Neuronal Transporters Regulate Glutamate Clearance, NMDA Receptor Activation, and Synaptic Plasticity in the Hippocampus

Abstract

In the mammalian brain, the specificity of excitatory synaptic transmission depends on rapid diffusion of glutamate away from active synapses and the powerful uptake capacity of glutamate transporters in astrocytes. The extent to which neuronal glutamate transporters influence the lifetime of glutamate in the extracellular space remains unclear. Here we show that EAAC1, the predominant neuronal glutamate transporter at excitatory synapses in hippocampal area CA1, buffers glutamate released during synaptic events and prolongs the time course of its clearance by astrocytes. EAAC1 does not significantly alter activation of receptors in the synaptic cleft. Instead, it reduces recruitment of perisynaptic/extrasynaptic NR2B-containing NMDARs, thereby facilitating induction of long-term potentiation by short burstsofhigh-frequencystimulation.WedescribenovelrolesofEAAC1inregulatingglutamatediffusionandproposethatNMDARsatdifferent subsynaptic locations can make distinct contributions to the regulation of synaptic strength.

Deriving the Time Course of Glutamate Clearance with a Deconvolution Analysis of Astrocytic Transporter Currents

“…The kinetics of the fSTC are similar to the kinetics of the synaptically-activated transporter current (STCs) isolated pharmacologically with high concentrations of the glutamate transporter antagonist TBOA (100 μM) 13 . For this reason and for simplicity, in previous work, we referred to the fSTC as STC 13,14 . Despite being currents with similar properties, fSTCs and STCs are not the same current.…”

“…Because the clearance waveform rises from and decays back to zero, when the method was first established, there was no specification made on how to set the integration window, and in fact this was done rather loosely 13 . One way around this issue is to limit the integration window to the time it takes to reach an arbitrary proportion of the FTC/fSTC peak, for example 10% 14 . This simple criterion allows being consistent when performing this analysis in different cells, improves the accuracy in the estimates of <t> when the clearance waveform does not decay perfectly to zero, and improves the sensitivity of the analysis to small differences in clearance waveforms that might occur in different groups of cells.…”

“…Both currents (stoichiometric and non-stoichiometric) can be recorded by obtaining whole-cell patch-clamp recordings from astrocytes, visually identified under Dodt illumination or infra-red differential interference contrast (IR-DIC) in acute brain slices 12 . The stoichiometric component of the current associated with glutamate transport across the membrane can be isolated by using CH 3 SO 3 -, or C 6 H 11 O 7 -based intracellular solutions and can be evoked by flash-uncaging glutamate on astrocytes 13,14 , or by activating glutamate release from neighboring synapses, either electrically 12 or with a targeted optogenetic control.…”

Dual Electrophysiological Recordings of Synaptically-evoked Astroglial and Neuronal Responses in Acute Hippocampal Slices

“…This is due to the filtering of GLT currents by factors such as the electrotonic properties of astrocytes or the asynchronous transmitter release, which distort their kinetics 13 . Methods extracting the temporal features of the filtering mechanisms have been developed and can be used to derive the actual glutamate clearance time course in physiological or pathological situations, as recenly performed 6,13,14 . Additionally, the simultaneous recording of the astroglial membrane depolarization, in current clamp, can provide insights into possible alterations of extracellular potassium transients.…”

EAAC1 Gene Deletion Alters Zinc Homeostasis and Exacerbates Neuronal Injury after Transient Cerebral Ischemia

“…However, several studies have shown that EAAC1 has a negligible effect on glutamate clearance from brain extracellular space and that this function is performed primarily by the astrocyte glutamate transporters GLT1 and GLAST (glutamate-aspartate transporter) (Tanaka et al, 1997;Watase et al, 1998). Recent studies indicate a role for EAAC1 in regulating synaptic glutamate uptake (Scimemi et al, 2009); however, EAAC1 also functions as a highaffinity, concentrative cysteine transporter (Zerangue and Kavanaugh, 1996;Chen and Swanson, 2003;Himi et al, 2003;Watabe et al, 2008), and it is diffusely distributed over nonsynaptic neuronal membranes (Coco et al, 1997;Shashidharan et al, 1997). EAAC1 uptake of cysteine into neurons provides cysteine substrate for the synthesis of glutathione, the major thiol antioxidant Li and Maret, 2009).…”