Activation of presynaptic group I metabotropic glutamate receptors enhances glutamate release in the rat spinal cord substantia gelatinosa

The activation of group I metabotropic glutamate receptors (mGluRs) produces a long-term potentiation of sensory transmission in the substantia gelatinosa (SG) region of the spinal cord (Prog. Brain Res. 129 (2000) 115). The mechanism(s) responsible for the induction of this potentiation is not known. Using rat spinal cord slice preparation and patch-clamp recordings, here we show, that the activation of the group I mGluRs by (S)-3,5-dihydroxyphenylglycine (DHPG, 1 microM), the mGluR1/5 agonist, increased the frequency of both activity-dependent spontaneous EPSCs, and activity-independent miniature EPSCs (mEPSCs). However, DHPG did not affect amplitude of mEPSCs. The effects of DHPG were not seen in the presence of the preferential mGluR1 antagonist CPCCOEt (10 microM). On the other hand, 2-methyl-6-(phenylethynyl)-pyridine (10 microM), a selective mGluR5 antagonist, blocked the DHPG facilitation present during the wash-out of the drug. This novel facilitating effect of the group I mGluR activation on glutamate release is the first report of a direct facilitatory action of both mGluR1 and mGluR5 subtypes on sensory transmission in the spinal cord SG region. These results indicate the potential contribution of synaptic activation of these facilitatory autoreceptors in plasticity of primary afferent neurotransmission.     

Excitation of cerebellar interneurons by group I metabotropic glutamate receptors

Cerebellar basket and stellate neurons (BSNs) provide feed-forward inhibition to Purkinje neurons (PNs) and thereby play a principal role in determining the output of the cerebellar cortex. During low-frequency transmission, glutamate released at parallel fiber synapses excites BSNs by binding to AMPA receptors; high-frequency transmission also recruits N-methyl-d-aspartate (NMDA) receptors. We find that, in addition to these ligand-gated receptors, a G-protein-coupled glutamate receptor subtype participates in exciting BSNs. Stimulation of metabotropic glutamate receptor 1alpha (mGluR1alpha) with the mGluR agonist (RS)-3,5-dihydroxyphenylglycine (DHPG) leads to an increase in spontaneous firing of BSNs and indirectly to an increase in the frequency of spontaneous inhibitory postsynaptic currents (sIPSCs) recorded in PNs. Under conditions in which ligand-gated glutamate receptors are blocked, parallel fiber stimulation generates a slow excitatory postsynaptic current (EPSC) in BSNs that is inhibited by mGluR1alpha-selective antagonists. This slow EPSC is capable of increasing BSN spiking and indirectly increasing sIPSCs frequency in PNs. Our findings reinforce the idea that distinct subtypes of glutamate receptors are activated in response to different patterns of activity at excitatory synapses. The results also raise the possibility that mGluR1alpha-dependent forms of synaptic plasticity may occur at excitatory inputs to BSNs.     

Glial glutamate transporter 1 regulates the spatial and temporal coding of glutamatergic synaptic transmission in spinal lamina II neurons

Glutamatergic synaptic transmission is a dynamic process determined by the amount of glutamate released by presynaptic sites, the clearance of glutamate in the synaptic cleft, and the properties of postsynaptic glutamate receptors. Clearance of glutamate in the synaptic cleft depends on passive diffusion and active uptake by glutamate transporters. In this study, we examined the role of glial glutamate transporter 1 (GLT-1) in spinal sensory processing. Excitatory postsynaptic currents (EPSCs) of substantia gelatinosa neurons recorded from spinal slices of young adult rats were analyzed before and after GLT-1 was pharmacologically blocked by dihydrokainic acid. Inhibition of GLT-1 prolonged the EPSC duration and the EPSC decay phase. The EPSC amplitudes were increased in neurons with weak synaptic input but decreased in neurons with strong synaptic input upon inhibition of GLT-1. We suggest that presynaptic inhibition, desensitization of postsynaptic AMPA receptors, and glutamate "spillover" contributed to the kinetic change of EPSCs induced by the blockade of GLT-1. Thus, GLT-1 is a key component in maintaining the spatial and temporal coding in signal transmission at the glutamatergic synapse in substantia gelatinosa neurons.     

Induction of homosynaptic long-term depression at spinal synapses of sensory a delta-fibers requires activation of metabotropic glutamate receptors

The synaptic strength between primary afferent Adelta-fibers, many of which convey pain-related information, and second order neurons in the spinal dorsal horn can be depressed for prolonged periods of time in a use- and N-methyl-D-aspartate receptor-dependent fashion. Here, we have used a transverse spinal cord slice-dorsal root preparation of young rat to characterize the nature of this form of long-term depression and the role of metabotropic glutamate receptors. Dorsal roots were bisected and intracellular recordings were made from lamina II neurons with independent excitatory synaptic inputs from both dorsal root halves. Conditioning stimulation of one dorsal root half (1 Hz, 900 pulses) induced long-term depression that was specific for the stimulated pathway, i. e. homosynaptic in nature. The induction of long-term depression was prevented by non-selective group I and group II mGluR antagonist (S)-alpha-methyl-4-carboxyphenylglycine, by selective group I receptor antagonist (S)-4-carboxyphenylglycine and by selective group II mGluR antagonist (RS)-alpha-methylserine-O-phosphate monophenyl ester. Group III mGluR antagonist (RS)-alpha-methylserine-O-phosphate was ineffective. Short-term depression was not affected by any of these antagonists.Thus, a homosynaptic form of long-term depression exists at putative nociceptive synapses in the spinal dorsal horn and its induction requires the activation of both group I and II metabotropic glutamate receptors.     

Activation of metabotropic glutamate receptor 1 inhibits glutamatergic transmission in the substantia nigra pars reticulata

The substantia nigra pars reticulata is a primary output nucleus of the basal ganglia motor circuit and is controlled by a fine balance between excitatory and inhibitory inputs. The major excitatory input to GABAergic neurons in the substantia nigra arises from glutamatergic neurons in the subthalamic nucleus, whereas inhibitory inputs arise mainly from the striatum and the globus pallidus. Anatomical studies revealed that metabotropic glutamate receptors (mGluRs) are highly expressed throughout the basal ganglia. Interestingly, mRNA for group I mGluRs are abundant in neurons of the subthalamic nucleus and the substantia nigra pars reticulata. Thus, it is possible that group I mGluRs play a role in the modulation of glutamatergic synaptic transmission at excitatory subthalamonigral synapses. To test this hypothesis, we investigated the effects of group I mGluR activation on excitatory synaptic transmission in putative GABAergic neurons in the substantia nigra pars reticulata using the whole cell patch clamp recording approach in slices of rat midbrain. We report that activation of group I mGluRs by the selective agonist (R,S)-3,5-dihydroxyphenylglycine (100 microM) decreases synaptic transmission at excitatory synapses in the substantia nigra pars reticulata. This effect is selectively mediated by presynaptic activation of the group I mGluR subtype, mGluR1. Consistent with these data, electron microscopic immunocytochemical studies demonstrate the localization of mGluR1a at presynaptic sites in the rat substantia nigra pars reticulata.From this finding that group I mGluRs modulate the major excitatory inputs to GABAergic neurons in the substantia nigra pars reticulata we suggest that these receptors may play an important role in basal ganglia functions. Studying this effect, therefore, provides new insights into the modulatory role of glutamate in basal ganglia output nuclei in physiological and pathophysiological conditions.     

Functions of glutamate transporters in cerebellar Purkinje cell synapses

Glutamate transporters play a critical role in the maintenance of low extracellular concentrations of glutamate, which prevents the overactivation of post‐synaptic glutamate receptors. Four distinct glutamate transporters, GLAST/EAAT1, GLT‐1/EAAT2, EAAC1/EAAT3 and EAAT4, are distributed in the molecular layer of the cerebellum, especially near glutamatergic synapses in Purkinje cells (PCs). This review summarizes the current knowledge about the differential roles of these transporters at excitatory synapses of PCs. Data come predominantly from electrophysiological experiments in mutant mice that are deficient in each of these transporter genes. GLAST expressed in Bergmann glia contributes to the clearing of the majority of glutamate that floods out of the synaptic cleft immediately after transmitter release from the climbing fibre (CF) and parallel fibre (PF) terminals. It is indispensable to maintain a one‐to‐one relationship in synaptic transmission at the CF synapses by preventing transcellular glutamate spillover. GLT‐1 plays a similar but minor role in the uptake of glutamate as GLAST. Although the loss of neither GLAST nor GLT‐1 affects cerebellar morphology, the deletion of both GLAST and GLT‐1 genes causes the death of the mutant animal and hinders the folium formation of the cerebellum. EAAT4 removes the low concentrations of glutamate that escape from uptake by glial transporters, preventing the transmitter from spilling over into neighbouring synapses. It also regulates the activation of metabotropic glutamate receptor 1 (mGluR1) in perisynaptic regions at PF synapses, which in turn affects mGluR1‐mediated events including slow EPSCs and long‐term depression. No change in synaptic function is detected in mice that are deficient in EAAC1.     

Modulation of AMPA excitatory postsynaptic currents in the spinal cord dorsal horn neurons by insulin

Glutamate AMPA receptors are critical for sensory transmission at the spinal cord dorsal horn (DH). Plasma membrane AMPA receptor endocytosis that can be induced by insulin may underlie long term modulation of synaptic transmission. Insulin receptors (IRs) are known to be expressed on spinal cord DH neurons, but their possible role in sensory transmission has not been studied. In this work the effect of insulin application on fast excitatory postsynaptic currents (EPSCs) mediated by AMPA receptors evoked in DH neurons was evaluated. Acute spinal cord slices from 6 to 10 day old mice were used to record EPSCs evoked in visually identified superficial DH neurons by dorsal root primary afferent stimulation. AMPA EPSCs could be evoked in all of the tested neurons. In 75% of the neurons the size of the AMPA EPSCs was reduced to 62.1% and to 68.9% of the control values when 0.5 or 10 μM insulin was applied. There was no significant change in the size of the AMPA EPSCs in the remaining 25% of DH neurons. The membrane permeable protein tyrosine kinase inhibitor, lavendustin A (10 μM), prevented the insulin induced AMPA EPSC depression. Our results suggest a possible role of the insulin pathway in modulation of sensory and nociceptive synaptic transmission in the spinal cord.     

Activation of group I metabotropic glutamate receptors modulates locomotor-related motoneuron output in mice

Fast glutamatergic transmission via ionotropic receptors is critical for the generation of locomotion by spinal motor networks. In addition, glutamate can act via metabotropic glutamate receptors (mGluRs) to modulate the timing of ongoing locomotor activity. In the present study, we investigated whether mGluRs also modulate the intensity of motor output generated by spinal motor networks. Application of the group I mGluR agonist (S)-3,5-dihydroxyphenylglycine (DHPG) reduced the amplitude and increased the frequency of locomotor-related motoneuron output recorded from the lumbar ventral roots of isolated mouse spinal cord preparations. Whole cell patch-clamp recordings of spinal motoneurons revealed multiple mechanisms by which group I mGluRs modulate motoneuron output. Although DHPG depolarized the resting membrane potential and reduced the voltage threshold for action potential generation, the activation of group I mGluRs had a net inhibitory effect on motoneuron output that appeared to reflect the modulation of fast, inactivating Na(+) currents and action potential parameters. In addition, group I mGluR activation decreased the amplitude of locomotor-related excitatory input to motoneurons. Analyses of miniature excitatory postsynaptic currents indicated that mGluRs modulate synaptic drive to motoneurons via both pre- and postsynaptic mechanisms. These data highlight group I mGluRs as a potentially important source of neuromodulation within the spinal cord that, in addition to modulating components of the central pattern generator for locomotion, can modulate the intensity of motoneuron output during motor behavior. Given that group I mGluR activation reduces motoneuron excitability, mGluRs may provide negative feedback control of motoneuron output, particularly during high levels of glutamatergic stimulation.     

Minocycline prevents impaired glial glutamate uptake in the spinal sensory synapses of neuropathic rats

Activation of glutamate receptors and glial cells in the spinal dorsal horn are two fundamental processes involved in the pathogenesis of various pain conditions, including neuropathic pain induced by injury to the peripheral or central nervous systems. Numerous studies have demonstrated that minocycline treatment attenuates allodynic and hyperalgesic behaviors induced by tissue inflammation or nerve injury. However, the synaptic mechanisms by which minocycline prevents hyperalgesia are not fully understood. We recently reported that deficient glutamate uptake by glial glutamate transporters (GTs) is key for the enhanced activation of N-methyl-d-aspartate (NMDA) receptors in the spinal sensory synapses of rats receiving partial sciatic nerve ligation (pSNL). In this study, we investigated how minocycline affects activation of NMDA receptors in the spinal sensory synapses in rats with pSNL by whole cell recordings of NMDA currents in spinal laminea I and II neurons from spinal slices. The effects of minocycline treatments on the dorsal horn expression of glial GTs and astrocyte marker glial fibrillary acidic protein (GFAP) were analyzed by immunohistochemistry. We demonstrated that normalized activation of NMDA receptors in synapses activated by both weak and strong peripheral input in the spinal dorsal horn is temporally associated with attenuated mechanical allodynia in rats with pSNL receiving intraperitoneal injection of minocycline. Minocycline ameliorated both the downregulation of glial GT expression and the activation of astrocytes induced by pSNL in the spinal dorsal horn. We further revealed that preventing deficient glial glutamate uptake at the synapse is crucial for preserving the normalized activation of NMDA receptors in the spinal sensory synapses in pSNL rats treated with minocycline. Our studies suggest that glial GTs may be a potential target for the development of analgesics.     

Group II metabotropic glutamate receptors inhibit glutamate release at thalamocortical synapses in the developing somatosensory cortex

Thalamocortical synapses provide a strong glutamatergic excitation to cortical neurons that is critical for processing sensory information. Unit recordings in vivo indicate that metabotropic glutamate receptors (mGluRs) reduce the effect of thalamocortical input on cortical circuits. However, it is not known whether this reduction is due to a reduction in glutamate release from thalamocortical terminals or from a decrease in cortical neuron excitability. To directly determine whether mGluRs act as autoreceptors on thalamocortical terminals, we examined the effect of mGluR agonists on thalamocortical synapses in slices. Thalamocortical excitatory postsynaptic currents (EPSCs) were recorded in layer IV cortical neurons in developing mouse brain slices. The activation of group II mGluRs with (2S,2'R,3'R)-2-(2',3'-dicarboxycyclopropyl)glycine (DCG IV) reduced thalamocortical EPSCs in both excitatory and inhibitory neurons, while the stimulation of group I or group III mGluRs had no effect on thalamocortical EPSCs. Consistent with a reduction in glutamate release, DCG IV increased the paired pulse ratio and the coefficient of variation of the EPSCs. The reduction induced by DCG IV was reversed by the group II mGluR antagonist, LY341495, and mimicked by another selective group II agonist, (2R,4R)-4-aminopyrrolidine-2,4-dicarboxylic acid (APDC). The mGluR2 subtype appears to mediate the reduction of thalamocortical EPSCs, since the selective mGluR3 agonist, N-acetylaspartylglutamate (NAAG), had no effect on the EPSCs. Consistent with this, we showed that mGluR2 is expressed in the barrels. Furthermore, blocking group II mGluRs with LY341495 reduced the synaptic depression induced by a short stimulus train, indicating that synaptically released glutamate activates these receptors. These results indicate that group II mGluRs modulate thalamocortical processing by inhibiting glutamate release from thalamocortical synapses. This inhibition provides a feedback mechanism for preventing excessive excitation of cortical neurons that could play a role in the plasticity and refinement of thalamocortical connections during this early developmental period.     

Inhibition of glutamate uptake in the spinal cord induces hyperalgesia and increased responses of spinal dorsal horn neurons to peripheral afferent stimulation

Glutamate is a primary excitatory neurotransmitter in the mammalian CNS. Glutamate released from presynaptic neurons is cleared from the synaptic cleft passively by diffusion and actively by glutamate transporters. In this study, the role of glutamate transporters in sensory processing in the spinal cord has been investigated in behavioral, in vivo and in vitro experiments. Intrathecal application of a non-selective glutamate transport inhibitor, L-trans-pyrrolidine-2,4-dicarboxylic acid (10 microl of 100 microM solution) induced hypersensitivity to peripheral mechanical and thermal stimuli. Topical application of L-trans-pyrrolidine-2,4-dicarboxylic acid (100 microM) onto the dorsal surface of the L3-L6 spinal cord increased spontaneous activities, innocuous and noxious stimulus-evoked responses and after-discharges of wide dynamic range neurons in the L4-5 spinal segments. Whole cell recordings made from superficial dorsal horn neurons in an isolated whole spinal cord from newborn rats (2-3 weeks old) revealed that bath-applied L-trans-pyrrolidine-2,4-dicarboxylic acid (100 microM) produced partial membrane depolarization, increased spontaneous action potentials with decreased neuronal membrane resistance and time constant, but without significant changes of capacitance. Finally, the amplitude and duration of primary afferent evoked-excitatory postsynaptic currents recorded from neurons in the substantia gelatinosa in the spinal slices from young adult rats (6-8 weeks old) were increased in the presence of L-trans-pyrrolidine-2,4-dicarboxylic acid (100 microM). This study indicates that glutamate transporters regulate baseline excitability and responses of dorsal horn neurons to peripheral stimulation, and suggests that dysfunction of glutamate transporters may contribute to certain types of pathological pain.     

Computational model of extracellular glutamate in the nucleus accumbens incorporates neuroadaptations by chronic cocaine

Chronic cocaine administration causes instability in extracellular glutamate in the nucleus accumbens that is thought to contribute to the vulnerability to relapse. A computational framework was developed to model glutamate in the extracellular space, including synaptic and nonsynaptic glutamate release, glutamate elimination by glutamate transporters and diffusion, and negative feedback on synaptic release via metabotropic glutamate receptors (mGluR2/3). This framework was used to optimize the geometry of the glial sheath surrounding excitatory synapses, and by inserting physiological values, accounted for known stable extracellular, extrasynaptic concentrations of glutamate measured by microdialysis and glutamatergic tone on mGluR2/3. By using experimental values for cocaine-induced reductions in cystine-glutamate exchange and mGluR2/3 signaling, and by predicting the down-regulation of glutamate transporters, the computational model successfully represented the experimentally observed increase in glutamate that is seen in rats during cocaine-seeking. This model provides a mathematical framework for describing how pharmacological or pathological conditions influence glutamate transmission measured by microdialysis.     

Reduction of excitatory postsynaptic responses by persistently active metabotropic glutamate receptors in the hippocampus

The release of glutamate from axon terminals is under the control of a variety of presynaptic receptors, including several metabotropic glutamate receptors (mGluRs). Synaptically released glutamate can activate mGluRs within the same synapse where it was released and also at a distance following its diffusion from the synaptic cleft. It is unknown, however, whether the release of glutamate is under the control of persistently active mGluRs. We tested the contribution of mGluR activation to the excitatory postsynaptic responses recorded from several types of GABAergic interneuron in strata oriens/alveus of the mouse hippocampus. The application of 1 microM (alphaS)-alpha-amino-alpha-[(1S,2S)-2-carboxycyclopropyl]xanthine-9-propanoic acid (LY341495), a broad-spectrum mGluR (subtypes 2/3/7/8) antagonist at this concentration, increased evoked-excitatory postsynaptic current (eEPSC) amplitudes by 60% (n = 33). On identified cell types, LY341495 had either no effect (7 of 14 basket and 7 of 13 oriens-lacunosum moleculare, O-LM cells) or resulted in a 32 +/- 30% (mean +/- SD) increase in EPSC amplitudes recorded from basket cells and a seven-times greater (216 +/- 102%) enhancement of EPSCs in O-LM cells. The enhancement of the first EPSC of a high-frequency train indicates persistent mGluR activation. During antagonist application, the relative increase in EPSC amplitude evoked by the second and subsequent pulses in the train was not larger than that of the first EPSC, showing no further receptor activation by the released transmitter. The effect of mGluR subtype selective agonists [3 microM L(+)-2-amino-4-phosphonobutyric acid (L-AP4): mGluR4/8; 600 microM L-AP4: mGluR4/7/8; 1 microM (2S,2'R,3'R)-2-(2',3'-dicarboxycyclopropyl)glycine (DCG-IU): mGluR2/3] and an antagonist (0.2 microM LY341495: mGluR2/3/8) suggests that persistently active mGluR2/3/8 control the excitability of hippocampal network.     

Regulation of main olfactory bulb mitral cell excitability by metabotropic glutamate receptor mGluR1

In the rodent main olfactory bulb (MOB), mitral cells (MCs) express high levels of the group I metabotropic glutamate receptor (mGluR) subtype, mGluR1. The significance of this receptor in modulating MC excitability is unknown. We investigated the physiological role of mGluR1 in regulating MC activity in rat and mouse MOB slices. The selective group I agonist (RS)-3,5-dihydroxyphenylglycine (DHPG), but not group II or III agonists, induced potent, dose-dependent, and reversible depolarization and increased firing of MCs. These effects persisted in the presence of blockers of fast synaptic transmission, indicating that they are due to direct activation of mGluRs on MCs. Voltage-clamp recordings showed that DHPG elicited a voltage-dependent inward current consisting of multiple components sensitive to potassium and calcium channel blockade and intracellular calcium chelation. MC excitatory responses to DHPG were absent in mGluR1 knockout mice but persisted in mGluR5 knockout mice. Broad-spectrum LY341495, MCPG, as well as preferential mGluR1 LY367385 antagonists blocked the excitatory effects of DHPG and also potently modulated MC spontaneous and olfactory nerve-evoked excitability. mGluR antagonists altered spontaneous membrane potential bistability, increasing the duration of the up and down states. mGluR antagonists also substantially attenuated MC responses to sensory input, decreasing the probability and increasing the latency of olfactory nerve-evoked spikes. These findings suggest that endogenous glutamate tonically modulates MC excitability and responsiveness to olfactory nerve input, and hence the operation of the MOB circuitry, via activation of mGluR1.     

Presynaptic modulation by group III metabotropic glutamate receptors (mGluRs) of the excitatory postsynaptic potential mediated by mGluR1 in rat cerebellar Purkinje cells

Purkinje neurons were recorded from rat cerebellar slices. Parallel fibres stimulation elicited a fast excitatory postsynaptic potential (EPSP) mediated by ionotropic glutamate (iGluR) -amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors followed by the inhibitory gamma-aminobutyric acidA (GABAA)-dependent postsynaptic potential. In the presence of antagonists for iGluRs and for GABAA receptors, brief tetanic activation evoked a slow metabotropic glutamate receptor (mGluR)-dependent EPSP (mGluR-EPSP). This mGluR-EPSP was blocked by the selective mGluR1 antagonists LY367385 and CPCCOEt, but not by the mGluR5 antagonist MPEP. Group II agonists affected neither iGluR-EPSP nor mGluR-EPSP. Conversely, L-AP4 and L-SOP, group III mGluR agonists, inhibited both iGluR- and mGluR-EPSPs. The depolarisations evoked by both AMPA and group I agonists were unaffected, indicating a presynaptic action of group III mGluRs. These data suggest that glutamate released by parallel fibres activates group III mGluR autoreceptors, depressing both iGluR- and mGluR1-mediated EPSPs.     

Suppressive effects of intrathecal granulocyte colony-stimulating factor on excessive release of excitatory amino acids in the spinal cerebrospinal fluid of rats with cord ischemia: role of glutamate transporters

Recently, the hematopoietic factor, granulocyte colony-stimulating factor (G-CSF), has been shown to exhibit neuroprotective effects in CNS injuries. Our previous study demonstrated that intrathecal (i.t.) G-CSF significantly improved neurological defects in spinal cord ischemic rats. Considerable evidence indicates that the release of excessive amounts of excitatory amino acids (EAAs) plays a critical role in neuron injury induced by ischemic insult. In the present study, we used a spinal cord ischemia-microdialysis model to examine whether i.t. G-CSF exerted antiexcitotoxicity effects in a rat model of spinal cord ischemia. I.t. catheters and a microdialysis probe were implanted in male Wistar rats. The results revealed that spinal cord ischemia-induced neurological defects were accompanied by a significant increase in the concentration of EAAs (aspartate and glutamate) in the spinal dialysates from 30 min to 2 days after reperfusion. I.t administration of G-CSF immediately after the performance of surgery designed to induce ischemia led to a significant reduction in ischemia-induced increases in the levels of spinal EAAs. Moreover, i.t. G-CSF also brought about a significant reduction in the elevation of spinal EAA concentrations induced by exogenous i.t. administration of glutamate (10 μl of 500 mM). I.t. G-CSF attenuated spinal cord ischemia-induced downregulation of expression of three glutamate transporters (GTs), glial transporter Glu–Asp transporter (GLAST), Glu transporter-1 (GLT-1), and excitatory amino acid carrier 1 (EAAC1) protein 48 h after spinal cord ischemic surgery. Immunohistofluorescent staining showed that i.t. G-CSF significantly upregulated expression of the three GTs in the gray matter of the lumbar spinal cord from 3 to 24 h after injection. We propose that i.t. G-CSF possesses an ability to reduce the extent of spinal cord ischemia-induced excitotoxicity by inducing the expression of glutamate transporters.     

Facilitated activation of metabotropic glutamate receptors in cerebellar Purkinje cells in glutamate transporter EAAT4-deficient mice

Around excitatory synapses in cerebellar Purkinje cells (PCs), GLAST and EAAT4 are expressed as predominant glial and neuronal glutamate transporters, respectively. EAAC1, another subtype of neuronal glutamate transporter, is also expressed in PCs. EAAT4 is co-localized with metabotropic glutamate receptors (mGluRs) at perisynaptic sites in excitatory synapses in PCs, and this neuronal transporter was reported to be involved in the regulation of mGluR activation induced by the stimulation of parallel fibers (PFs). However, it remains to be elucidated whether only EAAT4 is specifically involved in mGluR activation among the glutamate transporters expressed near excitatory synapses in PCs. Here we examined mGluR-mediated excitatory postsynaptic currents (mGluR-EPSCs) evoked by PF stimulation in cerebellar slices of mice deficient in EAAT4, EAAC1, or GLAST. PF-evoked mGluR-EPSCs showed larger amplitude and faster rising kinetics in EAAT4-deficient mice than in the wild-type mice. In contrast, there was no significant difference in either the amplitude or the rising kinetics of mGluR-EPSCs in GLAST- or EAAC1-deficient mice compared to wild-type mice. We conclude that EAAT4 is most closely involved in mGluR activation in PCs among the glutamate transporters.     

Presynaptic inhibition of corticothalamic feedback by metabotropic glutamate receptors

The thalamus relays sensory information to cortex, but this information may be influenced by excitatory feedback from cortical layer VI. The full importance of this feedback has only recently been explored, but among its possible functions are influences on the processing of sensory features, synchronization of thalamic firing, and transitions in response mode of thalamic relay cells. Uncontrolled, corticothalamic feedback has also been implicated in pathological thalamic rhythms associated with certain neurological disorders. We have found a form of presynaptic inhibition of corticothalamic synaptic transmission that is mediated by a Group II metabotropic glutamate receptor (mGluR) and activated by high-frequency corticothalamic activity. We tested putative retinogeniculate and corticogeniculate synapses for Group II mGluR modulation within the dorsal lateral geniculate nucleus of the ferret thalamus. Stimulation of optic-tract fibers elicited paired-pulse depression of excitatory postsynaptic currents (EPSCs), whereas stimulation of the optic radiations elicited paired-pulse facilitation. Paired-pulse responses were subsequently used to characterize the pathway of origin of stimulated synapses. Group II mGluR agonists (LY379268 and DCG-IV) applied to thalamic neurons under voltage-clamp conditions reduced the amplitude of corticogeniculate EPSCs. Stimulation with high-frequency trains produced a facilitating response that was reduced by Group II mGluR agonists, but was enhanced by the selective antagonist LY341495, revealing a presynaptic, mGluR-mediated reduction of high-frequency corticogeniculate feedback. Agonist treatment did not affect EPSCs from stimulation of the optic tract. NAAG (reported to be selective for mGluR3) was ineffective at the corticogeniculate synapse, implicating mGluR2 in the observed effects. Our data are the first to show a synaptically elicited form of presynaptic inhibition of corticothalamic synaptic transmission that is mediated by presynaptic action of mGluR2. This presynaptic inhibition may partially mute sensory feedback and prevent reentrant excitation from initiating abnormal thalamic rhythms.     

Immunohistochemical localization of group I and II metabotropic glutamate receptors in control and amyotrophic lateral sclerosis human spinal cord: upregulation in reactive astrocytes.

Excitotoxicity, which is mediated by the excessive activation of glutamate receptors, has been implicated in the pathogenesis of amyotrophic lateral sclerosis (ALS). There is substantial information about the distribution and function of ionotropic glutamate receptors in the spinal cord, although the role of metabotropic glutamate receptors (mGluRs) is poorly understood in this region of the brain, particularly under pathological conditions. We used immunocytochemistry to study the general distribution of group I and group II mGluR immunoreactivity in the human spinal cord, as well as the cell-specific expression of these receptors. We also investigated whether mGluR expression was altered in the spinal cord of patients with sporadic and familial ALS. Immunocytochemical analysis of control human spinal cord demonstrated that mGluR1alpha and mGluR5 (group I mGluRs) were highly represented in neuronal cells throughout the spinal cord. mGluR1alpha showed the highest relative level of expression in ventral horn neurons (laminae VIII and IX), whereas intense mGluR5 immunoreactivity was observed within the dorsal horn (superficial laminae I and II). Group II mGluRs (mGluR2/3) immunoreactivity was mainly concentrated in the inner part of the lamina II. With respect to specific neuronal populations, mGluR2/3 and mGluR5 appeared to be most frequently expressed in calbindin-containing and calretinin-containing cells, respectively. In control spinal cord only sparse astrocytes showed a weak to moderate mGluR immunoreactivity. Regional differences in immunoreactivity were apparent in ALS compared to control. In particular, mGluR expression was increased in reactive glial cells in both gray (ventral horn) and white matter of ALS spinal cord. Upregulation of mGluRs in reactive astrocytes may represent a critical mechanism for modulation of glial function and changes in glial-neuronal communication in the course of neurodegenerative diseases.     

Spatially distinct actions of metabotropic glutamate receptor activation in dorsal lateral geniculate nucleus

Thalamocortical neurons in the dorsal lateral geniculate nucleus (dLGN) dynamically communicate visual information from the retina to the neocortex, and this process can be modulated via activation of metabotropic glutamate receptors (mGluRs). Neurons within dLGN express different mGluR subtypes associated with distinct afferent synaptic pathways; however, the physiological function of this organization is unclear. We report that the activation of mGluR(5), which are located on presynaptic dendrites of local interneurons, increases GABA output that in turn produces an increased inhibitory activity on proximal but not distal dendrites of dLGN thalamocortical neurons. In contrast, mGluR(1) activation produces strong membrane depolarization in thalamocortical neurons regardless of distal or proximal dendritic locations. These findings provide physiological evidence that mGluR(1) appear to be distributed along the thalamocortical neuron dendrites, whereas mGluR(5)-dependent action occurs on the proximal dendrites/soma of thalamocortical neurons. The differential distribution and activation of mGluR subtypes on interneurons and thalamocortical neurons may serve to shape excitatory synaptic integration and thereby regulate information gating through the thalamus.