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Copyright © 1999, American Society for
Neurochemistry.
Bookshelf ID: NBK28252
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Siegel GJ, Agranoff BW, Albers RW, et al., editors. Basic Neurochemistry: Molecular, Cellular and Medical Aspects. 6th edition. Philadelphia: Lippincott-Raven; 1999.
Basic Neurochemistry: Molecular, Cellular and Medical Aspects. 6th edition.
Show detailsHistorically, the most compelling evidence that glutamate and aspartate function as neurotransmitters came from the observation that at low concentrations they excite virtually every neuron in the CNS. In the adult CNS, l-glutamate and l-aspartate are the most likely candidates for neurotransmitter action at excitatory amino acid receptors, and these amino acids are used by some of the most widely distributed neuronal types. Glutamate and aspartate are present in high concentrations in the CNS and are released in a Ca2+-dependent manner upon electrical stimulation in vitro. Both have powerful excitatory effects on neurons when iontophoresed in vivo. High-affinity uptake systems have been cloned and are located in nerve terminals and glial cells associated with many neuronal pathways (see below). Selective binding sites can be demonstrated by both autoradiographic immunohistochemical and pharmacological techniques in vitro. Although the unequivocal identification of glutamate and aspartate as neurotransmitter candidates has been hampered by their involvement in many other functions, they are accepted universally in this role.
Glutamate and aspartate are nonessential amino acids that do not cross the blood—brain barrier and, therefore, are synthesized from glucose and a variety of other precursors. Synthetic and metabolic enzymes for glutamate and aspartate have been localized to the two main compartments of the brain, neurons and glial cells (Fig. 15-7). Glutamic acid is in a metabolic pool with α-ketoglutaric acid and glutamine. A large fraction of the glutamate released from nerve terminals probably is taken up into glial cells, where it is converted into glutamine. Glutamine then cycles back to nerve terminals, where it participates in the replenishment of the transmitter pools of glutamate and GABA. At present, there is little evidence for the extracellular metabolism of glutamate in the CNS.
Figure 15-7
Metabolism of glutamate in synaptic structures. Glutamate is synthesized and stored within synaptic endings of nerve terminals. Synthesis of transmitter pools of glutamate is likely to involve two major synthetic pathways. The conversion of glutamine (more...)
The transmitter pool of glutamate is stored in synaptic vesicles
Electron-microscopic analysis of glutamate immunoreactive puncta within the cortex has shown that many of these structures are axon terminals. Most glutamate-positive axon terminals form asymmetrical synaptic contacts on small- and medium-caliber dendritic shafts and spines; glutamate-positive axon terminals on cell bodies are extremely rare.
Synaptic vesicles actively accumulate glutamate through a Mg2+/ATP-dependent process. This uptake mechanism is inhibited by substances that destroy the electrochemical gradient. The concentration of glutamate within synaptic vesicles is thought to be very high, in excess of 20 mM. Aspartate is neither an inhibitor nor a substrate for this vesicular uptake mechanism. A vesicular uptake mechanism for aspartate has not yet been demonstrated, somewhat weakening the case for considering aspartate to be a neurotransmitter. In hippocampal slices, however, reduction of the extracellular glucose concentration strongly reduces KCl-evoked glutamate release but enhances aspartate release, suggesting that when glutamate and aspartate metabolism via the tricarboxylic acid cycle is perturbed, aspartate may become preferentially available for release during synaptic transmission.
Glutamate-receptor activation underlies most fast excitatory synaptic transmission in the brain
Both AMPA and NMDA receptors are present on most CNS neurons, although there are a few notable exceptions. Accordingly, activation of AMPA and NMDA receptors appears to underlie the vast majority of “fast” synaptic transmission in the CNS. The concentration of glutamate required to half-maximally activate AMPA receptors is approximately 200 μM, about two orders of magnitude less than the concentration thought to be contained within synaptic vesicles; glutamate is much more potent on NMDA receptors, with an EC50 of 10 to 15 μM. Therefore, synaptically released glutamate has a high probability of transiently saturating its receptors on the adjacent cell. Considerable evidence suggests that on many cell types AMPA and NMDA receptors are clustered together within the same postsynaptic densities (Fig. 15-6A). Receptor clustering appears to be mediated by binding of the extreme C-terminal tail of certain glutamate-receptor subunits to a family of proteins that express the PDZ modular protein interaction domain (Fig. 15-2). These PDZ-containing proteins presumably form a scaffold or bridge between receptor and cytoskeletal proteins located near the postsynaptic membrane. This close apposition results in the simultaneous activation of these receptors. The integration of current flow through the two receptor systems endows the cell with a powerful means of regulating neuronal excitation. It should be remembered that aspartate has little or no affinity for AMPA receptors, so synaptically released aspartate will activate only NMDA receptors.
Both NMDA-and AMPA-receptor components of the EPSP are thought to be produced by a brief (1 msec) appearance of free transmitter in the synaptic cleft. Synaptically released glutamate thus results in a two-component excitatory postsynaptic current (EPSC) upon binding to AMPA and NMDA receptors at most central synapses (Fig. 15-6B). Activation of AMPA receptors mediates a component that has a rapid onset and decay, whereas the component mediated by NMDA receptor activation has a slower rise time and a decay lasting up to several hundred milliseconds (Fig. 15-6C). Rapid desensitization of AMPA receptors may control the time course of EPSPs at many synapses. The long time course of NMDA receptor activation, by contrast, provides more opportunities for temporal and spatial summation of multiple inputs. The resulting summed depolarization may allow other synaptic inputs or nonsynaptic membrane channels to initiate action potentials. The decay time of the NMDA receptor component is approximately 100 times longer than the mean open time of the channel. The more prolonged activation of NMDA receptors is thought to be due to the higher affinity of glutamate for NMDA than AMPA receptors; high affinity often results from a slow dissociation of the agonist from its receptor, which could result in multiple channel activations for each against binding event.
Ca2+ influx through N-methyl-d-aspartate and AMPA receptors mediates synaptic plasticity
Although activation of NMDA receptors results in appreciable current flow and tends to depolarize the cell membrane toward threshold for an action potential, this is unlikely to be the sole role of this receptor when activated at typical resting membrane potentials because NMDA receptors are highly permeable to Ca2+, as mentioned above. Likewise, AMPA receptors on some interneurons have no GluR2 subunits and are permeable to Ca2+ as described above. Thus, an important role of NMDA and some AMPA receptors may be to inject Ca2+ into the postsynaptic membrane.
The high permeability of NMDA receptor channels for divalent cations has many implications for cell function. Ca2+ concentration within the cell interior is heavily buffered to about 100 nM. The elevation of cytoplasmic Ca2+ by Ca2+ entry through NMDA-receptor channels may lead to the transient activation of a variety of Ca2+-activiated enzymes, including Ca2+/calmodulin-dependent protein kinase II, calcineurin, PKC, phospholipase A2, PI-PLC, nitric oxide synthase and a number of endonucleases. Activation of each of these enzymes occurs as a result of Ca2+ entry following glutamate-receptor activation. Although a variety of forms of synaptic plasticity have been found in the mammalian CNS, LTP and LTD of excitatory synaptic responses in hippocampal CA1 pyramidal cells have been characterized most extensively. LTP and LTD are activity-dependent alterations in synaptic efficacy that can last up to several weeks in vivo and are thought to be involved in the acquisition of spatial memories (Chap. 50).
Receptor knockouts reveal clues to ionotropic receptor functions
In most neurons, high expression of the edited GluR2 subunit ensures that synaptic AMPA receptors will permit only insignificant Ca2+ influx. However, mice engineered to harbor an editing-incompetent GluR2 gene expressed AMPA receptors with increased Ca2+ permeability [22]. These mice developed seizures and died by 3 weeks of age, demonstrating that GluR2 editing is essential for normal brain development. Deletion of the GluR2 editing enzyme RAD1 by gene targeting produces the same phenotype. Surprisingly, complete deletion of the GluR2 allele, which also increases Ca2+ permeability of AMPA receptors in targeted mouse neurons, neither induced seizures nor proved lethal in homozygous mice. Rather, Ca2+ entry through GluR2-lacking synaptic AMPA receptors could produce a form of LTP [23]. It is unclear why these two genetic manipulations had such different outcomes, although it is possible that a complete absence of GluR2 impairs AMPA receptor assembly because the density of synaptic AMPA receptors appeared to be low in the latter study.
Conventional gene targeting of the NMDA receptor NR1 subunit interferes with breathing and is lethal within a few hours of birth. However, mouse strains in which the NR1 gene knockout is restricted to the CA1 region of the hippocampus survive and grow normally [24]. This was accomplished by using a clever second-generation cre-loxP strategy for gene targeting. Temporal and spatial restriction of gene knockouts using the cre-loxP system should be applicable to any gene and may circumvent developmental problems associated with conventional strategies. With this approach, 34-bp loxP elements were inserted to flank the NR1 gene, rendering the gene susceptible to the action of the enzyme cre recombinase. Mice with a “floxed” NR1 allele were mated with transgenic mice that harbored a cre gene under the control of a calmodulin kinase II promoter, which restricted expression of cre to the CA1 pyramidal neurons, predominantly after postnatal day 21. In these mice, the NR1 gene was deleted by cre but only in CA1 pyramidal cells. Remarkably, LTP was impaired in the CA1 but not other hippocampal regions, and these mice exhibited impaired spatial memory in a water maze task. These findings demonstrate the essential role of NMDA receptors in LTP in the CA1 region and strongly suggest that LTP in the CA1 region is necessary for the acquisition of spatial memory.
- The transmitter pool of glutamate is stored in synaptic vesicles
- Glutamate-receptor activation underlies most fast excitatory synaptic transmission in the brain
- Ca2+ influx through N-methyl-d-aspartate and AMPA receptors mediates synaptic plasticity
- Receptor knockouts reveal clues to ionotropic receptor functions
- Glutamate and Aspartate Are the Major Excitatory Transmitters in the Brain - Bas...Glutamate and Aspartate Are the Major Excitatory Transmitters in the Brain - Basic Neurochemistry
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