Differential expression of group I metabotropic glutamate receptors in rat spinal cord somatic and autonomic motoneurons: possible implications for the pathogenesis of amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a neurodegenerative disorder characterized by the progressive loss of somatic, but not autonomic, motoneurons. The reason for this selective vulnerability is unknown. The pathogenesis of ALS is thought to involve glutamatergic excitotoxic mechanisms. While overactivation of ionotropic glutamate receptors may trigger excitotoxicity, we have previously shown that stimulation of group I metabotropic glutamate receptors (mGluRs) can exert neuroprotective effects on cultured motoneurons. Using in situ hybridization, we found a differential expression of group I mGluRs (mGluR1 and 5) in rat spinal cord. Autonomic motoneurons from the sacral parasympathetic Onuf's nucleus and thoracic sympathetic neurons, which are spared in ALS, express high levels of mGluR5, while somatic motoneurons do not. In addition, mGluR1 mRNA is found only in smaller somatic motoneurons, which seem to be less vulnerable in ALS. Thus, differential mGluR expression might provide a possible clue to the selective vulnerability of different motoneuronal subpopulations in ALS.     

Expression and cell distribution of group I and group II metabotropic glutamate receptor subtypes in taylor-type focal cortical dysplasia

PURPOSE: Focal cortical dysplasia (FCD) is known to be a major cause of intractable epilepsy. The cellular mechanism(s) underlying the epileptogenicity of FCD remain largely unknown. Because recent studies indicate that metabotropic glutamate receptor subtypes (mGluRs) play a role in epileptogenesis, we investigated the expression and cellular distribution pattern of mGluRs in FCD specimens. METHODS: Immunocytochemical expression of group I and group II mGluR subtypes was investigated in 15 specimens of human FCD obtained during epilepsy surgery. RESULTS: Strong mGluR1alpha and mGluR5 (group I mGluRs) immunoreactivity (IR) was observed in the majority of FCD specimens in dysplastic as well as in heterotopic neurons. mGluR1alpha was expressed in a subpopulation of neurons (mainly large dysplastic cells), whereas mGluR5 was represented in a higher percentage of dysplastic neuronal cells. Group II mGluRs (mGluR2/3) IR was observed less frequently than that in group I mGluRs and generally appeared in <10% of the dysplastic neurons. IR for all three mGluR subtypes was observed in balloon cells. mGluR2/3 appeared to be most frequently expressed in glial fibrillary acidic protein (GFAP)-positive balloon cells (glial type), and mGluR1alpha, in microtubule-associated protein (MAP)2-positive cells (neuronal type). mGluR5 was present in the majority of balloon cells. Occasionally glial mGluR1alpha IR was observed in bizarre glial cells with di- or multinuclei. Reactive astrocytes were intensively stained, mainly with mGluR5 and mGluR2/3. CONCLUSIONS: The cellular distribution of mGluR subtypes, with high expression of mGluR1alpha and mGluR5 in dysplastic neurons, suggests a possible contribution of group I mGluRs to the intrinsic and high epileptogenicity of dysplastic cortical regions.     

Expression of metabotropic glutamate receptors in rat meningeal and brain microvasculature and choroid plexus

This study investigated the distribution of metabotropic glutamate receptors (mGluRs) in meningeal and parenchymal microvasculature and in choroid plexus by means of Western blot analysis and immunohistochemistry. Western blot analysis demonstrated mGluR expression in both rat and human leptomeningeal tissues. In the rat, mGluR expression was developmentally regulated, with only mGluR2/3 showing expression at the embryonic day 19 developmental stage. In contrast, mGluR1 alpha, mGluR2/3, mGluR4a, and mGluR7 were expressed in leptomeninges from adult rats. Immunohistochemical analyses showed intense mGluR1 alpha immunoreactivity in the pia mater and blood vessels in the subarachnoid space and in the arachnoid layer of the meninges. mGluR2/3, mGluR4a, mGluR5, and mGluR7 were also expressed in meningeal microvasculature. In addition, the parenchymal microvasculature and choroid plexus were strongly immunoreactive for mGluR1 alpha, mGluR2/3, mGluR4a, mGluR5, and mGluR7. We used antibodies specific for phenotypic markers of microvascular and glial cells to characterize the cell type(s) immunopositive for mGluRs. Comparison of staining with anti-von Willebrand factor antibody and anti-mGluR antibodies revealed that mGluR immunoreactivity was present in cells that surrounded the luminal surface labeled by the endothelial cell marker. In these cells, smooth muscle actin and mGluR immunoreactivity overlapped, suggesting that, in addition to endothelial cells, pericytes within the microvasculature also express mGluRs. Furthermore, expression of mGluR1 alpha was also observed in pure pericyte cultures isolated from bovine retina. These data suggest that glutamate by means of activation of mGluRs may have a broad sphere of physiological influence in the brain which in addition to modulating synaptic transmission may also have a role in determining microvascular function and dysfunction.     

Group I metabotropic glutamate receptors at GABAergic synapses in monkeys.

Recent data showed that group I metabotropic glutamate receptors (mGluRs) are located perisynaptic to the postsynaptic specializations of asymmetric glutamatergic synapses in the cerebellum and hippocampus in rats. In the present study, we used immunogold labeling to elucidate the subsynaptic localization of group I mGluRs (mGluR1a and mGluR5) in the internal and external segments of the globus pallidus in monkeys. In contrast to hippocampal and cerebellar neurons, which receive massive glutamatergic inputs, dendrites of pallidal neurons are covered with GABAergic boutons from the striatum intermingled with a small proportion of glutamatergic terminals arising largely from the subthalamic nucleus. In line with previous data, mGluR1a and mGluR5 immunoreactivity was found at the edge of the postsynaptic specializations of asymmetric synapses established by subthalamic-like boutons in the monkey pallidum. However, a large proportion of gold particles were also seen in the main body of the postsynaptic specializations of symmetric synapses formed by striatal GABAergic terminals. These data raise questions about the possible sources of activation of these receptors and the potential roles of group I mGluRs in modulating GABAergic neurotransmission at striatopallidal synapses.     

Metabotropic glutamate receptors in superficial laminae of the rat dorsal horn.

Light and electron microscopic immunocytochemistry were employed here to show the distribution of metabotropic glutamate receptors (mGluRs) mGluR2/3 and mGluR5 in laminae I and II of the dorsal horn, to identify their pre- and postsynaptic location, and to test colocalization with gamma-aminobutyric acid (GABA). mGluR2/3 was mainly in the inner part of lamina II; mGluR5 was mainly in laminae I and II. Electron microscopy showed that both mGluR2/3 and mGluR5 were in perikarya, dendrites, and vesicle-containing profiles. Two main morphological types of primary afferent terminals can be distinguished in the superficial laminae: C1, likely to be endings of unmyelinated fibers, and C2, of small myelinated fibers. Quantitative data show that only a small fraction of C2s stained for either receptor; more common were immunopositive dendrites postsynaptic to these terminals, and most common were appositions between C2s and mGluR5 immunopositive dendrites. Vesicle-containing profiles were characteristically apposed to primary afferent terminals, mainly C2s. Immunopositivity for mGluRs, especially mGluR2/3, was present in vesicle-containing profiles apposed to C2, none to C1, and about half of the profiles immunostained for either receptor were also stained for GABA. The presence of presynaptic and postsynaptic mGluRs in both inhibitory and excitatory interneurons may contribute to complex processing of fast and slow responses to peripheral input in superficial laminae. As selective agonists of mGluRs may modulate GABA release, the present demonstration of mGluRs in GABAergic terminals of presumed interneurons suggests that facilitatory effects may involve a mechanism of disinhibition.     

Interaction between metabotropic and ionotropic glutamate receptors regulates neuronal network activity.

Experimental and computational techniques have been used to investigate the group I metabotropic glutamate receptor (mGluR)-mediated increase in the frequency of spinal cord network activity underlying locomotion in the lamprey. Group I mGluR activation potentiated the amplitude of NMDA-induced currents in identified motoneurons and crossed caudally projecting network interneurons. Group I mGluRs also potentiated NMDA-induced calcium responses. This effect was blocked by a group I mGluR-specific antagonist, but not by blockers of protein kinase A, C, or G. The effect of group I mGluRs activation was also tested on NMDA-induced oscillations known to occur during fictive locomotion. Activation of these receptors increased the duration of the plateau phase and decreased the duration of the hyperpolarizing phase. These effects were blocked by a group I mGluR antagonist. To determine its role in the modulation of NMDA-induced oscillations and the locomotor burst frequency, the potentiation of NMDA receptors by mGluRs was simulated using computational techniques. Simulating the interaction between these receptors reproduced the modulation of the plateau and hyperpolarized phases of NMDA-induced oscillations, and the increase in the frequency of the locomotor rhythm. Our results thus show a postsynaptic interaction between group I mGluRs and NMDA receptors in lamprey spinal cord neurons, which can account for the regulation of the locomotor network output by mGluRs.     

Effects of inflammation on the ultrastructural localization of spinal cord dorsal horn group I metabotropic glutamate receptors

Inflammatory pain is thought to induce functional plasticity of spinal dorsal horn neurons and may produce changes in glutamate receptor expression. Plasticity of group I metabotropic glutamate receptors (mGluR1 and mGluR5) is important in various neuronal systems, and these receptors are also known to modulate nociceptive neurotransmission in the spinal dorsal horn. The present study aimed at determining whether persistent inflammatory pain produces alterations in intracellular and plasma membrane-associated mGluR1alpha and mGluR5 in spinal cord dorsal horn. Persistent inflammation was induced in male Long Evans rats by a unilateral intraplantar injection of 100 muL of complete Freund's adjuvant (CFA). Three days after the CFA injection thermal withdrawal latencies were obtained prior to processing of transverse spinal cord sections for preembedding immunogold labeling after incubation in primary antibody for mGluR1alpha or mGluR5. Using electron microscopy, we quantified immunogold-labeled mGluR1alpha and mGluR5 profiles, located in lamina V and I-II, respectively, of both CFA-treated rats and untreated control rats. Compared to untreated rats, CFA-treated rats had a significant increase in the number of plasma membrane-associated mGluR5 immunogold-labeled particles in lamina I-II neurons of the spinal cord. Although no changes to mGluR1alpha expression were found in CFA-treated rats, plasma membrane-associated mGluR1alpha was significantly closer to the synapse. Therefore, in CFA-treated rats there was a specific increase in the ratio of plasma membrane-associated versus intracellular immunogold-labeled particles for mGluR5, and lateral movement of mGluR1alpha toward the synapse, indicating that peripheral inflammation-induced trafficking of group I mGluRs in spinal dorsal horn neurons may be an important factor in the development of plastic changes associated with inflammation-induced chronic pain.     

Perisynaptic Location of Metabotropic Glutamate Receptors mGluR1 and mGluR5 on Dendrites and Dendritic Spines in the Rat Hippocampus

Ionotropic and metabotropic (mGluR1a) glutamate receptors were reported to be segregated from each other within the postsynaptic membrane at individual synapses. In order to establish whether this pattern of distribution applies to the hippocampal principal cells and to other postsynaptic metabotropic glutamate receptors, the mGluR1a/b/c and mGluR5 subtypes were localized by immunocytochemistry. Principal cells in all hippocampal fields were reactive for mGluR5, the strata oriens and radiatum of the CA1 area being most strongly immunolabelled. Labelling for mGluR1b/c was strongest on some pyramids in the CA3 area, weaker on granule cells and absent on CA1 pyramids. Subpopulations of non-principal cells showed strong mGluR1 or mGluR5 immunoreactivity. Electron microscopic pre-embedding immunoperoxidase and both pre- and postembedding immunogold methods consistently revealed the extrasynaptic location of both mGluRs in the somatic and dendritic membrane of pyramidal and granule cells. The density of immunolabelling was highest on dendritic spines. At synapses, immunoparticles for both mGluR1 and mGluR5 were found always outside the postsynaptic membrane specializations. Receptors were particularly concentrated in a perisynaptic annulus around type I synaptic junctions, including the invaginations at 'perforated'synapses. Measurements of immunolabelling on dendritic spines showed decreasing levels of receptor as a function of distance from the edge of the synaptic specialization. We propose that glutamatergic synapses with an irregular edge develop in order to increase the circumference of synaptic junctions leading to an increase in the metabotropic to ionotropic glutamate receptor ratio at glutamate release sites. The perisynaptic position of postsynaptic metabotropic glutamate receptors appears to be a general feature of glutamatergic synaptic organization and may apply to other G-protein-coupled receptors.     

Induction of Presynaptic Reexpression of an Adhesion Protein in Lamina II after Dorsal Root Deafferentation in Adult Rat Spinal Cord

Limbic system-associated membrane protein (LAMP), a 64-kDa membrane protein, is an axon guidance adhesion molecule expressed by neurons in limbic system-related areas of the CNS. During development, LAMP is expressed on growing axons, growth cones, and their target neurons, but in adults it is restricted to membranes of somata and dendrites. In the adult spinal cord, LAMP immunoreactivity is found only on neurons of lamina II, lamina X, and the intermediolateral cell column and its ultrastructural localization is entirely postsynaptic. We studied changes in the expression of LAMP in lamina II of adult rat spinal cord after L1–S2 dorsal rhizotomy, a procedure that partially deafferents lamina II neurons and induces axonal sprouting by spared systems in lamina II. At the light microscopic level, LAMP immunoreactivity in lamina II was decreased in density at 3, 10, and 60 days postoperatively. This decrease in immunoreactivity suggests that LAMP expression by lamina II neurons may normally be regulated by specific afferent activity. Ultrastructurally, in control lamina II and after deafferentation in both control and deafferented lamina II at 3 and 60 days postoperatively, LAMP expression was restricted to postsynaptic membranes. Ten days after deafferentation, however, when axons are actively sprouting, LAMP was expressed on both axonal and postsynaptic membranes. The reexpression of LAMP on axonal profiles after deafferentation may identify axons that undergo sprouting in response to deafferentation.     

Comparative analysis of the subcellular and subsynaptic localization of mGluR1a and mGluR5 metabotropic glutamate receptors in the shell and core of the nucleus accumbens in rat and monkey

Group I metabotropic glutamate receptors (mGluRs) play critical roles in synaptic plasticity and drug addiction. To characterize potential sites whereby these receptors mediate their effects in the ventral striatum, we studied the subcellular and subsynaptic localization of mGluR1a and mGluR5 in the shell and core of the nucleus accumbens in rat and monkey. In both species, group I mGluRs are mainly postsynaptic in dendrites and spines, with rare presynaptic labeling in unmyelinated axons. Minor, yet significant, differences in proportions of specific immunoreactive elements were found between the accumbens shell and the accumbens core in monkey. At the subsynaptic level, significant differences were found in the proportion of plasma membrane-bound mGluR5 labeling between species. In dendrites, spines, and unmyelinated axons, a significantly larger proportion of mGluR5 labeling was bound to the plasma membrane in rats (50-70%) than in monkeys (30-50%). Conversely, mGluR1a displayed the same pattern of immunogold labeling in the two species. Electron microscopic colocalization studies revealed 30% colocalization of mGluR1a and mGluR5 in dendrites and as much as 50-65% in spines in both compartments of the rat accumbens. Both group I mGluRs were significantly expressed in D1-immunoreactive dendritic processes (60-75% colocalization) and spines (30-50%) of striatal projection neurons as well as dendrites of cholinergic (30-70%) and parvalbumin-containing (70-85%) interneurons. These findings highlight the widespread expression of group I mGluRs in projection neurons and interneurons of the shell and core of the nucleus accumbens, providing a solid foundation for regulatory and therapeutic functions of group I mGluRs in reward-related behaviors and drug addiction.     

Expression of metabotropic glutamate receptor mRNAs in the human spinal cord: implications for selective vulnerability of spinal motor neurons in amyotrophic lateral sclerosis

To elucidate the relevance of metabotropic glutamate receptors (mGluRs) to the selective vulnerability of motor neurons in the spinal cord in patients with amyotrophic lateral sclerosis (ALS), we investigated the distribution of mRNAs coding mGluR1-5 in the normal human spinal cord. The mRNAs for mGluR1, 4 and 5 were observed in the spinal gray matter, whereas mGluR2 mRNA was absent in the spinal cord and mGluR3 mRNA was displayed only on glial cells in the white matter. Signals for mGluR1 and mGluR5 were enriched in the dorsal horn, while mGluR4 mRNA was abundant in the ventral horn. Since agonists to group I mGluRs (mGluR 1 and 5) have been demonstrated to have neuroprotective effects on spinal motor neurons, less expression of mRNAs coding mGluR1 and mGluR5 in the ventral horn than in the dorsal horn may be implicated in the selective susceptibility of spinal motor neurons in ALS.     

High level of mGluR7 in the presynaptic active zones of select populations of GABAergic terminals innervating interneurons in the rat hippocampus

The release of neurotransmitters is modulated by presynaptic metabotropic glutamate receptors (mGluRs), which show a highly selective expression and subcellular location in glutamatergic terminals in the hippocampus. Using immunocytochemistry, we investigated whether one of the receptors, mGluR7, whose level of expression is governed by the postsynaptic target, was present in GABAergic terminals and whether such terminals targeted particular cells. A total of 165 interneuron dendritic profiles receiving 466 synapses (82% mGluR7a-positive) were analysed. The presynaptic active zones of most GAD-(77%) or GABA-positive (94%) synaptic boutons on interneurons innervated by mGluR7a-enriched glutamatergic terminals (mGluR7a-decorated) were immunopositive for mGluR7a. GABAergic terminals on pyramidal cells and most other interneurons in str. oriens were mGluR7a-immunonegative. The mGluR7a-decorated cells were mostly somatostatin- and mGluR1alpha-immunopositive neurons in str. oriens and the alveus. Their GABAergic input mainly originated from VIP-positive terminals, 90% of which expressed high levels of mGluR7a in the presynaptic active zone. Parvalbumin-positive synaptic terminals were rare on mGluR7a-decorated cells, but on these neurons 73% of them were mGluR7a-immunopositive. Some type II synapses innervating interneurons were immunopositive for mGluR7b, as were some type I synapses. Because not all target cells of VIP-positive neurons are known it has not been possible to determine whether mGluR7 is expressed in a target-cell-specific manner in the terminals of single GABAergic cells. The activation of mGluR7 may decrease GABA release to mGluR7-decorated cells at times of high pyramidal cell activity, which elevates extracellular glutamate levels. Alternatively, the presynaptic receptor may be activated by as yet unidentified endogenous ligands released by the GABAergic terminals or the postsynaptic dendrites.     

Vanilloid receptor VR1 is both presynaptic and postsynaptic in the superficial laminae of the rat dorsal horn.

Terminals in the rat spinal cord that express the vanilloid receptor VR1 are from small and medium dorsal root ganglion (DRG) neurons and appear prominent in lamina I and inner lamina II. Because primary afferents from these neurons can be myelinated or unmyelinated and their terminals in these laminae can be of various morphological and functional types, we undertook this study to identify the type(s) of VR1-positive afferent fibers and terminals. In the DRG, many small and medium-sized neurons are immunopositive. Under electron microscopy, dorsal root afferents that are immunopositive for VR1 are predominantly unmyelinated. Large numbers of VR1-positive terminals in lamina I are of the nonglomerular type and may contain dense core vesicles. VR1 immunoreactivity in terminals in lamina I is in good agreement with data on noxious, heat-sensitive neurons in the dorsal horn. Two types of glomerular afferent terminals in lamina II also are immunopositive for VR1. In both laminae, most VR1-positive terminals are distinct from substance P-positive terminals. However, the immunoreactivity in lamina II also is prominent in dendrites that are contacted by primary afferent endings. Because we also observed patchy immunostaining in cell bodies in lamina II, this unexpected result may reflect synthesis of VR1 by neurons in this lamina. However, because dorsal rhizotomy abolishes VR1 staining in both laminae I and II, it is suggested that the expression and intracellular dynamics of VR1 in lamina II neurons are controlled by presynaptic input.     

Ultrastructural localization of beta-arrestin-1 and -2 in rat lumbar spinal cord.

beta-arrestins play significant roles in agonist-mediated desensitization of G protein-coupled receptors. Although the presence of beta-arrestin subtypes, beta-arrestin-1 and(- 2) in rat brain has been studied extensively, their existence in the spinal cord has not been described. In the current study, we performed immunohistochemical analyses of beta-arrestins at both light and electron microscopic levels using rat lumbar 1-2 spinal cord segments. Intense immunoreactivity for beta-arrestin-1 was found in the motoneurons in lamina IX of the ventral horn and elongated cells in the dorsal nucleus of Clarke. Modest immunoreactivity was detected among the neurons of laminae V and VII/VIII, and weaker immunoreactivity in laminae III, IV, and X. beta-arrestin-2 immunoreactivity was also distributed through laminae III-X in the order of IX > dorsal nucleus of Clarke > V > VII/VIII > IV > III > X. Laminae I and II did not show immunoreactivity. At the electron microscopic level, both beta-arrestin-immunoreactive and nonimmunoreactive dendrites were observed, whereas axons and terminal boutons were devoid of immunoreactivity. In immunoreactive dendrites most beta-arrestin immunoreactivity was distributed throughout the cytoplasm, demonstrating their association with microtubules. In addition, strong immunoreactivity was often found at postsynaptic densities. Our results thus suggest beta-arrestins' possible involvement in both motor and sensory mechanisms at the postsynaptic level in rat lumbar spinal cord.     

Subcellular and subsynaptic localization of group I metabotropic glutamate receptors in the monkey subthalamic nucleus

Both subtypes of group I metabotropic glutamate receptor, mGluR1 and mGluR5, are expressed postsynaptically in neurons of the subthalamic nucleus (STN), and their activation induces different physiological responses. To test whether these effects could be explained by a differential localization of the two group I mGluRs, we analyzed the subcellular and subsynaptic distribution of mGluR1a and mGluR5 in the monkey STN. Double-immunofluorescence and light microscopic analyses revealed that both group I mGluR subtypes were strongly coexpressed in the neuropil and neuronal perikarya. Astrocytic perikarya exhibited intense mGluR1a, but no detectable mGluR5, immunoreactivity. At the electron microscopic level, immunoperoxidase labeling for both mGluR1a and mGluR5 was localized mainly in dendrites. A significant proportion of the total pool of mGluR1a-immunoreactive elements was accounted for by glial cell processes, whereas glial cell labeling was much less frequently encountered in sections immunostained for mGluR5. Preembedding immunogold labeling in STN dendrites revealed that 60-70% of the gold labeling for both mGluR subtypes was intracellular, whereas 30-40% was apposed to the plasma membrane. Of the plasma membrane-apposed particles, more than 90% were extrasynaptic; fewer than 10% were associated with symmetric or asymmetric synapses. Most of the synapse-associated labeling was found at the edges of both asymmetric and symmetric postsynaptic specializations. Some extrasynaptic gold particles were aggregated on parts of the plasma membrane tightly apposed by glial processes. These findings demonstrate that mGluR1a and mGluR5 exhibit a similar pattern of subsynaptic localization in monkey STN neurons, with both receptor subtypes exhibiting substantial extrasynaptic and perisynaptic localization.     

Modulation of calcium channels by group I and group II metabotropic glutamate receptors in dentate gyrus neurons from patients with temporal lobe epilepsy

PURPOSE: Metabotropic glutamate receptors (mGluRs) might be promising new drug targets for the treatment of epilepsy because the expression of certain mGluRs is regulated in epilepsy and because activation of mGluRs results in distinctive anti- and proconvulsant effects. Therefore, we examined how mGluR activation modulates high-voltage-activated (HVA) Ca2+ channels. METHODS: Whole-cell patch-clamp recordings were obtained from granule cells and interneuron-like cells acutely isolated from the dentate gyrus of patients with pharmacoresistent temporal lobe epilepsy. RESULTS: Agonists selective for either group I or group II mGluRs rapidly and reversibly reduced HVA currents in most dentate gyrus cells. These modulatory effects were inhibited by the respective group I and group II mGluR antagonists. The specific Ca2+ channel antagonists nifedipine and omega-conotoxin GVIA potently occluded the effects of group I and II mGluR agonists, respectively, indicating that group I mGluRs acted on L-type channels and group II mGluRs affected N-type channels. About two thirds of the responsive neurons were sensitive either to group I or group II mGluRs, whereas a minority of cells showed effects to agonists of both groups, indicating a variable mGluR expression pattern. CONCLUSIONS: Group I and group II mGluRs are expressed in human dentate gyrus neurons and modulate L- and N-type HVA channels, respectively. The data shed light on the possible cellular sequelae of the mGluR1 upregulation observed in human epileptic dentate gyrus as well as on possible mGluR-mediated anticonvulsant mechanisms.     

Glutamate inhibits thalamic reticular neurons.

Activation of metabotropic glutamate receptors (mGluRs) can result in long-lasting modulation of neuronal excitability. Multiple mGluR subtypes are localized within the rat thalamic reticular nucleus (TRN), and we have examined the effects of activating these different receptor subtypes on the excitability of these neurons using an in vitro slice preparation. Typical of most mGluR-sensitive preparations, the general mGluR agonist, (+/-)-1-aminocyclopentane-trans-1,3-dicarboxylic acid (ACPD) produced a robust, long-lasting excitatory response. Surprisingly, ACPD produced a membrane hyperpolarization in some neurons. Using selective mGluR agonists, we found that activation of group II mGluRs produces the hyperpolarization, whereas the depolarization is mediated by group I mGluRs. While the polarity of the postsynaptic response (hyperpolarization vs depolarization) was dependent on the mGluR subtype activated, both actions appear to result from modification of a linear K(+) conductance. The inhibitory action of Glutamate, via group II mGluRs, provides an avenue for a disinhibitory effect that could have interesting consequences upon a well-investigated, model neuronal circuit, turning its assumed functional role upside down.     

Colocalization of metabotropic glutamate receptors in rat dorsal root ganglion cells

Glutamate is the main excitatory transmitter in both central and peripheral nervous systems. Discovery of metabotropic glutamate receptors (mGluRs) made it clear that glutamate can have excitatory or inhibitory effects on neuronal function, with group I mGluRs enhancing cell excitability but group II and III mGluRs decreasing excitability. The present study investigated the colocalization of mGluR subtypes representing groups I, II, or III in rat L5 dorsal root ganglion (DRG) cells. The analyses show that group III has the highest expression, with 75.0% of DRG cells expressing mGluR8, followed by group II, with 51.6% expressing mGluR2/3, followed by group I, with only 6.8% expressing mGluR1alpha. mGluR8 is expressed by small, medium, and large diameter cells. In contrast, mGluR1alpha and mGluR2/3 are expressed by mainly small and medium cells. Approximately half of cells expressing group I mGluR1alpha also express either group II mGluR2/3 or group III mGluR8. These mGluR1alpha double-labeled populations are not likely to overlap since >1.0% of mGluR1alpha are triple-labeled. As expected from the high percentage of single-labeled mGluR2/3 and mGluR8 cells, there is a considerable population of double-labeled cells with approximately 30% of each population expressing both receptors. Due to the fact that the number of mGluR1alpha-expressing cells in the DRG is low, the percentage of triple-labeled cells is also low ( approximately 1-2%). The prevalence of groups II and III indicate that glutamate could have a substantial inhibitory effect of primary afferent function, reducing and/or fine-tuning sensory input before transmission to the spinal cord. These anatomical data highlight the potential inhibitory role glutamate may play in peripheral sensory transmission.     

Immunocytochemical localization of the mGluR1b metabotropic glutamate receptor in the rat hypothalamus

The mGluR1 metabotropic glutamate receptor is a G-protein-coupled receptor that exists as different C-terminal splice variants. When expressed in mammalian cells, the mGluR1 splice variants exhibit diverse transduction mechanisms and also slightly differ in their apparent agonist affinities. In the present study, we used an affinity-purified antiserum, specifically reactive to the mGluR1b splice variant, in combination with a highly sensitive preembedding immunocytochemical method for light microscopy to investigate the distribution of this receptor in the rat hypothalamus. An intense immunoreactivity for mGluR1b was observed in distinct hypothalamic nuclei. Thus, neuronal cell bodies and dendrites were stained in the preoptic area, suprachiasmatic nucleus, dorsal hypothalamus, lateral hypothalamus, dorsomedial nucleus, tuberomammilary nucleus, and lateral mammilary body. The ventromedial nucleus exhibited neuropil immunostaining but neuronal cell bodies were not labeled. Strong mGluR1b immunoreactivity was observed in magnocellular neurons of the neuroendocrine supraoptic, paraventricular, and arcuate nuclei. Also, neuronal cell bodies were heavily labeled in the retrochiasmatic nucleus, anterior commissural nucleus, and periventricular nucleus. These immunocytochemical observations, together with previous studies, suggest that mGluR1b is coexpressed with other class I mGluRs in some nuclei throughout the hypothalamus. However, mGluR1b is so far the only receptor of this class strongly expressed in the supraoptic, paraventricular, and arcuate nuclei, which might have relevant implications in the physiological control of the neuroendocrine hypothalamic-pituitary system. J. Comp. Neurol. 390:225–233, 1998. © 1998 Wiley-Liss, Inc.     

Synaptic relationship of the neurons containing a metabotropic glutamate receptor, MGluR5, with nociceptive primary afferent and GABAergic terminals in rat spinal superficial laminae

Recent pharmacological evidence showed that metabotropic glutamate receptors (mGluRs), particularly mGluRs1/5, had a potential role in spinal nociceptive processing. However, previous morphological studies on mGluRs have been limited mainly to their distribution in the spinal cord. In the present study, electron microscopic immunocytochemistry was employed to identify the synaptic relationship of the neurons containing mGluR5, with nociceptive primary afferent and gamma-aminobutyric acid-ergic (GABAergic) terminals in the superficial dorsal horn of the spinal cord. Nociceptive C- and A(delta)-primary afferent terminals selectively labeled with horseradish peroxidase conjugated to wheat-germ agglutinin were in asymmetric synaptic contacts with or in direct apposition to mGluR5 positive dendritic profiles. The double-labeling studies revealed that mGluR5 immunoreactive dendrites also received symmetric synaptic contacts from axon terminals labeled with immunogold particles indicating GABA. The present demonstration of mGluR5 neurons receiving inputs from both nociceptive primary afferents and GABAergic terminals of presumed interneurons further supports the involvement of mGluR5 in the transmission and modulation of nociceptive information in the spinal cord.