Metabotropic Receptor mGluR1 Research Articles

Lack of mGluR6-related cascade elements leads to retrograde trans-synaptic effects on rod photoreceptor synapses via matrix-associated proteins.

Heterotrimeric G-proteins couple metabotropic receptors to downstream effectors. In retinal ON bipolar cells, Go couples the metabotropic receptor mGluR6 to the TRPM1 channel and closes it in the dark, thus hyperpolarizing the cell. Light, via GTPase-activating proteins, deactivates Go , opens TRPM1 and depolarizes the cell. Go comprises Gαo1 , Gβ3 and Gγ13; all are necessary for efficient coupling. In addition, Gβ3 contributes to trafficking of certain cascade proteins and to maintaining the synaptic structure. The goal of this study was to determine the role of Gαo1 in maintaining the cascade and synaptic integrity. Using mice lacking Gαo1 , we quantified the immunostaining of certain mGluR6-related components. Deleting Gαo1 greatly reduced staining for Gβ3, Gγ13, Gβ5, RGS11, RGS7 and R9AP. Deletion of Gαo1 did not affect mGluR6, TRPM1 or PCP2. In addition, deleting Gαo1 reduced the number of rod bipolar dendrites that invaginate the rod terminal, similar to the effect seen in the absence of mGluR6, Gβ3 or the matrix-associated proteins, pikachurin, dystroglycan and dystrophin, which are localized presynaptically to the rod bipolar cell. We therefore tested mice lacking mGluR6, Gαo1 and Gβ3 for expression of these matrix-associated proteins. In all three genotypes, staining intensity for these proteins was lower than in wild type, suggesting a retrograde trans-synaptic effect. We propose that the mGluR6 macromolecular complex is connected to the presynaptic rod terminal via a protein chain that includes the matrix-associated proteins. When a component of the macromolecular chain is missing, the chain may fall apart and loosen the dendritic tip adherence within the invagination.

An Ultrastructural and Immunohistochemical Analysis of the Outer Plexiform Layer of the Retina of the European Silver Eel (Anguilla anguilla L)

Here we studied the ultrastructural organization of the outer retina of the European silver eel, a highly valued commercial fish species. The retina of the European eel has an organization very similar to most vertebrates. It contains both rod and cone photoreceptors. Rods are abundantly present and immunoreactive for rhodopsin. Cones are sparsely present and only show immunoreactivity for M-opsin and not for L-, S- or UV-cone opsins. As in all other vertebrate retinas, Müller cells span the width of the retina. OFF-bipolar cells express the ionotropic glutamate receptor GluR4 and ON-bipolar cells, as identified by their PKCα immunoreactivity, express the metabotropic receptor mGluR6. Both the ON- and the OFF-bipolar cell dendrites innervate the cone pedicle and rod spherule. Horizontal cells are surrounded by punctate Cx53.8 immunoreactivity indicating that the horizontal cells are strongly electrically coupled by gap-junctions. Connexin-hemichannels were found at the tips of the horizontal cell dendrites invaginating the photoreceptor synapse. Such hemichannels are implicated in the feedback pathway from horizontal cells to cones. Finally, horizontal cells are surrounded by tyrosine hydroxylase immunoreactivity, illustrating a strong dopaminergic input from interplexiform cells.

Glutamate Receptors in Human Caudate Nucleus in Normal Aging and Alzheimer’s Disease

Neostriatum is one of the brain areas that are not primarily affected in Alzheimer's disease, according to classic regard of the disease. However, recent data emphasize the involvement of neostriatum, especially the head of the caudate nucleus, in the emergence of characteristic symptoms of the disease. Glutamatergic neurotransmission is a key component of striatal pathways. The present study is focused on glutamate receptors of striatal neurons on human caudate nucleus in normal aging and Alzheimer's disease. Immunohistochemical studies were carried out for N-methyl-D-aspartate receptor subunit 1 (NMDAR1), α -amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor subunit 2 (GluR2) and metabotropic glutamate receptor 5 (mGluR5). Ionotropic receptors (NMDAR1 and GluR2) were found to be expressed by 82% - 93% of striatal neurons with no significant alterations in aging and Alzheimer's disease. On the other hand, metabotropic receptor mGluR5 was found to be expressed by just 40% of striatal neurons in young individuals, with significant intensity variations among the neurons. This percent was increased in elderly individuals and Alzheimer's disease patients to 80% and 92% of striatal neurons, respectively. The up-regulation of mGluR5 both in normal aging and Alzheimer's disease is possibly associated with reorganization of neuronal connections, indicates the complexity of this receptor function and renders quite unpredictable the intervention and treatment of dementia with mGluR5 inhibitors or modulators.

Preferential Localization of Muscarinic M1Receptor on Dendritic Shaft and Spine of Cortical Pyramidal Cells and Its Anatomical Evidence for Volume Transmission

Acetylcholine (ACh) plays important roles for higher brain functions, including arousal, attention, and cognition. These effects are mediated largely by muscarinic acetylcholine receptors (mAChRs). However, it remains inconclusive whether the mode of ACh-mAChR signaling is synaptic, so-called "wired," transmission mediated by ACh released into the synaptic cleft, or nonsynaptic, so-called "volume," transmission by ambient ACh. To address this issue, we examined cellular and subcellular distribution of M(1), the most predominant mAChR subtype in the cerebral cortex and hippocampus, and pursued its anatomical relationship with cholinergic varicosities in these regions of adult mice. M(1) was highly expressed in glutamatergic pyramidal neurons, whereas it was low or undetectable in various GABAergic interneuron subtypes. M(1) was preferentially distributed on the extrasynaptic membrane of pyramidal cell dendrites and spines. Cholinergic varicosities often made direct contact to pyramidal cell dendrites and synapses. At such contact sites, however, synapse-like specialization was infrequent, and no particular accumulation was found at around contact sites for both M(1) and presynpatic active zone protein Bassoon. These features contrasted with those of the glutamatergic system, in which AMPA receptor GluA2 and metabotropic receptor mGluR5 were recruited to the synaptic or perisynaptic membrane, respectively, and Bassoon was highly accumulated in the presynaptic terminals. These results suggest that M(1) is so positioned to sense ambient ACh released from cholinergic varicosities at variable distances, and to enhance the synaptic efficacy and excitability of pyramidal cells. These molecular-anatomical arrangements will provide the evidence for volume transmission, at least in M(1)-mediated cortical cholinergic signaling.

Non-NMDA receptors in frog retina: An immunocytochemical study

Glutamate is one of the main neurotransmitters in the retina. Its effects are mediated by a large number of ionotropic and metabotropic membrane receptors. The distribution of ionotropic AMPA receptor subunits GluR1-4, kainate receptor subunits GluR5-7 and KA2, delta receptors 1-2, as well as the metabotropic receptor mGluR6 were studied in the frog retina. Indirect immunofluorescence was used to localize the different receptor subunits. Results showed that all subunits, with the exception of GluR1 and GluR5, are widely distributed in the retina. They are mainly located in both plexiform layers: the outer (OPL) and the inner one (IPL), where punctate labelling, a sign of synaptic localization, is observed. The metabotropic receptor mGluR6 is localised only in the OPL. The AMPA receptor subunit GluR4 is localised on the glial Müller cells of the retina. The vast majority of the subunits possess specific patterns of labelling that indicate that they are involved with different retinal functions. The significance of the AMPA receptors and involvement of glia in modulation of synaptic transmission are discussed.

High-Density Presynaptic Transporters Are Required for Glutamate Removal from the First Visual Synapse

Reliable synaptic transmission depends not only on the release machinery and the postsynaptic response mechanism but also on removal or degradation of transmitter from the synaptic cleft. Accumulating evidence indicates that postsynaptic and glial excitatory amino acid transporters (EAATs) contribute to glutamate removal. However, the role of presynaptic EAATs is unclear. Here, we show in the mouse retina that glutamate is removed from the synaptic cleft at the rod to rod bipolar cell (RBC) synapse by presynaptic EAATs rather than by postsynaptic or glial EAATs. The RBC currents evoked by electrical stimulation of rods decayed slowly after pharmacological blockade of EAATs. Recordings of the evoked RBC currents from EAAT subtype-deficient mice and the EAAT-coupled anion current reveal that functional EAATs are localized to rod terminals. Model simulations suggest that rod EAATs are densely packed near the release site and that rods are equipped with an almost self-sufficient glutamate recollecting system.

Regulation of the Retinal Bipolar Cell mGluR6 Pathway by Calcineurin

Glutamate produces a hyperpolarizing postsynaptic potential in ON bipolar cells by binding to the metabotropic receptor mGluR6 and subsequently closing a cation-selective channel. It has been proposed that Ca(2+) influx through the cation channel triggers a depression of the synaptic potential. Here we report that this Ca(2+)-mediated depression requires activation of calcineurin, a Ca(2+)/calmodulin-regulated phosphatase. We measured glutamate-evoked currents (I(glu)) with whole cell recordings of ON bipolar cells in light-adapted retinal slices. Depression of I(glu) by Ca(2+) was prevented by inhibitors of calcineurin or by tightly buffering Ca(2+) with bis-(o-aminophenoxy)-N,N,N',N'-tetraacetic acid (BAPTA). However, when cells were dialyzed with BAPTA and a Ca(2+)-independent form of calcineurin (CaN420), depression of I(glu) was restored. Similarly, CaN420 induced depression of I(glu) during continuous glutamate application, a protocol that ordinarily prevents depression. Analysis of changes in the amplitude of the cation-selective current (I(cat)) of cells that were dialyzed with high Ca(2+) (1 microM), or with BAPTA and CaN420, indicates that Ca(2+) depresses I(glu) by reducing I(cat) and that calcineurin acts via the same mechanism. Ca(2+)-mediated depression of I(glu) was not found to involve CaMKII, as inhibitors of CaMKII did not prevent this depression nor did they affect the sensitivity of the response to small changes in the concentration of mGluR6 agonist. Our data suggest that Ca(2+) and calcineurin may play an adaptive role at the synapse between photoreceptor and ON bipolar cells, closing postsynaptic cation channels that are opened by a drop in synaptic glutamate levels during prolonged photoreceptor illumination.

The Metabotropic Receptor mGluR6 May Signal Through Go, But Not Phosphodiesterase, in Retinal Bipolar Cells

Bipolar cells are retinal interneurons that receive synaptic input from photoreceptors. Glutamate, the photoreceptor transmitter, hyperpolarizes On bipolar cells by closing nonselective cation channels, an effect mediated by the metabotropic receptor mGluR6. Previous studies of mGluR6 transduction have suggested that the receptor couples to a phosphodiesterase (PDE) that preferentially hydrolyzes cGMP, and that cGMP directly gates the nonselective cation channel. This hypothesis was tested by dialyzing On bipolar cells with nonhydrolyzable analogs of cGMP. Whole-cell recordings were obtained from On bipolar cells in slices of larval tiger salamander retina. Surprisingly, On bipolar cells dialyzed with 8-(4-chlorophenylthio)-cyclic GMP (8-pCPT-cGMP), or 8-bromo-cyclic GMP (8-Br-cGMP) responded normally to glutamate or L-2-amino-4-phosphonobutyrate (L-APB). Response amplitudes and kinetics were not significantly altered compared with cells dialyzed with cGMP alone. Comparable results were obtained with the PDE inhibitor 3-isobutyl-1-methyl-xanthine (IBMX) or with 8-pCPT-cGMP and IBMX together, indicating that PDE is not required for mGluR6 signal transduction. Addition of the G-protein subunit G(o)alpha to the pipette solution suppressed the cation current and occluded the glutamate response, whereas dialysis with G(i)alpha or with transducin Gbetagamma had no significant effect on either the cation current or the response. Dialysis of an antibody directed against G(o)alpha also reduced the glutamate response, indicating a functional role for endogenous G(o)alpha. These results indicate that mGluR6 may signal through G(o), rather than a transducin-like G-protein.

Neuroadaptations in Ionotropic and Metabotropic Glutamate Receptor mRNA Produced by Cocaine Treatment

The expression of glutamate receptor/subunit mRNAs was examined 3 weeks after discontinuing 1 week of daily injections of saline or cocaine. The level of mRNA for GluR1-4, NMDAR1, and mGluR5 receptors was measured with in situ hybridization and RT-PCR. In nucleus accumbens, acute cocaine treatment significantly reduced the mRNA level for GluR3, GluR4, and NMDAR1 subunits, whereas repeated cocaine reduced the level for GluR3 mRNA. Acute cocaine treatment also reduced the NMDAR1 mRNA level in dorsolateral striatum and ventral tegmental area. In prefrontal cortex, repeated cocaine treatment significantly increased the level of GluR2 mRNA. The GluR2 mRNA level was not changed by acute or repeated cocaine in any other brain regions examined. Repeated cocaine treatment also significantly increased mGluR5 mRNA levels in nucleus accumbens shell and dorsolateral striatum. Functional properties of the ionotropic glutamate receptors are determined by subunit composition. In addition, metabotropic glutamate receptors can modulate synaptic transmission and the response to stimulation of ionotropic receptors. Thus, the observed changes in levels of AMPA and NMDA receptor subunits and the mGluR5 metabotropic receptor may alter excitatory neurotransmission in the mesocorticolimbic dopamine system, which could play a significant role in the enduring biochemical and behavioral effects of cocaine.

Aminobicyclo[2.2.1.]heptane dicarboxylic acids (ABHD), rigid analogs of ACPD and glutamic acid: synthesis and pharmacological activity on metabotropic receptors mGluR1 and mGluR2

Isomeric norbornane-derived rigid analogs mimicking different potential conformations of ACPD (1-aminocyclopentane-1,3-dicarboxylic acid) and glutamic acid have been synthesized, via the hydantoin route, to be used as conformational probes for bioactive conformations at the glutamatergic receptors of the central nervous system. Activities on metabotropic receptors mGluR1 and mGluR2 are reported and discussed.

Regulation of cGMP-dependent current in On bipolar cells by calcium/calmodulin-dependent kinase.

The metabotropic receptor mGluR6 is localized to the dendrites of On bipolar cells and mediates synaptic input from photoreceptors. The binding of glutamate to the receptor activates a phosphodiesterase (PDE), which then hydrolyzes cGMP. A nonselective cationic conductance, believed to be gated directly by cGMP, is turned off as a result of the fall in cGMP levels, and the cell hyperpolarizes. Here we present evidence for regulation of the conductance by an additional mechanism that it is independent of cGMP. Whole-cell recordings were obtained from On bipolar cells in slices of tiger salamander retina. Dialysis of cells with 1 microM KN-62 or 10 microM KN-93, two inhibitors of type II calmodulin-dependent protein kinase (CaMKII), depressed cGMP-dependent currents. This depression persisted when hydrolysis of cGMP was prevented with IBMX, a broad-spectrum PDE inhibitor, suggesting that CaMKII acts downstream from the PDE in the cascade. The depression of cGMP-dependent currents was probably not due to a direct interaction of the inhibitors with the channels as neither 1 microM KN-62 or 10 microM KN-93 was found to have any effect on cyclic nucleotide-gated channels when applied directly to excised patches of rod outer segments. We propose that phosphorylation by CaMKII may be an important mechanism for regulating the cGMP-dependent conductance of On bipolar cells.

ON cone bipolar cells in rat express the metabotropic receptor mGluR6.

The rod bipolar cell and about five types of ON cone bipolar cells depolarize to light by employing a sign-reversing metabotropic glutamate receptor. Glutamate responses are similar in both rod bipolar and cone bipolar cells, but the receptor mediating this response (mGluR6) was so far demonstrated only in rod bipolar cells. To test if ON cone bipolar cells also express mGluR6, we immunoreacted rat retina with an antibody specific for mGluR6, and studied the staining from serial ultrathin sections. We demonstrate that mGluR6 is indeed expressed in the dendritic tips of cone bipolar cells, the majority of which receive a ribbon synapse, and thus probably are ON cone bipolar cells. We further show that half of the dendritic tips contacting the cones stain for mGluR6, thus implying that all ON cone bipolar cell types express mGluR6.