Standard Amblyopia Therapy in Adults with Longstanding Amblyopia Improves Visual Acuity and Contrast Sensitivity.

Purpose: Perceptual learning or dichoptic training may result in improved acuity in adult amblyopes. However, for amblyopic children (<18 years), most clinicians recommend standard part-time patching. The purpose of this study was to determine if standard amblyopia therapy results in an enhancement in vision in the amblyopic eye of adults. Patients and Methods: Fifteen amblyopes (20/30 or worse) were recruited and nine (age (SD) 32.9 (16.31)) with anisometropia or anisometropia and strabismus (ie, combined mechanism amblyopia) completed the study. Previous therapy did not exclude subjects. The subjects received a comprehensive eye exam and wore their best correction for at least four weeks prior to baseline testing. The non-amblyopic eye was patched for 2 hours per day (Amblyopia iNET training for 30 minutes and near/distance activities for 1.5 hours). The subjects had a baseline amblyopia evaluation followed by one visit per week for 12 weeks. At 12 weeks, the treatment was tapered off over one month and the subjects had a final amblyopia evaluation at 24 weeks. Contrast sensitivity was measured at baseline and 12 weeks with the Quick CSF system. Results: The subjects had a significant improvement in visual acuity across the weeks (p < 0.001). At baseline, weeks 12 and 24, the average logMAR acuities (SE) were 0.55 (0.09), 0.41 (0.08), and 0.38 (0.09), respectively. Weeks 4 to 24 were significantly different (p < 0.001) from baseline. The average acuity improvement over the 24 weeks was 1.7 logMAR lines. There was a significant increase in the area under the log contrast sensitivity function (p = 0.002) and its estimated acuity (p = 0.036) from baseline to 12 weeks. Conclusion: Standard amblyopia treatment can result in an improvement in visual acuity and contrast sensitivity in adults with longstanding anisometropic or combined mechanism amblyopia even if they had prior therapy.

Inhibitors of the Neutral Amino Acid Transporters ASCT1 and ASCT2 Are Effective in In Vivo Models of Schizophrenia and Visual Dysfunction.

The N-methyl-d-aspartate receptor coagonist d-serine is a substrate for the neutral amino acid transporters ASCT1 and ASCT2, which may regulate its extracellular levels in the central nervous system (CNS). We tested inhibitors of ASCT1 and ASCT2 for their effects in rodent models of schizophrenia and visual dysfunction, which had previously been shown to be responsive to d-serine. L-4-fluorophenylglycine (L-4FPG), L-4-hydroxyPG (L-4OHPG), and L-4-chloroPG (L-4ClPG) all showed high plasma bioavailability when administered systemically to rats and mice. L-4FPG showed good brain penetration with brain/plasma ratios of 0.7-1.4; however, values for L-4OHPG and L-4ClPG were lower. Systemically administered L-4FPG potently reduced amphetamine-induced hyperlocomotion in mice, whereas L-4OHPG was 100-fold less effective and L-4ClPG inactive at the doses tested. L-4FPG and L-4OHPG did not impair visual acuity in naive rats, and acute systemic administration of L-4FPG significantly improved the deficit in contrast sensitivity in blue light-treated rats caused by retinal degeneration. The ability of L-4FPG to penetrate the brain makes this compound a useful tool to further evaluate the function of ASCT1 and ASCT2 transporters in the CNS.

Phenylglycine analogs are inhibitors of the neutral amino acid transporters ASCT1 and ASCT2 and enhance NMDA receptor-mediated LTP in rat visual cortex slices.

The N-methyl-d-aspartate receptor (NMDA) co-agonist d-serine is a substrate for the neutral amino acid transporters ASCT1 (SLC1A4) and ASCT2 (SLC1A5). We identified l-phenylglycine (PG) and its analogs as inhibitors of ASCT1 and ASCT2. PG analogs were shown to be non-substrate inhibitors of ASCT1 and ASCT2 with a range of activities relative to other amino acid transport systems, including sodium-dependent glutamate transporters, the sodium-independent d-serine transporter asc-1 and system L. L-4-chloroPG was the most potent and selective ASCT1/2 inhibitor identified. The PG analogs facilitated theta-burst induced long-term potentiation in rat visual cortex slices in a manner that was dependent on extracellular d-serine. For structurally-related PG analogs, there was an excellent correlation between ASCT1/2 transport inhibition and enhancement of LTP which was not the case for inhibition of asc-1 or system L. The ability of PG analogs to enhance LTP is likely due to inhibition of d-serine transport by ASCT1/2, leading to elevated extracellular levels of d-serine and increased NMDA receptor activity. These results suggest that ASCT1/2 may play an important role in regulating extracellular d-serine and NMDA receptor-mediated physiological effects and that ASCT1/2 inhibitors have the potential for therapeutic benefit.

Restoration of visual performance by d-serine in models of inner and outer retinal dysfunction assessed using sweep VEP measurements in the conscious rat and rabbit.

The NMDA subtype of glutamate receptor and its co-agonist d-serine play a key role in synaptic function in the central nervous system (CNS), including visual cortex and retina. In retinal diseases such as glaucoma and macular degeneration, a loss of vision arises from malfunction of retinal cells, resulting in a glutamate hypofunctional state along the visual pathway in the affected parts of the visual field. An effective strategy to remedy this loss of function might be to increase extracellular levels of d-serine and thereby boost synaptic NMDA receptor-mediated visual transmission and/or plasticity to compensate for the impairment. We tested this idea in brain slices of visual cortex exhibiting long-term potentiation, and in rodent models of visual dysfunction caused by retinal insults at a time when the injury had stabilized to look for neuroenhancement effects. An essential aspect of the in vivo studies involved adapting sweep VEP technology to conscious rats and rabbits and combining it with intracortical recording while the animals were actively attending to visual information. Using this technology allowed us to establish complete contrast sensitivity function curves. We found that systemic d-serine dose-dependently rescued the contrast sensitivity impairment in rats with blue light-induced visual dysfunction. In rabbits with inner retinal dysfunction, both systemic and intravitreal routes of d-serine provided a rescue of visual function. In sum, we show that co-agonist stimulation of the NMDA receptor via administration of exogenous d-serine might be an effective therapeutic strategy to enhance visual performance and compensate for the loss of vision resulting from retinal disease.

D-Serine Is a Substrate for Neutral Amino Acid Transporters ASCT1/SLC1A4 and ASCT2/SLC1A5, and Is Transported by Both Subtypes in Rat Hippocampal Astrocyte Cultures.

N-methyl-D-aspartate (NMDA) receptors play critical roles in synaptic transmission and plasticity. Activation of NMDA receptors by synaptically released L-glutamate also requires occupancy of co-agonist binding sites in the tetrameric receptor by either glycine or D-serine. Although D-serine appears to be the predominant co-agonist at synaptic NMDA receptors, the transport mechanisms involved in D-serine homeostasis in brain are poorly understood. In this work we show that the SLC1 amino acid transporter family members SLC1A4 (ASCT1) and SLC1A5 (ASCT2) mediate homo- and hetero-exchange of D-serine with physiologically relevant kinetic parameters. In addition, the selectivity profile of D-serine uptake in cultured rat hippocampal astrocytes is consistent with uptake mediated by both ASCT1 and ASCT2. Together these data suggest that SLC1A4 (ASCT1) may represent an important route of Na-dependent D-serine flux in the brain that has the ability to regulate extracellular D-serine and thereby NMDA receptor activity.

Activity of the enantiomers of erythro-3-hydroxyaspartate at glutamate transporters and NMDA receptors.

The enantiomers of erythro-3-hydroxyaspartate were tested for activity at glutamate transporters and NMDA receptors. Both enantiomers inhibited glutamate transporters in rat hippocampal crude synaptosomes and elicited substrate-like activity at excitatory amino acid transporter 1, 2, and 3 as measured by voltage clamp in the Xenopus oocyte expression system. The enantiomers had similar affinities, but the D-enantiomer showed a lower maximal effect at excitatory amino acid transporter 1, 2, and 3 than the L-enantiomer. Surprisingly, D-erythro-3-hydroxyaspartate was a potent NMDA receptor agonist with an EC50 value in rat hippocampal neurons of 320 nM, whereas the L-enantiomer was 100-fold less potent. L-erythro-3-hydroxyaspartate showed activity at both glutamate transporters and NMDA receptors at concentrations that are reported to inhibit serine racemase, indicating a lack of selectivity. This enantiomeric pair may assist in shedding further light on the structural requirements for substrate activity at glutamate transporters and for agonist activity at NMDA receptors. The erythro enantiomers of 3-hydroxyaspartate had interesting and surprising effects on glutamate neurotransmitter systems. L-erythro-3-hydroxyaspartate had activity at both glutamate transporters (EAAT1/2/3) and NMDA receptors. D-erythro-3-hydroxyaspartate acted on EAATs, but was also identified as a highly potent NMDA receptor agonist. These enantiomers shed further light on the structural requirements for activity at EAATs and NMDA receptors.

Effects of 3-aminoglutarate, a "silent" false transmitter for glutamate neurons, on synaptic transmission and epileptiform activity.

Pharmacological tools that interact with the mechanisms that regulate vesicular filling and release of the neurotransmitter L-glutamate would be of enormous value. In this study, we provide physiological evidence that the glutamate analog, 3-aminoglutarate (3-AG), acts as a false transmitter to reduce presynaptic glutamate release. 3-AG inhibits glutamate-mediated neurotransmission both in primary neuronal cultures and in brain slices with more intact neural circuits. When assayed with the low affinity glutamate receptor antagonist γ-DGG, we demonstrate that 3-AG significantly reduces the synaptic cleft glutamate concentration, suggesting that 3-AG may act as a false transmitter to compete with glutamate during vesicle filling. Furthermore, using three different epileptic models (Mg(2+)-free, 4-AP, and high K(+)), we demonstrate that 3-AG is capable of suppressing epileptiform activity both before and after its induction. Our studies, along with those of the companion paper by Foster et al. (2015) indicate that 3-AG is a "silent" false transmitter for glutamate neurons that is a useful pharmacological tool to probe the mechanisms governing vesicular storage and release of glutamate under both physiological and pathophysiological conditions. 3-AG may have potential therapeutic value in conditions where the glutamate neurotransmitter system is pathologically overactive.

3-Aminoglutarate is a "silent" false transmitter for glutamate neurons.

Understanding the storage and release of the excitatory neurotransmitter, L-glutamate by synaptic vesicles has lagged behind receptor characterizations due to a lack of pharmacological agents. We report that the glutamate analog, 3-aminoglutarate (3-AG) is a "silent" false transmitter for glutamate neurons that may be a useful tool to study storage and release mechanisms. Like L-glutamate itself, 3-AG is a high-affinity substrate for both the plasma membrane (EAATs) and vesicular (vGLUT) glutamate transporters. As such, EAATs facilitate 3-AG entry into neuronal cytoplasm allowing 3-AG to compete with L-glutamate for transport into vesicles thus reducing glutamate content. In a synaptosomal preparation, 3-AG inhibited calcium-dependent endogenous L-glutamate release. Unlike L-glutamate, 3-AG had low affinity for both ionotropic (NMDA and AMPA) and G-protein coupled (mGlu1-8) receptors. Consequently, 3-AG behaves as a "silent" false transmitter that may be used in physiological experiments to probe synaptic vesicle storage and release mechanisms for L-glutamate. The companion paper by Wu et al. (2015) describes initial experiments that explore the effects of 3-AG on glutamate synaptic transmission under physiological and pathophysiological conditions.

GABA(B) receptor-mediated presynaptic inhibition has history-dependent effects on synaptic transmission during physiologically relevant spike trains.

Presynaptic inhibition is a form of neuromodulation that interacts with activity-dependent short-term plasticity so that the magnitude, and sometimes even the polarity, of that activity-dependent short-term plasticity is changed. However, the functional consequences of this interaction during physiologically relevant spike trains are poorly understood. We examined the effects of presynaptic inhibition on excitatory synaptic transmission during physiologically relevant spike trains, using the GABA(B) receptor (GABA(B)R) agonist baclofen to engage presynaptic inhibition and field EPSPs (fEPSPs) in hippocampal slices to monitor synaptic output. We examined the effects of baclofen on the relationship between an fEPSP during the spike train and the timing of spikes preceding that fEPSP, a relationship that we refer to as the history dependence of synaptic transmission. Baclofen alters this history dependence by causing no inhibition during short interspike intervals (ISIs) in the spike train but a maximal inhibition during long ISIs. This effect strengthens the dependence of the fEPSP on the first ISI preceding it. One consequence of this effect is that the apparent affinity of baclofen is strongly reduced during physiologically relevant spike trains when compared with conventional stimulus paradigms, and a second consequence is that the overall inhibition experienced by a synapse will vary considerably during repeated trials of a behavioral task. We conclude that GABA(B)R-mediated presynaptic inhibition is more accurately described as a high-pass filter than as a simple inhibition, and that this filtering must be taken into account to accurately assess the effects of presynaptic inhibition under physiologically relevant conditions.

Presynaptic kainate receptors play different physiological roles in mossy fiber and associational-commissural synapses in CA3 of hippocampus from adult rats.

Mossy fiber (MF) and Associational-commissural (Assoc-com) synaptic responses were recorded simultaneously in CA3 from the same hippocampal slice. Low concentrations of Kainate (KA) (50 and 100 nM) reversibly increased the synaptic response and decreased the paired-pulse facilitation (PPF) in the MF synapse, while reversibly decreasing the synaptic response and increasing the PPF in the Assoc-com synapse. The same concentration of KA had no effect on synaptic transmission or the induction and expression of long-term potentiation (LTP) in area CA1. MF LTP partially occluded the effect of KA on MF responses. These results suggest that KARs function as presynaptic autoreceptors in both MF and Assoc-com synapses but play different roles.