Excitatory amino acid transporter 5, a retinal glutamate transporter coupled to a chloride conductance

Abstract

Although a glutamate-gated chloride conductance with the properties of a sodium-dependent glutamate transporter has been described in vertebrate retinal photoreceptors and bipolar cells, the molecular species underlying this conductance has not yet been identified. We now report the cloning and functional characterization of a human excitatory amino acid transporter, EAAT5, expressed primarily in retina. Although EAAT5 shares the structural homologies of the EAAT gene family, one novel feature of the EAAT5 sequence is a carboxy-terminal motif identified previously in N-methyl-D-aspartate receptors and potassium channels and shown to confer interactions with a family of synaptic proteins that promote ion channel clustering. Functional properties of EAAT5 were examined in the Xenopus oocyte expression system by measuring radiolabeled glutamate f lux and twoelectrode voltage clamp recording. EAAT5-mediated Lglutamate uptake is sodium-and voltage-dependent and chloride-independent. Transporter currents elicited by glutamate are also sodium-and voltage-dependent, but ion substitution experiments suggest that this current is largely carried by chloride ions. These properties of EAAT5 are similar to the glutamate-elicited chloride conductances previously described in retinal neurons, suggesting that the EAAT5-associated chloride conductance may participate in visual processing.The uptake of glutamate and other excitatory amino acids is mediated by a gene family of high affinity sodium-dependent transporters that includes four known mammalian subtypes; in humans, we have termed these glutamate transporters excitatory amino acid transporter (EAAT) 1 through 4 (1, 2). The transport of glutamate is driven by the cotransport of sodium ions and the countertransport of potassium ions down their electrochemical gradients, and recent studies suggest that this complex process involves the cotransport of protons as well (3-6). Because there is net inward movement of positive charge with the transport of each molecule of glutamate, the transport process is readily studied in the Xenopus oocyte expression system by observing the associated current. In addition to these transport currents, however, we have found that application of substrate to the transporter also gates an uncoupled, passive flux of chloride ions (2, 7). The relative magnitude of this associated chloride conductance varies with each cloned EAAT subtype; for EAAT1-EAAT3, the magnitude of the chloride current at physiological membrane potentials is similar to that of the electrogenic cotransport current, but the currents generated by EAAT4 are almost entirely due to the flux of chloride ions. In vivo, a glutamatedependent current that has a transporter-like pharmacology is carried largely by chloride ions in retinal cone (8) and rod (9) photoreceptors and bipolar cells (10). In bipolar cells, this chloride current has been proposed to mediate the cone component of the ON bipolar cell light response (10). Although the properties of EAAT4 are similar to the glu...

Glutamate uptake in retinal glial cells during diabetes

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Glutamate transporters in platelets: EAAT1 decrease in aging and in Alzheimer’s disease

“…Their human counterparts have 95% homology in amino acid sequence and were respectively called EAAT1, EAAT2 and EAAT3 [2]. More recently, two new transporters, which appear linked to chloride fluxes, were cloned from human cerebellum [16] and retina [1]. The first three transporters are involved in high affinity, sodium-dependent glutamate uptake and knock-out models indicate a specific role of each transporter in preventing excitotoxicity and neuronal degeneration [46,54].…”

Inhibition of glutamate uptake by a polypeptide toxin (phoneutriatoxin 3-4) from the spider Phoneutria nigriventer

“…Several isoforms of amino acid transporters, named EAAT1 (GLAST), EAAT2 (GLT-1), EAAT3 (EAAC1), EAAT4 and EAAT5, have been identified in the central and peripheral nervous systems of different species [3][4][5][6][7][8]. Their function relies on the Na + gradient across the plasma membrane and on the countertransport of K + and OH − [1].…”