D-Serine as a glial modulator of nerve cells

Until the last decade, it was widely accepted that D-amino acids had no functional role in higher organisms, but that they were restricted to lower organisms, such as bacteria, where they are integrated into the proteoglycans of the cell wall. However, D-serine proved to be an effective coagonist at the "glycine-binding" site of the N-methyl-D-aspartate (NMDA) glutamate receptors, and this observation led to chemical analyses that have now revealed the presence of high levels of D-serine in the central nervous system, including many regions of the brain and retina. D-Serine has been localized to astrocytes and can be released by glutamate through stimulation of AMPA receptors. A new enzyme, serine racemase has been localized to glial cells and converts L-serine to D-serine. Degradation of D-serine takes place through D-amino acid oxidase, an enzyme once thought to metabolize D-amino acids from external sources. Although the "glycine-binding" site of NMDA receptors was initially regarded as a saturated site, evidence in many brain regions has established that this site is not saturated and is therefore modulated by interactions between glial cells and neurons. In some, but not all, studies, D-serine enhances NMDA-mediated currents; a light-evoked enhancement to NMDA currents has been reported in the retina. D-serine also plays a role in synaptic and cellular development, particularly in the cerebellum, where the normal developmental sequences underlying synapse formation onto Purkinje cells and the migration of granule cells are dependent on NMDA receptors during a time when high levels of D-serine are expressed in the Bergmann glia and other cerebellar astrocytes. D-serine must be added to the list of agents through which glial cells participate in controlling the excitability of neurons.     

Extrasynaptic GABA and glutamate receptors in epilepsy

Epilepsy is a neurological disorder in which normal brain function is disrupted as a consequence of intensive and synchronous burst activity from neuron assemblies. Epilepsies result from long-lasting plastic changes in the brain affecting neurotransmitter release, the properties of receptors and channels, synaptic reorganization and astrocyte activity. There is considerable evidence for alterations in glutamatergic and GABAergic synaptic transmission in the origin of the paroxysmal depolarization shifts that initiate epileptic activity. However, recent studies on non-synaptic transmission, receptor mobility and glia-neuron signaling pathways suggest that extrasynaptic GABA and glutamate receptors may play an important role in seizure initiation, maintenance and arrest. Extracellular aminoacids such as glutamate, aspartate, glycine and GABA seem to communicate neurons and glial cells acting primarily on extrasynaptic receptors. Synaptic and extrasynaptic glutamate and GABA receptors have been show to play different roles in neuronal excitability. NMDA and GABAA receptors expressed in a single neuron can be differentially regulated based on subcellular localization, and it has been proposed that distinct regulation of synaptic versus extrasynaptic receptors provides a mechanism for receptor adaptation in response to a variety of stimuli. Furthermore, glutamate and GABA receptors are highly mobile, and the number and composition of extrasynaptic receptors can be modulated by several factors. This review addresses recent advances in our understanding of the role of extrasynaptic receptors in epilepsy, suggesting that extrasynaptic receptors and their mechanisms of regulation are expected to be important pharmacological targets.     

Glutamate-mediated glial injury: mechanisms and clinical importance

Primary and/or secondary glial cell death can cause and/or aggravate human diseases of the central nervous system (CNS). Like neurons, glial cells are vulnerable to glutamate insults. Astrocytes, microglia, and oligodendrocytes express a wide variety of glutamate receptors and transporters that mediate many of the deleterious effects of glutamate. Astrocytes are responsible for most glutamate uptake in synaptic and nonsynaptic areas and consequently, are the major regulators of glutamate homeostasis. Microglia in turn may secrete cytokines, which can impair glutamate uptake and reduce the expression of glutamate transporters. Finally, oligodendrocytes, the myelinating cells of the CNS, are very sensitive to excessive glutamate signaling, which can lead to the apoptosis or necrosis of these cells. This review aims at summarizing the mechanisms leading to glial cell death as a consequence of alterations in glutamate signaling, and their clinical relevance. A thorough understanding of these events will undoubtedly lead to better therapeutic strategies to treat CNS diseases affecting glia and in particular, those that involve damage to white matter tracts.     

The role of glutamate and its receptors in the proliferation,migration, differentiation and survival of neural progenitor cells

The mammalian central nervous system derives from multipotent neural progenitor cells (NPCs) of the developing brain. During development the progenitor cells have enormous potential. They proliferate actively and differentiate into all the three main cell types, i.e., neurons, astrocytes and oligodendrocytes, of the adult brain through a tightly regulated process that coordinates cell proliferation, survival, migration, differentiation and apoptosis. This process is regulated by multiple extracellular signals including neurotrophic factors, chemoattractants and neurotransmitters in a coordinated manner. The main excitatory neurotransmitter glutamate is involved in promoting and/or inhibiting the proliferation, survival, migration and differentiation of NPCs acting via ionotropic or metabotropic receptors. The role of glutamate in the regulation of cortical NPCs has been most extensively studied. Glutamate appears to have a similar role in hippocampal, striatal as well as adult neural progenitors. Ionotropic α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)/kainate (KA) receptors and metabotropic glutamate receptor 5 (mGluR5) are expressed early during embryonic development as well as in the neurogenic zones of the adult brain. Ca²⁺-permeable AMPA/KA receptors are initially of importance for cell proliferation and neuronal motility. At later stages of development N-methyl-D-aspartate (NMDA) receptors have a more prominent role. MGluR5, which is the main metabotropic glutamate receptor during early development, is expressed in early progenitors and radial glial cells. Activation of this receptor promotes the proliferation and survival of NPCs. MGluR5 is involved in the extension of radial glial processes and in regulation of the migration of early cortical neurons.     

The role of glutamate and its receptors in multiple sclerosis

Glutamate is an excitatory neurotransmitter of the central nervous system, which has a central role in a complex communication network established between neurons, astrocytes, oligodendrocytes, and microglia. Multiple abnormal triggers such as energy deficiency, oxidative stress, mitochondrial dysfunction, and calcium overload can lead to abnormalities in glutamate signaling. Thus, the disturbance of glutamate homeostasis could affect practically all physiological functions and interactions of brain cells, leading to excitotoxicity. Excitotoxicity is the pathological process by which nerve cells are damaged or killed by excessive stimulation by glutamate. Although neuron degeneration and death are the ultimate consequences of multiple sclerosis (MS), it is now widely accepted that alterations in the function of surrounding glial cells are key features in the progression of the disease. The present knowledge raise the possibility that the modulation of glutamate release and transport, as well as receptors blockade or glutamate metabolism modulation, might be relevant targets for the development of future therapeutic interventions in MS.     

Glycine- and GABA-activated Currents in Identified Glial Cells of the Developing Rat Spinal Cord Slice

In the neonatal rat spinal cord, four types of glial cells, namely astrocytes, oligodendrocytes and two types of precursor cells, can be distinguished based on their membrane current patterns and distinct morphological features. In the present study, we demonstrate that these cells respond to the inhibitory neurotransmitters glycine and GABA, as revealed with the whole-cell recording configuration of the patch-clamp technique. All astrocytes and glial precursor cells and a subpopulation of oligodendrocytes responded to glycine. The involvement of glycine receptors was inferred from the observation that the response was blocked by strychnine and that the induced current reversed close to the Cl^- equilibrium potential. GABA induced large membrane currents in astrocytes and precursor cells while oligodendrocytes showed only small responses. The GABA-activated current was due to the activation of GABA_A receptors since muscimol mimicked and bicuculline blocked the response; moreover, the reversal potential was close to the Cl^- equilibrium potential. Besides the increase in a Cl^- conductance, GABA^A receptor activation also induced a block of the resting K⁺ conductance, as observed previously in Bergmann glial cells. Our experiments show that while glial GABA_A receptors are found in many brain regions and the spinal cord, glial glycine receptors have so far been detected only in the spinal cord. The restricted coexpression of glial and neuronal glycine receptors in a defined central nervous system grey matter area implies that such glial receptors may be involved in synaptic transmission.     

Targeting the NMDA receptor subunit NR2B for the treatment of neuropathic pain

Neuropathic pain is generally defined as a chronic pain state resulting from peripheral or central nerve injury, or both. An effective treatment for neuropathic pain is still lacking. The NMDA receptor, one type of the ionotropic glutamate receptors, is known to be important for triggering long-lasting changes in synapses. NMDA receptor-dependent synaptic plasticity plays roles not only in physiological functions such as learning and memory, but also in unwanted pathological conditions such as chronic pain. This review addresses recent progress on NMDA receptors in neuropathic pain, with particular emphasis on the NR2B-subunit-containing receptors. The expression and function of NMDA receptors in synaptic plasticity in the pain transmission pathway from dorsal root ganglia to the anterior cingulate cortex is reviewed, and preclinical and clinical investigations of selective NMDA receptor in neuropathic pain are discussed. The NMDA receptors, in particular NR2B-containing NMDA receptors, serve as promising targets for treatment of neuropathic pain.     

On the hypes and falls in neuroprotection: targeting the NMDA receptor.

Activation of the NMDA (N-methyl-D-aspartate) responsive subclass of glutamate receptors is an important mechanism of excitatory synaptic transmission. Moreover, NMDA receptors are widely involved in many forms of synaptic plasticity such as long-term potentiation (LTP) and long-term depression (LTD), which are thought to underlie complex tasks, including learning and memory. Dysfunction of these ligand-gated cation channels has been identified as an underlying molecular mechanism in neurological disorders ranging from acute stroke to chronic neurodegeneration in amyotrophic lateral sclerosis. Excessive glutamate levels have been detected following brain trauma and cerebral ischemia, resulting in an unregulated stimulation of NMDA receptors. These conditions are thought to elicit a cascade of excitation-mediated neuronal damage where massive increases in intracellular calcium concentrations finally trigger neuronal damage and apoptosis. Consistent with the hypothesis of NMDA receptors as essential mediators of excitotoxicity, the different functional domains of these ion channels have been identified as potential targets for neuroprotective agents. Following an initial hype on potential NMDA receptor therapeutics, the authors currently see a period of skepticism that, in reverse, appears to neglect the therapeutic potential of this receptor class. This review attempts a reappraisal of this important class of neurotransmitter receptors, with a focus on NMDA receptor heterogeneity, ligand binding domains, and candidate diseases for a potential neuroprotective therapy.     

Analysis of ion channel expression by astrocytes in red nucleus brain stem slices of the rat

The red nucleus (RN) has been widely used to study the formation and remodeling of synaptic connections during development and in post-lesion plasticity. Since glial cells are thought to contribute to synaptic plasticity, and information on functional properties of brain stem glia is missing, we analyzed voltage-gated ion channels as well as glutamate receptors expressed by glial cells of the RN. The patch-clamp technique was applied to identified cells in acute brain stem slices of 5- to 12-day-old rats. Based on their pattern of membrane currents, two types of glial cells could be distinguished. A first type was characterized by passive, symmetrical currents. The second population of cells, which was the focus of the present study, expressed a complex pattern of voltage-gated channels. These cells could be labeled with antibodies against glutamine synthetase and S100β, suggesting an astroglial origin. Depolarizing voltage steps activated transient and delayed rectified K⁺ currents as well as Na⁺ currents. In addition, a subset of cells expressed Ba²⁺ sensitive inward rectifier K⁻ currents activated by hyperpolarization. All “complex” glial cells analyzed possessed ionotropic glutamate receptors of the α-amino-3-hydroxy-5-methyl-4-isoxazoleprorionic acid (AMPA) subtype, while functional kainate and N-methyl-D-aspartate (NMDA) receptors could not be detected. Receptor activation blocked outward rectifying K⁺ currents, similar to previous observations in glial cells of the hippocampus and the corpus callosum. GLIA 19:234–246, 1997. © 1997 Wiley-Liss, Inc.     

Glutamate metabolism, release, and quantal transmission at central excitatory synapses: implications for neural plasticity.

Glutamate is thought to act as a neurotransmitter at several excitatory synapses in the brain. Available knowledge reveals complex intercompartmental dynamics of glutamate, which is synthesized, accumulated and released by presynaptic elements, activates postsynaptic receptors, and is eventually re-uptaken and interconverted to glutamine by the participation of surrounding glial cells. The postsynaptic reactions to physiological release of glutamate during neurotransmission are considered in relation to the quantal approach, which is revealing unsuspected complexity in central synaptic mechanisms. This issue is particularly important in view of its implications in the study of long-term synaptic modifications.     

Differential mechanisms of Ca2+ responses in glial cells evoked by exogenous and endogenous glutamate in rat hippocampus

The mechanisms of Ca2+ responses evoked in hippocampal glial cells in situ, by local application of glutamate and by synaptic activation, were studied in slices from juvenile rats using the membrane permeant fluorescent Ca2+ indicator fluo-3AM and confocal microscopy. Ca2+ responses induced by local application of glutamate were unaffected by the sodium channel blocker tetrodotoxin and were therefore due to direct actions on glial cells. Glutamate-evoked responses were significantly reduced by the L-type Ca2+ channel blocker nimodipine, the group I/II metabotropic glutamate receptor antagonist (S)-alpha-methyl-4-carboxyphenylglycine (MCPG), and the N-methyl-D-aspartate (NMDA) receptor antagonist (+/-)2-amino-5-phosphonopentanoic acid (APV). However, glutamate-induced Ca2+ responses were not significantly reduced by the non-NMDA receptor antagonist 6-cyano-7-nitro-quinoxaline-2,3-dione (CNQX). These results indicate that local application of glutamate increases intracellular Ca2+ levels in glial cells via the activation of L-type Ca2+ channels, NMDA receptors, and metabotropic glutamate receptors. Brief (1 s) tetanization of Schaffer collaterals produced increases in intracellular Ca2+ levels in glial cells that were dependent on the frequency of stimulation (> or =50 Hz) and on synaptic transmission (abolished by tetrodotoxin). These Ca2+ responses were also antagonized by the L-type Ca2+ channel blocker nimodipine and the metabotropic glutamate receptor antagonist MCPG. However, the non-NMDA receptor antagonist CNQX significantly reduced the Schaffer collateral-evoked Ca2+ responses, while the NMDA antagonist APV did not. Thus, these synaptically mediated Ca2+ responses in glial cells involve the activation of L-type Ca2+ channels, group I/II metabotropic glutamate receptors, and non-NMDA receptors. These findings indicate that increases in intracellular Ca2+ levels induced in glial cells by local glutamate application and by synaptic activity share similar mechanisms (activation of L-type Ca2+ channels and group I/II metabotropic glutamate receptors) but also have distinct components (NMDA vs. non-NMDA receptor activation, respectively). Therefore, neuron-glia interactions in rat hippocampus in situ involve multiple, complex Ca2+-mediated processes that may not be mimicked by local glutamate application.     

Organization of postsynaptic density proteins and glutamate receptors in axodendritic and dendrodendritic synapses of the rat olfactory bulb

Glutamate neurotransmission in the olfactory bulb involves both axodendritic synapses and dendrodendritic reciprocal synapses and possibly also extrasynaptic receptors. By using a sensitive immunogold procedure, we have investigated the organization of two synaptic scaffolding molecules, PSD-95 and PSD-93, as well as N-methyl-D-aspartate (NMDA) and alpha-amino-3-hydroxy-5-methylisoxazole-4-proprionic acid (AMPA) receptors, at these heterogeneous glutamate signaling sites. Immunolabeling for PSD-95 and PSD-93 was present in all major types of putative glutamatergic synapse, suggesting that these proteins are essential components of the synaptic signaling apparatus. The linear density and the subsynaptic distribution of PSD-95/PSD-93 gold particles did not differ significantly between axodendritic and dendrodendritic synapses. Antibodies recognizing NMDA and AMPA receptor subunits also labeled asymmetric synapses throughout the olfactory bulb. Immunolabeling for the AMPA receptor subunits GluR2/3 was similar in all types of synapse. In contrast, immunogold signals for the NR1 subunit of NMDA receptors varied significantly among different synapse populations, with olfactory nerve synapses in the glomerular layer showing the lowest labeling intensity. Although the lateral dendrites of mitral and tufted cells have been reported to respond to glutamate, they did not display significant plasma membrane labeling for the NR1 subunit or for PSD-95, suggesting that the physiological effects of glutamate at these sites are mediated by NMDA autoreceptors that are not clustered and occur only at a low density on the dendritic surface. Our quantitative analysis of olfactory bulb synapses indicates that the density of NMDA receptors is not determined by the complement of PSD-95/PSD-93. The latter molecules appear to be expressed in an all-or-none fashion and may form a standard lattice common to different types of glutamatergic synapse.     

Presynaptic NMDA receptors: Are they dendritic receptors in disguise?

The N-methyl-d-aspartate (NMDA) receptor plays an essential role in excitatory transmission, synaptic integration, and learning and memory. In the classical view, postsynaptic NMDA receptors act as canonical coincidence detectors providing a ‘molecular switch’ for the induction of various forms of short- and long-term synaptic plasticity. Over the past twenty years there has been accumulating evidence to suggest that NMDA receptors are also expressed presynaptically and are involved in the regulation of synaptic transmission and specific forms of activity-dependent plasticity in developing neural circuits. However, the existence of presynaptic NMDA receptors remains a contentious issue. In this review, I will discuss the criteria required for identifying functional presynaptic receptors, novel methods for probing NMDA receptor function, and recent evidence to suggest that NMDA receptors are expressed at presynaptic sites in a target-specific manner.     

NMDA-receptor trafficking and targeting: implications for synaptic transmission and plasticity

Dynamic regulation of synaptic efficacy is thought to play a crucial role in formation of neuronal connections and in experience-dependent modification of neural circuitry. The molecular and cellular mechanisms by which synaptic changes are triggered and expressed are the focus of intense interest. This articles reviews recent evidence that NMDA receptors undergo dynamically regulated targeting and trafficking, and that the physical transport of NMDA receptors in and out of the synaptic membrane contributes to several forms of long-lasting synaptic plasticity. The identification of targeting and internalization sequences in NMDA-receptor subunits has begun the unraveling of some mechanisms that underlie activity-dependent redistribution of NMDA receptors. Given that NMDA receptors are widely expressed throughout the CNS, regulation of NMDA-receptor trafficking provides a potentially important way to modulate efficacy of synaptic transmission.     

Glutamate receptors and glutamatergic signalling in the peripheral nerves

In the peripheral nervous system, the vast majority of axons are accommodated within the fibre bundles that constitute the peripheral nerves. Axons within the nerves are in close contact with myelinating glia, the Schwann cells that are ideally placed to respond to, and possibly shape, axonal activity. The mechanisms of intercellular communication in the peripheral nerves may involve direct contact between the cells, as well as signalling via diffusible substances. Neurotransmitter glutamate has been proposed as a candidate extracellular molecule mediating the cross-talk between cells in the peripheral nerves. Two types of experimental findings support this idea: first, glutamate has been detected in the nerves and can be released upon electrical or chemical stimulation of the nerves; second, axons and Schwann cells in the peripheral nerves express glutamate receptors. Yet, the studies providing direct experimental evidence that intercellular glutamatergic signalling takes place in the peripheral nerves during physiological or pathological conditions are largely missing. Remarkably, in the central nervous system, axons and myelinating glia are involved in glutamatergic signalling. This signalling occurs via different mechanisms, the most intriguing of which is fast synaptic communication between axons and oligodendrocyte precursor cells. Glutamate receptors and/or synaptic axon-glia signalling are involved in regulation of proliferation, migration, and differentiation of oligodendrocyte precursor cells, survival of oligodendrocytes, and re-myelination of axons after damage. Does synaptic signalling exist between axons and Schwann cells in the peripheral nerves? What is the functional role of glutamate receptors in the peripheral nerves? Is activation of glutamate receptors in the nerves beneficial or harmful during diseases? In this review, we summarise the limited information regarding glutamate release and glutamate receptors in the peripheral nerves and speculate about possible mechanisms of glutamatergic signalling in the nerves. We highlight the necessity of further research on this topic because it should help to understand the mechanisms of peripheral nervous system development and nerve regeneration during diseases.     

Metabotropic glutamate receptor subtypes as targets for neuroprotective drugs.

Metabotropic glutamate (mGlu) receptors have been considered as potential targets for neuroprotective drugs, but the lack of specific drugs has limited the development of neuroprotective strategies in experimental models of acute or chronic central nervous system (CNS) disorders. The advent of potent and centrally available subtype-selective ligands has overcome this limitation, leading to an extensive investigation of the role of mGlu receptor subtypes in neurodegeneration during the last 2 years. Examples of these drugs are the noncompetitive mGlu1 receptor antagonists, CPCCOEt and BAY-36-7620; the noncompetitive mGlu5 receptor antagonists, 2-methyl-6-(phenylethynyl)pyridine, SIB-1893, and SIB-1757; and the potent mGlu2/3 receptor agonists, LY354740 and LY379268. Pharmacologic blockade of mGlu1 or mGlu5 receptors or pharmacologic activation of mGlu2/3 or mGlu4/7/8 receptors produces neuroprotection in a variety of in vitro or in vivo models. MGlu1 receptor antagonists are promising drugs for the treatment of brain ischemia or for the prophylaxis of neuronal damage induced by synaptic hyperactivity. MGlu5 receptor antagonists may limit neuronal damage induced by a hyperactivity of N-methyl-d-aspartate (NMDA) receptors, because mGlu5 and NMDA receptors are physically and functionally connected in neuronal membranes. A series of observations suggest a potential application of mGlu5 receptor antagonists in chronic neurodegenerative disorders, such as amyotrophic lateral sclerosis and Alzheimer disease. MGlu2/3 receptor agonists inhibit glutamate release, but also promote the synthesis and release of neurotrophic factors in astrocytes. These drugs may therefore have a broad application as neuroprotective agents in a variety of CNS disorders. Finally, mGlu4/7/8 receptor agonists potently inhibit glutamate release and have a potential application in seizure disorders. The advantage of all these drugs with respect to NMDA or AMPA receptor agonists derives from the evidence that mGlu receptors do not "mediate," but rather "modulate" excitatory synaptic transmission. Therefore, it can be expected that mGlu receptor ligands are devoid of the undesirable effects resulting from the inhibition of excitatory synaptic transmission, such as sedation or an impairment of learning and memory.     

NMDA-R1 subunit of the cerebral cortex co-localizes with neuronal nitric oxide synthase at pre- and postsynaptic sites and in spines

The majority of nitric oxide's (NO) physiologic and pathologic actions in the brain has been linked to NMDA receptor activation. In order to determine how the NO-synthesizing enzyme within brain, neuronal NO synthase (nNOS), and NMDA receptors are functionally linked, previous studies have used in situ hybridization techniques in combination with light microscopic immunocytochemistry to show that the two are expressed within single neurons. However, this light microscopic finding does not guarantee that NMDA receptors are distributed sufficiently close to nNOS within single neurons to allow direct interaction of the two. Thus, in this study, dual immuno-electron microscopy was performed to determine whether nNOS and NMDA receptors co-exist within fine neuronal processes. We show that nNOS and the obligatory subunit of functional NMDA receptors, i.e. the NMDA-R1, co-exist within dendritic shafts, spines and terminals of the adult rat visual cortex. Axon terminals form asymmetric synaptic junctions with the dually labeled dendrites, suggesting that the presynaptic terminals release glutamate. Axons and dendrites expressing one without the other also are detected. These results indicate that it is possible for the generation of NO to be temporally coordinated with glutamatergic synaptic transmission at axo-dendritic and axo-axonic junctions and that NO may be generated independently of glutamatergic synaptic transmission. Together, our observations point to a greater complexity than previously recognized for glutamatergic neurotransmission, based on the joint versus independent actions of NO relative to NMDA receptors at pre- versus postsynaptic sites.     

Neurons derived from embryonic stem (ES) cells resemble normal neurons in their vulnerability to excitotoxic death

We determined whether embryonic stem (ES) cells could provide a model system for examining neuronal death mediated by glutamate receptors. Although limited evidence indicates that normal neurons can be derived from mouse ES cells, there have been no studies examining pathophysiological responses in mouse ES cell systems. Mouse ES cells, induced down a neural lineage by retinoic acid (RA), were found to have enhanced long-term survival when plated onto a layer of cultured mouse cortical glial cells. In these conditions, the ES cells differentiated into neural cells that appeared normal morphologically and displayed normal features of immunoreactivity when tested for neuron-specific elements. Varying the culture medium generated cultures of mixed neuronal/glial cells or enriched in oligodendrocytes. These cultures were viable for at least four weeks. Real-time PCR analysis of N-methyl-D-aspartate (NMDA) receptor subunits revealed an appropriate age-in-vitro dependent pattern of expression. Neurons derived from ES cells were vulnerable to death induced by a 24-h exposure to the selective glutamate receptor agonists NMDA, kainate, and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA). This vulnerability to agonist-induced death increased with age in vitro, and related closely to expression of receptor subunits, as it does in cultured primary neurons. Experiments with selective receptor antagonists showed that glutamate receptors mediated the NMDA- and kainate-induced death. Neuronal differentiated ES cells therefore exhibited an excitotoxic response resembling that displayed by central nervous system (CNS) neurons. Thus, ES cells, which are very amenable to genetic manipulation, provide a valid system for studying glutamate receptor-mediated toxicity at the molecular level.     

Ionotropic glutamate receptors: still a target for neuroprotection in brain ischemia? Insights from in vitro studies

Although experimental studies have widely shown that the pharmacological blockade of ionotropic glutamate receptors reduces ischemic damage, clinical trials with classical AMPA and NMDA glutamate receptor antagonists have provided negative results. To address the involvement of ionotropic glutamate receptors in ischemic damage, corticostriatal brain slices were prepared from adult rats. Extracellular recordings were performed in the striatum after stimulation of the glutamatergic corticostriatal fibres. In vitro ischemia was induced for a 10-min period by omitting oxygen and glucose from the external medium. Under control conditions, ischemia produced an irreversible loss of the corticostriatal field potential amplitude, AP5, a competitive NMDA receptor antagonist, induced a slight rescue of the potential, while ifenprodil, a positive modulator of the proton sensor of the NMDA receptors, allowed a complete recovery from the ischemic insult. Similar neuroprotection was achieved by utilizing either CNQX, a broad spectrum AMPA receptors antagonist, or Joro spider toxin, a selective blocker of calcium permeable AMPA receptors. Interestingly, while CNZX also fully suppressed physiological excitatory transmission, Joro spider toxin was ineffective on this parameter. Finally, lamotrigine and remacemide, two antiepileptic drugs that differentially affect excitatory transmission, exerted neuroprotective effects against ischemia. Noticeably, a combination of low concentrations of these two drugs exerted a stronger neuroprotection than a single drug given in isolation. Thus, it might be possible to reach a neuroprotective action by utilizing doses of these compounds low enough to avoid side effects. Our experimental data still support the idea that a negative modulation of excitatory transmission can be neuroprotective against ischemia. In addition, our findings support the concept that it is possible to produce a significant neuroprotective action in the absence of a relevant interference with normal synaptic transmission.