Fast glutamate uptake via EAAT2 shapes the cone-mediated light offset response in bipolar cells

Abstract

Excitatory amino acid transporters (EAATs) are responsible for extracellular glutamate uptake within the retina, and are expressed by retinal neurons and Müller cells. Their role within glutamatergic synapses is not completely understood. In the salamander retina, five distinct EAAT-encoding genes have been cloned, making the amphibian retina an excellent system to study EAAT function. This study focused on sEAAT2, which is expressed in photoreceptor terminals and Off-bipolar cells in two isoforms, sEAAT2A and sEAAT2B. Using whole-cell patch-clamp recording, florescence imaging and antibody labelling methods, we systematically studied the functions of these two isoforms at the synapse between photoreceptors and bipolar cells, both in dark and with photic stimulation. Both sEAAT2A and sEAAT2B were sensitive to dihydrokainic acid (DHKA), a known EAAT2-specific inhibitor. Each isoform of sEAAT2 was found to play a role in tonic glutamate uptake at the cone synapse in darkness. Furthermore, presynaptic sEAAT2A strongly suppressed the rapid, transient glutamate signal from cones following light-offset. This was achieved by quickly binding exocytosed glutamate, which subsequently limited glutamate spillover to adjacent receptors at postsynaptic sites. Since the intensity and duration of photic stimulation determine the magnitude of these cone transient signals, we postulate that presynaptic cone EAATs contribute to the encoding of contrast sensitivity in cone vision. Glutamate is the main excitatory neurotransmitter in the retina and throughout the CNS, where its release from nerve terminals is essential for relaying signals between nerve cells. However, if allowed to accumulate in the synaptic cleft, it can result in prolonged activation of target cells, depletion within glutamatergic neurons, and neurotoxicity. These consequences are precluded by the presence of excitatory amino acid transporters (EAATs), which are responsible for the uptake of this amino acid into the neurons from which it is discharged. Using multiple techniques, we studied the functions of EAAT2, a subtype of glutamate transporter, in the synapse between photoreceptors and bipolar cells of the salamander retina. We found that presynaptic EAAT2A rapidly bound exocytosed glutamate, thereby limiting glutamate spillover to adjacent receptors at postsynaptic sites, and suppressing the rapid, transient glutamate signal from cones following light termination. Thus, EAAT2 preserves the cone signalling pathway in the distal retina. © 2010 The Authors. Journal compilation © 2010 The Physiological Society.