GluR2 mRNA Research Articles

Roles of YB-1 under arsenite-induced stress: Translational activation of HSP70 mRNA and control of the number of stress granules

BackgroundWhen cells become stressed, they form stress granules (SGs) and show an increase of the molecular chaperone HSP70. The translational regulator YB-1 is a component of SGs, but it is unclear whether it contributes to the translational induction of HSP70 mRNA. Here we examined the roles of YB-1 in SG assembly and translational regulation of HSP70 mRNA under arsenite-induced stress. MethodUsing arsenite-treated NG108-15 cells, we examined whether YB-1 was included in SGs with GluR2 mRNA, a target of YB-1, and investigated the interaction of YB-1 with HSP70 mRNA and its effect on translation of the mRNA. We also investigated the distribution of these mRNAs to SGs or polysomes, and evaluated the role of YB-1 in SG assembly. ResultsArsenite treatment reduced the translation level of GluR2 mRNA; concomitantly, YB-1-bound HSP70 mRNA was increased and its translation was induced. Sucrose gradient analysis revealed that the distribution of GluR2 mRNA was shifted from heavy-sedimenting to much lighter fractions, and also to SG-containing non-polysomal fractions. Conversely, HSP70 mRNA was shifted from the non-polysomal to polysome fractions. YB-1 depletion abrogated the arsenite-responsive activation of HSP70 synthesis, but SGs harboring both mRNAs were still assembled. The number of SGs was increased by YB-1 depletion and decreased by its overexpression. ConclusionIn arsenite-treated cells, YB-1 mediates the translational activation of HSP70 mRNA and also controls the number of SGs through inhibition of their assembly. General significanceUnder stress conditions, YB-1 exerts simultaneous but opposing actions on the regulation of translation via SGs and polysomes.

AMPA receptors mediate passive avoidance deficits induced by sleep deprivation

The present study addressed the effects of sleep deprivation (SD) on AMPA receptor (AMPAR) binding in brain regions associated with learning and memory, and investigated whether treatment with drugs acting on AMPAR could prevent passive avoidance deficits in sleep deprived animals. [3H]AMPA binding and GluR1 in situ hybridization signals were quantified in different brain regions of male Wistar rats either immediately after 96h of sleep deprivation or after 24h of sleep recovery following 96h of sleep deprivation. Another group of animals were sleep deprived and then treated with either the AMPAR potentiator, aniracetam (25, 50 and 100mg/kg, acute administration) or the AMPAR antagonist GYKI-52466 (5 and 10mg/kg, acute and chronic administration) before passive avoidance training. Task performance was evaluated 2h and 24h after training. A significant reduction in [3H]AMPA binding was found in the hippocampal formation of SD animals, while no alterations were observed in GluR1 mRNA levels. The highest dose of aniracetam (100mg/kg) reverted SD-induced impairment of passive avoidance performance in both retention tests, whereas GYKI-52466 treatment had no effect. Pharmacological enhancement of AMPAR function may revert hippocampal-dependent learning impairments produced after SD. We argue that such effects might be associated with reduced AMPAR binding in the hippocampus of sleep deprived animals.

Depletion of serotonin in the basolateral amygdala elevates glutamate receptors and facilitates fear-potentiated startle.

Our previous experiments demonstrated that systemic depletion of serotonin (5-hydroxytryptamine, 5-HT), similar to levels reported in patients with emotional disorders, enhanced glutamateric activity in the lateral nucleus of the amygdala (LA) and potentiated fear behaviors. However, the effects of isolated depletion of 5-HT in the LA, and the molecular mechanisms underlying enhanced glutamatergic activity are unknown. In the present study, we tested the hypothesis that depletion of 5-HT in the LA induces increased fear behavior, and concomitantly enhances glutamate receptor (GluR) expression. Bilateral infusions of 5,7-dihydroxytryptamine (4 μg per side) into the LA produced a regional reduction of serotonergic fibers, resulting in decreased 5-HT concentrations. The induction of low 5-HT in the LA elevated fear-potentiated startle, with a parallel increase in GluR1 mRNA and GluR1 protein expression. These findings suggest that low 5-HT concentrations in the LA may facilitate fear behavior through enhanced GluR-mediated mechanisms. Moreover, our data support a relationship between 5-HT and glutamate in psychopathologies.

An EJC Factor RBM8a Regulates Anxiety Behaviors

Neuroplasticity depends on the precise timing of gene expression, which requires accurate control of mRNA stability and rapid elimination of abnormal mRNA. Nonsense-mediated mRNA decay (NMD) is an RNA surveillance mechanism that ensures the speedy degradation of mRNAs carrying premature termination codons (PTCs). This mechanism relies on several key Exon Junction Complex (EJC) factors to distinguish PTCs from normal stop codons. NMD degrades not only aberrant transcripts carrying PTCs, but also normal transcripts harboring a normal stop codon [1]. Intriguingly, mutations in an NMD factor, Upf3b, have been found in patients with autism [2, 3]. A binding partner of Upf3b, RBM8a, is located in the 1q21.1 copy-number variation (CNV) associated with mental retardation, autism [4], schizophrenia [5], and microcephaly [6]. However, the functions of EJC factors and their roles in behavioral regulation are still elusive. RBM8a protein is a core component of the EJC that plays an important role in NMD. Recent genetic study indicated that RBM8a gain-of-function significantly associated with intellectual disability [7]. In this study we investigated the effect of RBM8a overexpression on affective behaviors in mice. Lentivirus expressing RBM8a was infused into the hippocampus of adult mice to conduct behavioral studies including social interaction, open field, elevated plus maze, and forced swimming tests. Our results showed that overexpression of RBM8a in the mouse dentate gyrus (DG) leads to increased anxiety-like behavior, abnormal social interaction and decreased immobile time in forced swimming test (FST). To examine the underlying mechanism, we found that overexpressing RBM8a in cultured primary neurons lead to significant higher frequency of miniature excitatory postsynaptic currents (mEPSCs). To explore the underlying mechanism of RBM8a mediated behavioral changes, RNA-immunoprecipitation (RNA-IP) detected that RBM8a binds to CaMK2, GluR1 and Egr1 mRNA, suggesting that RBM8a may target neuronal genes to regulate behaviors. This is the first study that demonstrates the key role of RBM8a on the emotional behaviors in mice. These results reveal new neural mechanisms by which NMD modulates behaviors and potentially provide a better understanding of pathophysiology underlying psychiatric disorders.

Increased excitability and molecular changes in adult rats after a febrile seizure

Both early life inflammation and prolonged febrile seizures have been associated with increased excitation in the adult brain. We hypothesized this may be due in part to changes in the cation-chloride cotransporter system. Rat pups received saline or lipopolysaccharide/kainic acid (LPS/KA) resulting in inflammation, followed by a behavioral febrile seizure (FS) in approximately 50% of rats. Adult animals from the saline, inflammation, or inflammation+FS groups underwent the following: (1) in vitro electrophysiologic studies; (2) Western blotting or polymerase chain reaction; or (3) application of the Na-K-Cl cotransporter 1 (NKCC1) blocker bumetanide to determine its effect on reversing increased excitability in vitro. The inflammation and inflammation+FS groups demonstrated increased excitability in vitro and increased hippocampal protein expression of NR2B and GABAA α5 receptor subunits and mRNA expression of NKCC1. The inflammation+FS group also had decreased protein expression of GluR2 and GABAA α1 receptor subunits and mRNA and protein expression of KCC2. Bumetanide decreased in vitro 4-aminopyridine-induced inter-ictal activity in the inflammation and inflammation+FS groups. The results demonstrate early-life inflammation with or without a behavioral FS can lead to long-lasting molecular changes and increased excitability in the adult rat hippocampus, although some changes are more extensive when inflammation is accompanied by behavioral seizure activity. Bumetanide is effective in reversing increased excitability in vitro, providing evidence for a causal role for cation-chloride cotransporters and suggesting this drug may prove useful for treating epilepsy that develops after a FS.

Effects of the chinese traditional prescription xiaoyaosan decoction on chronic immobilization stress-induced changes in behavior and ultrastructure in rat hippocampus.

Xiaoyaosan (XYS) decoction has been widely used as a traditional medicine for treating stress and depression-related disorders in China for thousands of years. Aim of the Study. To observe the potential mechanism of XYS decoction's antidepressant-like effect in α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors related to synaptic plasticity in the hippocampus rats induced by chronic immobilization stress (CIS). Materials and Methods. Animals were randomly divided into five groups: (1) control group; (2) sham-operated group; (3) CIS group, in which rats were conducted CIS for 21 days; (4) XYS decoction treatment group; (5) 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX) positive group, in which the amygdala of CIS rats was unilaterally microinjected with a competitive glutamate receptor antagonist, CNQX. After CIS for 21 days, the open field test (OPT) and elevated plus-maze test (EPM) were measured, the ultrastructure of hippocampus CA1 subregion was observed by the electron microscopy; both the GluR1 and GluR2 mRNA level of AMPA receptor subunits in hippocampus CA1 subregion were detected by real-time qPCR. Results. Rats subjected to CIS exhibited increases in time in central zone and decreases in total distance traveled in the OPT. In the EPM, they also showed decreases in center zone time and entries, open arm time and entries, and an increase in close arm time. Ultrastructural damage in the hippocampus CA1 was also observed. XYS decoction and CNQX showed significant improvement behavioral changes and the ultrastructural damage of the hippocampus CA1; XYS decoction also reversed CIS-induced decreases in GluR2 mRNA and increases in GluR1 mRNA in the hippocampus CA1 as well as CNQX. Conclusions. XYS decoction may effectively produce an antidepressant-like effect, which appears to be involved AMPA receptors related synaptic plasticity of hippocampus.

Connexin and AMPA receptor expression changes over time in the rat olfactory bulb

Circadian rhythms affect olfaction by an unknown molecular mechanism. Independent of the suprachiasmatic nuclei, the mammalian olfactory bulb (OB) has recently been identified as a circadian oscillator. The electrical activity in the OB was reported to be synchronized to a daily rhythm and the clock gene, Period1, was oscillatory in its expression pattern. Because gap junctions composed of connexin36 and α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs) have been reported to work together to synchronize firing of action potentials in the OB, we hypothesized that circadian electrical oscillations could be synchronized by daily changes in the expression of connexins and AMPAR subunits (GluR1–4). We examined the OB for the presence of clock genes by polymerase chain reaction (PCR) and whether Period2, connexins, and AMPARs fluctuated across the light/dark cycle by quantitative PCR or SDS–PAGE/Western blot analysis. We observed significant changes in the messenger RNA and protein expression of our targets across 24 or 48h. Whereas most targets were rhythmic by some measures, only GluR1 mRNA and protein were both rhythmic by the majority of our tests of rhythmicity across all time scales. Differential expression of these synaptic proteins over the light/dark cycle may underlie circadian synchronization of action potential firing in the OB or modify synaptic interactions that would be predicted to impact olfactory coding, such as alteration of granule cell inhibition, increased number of available AMPARs to bind glutamate, or an increased gap junction conductance between mitral/tufted cells.

Aestivation and hypoxia-related events share common silent neuron trafficking processes

BackgroundThe availability of oxygen is a limiting factor for neuronal survival since low levels account not only for the impairment of physiological activities such as sleep-wake cycle, but above all for ischemic-like neurodegenerative disorders. In an attempt to improve our knowledge concerning the type of molecular mechanisms operating during stressful states like those of hypoxic conditions, attention was focused on eventual transcriptional alterations of some key AMPAergic silent neuronal receptor subtypes (GluR1 and GluR2) along with HSPs and HIF-1α during either a normoxic or a hypoxic aestivation of a typical aquatic aestivator, i.e. the lungfish (Protopterus annectens).ResultsThe identification of partial nucleotide fragments codifying for both AMPA receptor subtypes in Protopterus annectens displayed a putative high degree of similarity to that of not only fish but also to those of amphibians, birds and mammals. qPCR and in situ hybridization supplied a very high (p < 0.001) reduction of GluR1 mRNA expression in diencephalic areas after 6 months of aerial normoxic aestivation (6mAE). Concomitantly, high (p < 0.01) levels of HSP70 mRNAs in hypothalamic, mesencephalic and cerebellar areas of both 6mAE and after 6 months of mud hypoxic aestivation (6mMUD) were detected together with evident apoptotic signals. Surprisingly, very high levels of GluR2 mRNAs were instead detected in thalamic along with mesencephalic areas after 6 days of normoxic (6dAE) and hypoxic (6dMUD) aestivation. Moreover, even short- and long-term hypoxic states featured high levels of HIF-1α and HSP27 transcripts in the different brain regions of the lungfish.ConclusionsThe distinct transcriptional variations of silent neurons expressing GluR1/2 and HSPs tend to corroborate these factors as determining elements for the physiological success of normoxic and hypoxic aestivation. A distinct switching among these AMPA receptor subtypes during aestivation highlights new potential adaptive strategies operating in key brain regions of the lungfish in relation to oxygen availability. This functional relationship might have therapeutic bearings for hypoxia-related dysfunctions, above all in view of recently identified silent neuron-dependent motor activity ameliorations in mammals.

Mechanism of YB-1-mediated translational induction of GluR2 mRNA in response to neural activity through nAChR

BackgroundWe have reported previously that YB-1 induces translation of GluR2 mRNA in response to neural activity, and that HSP60 affects the association of YB-1 with polysomes. Here we examined the mechanism of YB-1-mediated translational activation of GluR2 mRNA through the nAChR. MethodsExpression of nAChRs in NG108-15 cells was verified. Translation of GluR2 mRNA and YB-1/HSP60 interaction were examined in nicotine-treated NG108-15 cells. Effects of inhibition of α7-nAChR and the PI3K/Akt pathway were investigated. The ratios of YB-1 to GluR2 mRNA and to HSP60 were explored in polysomal and non-polysomal fractions, respectively, and the role of HSP60 in cytoplasmic retention of YB-1 was evaluated. ResultsNicotine treatment transiently induced translation of GluR2 mRNA and Akt phosphorylation with a concomitant increase of YB-1/HSP60 interaction. Both α-bungarotoxin and LY294002 abolished the effects of nicotine. On a sucrose gradient, nicotine treatment shifted the distribution of YB-1 to much heavier-sedimenting polysome fractions. In these fractions, the ratio of YB-1 to its binding GluR2 mRNA was decreased, and ribosome association with the YB-1-bound GluR2 mRNA was increased. HSP60 was distributed only in the non-polysomal fractions as its binding to YB-1 increased. In HSP60-depleted cells, nicotine treatment induced nuclear localization of YB-1. ConclusionYB-1 is released from GluR2 mRNA during α7-nAChR-mediated neurotransmission, causing the PI3K/Akt pathway to recruit ribosomes into the translational machinery, and HSP60 is involved in cytoplasmic retention of polysome-free YB-1. General significanceActivation of the PI3K/Akt pathway through the α7-nAChR and YB-1/HSP60 interaction are important for YB-1-mediated translational activation of GluR2 mRNA.

Chromatin structure and expression of the AMPA receptor subunit Glur2 in human glioma cells: Major regulatory role of REST and Sp1

It has been suggested that reduced glutamate receptor expression protects glioma cells from glutamate toxicity. GluR2 is the critical subunit of the GluR2 subtype of AMPA glutamate receptors as this subunit determines the Ca(2+) permeability of the receptor. The gene encoding the GluR2 subtype of AMPA receptors has been described as a target gene for the transcription repressor REST. However, we recently showed that the GluR2 gene is not regulated by REST in several neuronal and neuroendocrine cell lines, due to a repressive chromatin environment. Here, we show that the GluR2 gene has an open chromatin configuration in human glioma cells. Overexpression of REST reduced GluR2 mRNA levels while shRNA-mediated depletion of REST or expression of a REST mutant, that contained a transcriptional activation domain, enhanced GluR2 expression. Incubation with trichostatin A (TSA), a histone deacetylase inhibitor, induced acetylation of histone 4 of the GluR2 locus in glioma cells, leading to an upregulation of GluR2 expression. Together, these data suggest that REST is responsible for the reduced expression of GluR2 in glioma cells. The transcription factor Sp1 additionally binds under physiological conditions to the GluR2 gene in human glioma cells and expression of a dominant-negative mutant of Sp1 reduced expression of GluR2. Thus, the regulation via Sp1 represents a further control point for GluR2 expression in glioma cells. Together, we show that the GluR2 gene is embedded into an open chromatin configuration in glioma cells and expression of GluR2 is controlled by REST and Sp1.

Commentary ( Two Hits with One Shot – A Possibility of Simultaneous Targeting Motor Neuron Loss and Depression in ALS by Upregulating ADAR2 )

Commentary ( Two Hits with One Shot – A Possibility of Simultaneous Targeting Motor Neuron Loss and Depression in ALS by Upregulating ADAR2 ) pp.863-863 (1) Author: Laszlo Siklos Abstract: A leading hypothesis for the selective death of motor neurons in sporadic amyotrophic lateral sclerosis (ALS) is excitotoxicity mediated by Ca2+-permeable α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptors. The AMPA receptors are assembled from 4 subunits and their permeability for Ca2+ is determined by the absence of the glutamate receptor subunit type 2 (GluR2), associated with an increased portion of unedited GluR2 mRNA at the glutamine/arginine (Q/R) site. RNA editing at this location is predominantly catalyzed by adenosine deaminase acting on RNA 2 (ADAR2). Thus, the observation that a substantial portion of Q/R site unedited GluR2 mRNA is present in postmortem spinal cord tissue of sporadic ALS patients suggests inefficiency of ADAR2 to be a disease-specific molecular dysfunction in ALS. To decide whether this malfunction could be a cause of motor neuron death in vivo, Hideyama and colleagues developed a conditional ADAR2 knock-out mouse line (designated as AR2), which carries gene-targeted floxed ADAR2 alleles that become functionally ablated by Cre recombinase expressed from a transgene (VAChT-Cre.Fast) in approximately 50% of motor neurons. They demonstrated that only the ADAR2-lacking spinal motor neurons degenerated in these animals, and this could be prevented by generating Q/R site edited GluR2 in the absence of ADAR2 by a point mutation in the endogenous GluR2 alleles. These results convincingly demonstrate that the inefficient GluR2 Q/R site editing found in the spinal cords of patients with sporadic ALS proposes the decreased level of ADAR2 mRNA to be a direct cause of death of motor neurons in mice, which accentuates the detrimental role of elevated Ca2+ influx in motor neurons as a consequence of increased Ca2+-permeability of AMPA receptors. From therapeutic point of view, an interesting aspect of the Ca2+-mediated cell death in ALS is connected with certain emotional reactions of the patients such as depression and anxiety, which has received particular attention in ALS. Although many researchers with experience with ALS challenge the lay opinion of an increased risk of developing major depressive disorder in ALS, studies using standardized methodology assessing nonphysical symptoms reported an occurrence exceeding the prevalence of that in the general population. There is growing evidence that besides the monoaminergic system the glutamatergic system, particularly N-methyl-D-aspartate receptor mediated Ca2+-entry plays an important role in the pathobiology of major depressive disorder. It is thus intriguing that conventional tricyclic antidepressants, specific serotonin reuptake inhibitors and serotonin/norepinephrine reuptake inhibitors not only down-regulate N-methyl-D-aspartate receptors but – at least in an in vitro model system – have the potency to enhance GluR2 Q/R site-editing either by up-regulating ADAR2 mRNA expression level, or through other unidentified mechanisms. It will now be important to determine whether these drugs exert a similar protective effect on motor neurons in in vivo models of ALS, and thus constitute a new therapeutic approach for this disease.

P-chlorophenylalanine increases glutamate receptor 1 transcription in rat amygdala

The amygdala is a key limbic structure strongly implicated in both epilepsy and anxiety disorders. Epilepsy-like mechanisms involve an increased glutamatergic activity, whereas disturbances in serotonin [5-hydroxytryptamine (5-HT)] systems are associated with anxiety-like behavior. Previous studies suggest that low 5-HT increases amygdala excitability, but the molecular mechanisms are not well characterized. Herein we explore the ability of low serotonin to increase glutamate receptor transcription. Using quantitative reverse transcriptase-polymerase chain reaction, we found that rats treated with P-chlorophenylalanine, an inhibitor of tyrosine-5-hydroxylase, resulted in a 21-fold increase in glutamate receptor 1 (GluR1) mRNA expression in the amygdala. These results suggest that low 5-HT induces hyperexcitability of amygdala neurons by increasing GluR1 transcription, and the upregulation of amygdala GluR1 may be important in the pathophysiology of anxiety disorders.

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.

Exposure of neurons to excitotoxic levels of glutamate induces cleavage of the RNA editing enzyme, adenosine deaminase acting on RNA 2, and loss of GLUR2 editing

AMPA receptors are glutamate receptors that are tetramers of various combinations of GluR1-4 subunits. AMPA receptors containing GluR1, 3 and 4 are Ca2+ permeable, however, AMPA receptors containing even a single subunit of GluR2 are Ca2+ impermeable. Most AMPA receptors are Ca2+ impermeable due to the presence of GluR2. GluR2 confers special properties on AMPA receptors through the presence of arginine at the pore apex; other subunits (GluR1, 3, 4) contain glutamine at the pore apex and allow Ca2+ influx. Normally, an RNA editing step changes DNA-encoded glutamine to arginine, introduces arginine in the GluR2 pore apex. GluR2 RNA editing is carried out by an RNA-dependent adenosine deaminase (ADAR2). Loss of GluR2 editing leads to the formation of highly excitotoxic AMPA channels [Mahajan and Ziff (2007) Mol Cell Neurosci 35:470-481] and is shown to contribute to loss of motor neurons in amyotrophic lateral sclerosis (ALS). Relatively higher levels of Ca2+-permeable AMPA receptors are found in motor neurons and this has been correlated with lower GluR2 mRNA levels. However, the reason for loss of GluR2 editing is not known. Here we show that exposure of neurons to excitotoxic levels of glutamate leads to specific cleavage of ADAR2 that leads to generation of unedited GluR2. We demonstrate that cleaved ADAR2 leads to a decrease or loss of GluR2 editing, which will further result in high Ca2+ influx and excitotoxic neuronal death.

Functional α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptors in differentiating embryonic neural progenitor cells

Glutamate-responsive α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptors are considered to play a significant role in neurogenesis. We have studied the functional expression of these receptors in migrating embryonic neural progenitor cells (NPCs). The majority of neurosphere-derived NPCs express AMPA receptors already during the first day of differentiation, based on mRNA quantification, immunocytochemistry, and Ca²+ imaging. The expression of GluR1 mRNA was significantly increased at 5 days of differentiation. The AMPA receptor subunits coexpressed with neuronal markers and were present in all cells at the outer periphery of the migration zone. In migrating NPCs, most of the AMPA receptors were philantotoxin sensitive and Ca²+-permeable, suggesting that in addition to their role in plasticity, the receptors are of importance in NPC differentiation.

Exposure to cyclic intermittent hypoxia increases expression of the AMPA receptor subunit GluR1 in the carotid body

We previously reported that: 1) N‐methyl‐D‐aspartate (NMDA) receptor subunits NR1 and NR2A/2B are expressed in rat carotid body; 2) exposure of rats to cyclic intermittent hypoxia (CIH) enhances carotid body NR1 levels, and 3) administration of the NR1 blocker, MK801, reduces endothelin‐1 induced carotid sinus nerve activity in CIH‐exposed rats. To further explore glutamatergic signaling in the modulation of chemosensitivity after CIH exposure we assessed carotid bodies from Sprague‐Dawley (SD) rats for non‐NMDA glutamate receptors, including α‐amino‐3‐hydroxy‐5‐methyl‐4‐isoxazolepropionic acid (AMPA) receptors and Kainate receptors using RT‐PCR, immunohistochemistry, immunofluorescence and western blot analysis. mRNA of the AMPA receptor subunit GluR1 is strongly expressed in the carotid body while there is weak expression of mRNA from GluR2, GluR3, GluR6 and KA2. Furthermore, the intense GluR1 immunoreactivity in the carotid body co‐localizes with tyrosine hydroxylase in glomus cells. To examine the effect of CIH on the GluR1 level in the carotid body, we next exposed male SD rats to CIH or to room air sham for 8 h/day for 3 weeks. CIH significantly enhanced the expression of both GluR1 mRNA (206.50 ± 13.35% of sham) and GluR1 protein (174.27 ± 9.12% of sham). These findings further suggest glutamatergic signaling may contribute to changes in carotid chemoreflex sensitivity induced by CIH exposure.

Upregulation of AMPA receptor GluR2 (GluA2) subunits in subcortical ischemic vascular dementia is repressed in the presence of Alzheimer's disease

Glutamatergic AMPA receptors are of clinical significance in dementia because of their roles in mediating fast excitatory neurotransmission and other synaptic events relevant to cognition. Reductions in the AMPA receptor GluR2 subunit are well-established in Alzheimer's disease (AD), but the status of GluR2 in subcortical ischemic vascular dementia (SIVD) and mixed AD/SIVD (MIX) has not been investigated. In this study we measured GluR2 immunoreactivity and mRNA levels in the postmortem neocortex of a longitudinally assessed cohort of SIVD and MIX, together with age-matched controls. We found that GluR2 immunoreactivity and mRNA were up-regulated in SIVD, but remained unchanged in MIX. Furthermore, higher GluR2 immunoreactivity was associated with milder cognitive impairment and lower concentrations of Aβ42 peptide and phosphorylated tau. Our study suggests that GluR2 up-regulation may be an adaptive process in SIVD, and that this process is repressed in the presence of concomitant AD in mixed dementia.

Protective effect of the daming capsule on impaired baroreflexes in STZ-induced diabetic rats with hyperlipoidemia

BackgroundThe Daming capsule (DMC) is a traditional Chinese medicine used to treat hyperlipoidemia. Both clinic trials and studies on animal models have demonstrated that DMC is beneficial against diabetic symptoms. Impairment of the baroreflex can cause life-threatening arrhythmias and sudden cardiac death in patients with diabetes mellitus (DM). This study was designed to elucidate the effects of DMC on baroreflexes in streptozocin (STZ)-induced diabetic rats with hyperlipoidemia.MethodsWistar rats were randomly divided into three groups: untreated controls, rats pretreated STZ and high lipids (a diabetes model or DM rats), and DM rats treated with DMC. The baroreflex sensitivity was examined during intravenous injection of phenylephrine (PE) or sodium nitroprusside (SNP) and quantified by the change in heart rate over the change in mean arterial blood pressure (ΔHR/ΔMABP). Morphological remodeling of baroreceptors was analyzed by transmission electron microscopy (TEM). The mRNA levels and expression of GluR2 and a GABAA receptor subunit were measured by quantitative RT-PCR and Western blotting.ResultsCompared to untreated DM rats, DMC significantly elevated the ratio of ΔHR/ΔMABP by enhancing the compensatory reduction in HR (-ΔHR) in response to PE-induced hypertension (+ΔMABP) (P < 0.05). In the presence of SNP, DMC increased the ΔMABP (P < 0.05). In addition, DMC markedly shortened the duration of blood pressure changes elicited by PE or SNP in DM rats compared to the untreated DM group (P < 0.05). Electron microscopy revealed disrupted myelin sheaths, swollen ER, and lysed mitochondria in the nucleus ambiguous (NAm) DM rats. These signs of neuropathology were largely prevented by treatment with DMC for 30 days. Treatment with DMC elevated both mRNA and protein level of GluR2 in the NAm of DM rats, but had no effect on GABAA receptor expression.ConclusionThe Daming capsule partially reversed the parasympathetic baroreflex impairment observed in STZ-induced diabetic rats with hyperlipoidemia. Treatment with DMC also prevented the degeneration of neurons and myelinated axons in the brain stem NAm and reversed the down-regulation of GluR2 mRNA. Rescue of NAm function may contribute to the medicinal properties of DMC in diabetic rats.

YB-1 binds to GluR2 mRNA and CaM1 mRNA in the brain and regulates their translational levels in an activity-dependent manner.

The translational regulator YB-1 binds to mRNAs. In the brain, YB-1 is prominently expressed from the prenatal stage until the first week after birth, being associated with polysomes and distributed in neuronal dendrites, but its expression declines to a much lower level thereafter. It is therefore of interest to identify the mRNAs whose translation is controlled by YB-1 in the postnatal growing brain. In this study we found that YB-1 interacted with the mRNAs for glutamate receptor subunit 2 (GluR2) and calmodulin1 (CaM1) in both brain and NG108-15 cells. Overexpression or knockdown of YB-1 altered the levels of these proteins significantly in cultured cells without any change in their mRNA levels. When the cells were treated with neurotransmitters, translation of these proteins was induced within a short time, and a change in the amount of YB-1 on its target mRNAs was observed in the heavy-sedimenting polysome fractions on a sucrose gradient. Depletion of YB-1 expression by siRNA abrogated the translational activation. Furthermore, in the brain of kainic acid-treated mice, the distribution of YB-1 was shifted to much heavier fractions associated with polysomes within 30min to 1h after the treatment, and the distribution returned to lighter fractions within the following 2h. The protein levels of GluR2 and CaM1 were also increased transiently when the distribution of YB-1 on the gradient changed. These results suggest that in the brain of growing mice, YB-1 binds to GluR2 and CaM1 mRNAs and regulates their translation in an activity-dependent manner.

AMPA receptor-mediated neuronal death in sporadic ALS

alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptor-mediated excitotoxicity has been proposed to play a role in death of motor neurons in amyotrophic lateral sclerosis (ALS). We demonstrated that RNA editing of GluR2 mRNA at the glutamine/arginine (Q/R) site was decreased in autopsy-obtained spinal motor neurons, but not in cerebellar Purkinje cells, of patients with sporadic ALS. This molecular change occurs in motor neurons of sporadic ALS cases with various phenotypes, but not in degenerating neurons of patients with other neurodegenerative diseases, including SOD1-associated familial ALS. Because GluR2 Q/R site-editing is specifically catalyzed by adenosine deaminase acting on RNA 2 (ADAR2), it is likely that regulatory mechanism of ADAR2 activity does not work well in the motor neurons of sporadic ALS. Indeed, ADAR2 expression level was significantly decreased in the spinal ventral gray matter of sporadic ALS as compared to normal control subjects. It is likely that ADAR2 underactivity selective in motor neurons induced deficient GluR2 Q/R site-editing, which results in the neuronal death of sporadic ALS. Thus, among multiple different molecular mechanisms underlying death of motor neurons, it is likely that an increase of the proportion of Q/R site-unedited GluR2-containing Ca(2+)-permeable AMPA receptors initiates the death of motor neurons in sporadic ALS. To this end, normalization of ADAR2 activity in motor neurons may become a therapeutic strategy for sporadic ALS.