The many faces of the EAAT transporter family, or- the controllers of “OFF”

Overview

• Introduction
• Transport of glutamate and anions
• Distributions of the EAATs
• Products
• Literature

Introduction

Glutamate is the most important excitatory neurotransmitter in the brain. Many different ionotropic and metabotropic receptors mediate its stimulating signal to the neurons. But, in a switch, the “OFF” signal is as important as the “ON” signal. The family of excitatory amino acid transporters (EAATs) rapidly removes glutamate from the synaptic cleft and the surrounding extracellular space by recycling the glutamate back to the neurons and astrocytes, and so help to terminate the excitatory signal of glutamate. In this way, the EAATs manage:

• to maintain the temporal fidelity of the glutamatergic signal. By removing the glutamate from the synaptic cleft, they prevent excessive stimulation of the post-synaptic receptors after the initial signal has been sent, impairing the detection of the next signal that arrives.
• to maintain the spatial fidelity of the signal and prevent glutamate from “escaping” from the synapse into which it has been released and stimulating unintended extrasynaptic or synaptic glutamate receptors.
• to protect neurons against the toxic actions of excessive glutamate in the extracellular space. Exacerbated or prolonged activation of glutamate receptors starts a cascade of neurotoxicity that ultimately leads to the loss of neuronal function and cell death, called excitotoxicity.

Transport of glutamate and anions

EAATs work as symporters by transporting one K⁺ ion out of the cell and simultaneously taking up a glutamate neurotransmitter/molecule (aspartate can also be transported) and three Na⁺ ions as well as an H⁺ into the cell in exchange (Alleva et al., 2022), see Fig. 1 (adapted from Freidman et al., 2020). This transport is dependent on an electrochemical gradient of sodium ions, and is facilitated by homo- or heterotrimers of the EAATs at the cell membrane (Kovermann et al., 2022).
EAATs are not only glutamate transporters, but also anion channels (Cl⁺) that open in response to transitions within the glutamate transport cycle (Otis & Jahr, 1998).

Recently published data suggest that EAAT5, a retina specific member of the EAAT family, functions not only as a glutamate importer but also as a glutamate-gated Cl^--channel, particularly in cone photoreceptors (Lukasiewcz et al., 2021).
When glutamate is taken up into glial cells by the EAATs, it is converted to glutamine and subsequently transported back into the presynaptic neuron, converted back into glutamate, and taken up into synaptic vesicles by action of the VGLUTs. This process is named the glutamate–glutamine cycle (Andersen & Schousboe, 2023), see Figure 2.

Distribution of the EAATs

The EAAT family consists so far of 5 members, EAAT1 to EAAT5, all with a different glutamate uptake kinetic and a different degree of chloride permeability and distribution (Todd & Harding, 2020), see Table 1.

EAAT1:
EAAT1, also referred to as GLAST-1, is expressed throughout the CNS, and is highly expressed in astrocytes and Bergmann glia in the cerebellum. In the retina, EAAT1 is expressed in Müller cells.
Robust EAAT expression, in particular EAAT1, is a widely used marker of adult neural stem cell (NSC) phenotype (Rieskamp et al., 2023).
We offer several KO-validated antibodies for the detection of EAAT1.

EAAT2:

EAAT2, also named GLT-1, is the most abundant of the EAATs in the brain. It is predominantly localized in the astrocytic branches and is highly expressed in the cerebellum and hippocampus, respectively (Yeung et al., 2021). Recently, a reduction in EAAT2 has been found in several neurodegenerative diseases such as Alzheimer’s Disease, Multiple Sclerosis and Amyotrophic Lateral Sclerosis (ALS), reviewed in Dahlmanns et al., 2023. In this context, it is interesting to note that presenilin 1 (PS1), the active subunit of gamma-secretase, interacts directly with EAAT2 and influences the cell surface localization of the transporter (Perrin et al., 2024).
For the detection of EAAT2 we can provide excellent, KO-validated, polyclonal rabbit and guinea pig antibodies. And additionally a mouse monoclonal antibody that display superior results in WB, ICC, IHC and FFPE (IHC-P) applications.

EAAT3:

EAAT3, also known as EAAC1 or SLC1A1, is the “neuronal” glutamate transporter in the brain, mainly localized at the axonal terminal and the dendrites of the cerebral cortex, hippocampus, striatum, and basal ganglia (Escobar et al., 2019). The latest research results postulate a link between EAAT3 expression and obsessive-compulsive disorder (OCD), reviewed in Escobar et al., 2019. Outside of the CNS EAAT3 can be found in the lung, small intestine, skeletal muscle, renal outer medulla, medullary ray and cortex (Todd & Hardingham, 2020).
We offer 2 outstanding polyclonal rabbit antibodies in our portfolio.

EAAT4:

EAAT4, also referred to as SLC1A6, is a predominantly neuronal glutamate importer. It is localized primarily on Purkinje cells of the cerebellum, with some sparse expression in certain subregions of the forebrain and midbrain (Massie et al., 2008). Recent data showed that the expression of EAAT4 follows a parasagittal banding pattern similar to the expression of aldolase C (zebrin), creating microzones of molecularly diverse Purkinje cells with high and low levels of EAAT4 (Malhotra et al., 2021). The glutamate transport capacity of EAAT4 is rather low compared to EAAT1 and EAAT2, so that some researchers postulate that EAAT4 acts physiologically mainly as anion channel (Suslova et al., 2023).
New in our portfolio are now 2 excellent rabbit polyclonal antibodies against EAAT4.

EAAT5:

EAAT5 protein, sometimes also called AAAT, can be predominantly found in the vertebrate retina, where it transports (together with EAAT2) glutamate into rod cells, cone cells and rod bipolar cells. EAAT5 transporters are clustered just beneath synaptic ribbons in the presynaptic active zone of rods and cones perfectly positioned to capture recently released glutamate. Inhibition of EAAT5 has shown that this protein is important for the temporal signal resolution of glycinergic amacrine cells (AII amacrine cells) in the retina (Tang et al., 2022). Outside the CNS, EAAT5 is also expressed in liver, kidney, intestine, heart, lung and muscle. We offer a polyclonal guinea pig KO-validated antibody against EAAT5.

Products

Literature

Alleva et al., 2022: Molecular Basis of Coupled Transport and Anion Conduction in Excitatory Amino Acid Transporters. PMID: 33587237

Andersen & Schousboe, 2023: Glial Glutamine Homeostasis in Health and Disease. PMID: 36322369

Dahlmanns et al., 2023: Glial Glutamate Transporter-Mediated Plasticity: System xc-/xCT/SLC7A11 and EAAT1/2 in Brain Diseases. PMID: 37005761

Escobar et al., 2019: The Neuronal Glutamate Transporter EAAT3 in Obsessive-Compulsive Disorder. PMID: 31803055

Freidman et al., 2020: Amino Acid Transporters and Exchangers from the SLC1A Family: Structure, Mechanism and Roles in Physiology and Cancer. PMID: 31981058

Kovermann et al., 2022: Cellular Physiology and Pathophysiology of EAAT Anion Channels. PMID: 35087380

Lukasiewcz et al., 2021: EAAT5 Glutamate Transporter-Mediated Inhibition in the Vertebrate Retina. PMID: 34025361

Malhotra et al., 2021: Climbing Fiber-Mediated Spillover Transmission to Interneurons Is Regulated by EAAT4. PMID: 34400517

Massie et al., 2008: High-affinity Na+/K+-dependent glutamate transporter EAAT4 is expressed throughout the rat fore- and midbrain. PMID: 18770868

Otis & Jahr, 1998: Anion currents and predicted glutamate flux through a neuronal glutamate transporter. PMID: 9736633

Perrin et al., 2024: Identification of PS1/gamma-secretase and glutamate transporter GLT-1 interaction site. PMID: 38499151

Rieskamp et al., 2023: Excitatory amino acid transporter 1 supports adult hippocampal neural stem cell self-renewal. PMID: 37534178

Suslova et al., 2023: Apo state pore opening as functional basis of increased EAAT anion channel activity in episodic ataxia 6. PMID: 37538371

Tang et al., 2022: Glutamate Transporters EAAT2 and EAAT5 Differentially Shape Synaptic Transmission from Rod Bipolar Cell Terminals. PMID: 35523583

Todd & Hardingham, 2020: The Regulation of Astrocytic Glutamate Transporters in Health and Neurodegenerative Diseases. PMID: 33348528

Yeung et al., 2021: EAAT2 Expression in the Hippocampus, Subiculum, Entorhinal Cortex and Superior Temporal Gyrus in Alzheimer’s Disease. PMID: 34588956