor iGluSnFR, and restricted its expression to either CA3 or CA1 neurons to visualize glutamate dynamics at pre- and postsynaptic microenvironments, respectively. We demonstrate that inhibition of the primarily astrocytic glutamate transporter-1 (GLT-1) slows glutamate clearance to a greater extent at presynaptic compared to postsynaptic membranes. GLT-1 expression was reduced early in a mouse model of AD, resulting in slower glutamate clearance rates at presynaptic but not postsynaptic membranes that opposed presynaptic short-term plasticity. Overall, our data demonstrate that the presynapse is particularly vulnerable to GLT-1 dysfunction and may have implications for presynaptic impairments in a variety of brain diseases.

show abstract

Pharmacological evidence for the concept of spare glutamate transporters

Cited by 3 publications

References 41 publications

Asymmetric dysregulation of glutamate dynamics across the synaptic cleft in a mouse model of Alzheimer’s disease

Asymmetric dysregulation of glutamate dynamics across the synaptic cleft in a mouse model of Alzheimer’s disease

Impedance-Based Phenotypic Readout of Transporter Function: A Case for Glutamate Transporters

Asymmetric dysregulation of glutamate dynamics across the synaptic cleft in a mouse model of Alzheimer disease

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