AMPA Glutamate Receptor Subunits GluR1 Research Articles

Reinforcement-Related Regulation of AMPA Glutamate Receptor Subunits in the Ventral Tegmental Area Enhances Motivation for Cocaine

Chronic cocaine use produces numerous biological changes in brain, but relatively few are functionally associated with cocaine reinforcement. Here we show that daily intravenous cocaine self-administration, but not passive cocaine administration, induces dynamic upregulation of the AMPA glutamate receptor subunits GluR1 and GluR2 in the ventral tegmental area (VTA) of rats. Increases in GluR1 protein and GluR1(S845) phosphorylation are associated with increased GluR1 mRNA in self-administering animals, whereas increased GluR2 protein levels occurred despite substantial decreases in GluR2 mRNA. We investigated the functional significance of GluR1 upregulation in the VTA on cocaine self-administration using localized viral-mediated gene transfer. Overexpression of GluR1(WT) in rat VTA primarily infected dopamine neurons (75%) and increased AMPA receptor-mediated membrane rectification in these neurons with AMPA application. Similar GluR1(WT) overexpression potentiated locomotor responses to intra-VTA AMPA, but not NMDA, infusions. In cocaine self-administering animals, overexpression of GluR1(WT) in the VTA markedly increased the motivation for cocaine injections on a progressive ratio schedule of cocaine reinforcement. In contrast, overexpression of protein kinase A-resistant GluR1(S845A) in the VTA reduced peak rates of cocaine self-administration on a fixed ratio reinforcement schedule. Neither viral vector altered sucrose self-administration, and overexpression of GluR1(WT) or GluR1(S845A) in the adjacent substantia nigra had no effect on cocaine self-administration. Together, these results suggest that dynamic regulation of AMPA receptors in the VTA during cocaine self-administration contributes to cocaine addiction by acting to facilitate subsequent cocaine use.

Expression of glutamate receptors and potassium channels in vestibular nuclei neurons during development of signal processing

The principal cells of the chick tangential vestibular nucleus offer a simple neuron model to study signal processing in second-order, vestibular reflex projection neurons. The principal cells represent a relatively uniform population of vestibular nuclei neurons which receive a major input from the primary vestibular fibers and send axons to targets mainly involved in the vestibuloocular reflexes. Here, studies performed on ion channels involved in the emergence and establishment of signal processing in this morphologically-identified subset of vestibular nuclei neurons are reviewed, including the AMPA glutamate receptor subunits GluR1, GluR2, GluR3, and GluR4 and the potassium channel subunits Kv1.1 and Kv1.2.

Nicotine Regulates Multiple Synaptic Proteins by Inhibiting Proteasomal Activity

Ubiquitination regulates the degradation, membrane trafficking, and transcription of proteins. At mammalian synapses, the ubiquitin-proteasome system (UPS) influences synaptic transmission and plasticity. Nicotine also has the ability to affect synaptic function via mechanisms that remain partially unknown. We found that nicotine, at concentrations achieved by smokers, reduced proteasomal activity, produced accumulation of ubiquitinated synaptic proteins, and increased total protein levels. In particular, a 24 h exposure to nicotine decreased proteasome-dependent degradation of the alpha7 nicotinic acetylcholine receptor (nAChR) subunit, as indicated by the accumulation of ubiquitinated alpha7. The same nicotine treatment increased the levels of the AMPA glutamate receptor subunit GluR1, the NMDA receptor subunit NR2A, the metabotropic receptor mGluR1alpha, the plasticity factor Homer-1A, and the scaffolding postsynaptic density protein PSD-95, whereas the levels of another scaffolding protein, Shank, were reduced. These changes were associated with an inhibition of proteasomal chymotrypsin-like activity by nicotine. The nAChR antagonist mecamylamine was only partially able to block the effects of nicotine on the UPS, indicating that nAChR activation does not completely explain nicotine-induced inhibition of proteasomal catalytic activity. A competition binding assay suggested a direct interaction between nicotine and the 20S proteasome. These results suggest that the UPS might participate in nicotine-dependent synaptic plasticity.

Glutamate receptor subunit expression in the rhesus macaque locus coeruleus

The locus coeruleus (LC) is a major noradrenergic brain nucleus that regulates states of arousal, optimizes task-oriented decision making, and may also play an important role in modulating the activity of the reproductive neuroendocrine axis. Rodent studies have shown that the LC is responsive to glutamate receptor agonists, and that it expresses various glutamate receptor subunits. However, glutamate receptor subunit expression has not been extensively examined in the primate LC. We previously demonstrated expression of the NR1 NMDA glutamate receptor subunit in the rhesus macaque LC and now extend this work by also examining the expression of non-NMDA (AMPA and kainate) ionotropic glutamate receptor subunits. Using in situ hybridization histochemistry and immunohistochemistry, we confirmed the presence of the obligatory NR1 subunit in the LC. In addition, we demonstrated expression of the AMPA glutamate receptor subunits GluR1, GluR2, and GluR3. More extensive receptor profiling, using rhesus monkey gene microarrays (Affymetrix GeneChip®), further corroborated the histological findings and showed expression of mRNA encoding ionotropic glutamate receptor subunits NR2A, NR2D, GluR4, and GluR6, as well as the metabotropic glutamate receptor subunits mGluR1, mGluR3, mGluR4, mGluR5, and mGluR7. These data provide a foundation for future examination of how changes in glutamate receptor composition contribute to the control of primate physiology.

Optineurin Negatively Regulates TNFα- Induced NF-κB Activation by Competing with NEMO for Ubiquitinated RIP

NF-kappaB essential modulator (NEMO), the regulatory subunit of the IkappaB kinase (IKK) that activates NF-kappaB, is essential for NF-kappaB activation. NEMO was recently found to contain a region that preferentially binds Lys (K)63-linked but not K48-linked polyubiquitin (polyUb) chains, and the ability of NEMO to bind to K63-linked polyUb RIP (receptor-interacting protein) is necessary for efficient tumor necrosis factor alpha (TNFalpha)-induced NF-kappaB activation. Optineurin is a homolog of NEMO, and mutations in the optineurin gene are found in a subset of patients with glaucoma, a neurodegenerative disease involving the loss of retinal ganglion cells. Although optineurin shares considerable homology with NEMO, in resting cells, it is not present in the high-molecular-weight complex containing IKKalpha and IKKbeta, and optineurin cannot substitute for NEMO in lipopolysaccharide (LPS)-induced NF-kappaB activation. On the other hand, the overexpression of optineurin blocks the protective effect of E3-14.7K on cell death caused by the overexpression of TNFalpha receptor 1 (TNFR1). Here we show that optineurin has a K63-linked polyUb-binding region similar to that of NEMO, and like NEMO, it bound K63- but not K48-linked polyUb. Optineurin competitively antagonized NEMO's binding to polyUb RIP, and its overexpression inhibited TNFalpha-induced NF-kappaB activation. This competition occurs at physiologic protein levels because microRNA silencing of optineurin resulted in markedly enhanced TNFalpha-induced NF-kappaB activity. These results reveal a physiologic role for optineurin in dampening TNFalpha signaling, and this role might provide an explanation for its association with glaucoma.

Regulation of Drug Reward by cAMP Response Element-Binding Protein: Evidence for Two Functionally Distinct Subregions of the Ventral Tegmental Area

The transcription factor cAMP response element binding protein (CREB) is implicated in the actions of drugs of abuse in several brain areas, but little information is available about a role for CREB in the ventral tegmental area (VTA), one of the key reward regions of the brain. Here, we demonstrate that chronic exposure to drugs of abuse induces CREB activity throughout the VTA. Using viral-mediated gene transfer, we expressed green fluorescent protein (GFP)-tagged CREB or mCREB (a dominant-negative form of CREB) in the VTA and, using a conditioned place-preference paradigm, found that CREB activation within the rostral versus caudal subregions of the VTA produces opposite effects on drug reward. We identified VTA subregion-specific differences in the proportion of dopaminergic and GABAergic neurons and in the dopaminergic projections to the nucleus accumbens, another brain region implicated in drug reward, and suggest that this may contribute to behavioral differences in this study. We also measured expression levels of tyrosine hydroxylase and the AMPA glutamate receptor subunit GluR1, both of which are known to contribute to drug reward in the VTA, and found that both of these genes are upregulated following the expression of CREB-GFP and downregulated following expression of mCREB-GFP, raising the possibility that CREB may exert its effects on drug reward, in part, via regulation of these genes. These results suggest a novel role for CREB in mediating drug-induced plasticity in the VTA and establish two functionally distinct subregions of the VTA in which CREB differentially regulates drug reward.

The Hypothalamic Neuropeptide Melanin-Concentrating Hormone Acts in the Nucleus Accumbens to Modulate Feeding Behavior and Forced-Swim Performance

Melanin-concentrating hormone (MCH) is a hypothalamic neuropeptide with a prominent role in feeding and energy homeostasis. The rodent MCH receptor (MCH1R) is highly expressed in the nucleus accumbens shell (AcSh), a region that is important in the regulation of appetitive behavior. Here we establish a role for MCH and MCH1R in mediating a hypothalamic-limbic circuit that regulates feeding and related behaviors. Direct delivery of an MCH1R receptor antagonist to the AcSh blocked feeding and produced an antidepressant-like effect in the forced swim test, whereas intra-AcSh injection of MCH had the opposite effect. Expression studies demonstrated that MCH1R is present in both the enkephalin- and dynorphin-positive medium spiny neurons of the AcSh. Biochemical analysis in AcSh explants showed that MCH signaling blocks dopamine-induced phosphorylation of the AMPA glutamate receptor subunit GluR1 at Ser845. Finally, food deprivation, but not other stressors, stimulated cAMP response element-binding protein-dependent pathways selectively in MCH neurons of the hypothalamus, suggesting that these neurons are responsive to a specific set of physiologically relevant conditions. This work identifies a novel hypothalamic-AcSh circuit that influences appetitive behavior and mediates the antidepressant activity of MCH1R antagonists.

Structurally dissimilar antimanic agents modulate synaptic plasticity by regulating AMPA glutamate receptor subunit GluR1 synaptic expression.

A growing body of data from clinical and preclinical studies suggests that the glutamatergic system may represent a novel therapeutic target for severe recurrent mood disorders. Since synapse-specific glutamate receptor expression/localization is known to play critical roles in synaptic plasticity, we investigated the effects of mood stabilizers on AMPA receptor expression. Rats were treated chronically with lithium or valproate, hippocampal synaptosomes were isolated, and GluR1 levels were determined. Additionally, hippocampal neurons were prepared from E18 rat embryos and treated with lithium or valproate. Surface expression of GluR1 was determined using a biotinylation assay, and double-immunostaining with anti-GluR1 and anti-synaptotagmin antibodies was used to determine synaptic GluR1 levels. The AMPA receptor subunit GluR1 expression in hippocampal synaptosomes was significantly reduced by both chronic lithium and valproate. Overall, these studies show that AMPA receptor subunit GluR1 is a common target for two structurally highly dissimilar, but highly efficacious, mood stabilizers, lithium and valproate. These studies suggest that regulation of glutamatergically mediated synaptic plasticity may play a role in the treatment of mood disorders, and raise the possibility that agents more directly affecting synaptic GluR1 may represent novel therapies for this devastating illness.

Tumor Necrosis Factor α Increases Neuronal Vulnerability to Excitotoxic Necrosis by Inducing Expression of the AMPA–Glutamate Receptor Subunit GluR1 via an Acid Sphingomyelinase- and NF-κB-Dependent Mechanism

Acid sphingomyelinase (ASMase) and NF-κB participate in tumor necrosis factor α (TNFα) signal transduction. Mice in which the genes encoding ASMase or the p50 subunit of NF-κB are disrupted have been reported to be less vulnerable than wild-type mice to focal brain ischemia. We now demonstrate selective diminution in expression of GluR1, an α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate-type glutamate receptor (AMPA-GluR) protein subunit, in these two groups of knockout mice. To confirm that neuronal GluR1 expression is regulated by ASMase and NF-κB, and to learn whether this regulation has pathophysiological significance, we treated cultured human NT2-N neurons with TNFα. This induced GluR1 expression and increased susceptibility of the neurons to kainate necrosis. Both induction of GluR1 and heightened vulnerability to kainate were blocked by inhibiting ASMase or by antisense knockdown of NF-κB p50. We conclude that TNFα can sensitize neurons to excitotoxic necrosis by inducing expression of GluR1 via an ASMase- and NF-κB-dependent mechanism. TNFα levels are frequently elevated during ischemia and other CNS diseases in which excitotoxicity contributes to neuronal loss. Our results suggest that inhibiting TNFα signal transduction will diminish neuronal necrosis in these diseases.

Development of glutamatergic synapses in the rat retina: the postnatal expression of ionotropic glutamate receptor subunits.

We examined the distribution of the AMPA glutamate receptor subunits GluR1 to GluR4, of the kainate receptor subunits GluR6/7 and KA2, and of the glutamate receptor subunits delta1/2, during postnatal development of the rat retina by immunocytochemistry and light microscopy using receptor subunit specific antisera. The various ionotropic glutamate receptor subunits were expressed early in postnatal rat retina, and most of the subunits, with the exception of delta1/2. were found in both synaptic layers of rat retina. The glutamate receptor subunits studied showed differences in their time of appearance, their spatial distribution patterns, and in their expression levels in the developing rat retina. Interestingly, most of the AMPA receptor subunits were expressed earlier than the kainate receptor subunits in the two synaptic layers of the retina, indicating that AMPA glutamate receptors play an important role in early postnatal glutamatergic synaptic transmission. We also studied the ultrastructural localization of the AMPA glutamate receptor subunits GluR1 to GluR4 by immunocytochemistry and electron microscopy in the inner plexiform layer of the mature rat retina. Most of the subunits were found postsynaptic to the ribbon synapses of OFF-cone, ON-cone, and rod bipolar cells. The results of this study suggest an involvement of ionotropic glutamate receptors in processes of synaptic maturation and the formation of synaptic circuitries in the developing plexiform layers of the retina. Furthermore, AMPA and kainate receptors play a role in synaptic processing and in the development of both the scotopic and photopic pathways in the rat retina.

An alternative pathway for rod signals in the rodent retina: rod photoreceptors, cone bipolar cells, and the localization of glutamate receptors.

In the mammalian retina, extensive processing of spatiotemporal and chromatic information occurs. One key principle in signal transfer through the retina is parallel processing. Two of these parallel pathways are the ON- and OFF-channels transmitting light and dark signals. This dual system is created in the outer plexiform layer, the first relay station in retinal signal transfer. Photoreceptors release glutamate onto ON- and OFF-type bipolar cells, which are functionally distinguished by their postsynaptic expression of different types of glutamate receptors, namely ionotropic and metabotropic glutamate receptors. In the current concept, rod photoreceptors connect only to rod bipolar cells (ON-type) and cone photoreceptors connect only to cone bipolar cells (ON- and OFF-type). We have studied the distribution of (RS)-alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) glutamate receptor subunits at the synapses in the outer plexiform layer of the rodent retina by immunoelectron microscopy and serial section reconstruction. We report a non-classical synaptic contact and an alternative pathway for rod signals in the retina. Rod photoreceptors made synaptic contact with putative OFF-cone bipolar cells that expressed the AMPA glutamate receptor subunits GluR1 and GluR2 on their dendrites. Thus, in the retina of mouse and rat, an alternative pathway for rod signals exists, where rod photoreceptors bypass the rod bipolar cell and directly excite OFF-cone bipolar cells through an ionotropic sign-conserving AMPA glutamate receptor.