AMPA Type Research Articles

Transcriptomic expression of AMPA receptor subunits and their auxiliary proteins in the human brain

Receptors to glutamate of the AMPA type (AMPARs) serve as the major gates of excitation in the human brain, where they participate in fundamental processes underlying perception, cognition and movement. Due to their central role in brain function, dysregulation of these receptors has been implicated in neuropathological states associated with a large variety of diseases that manifest with abnormal behaviors. The participation of functional abnormalities of AMPARs in brain disorders is strongly supported by genomic, transcriptomic and proteomic studies. Most of these studies have focused on the expression and function of the subunits that make up the channel and define AMPARs (GRIA1-GRIA4), as well of some accessory proteins. However, it is increasingly evident that native AMPARs are composed of a complex array of accessory proteins that regulate their trafficking, localization, kinetics and pharmacology, and a better understanding of the diversity and regional expression of these accessory proteins is largely needed. In this review we will provide an update on the state of current knowledge of AMPA receptors subunits in the context of their accessory proteins at the transcriptome level. We also summarize the regional expression in the human brain and its correlation with the channel forming subunits. Finally, we discuss some of the current limitations of transcriptomic analysis and propose potential ways to overcome them.

Pharmacological modulation of AMPA receptors rescues specific impairments in social behavior associated with the A350V Iqsec2 mutation

In this study we tested the hypothesis that pharmacological modulation of glutamatergic neurotransmission could rescue behavioral deficits exhibited by mice carrying a specific mutation in the Iqsec2 gene. The IQSEC2 protein plays a key role in glutamatergic synapses and mutations in the IQSEC2 gene are a frequent cause of neurodevelopmental disorders. We have recently reported on the molecular pathophysiology of one such mutation A350V and demonstrated that this mutation downregulates AMPA type glutamatergic receptors (AMPAR) in A350V mice. Here we sought to identify behavioral deficits in A350V mice and hypothesized that we could rescue these deficits by PF-4778574, a positive AMPAR modulator. Using a battery of social behavioral tasks, we found that A350V Iqsec2 mice exhibit specific deficits in sex preference and emotional state preference behaviors as well as in vocalizations when encountering a female mouse. The social discrimination deficits, but not the impaired vocalization, were rescued with a single dose of PF-4778574. We conclude that social behavior deficits associated with the A350V Iqsec2 mutation may be rescued by enhancing AMPAR mediated synaptic transmission.

Inhibition of AKT/GSK3β/CREB Pathway Improves the Responsiveness to AMPA Receptor Antagonists by Regulating GRIA1 Surface Expression in Chronic Epilepsy Rats.

α-Amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor (AMPAR) has been reported as one of the targets for treatment of epilepsy. Although maladaptive regulation of surface expression of glutamate ionotropic receptor AMPA type subunit 1 (GRIA1) subunit is relevant to the responsiveness to AMPAR antagonists (perampanel and GYKI 52466) in LiCl-pilocarpine-induced chronic epilepsy rats, the underlying mechanisms of refractory seizures to AMPAR antagonists have yet been unclear. In the present study, we found that both AMPAR antagonists restored the up-regulations of GRIA1 surface expression and Src family-mediated glycogen synthase kinase 3β (GSK3β)-Ca2+/cAMP response element-binding protein (CREB) phosphorylations to control levels in responders (whose seizure activities were responsive to AMPAR) but not non-responders (whose seizure activities were uncontrolled by AMPAR antagonists). In addition, 3-chloroacetyl indole (3CAI, an AKT inhibitor) co-treatment attenuated spontaneous seizure activities in non-responders, accompanied by reductions in AKT/GSK3β/CREB phosphorylations and GRIA1 surface expression. Although AMPAR antagonists reduced GRIA2 tyrosine (Y) phosphorylations in responders, they did not affect GRIA2 surface expression and protein interacting with C kinase 1 (PICK1) protein level in both responders and non-responders. Therefore, our findings suggest that dysregulation of AKT/GSK3β/CREB-mediated GRIA1 surface expression may be responsible for refractory seizures in non-responders, and that this pathway may be a potential target to improve the responsiveness to AMPAR antagonists.

Механизмы ингибирования AMPA рецепторов диминазеном

Diminazene is an anti-infective medication from the diarylamidine group. In this work we have shown for the first time that it inhibits AMPA type ionotropic glutamate receptors. Experiments were conducted on isolated Wistar rat neurons. Calcium-permeable AMPA receptors were studied on giant striatum cholinergic interneurons, while calcium-impermeable AMPA receptors – on hippocampal CA1 pyramidal neurons. Cells were isolated from slices by vibrodissociation. Currents were recorded by whole-cell patch-clamp method. Diminazene inhibited kainate-induced currents in both types of neurons in a concentration-dependent manner. IC50 values for calcium-permeable and calcium-impermeable AMPA receptors were 60±11 and 160±30 µM, respectively. It is interesting that the degree of inhibition by fixed diminazene concentration was increased with the increase of agonist concentration. The voltage-dependence curve for the inhibition of calcium-permeable AMPA receptors by fixed diminazene concentration was biphasic: minimal inhibition was observed at positive holding potentials, maximal – at -40 - -60 mV, while further hyperpolarization resulted in the smooth decrease of block. All these features strongly suggest that channel block with the ability to permeate into the cytoplasm is the main molecular mechanism of AMPA receptor inhibition by diminazene.

Src/CK2/PTEN-Mediated GluN2B and CREB Dephosphorylations Regulate the Responsiveness to AMPA Receptor Antagonists in Chronic Epilepsy Rats.

Both α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor (AMPAR) and N-methyl-D-aspartate receptor (NMDAR) have been reported as targets for treatment of epilepsy. To investigate the roles and interactions of AMPAR and NMDAR in ictogenesis of epileptic hippocampus, we analyzed AMPAR antagonists (perampanel and GYKI 52466)-mediated phosphatase and tensin homolog deleted on chromosome 10 (PTEN) regulation and glutamate ionotropic receptor NMDA type subunit 2B (GluN2B) tyrosine (Y) 1472 phosphorylation in epilepsy rats. Both perampanel and GYKI 52466 increased PTEN expression and its activity (reduced phosphorylation), concomitant with decreased activities (phosphorylations) of Src family-casein kinase 2 (CK2) signaling pathway. Compatible with these, they also restored the upregulated GluN2B Y1472 and Ca2+/cAMP response element-binding protein (CREB) serine (S) 133 phosphorylations and surface expression of glutamate ionotropic receptor AMPA type subunit 1 (GRIA1) to basal level in the epileptic hippocampus. These effects of perampanel and GYKI 52466 are observed in responders (whose seizure activities are responsive to AMPAR antagonists), but not non-responders (whose seizure activities were uncontrolled by AMPAR antagonists). Therefore, our findings suggest that Src/CK2/PTEN-mediated GluN2B Y1472 and CREB S133 regulations may be one of the responsible signaling pathways for the generation of refractory seizures in non-responders to AMPAR antagonists.

Genetic Variations of Ionotropic Glutamate Receptor Pathways on Interferon-α-induced Depression in Patients with Hepatitis C Viral Infection

ImportanceThe most supportive evidence of the inflammation theory of depression is that up to one-third of patients with Hepatitis C virus infection (HCV) develop clinical depressive episodes during interferon-α (IFN-α) therapy. As glutamate-mediated excitotoxicity has been found to be a consequence of excessive inflammation and a pathogenic mechanism of depression, it is plausible to investigate genes on ionotropic glutamate receptor pathways. ObjectiveTo identify the at-risk genetic variations on ionotropic glutamate receptor pathways for interferon-α-induced depression. MethodWe assessed 291 patients with chronic HCV undergoing IFN-α therapy and analyzed the single nucleotide polymorphisms (SNPs) in genes related to ionotropic glutamate receptors in this prospective case-control study. Patients who developed IFN-α-induced depression anytime during the treatment were defined as the case group, while those who did not were defined as the control group. Genomic DNA was extracted from peripheral blood and analyzed by Affymetrix TWB array. Allelic and haplotype association tests were conducted using χ2 tests to assess the difference in allele and haplotype frequencies between cases and controls. Additionally, we performed 5000 permutations to control gene-wide family-wise error rates and create empirical p-values. Stratified analyses were then done to control for confounders and adjust odds ratios for our significant SNPs. We also did an additional stratified analysis to re-assess genes with near-significant SNPs (empirical p-value=0.05-0.10), employing Bonferroni correction with the effective number of independent tests to control gene-wide family-wise error rates. ResultsThe minor and major allele frequencies of rs7542 (empirical p-value=0.0310) in MAPK3, rs3026685 (empirical p-value=0.0378) in PICK1, rs56005409 (empirical p-value=0.0332) in PRKCA, rs12914792 (empirical p-value=0.0096), rs17245773 (empirical p-value=0.0340) in RASGRF1, and rs78387863 (empirical p-value=0.0086), rs74365480 (empirical p-value=0.0200) in RASGRF2 were found significantly different between cases and controls. Haplotype association tests also revealed one significant haplotype in PRKCA (empirical p-value=0.0200) and one in RASGRF1 (empirical p-value=0.0048). Stratified analyses showed no signs of confounders for most of our significant SNPs, except for rs78387863 in RASGRF2. After a re-assessment of our near-significant genes by stratified analyses, two SNPs in GRIN2B turned significant. ConclusionsThis study provided supportive evidence of the involvement of the RAS/RAF/mitogen-activated protein kinase (MAPK) signaling pathway and glutamate ionotropic receptor AMPA type subunit 2(GluR2) transportation in the pathogenesis of IFN-α-induced depression.

Association of GRIA1 polymorphisms with ovarian response to human menopausal gonadotropin in Iranian women

ObjectiveGlutamate ionotropic receptor AMPA type subunit 1 (GRIA1) is a subunit of a ligand-gated ion channel that regulates the secretion of follicle-stimulating hormone (FSH) and luteinizing hormone (LH) by controlling the release of gonadotropin-releasing hormone. Few studies have investigated the association between the GRIA1 gene and human infertility. This study evaluated the association of the GRIA1 rs548294 C > T and rs2195450 G > A polymorphisms with the ovarian response to human menopausal gonadotropin (HMG) in Iranian women.MethodsOne hundred women with histories of at least 1 year of infertility were included. On the second day of menstruation, patients were injected with HMG; on the third day, blood samples were collected. After hormonal analysis, the GRIA1 rs548294 C > T and rs2195450 G > A genotypes of samples were identified via the restriction fragment length polymorphism method, and on day 9, the number of follicles was assessed via ultrasound.ResultsFor the GRIA1 rs548294 C > T and rs2195450 G > A single nucleotide polymorphisms, the subjects with CT and GG genotypes, respectively, displayed the highest mean FSH level, LH level, and number of follicles on day 9 of the menstrual cycle (p < 0.05). Significant positive correlations were observed between LH and FSH (p < 0.01), LH and follicle count (p < 0.01), FSH and age (p < 0.05), follicle count and age (p = 0.048), and FSH and follicle count (p < 0.01).ConclusionThis study showed a significant relationship between GRIA1 polymorphisms and ovarian response to the induction of ovulation. Therefore, determining patients’ GRIA1 genotype may be useful for improving treatment and prescribing suitable doses of ovulation-stimulating drugs.

PTEN Is Required for The Anti-Epileptic Effects of AMPA Receptor Antagonists in Chronic Epileptic Rats.

α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor (AMPAR) is one of the ligand-gated ion channels for glutamate, which is an important player in the generation and spread of seizures. The efficacy of AMPAR functionality is regulated by the trafficking, synaptic targeting, and phosphorylation. Paradoxically, AMPAR expression and its phosphorylation level are decreased in the epileptic hippocampus. Therefore, the roles of AMPAR in seizure onset and neuronal hyperexcitability in ictogenesis remain to be elucidated. In the present study, we found that AMPAR antagonists (perampanel and GYKI 52466) decreased glutamate ionotropic receptor AMPA type subunit 1 (GRIA1) surface expression in the epileptic rat hippocampus. They also upregulated phosphatase and tensin homolog deleted on chromosome 10 (PTEN) expression and restored to basal levels the upregulated phosphoinositide 3-kinase (PI3K)/AKT1 phosphorylations. Dipotassium bisperoxovanadium(pic) dihydrate (BpV(pic), a PTEN inhibitor) co-treatment abolished the anti-epileptic effects of perampanel and GYKI 52466. Therefore, our findings suggest that PTEN may be required for the anti-epileptic effects of AMPAR antagonists.

AMPA receptor nanoscale dynamic organization and synaptic plasticities.

The emergence of new imaging techniques and molecular tools has refreshed our understanding of the principles of synaptic transmission and plasticity. Superresolution imaging and biosensors for measuring enzymatic activities in live neurons or neurotransmitter levels in the synaptic cleft are giving us an unprecedented integrated and nanoscale view on synaptic function. Excitatory synapses are now conceptualized as organized in subdomains, enriched with specific scaffolding proteins and glutamate receptors, molecularly organized with respect to the pre-synaptic source of glutamate. This new vision of basic synaptic transmission changes our understanding of the molecular modifications which sustain synaptic plasticities. Long-term potentiation can no longer be explained simply by an increase in receptor content at the synapse. We review here the latest data on the role of nanoscale and dynamic organization of AMPA type glutamate receptors on synaptic transmission at both basal state and during short and long-term plasticities.

Proteomic analysis reveals a biosignature of decreased synaptic protein in cerebrospinal fluid of major depressive disorder

Major depressive disorder (MDD) is a leading cause of morbidity with a lifetime prevalence of 10%. There is increasing evidence suggesting synaptic dysfunction and impaired integrity of certain brain circuits in MDD. Here we investigate the cerebrospinal fluid proteome of psychiatric patients focusing on MDD by deep proteomic profiling approach combined with a further validation step using targeted mass spectrometry. We demonstrate profound CSF proteomic changes during on-going depression episodes in MDD patients (n = 40) in comparison to controls (n = 27), schizophrenia spectrum disorder (n = 13), and bipolar disorder patients (n = 11). The discovery analysis with isobaric tags for relative and absolute quantitation (iTRAQ) reveals changes in proteins associated with synaptic transmission, myelination, and Wnt signaling in CSF of MDD. The multiple reaction monitoring (MRM) validation analysis confirms significantly decreased levels of eight proteins including the membrane synaptic proteins neurexin 3 (NRXN3), contactin-associated protein-like 4 (CNTNAP4), and glutamate ionotropic receptor AMPA type subunit 4 (GRIA4) in the CSF of MDD patients in comparison to the controls. Overall, the study demonstrates proteins that constitute an MDD biosignature for further validation studies and provides insight into the pathophysiology of MDD and other psychiatric disorders.

PD24-01 NITRIC OXIDE CENTRALLY INDUCES FREQUENT URINATION VIA BRAIN GLUTAMATERGIC RECEPTORS IN RATS

You have accessJournal of UrologyBladder & Urethra: Anatomy, Physiology & Pharmacology II (PD24)1 Apr 2020PD24-01 NITRIC OXIDE CENTRALLY INDUCES FREQUENT URINATION VIA BRAIN GLUTAMATERGIC RECEPTORS IN RATS Hideaki Ono, Takahiro Shimizu*, Suo Zou, Masaki Yamamoto, Yohei Shimizu, Yurika Hata, Shogo Shimizu, Youichirou Higashi, Takaaki Aratake, Tomoya Hamada, Yoshiki Nagao, Masashi Honda, and Motoaki Saito Hideaki OnoHideaki Ono More articles by this author , Takahiro Shimizu*Takahiro Shimizu* More articles by this author , Suo ZouSuo Zou More articles by this author , Masaki YamamotoMasaki Yamamoto More articles by this author , Yohei ShimizuYohei Shimizu More articles by this author , Yurika HataYurika Hata More articles by this author , Shogo ShimizuShogo Shimizu More articles by this author , Youichirou HigashiYouichirou Higashi More articles by this author , Takaaki AratakeTakaaki Aratake More articles by this author , Tomoya HamadaTomoya Hamada More articles by this author , Yoshiki NagaoYoshiki Nagao More articles by this author , Masashi HondaMasashi Honda More articles by this author , and Motoaki SaitoMotoaki Saito More articles by this author View All Author Informationhttps://doi.org/10.1097/JU.0000000000000881.01AboutPDF ToolsAdd to favoritesDownload CitationsTrack CitationsPermissionsReprints ShareFacebookLinked InTwitterEmail Abstract INTRODUCTION AND OBJECTIVE: In the lower urinary tract, nitric oxide (NO) is well known as a relaxation factor in the urethra. On the other hand, in the central nervous system, NO is reported to have both inhibitory and facilitatory effects on the micturition reflex. We previously reported that intracerebroventricularly (icv) administered SIN-1, a NO donor, reduced intercontraction intervals (ICI), single voided volume and bladder capacity without altering post-voided residual urine volume, indicating that SIN-1 centrally induced frequent urination in rats. In this study, we examined central mechanisms for the SIN-1-induced frequent urination focusing on glutamatergic receptors (NMDA and AMPA types) in rats. METHODS: Urethane anesthetized (0.8 g/kg, ip) male Wistar rats (300-450 g) were used. A catheter was inserted into the bladder from the dome to perform cystometrograms (CMG, 12 ml/h saline infusion). Three hours after the surgery, SIN-1 (100 or 250 μg/rat) or vehicle was icv administered. In some rats, carboxy-PTIO (PTIO, NO scavenger, 750 μg/rat), MK-801 (NMDA type glutamatergic receptor antagonist, 10 or 30 nmol/rat) or DNQX (AMPA type glutamatergic receptor antagonist, 3 nmol/rat) was administered 30 min before SIN-1 administration (250 μg/rat, icv). CMG and evaluation of ICI and maximal voiding pressure (MVP) were started 60 min before the first icv administration. RESULTS: Icv administered SIN-1 dose-dependently reduced ICI compared with the vehicle-administered control group without altering MVP (Fig. 1). The SIN-1-induced ICI reduction was significantly attenuated by central pretreatment with PTIO or MK-801 (Fig. 2A and 2B), but not by DNQX (data not shown). CONCLUSIONS: NO centrally induces frequent urination via brain NMDA, but not AMPA, type glutamatergic receptors in rats. Thus, the brain nitrergic pathway that is directly involved in the control of micturition could be a new target for the treatment of bladder dysfunction. Source of Funding: JSPS KAKENHI (#17K09303), Narishige Neuroscience Research Foundation in Japan, Takeda Science Foundation, The Smoking Research Foundation in Japan © 2020 by American Urological Association Education and Research, Inc.FiguresReferencesRelatedDetails Volume 203Issue Supplement 4April 2020Page: e532-e532 Advertisement Copyright & Permissions© 2020 by American Urological Association Education and Research, Inc.MetricsAuthor Information Hideaki Ono More articles by this author Takahiro Shimizu* More articles by this author Suo Zou More articles by this author Masaki Yamamoto More articles by this author Yohei Shimizu More articles by this author Yurika Hata More articles by this author Shogo Shimizu More articles by this author Youichirou Higashi More articles by this author Takaaki Aratake More articles by this author Tomoya Hamada More articles by this author Yoshiki Nagao More articles by this author Masashi Honda More articles by this author Motoaki Saito More articles by this author Expand All Advertisement PDF downloadLoading ...

Fisetin Prevents HT22 Cells From High Glucose-Induced Neurotoxicity via PI3K/Akt/CREB Signaling Pathway.

Hyperglycemia has been widely considered as a key risk factor for diabetic encephalopathy which can cause neuronal apoptosis and cognitive deficits. The flavonoid compound, fisetin, possesses potential neuroprotective effects and also enhances learning and memory. However, the role of fisetin in hyperglycemia-induced neuronal cytotoxicity has not been fully elucidated. In the present study, HT22 murine hippocampal neuronal cell line was used to establish the injured cell model. Cell proliferation and cytotoxicity assay, Hoechst 33258 staining, qRT-PCR, western blot analysis, and specific inhibitor were used to investigate the effect and molecular mechanisms of fisetin on high glucose (HG)-induced neurotoxicity in HT22 cells. Our results showed that 125 μM and 48 h of treatment was identified as optimal damage parameter of HG. Fisetin significantly improved HG-inhibited cell viability. The levels of LDH, malondialdehyde (MDA), and superoxide dismutase (SOD) were noticeably modulated by fisetin, which alleviated HG-induced HT22 cell oxidative damage. Besides, the apoptosis of HT22 cells was rescued by fisetin pretreatment. In addition, fisetin also prevented HG-induced downregulation of the mRNA expression of Bdnf, Gdnf, synaptophysin (Syp), and glutamate ionotropic receptor AMPA type subunit 1 (Gria1) in cells. More importantly, the decreased phosphorylation of phosphoinositide 3 kinase (PI3K), Akt, and cAMP-response element binding protein (CREB) was rescued by fisetin treatment and that neuroprotective effect of fisetin was partially blocked by PI3K inhibitor, LY294002. These findings indicate that fisetin has potent neuroprotective effect and prevents HG-induced neurotoxicity by activation of PI3K/Akt/CREB pathway.

CircCBFB-mediated miR-28-5p facilitates abdominal aortic aneurysm via LYPD3 and GRIA4

Heading aimsAbdominal aortic aneurysm (AAA) is featured by the growth impediment and apoptosis surge of VSMCs (vascular smooth muscle cells). MicroRNAs (miRNAs) are suggested to affect cellular behaviors including cell growth and apoptosis. This study concentrated on unraveling the emerging role of miR-28-5p in abdominal aortic aneurysm. Materials and methodsPreviously, miR-28-5p was reported to be highly expressed in AAA. Functional assays were utilized to determine the role of miR-28-5p in VSMC apoptosis. To narrow down the downstream mRNAs, bioinformatics methods were utilized. The interaction between miR-28-5p and GRIA4 (glutamate ionotropic receptor AMPA type subunit 4) or LYPD3 (LY6/PLAUR domain containing 3) was explored. Candidate circRNAs (circular RNAs) of miR-28-5p were identified. Rescue analyses validated function of circCBFB (core-binding factor subunit beta)/miR-28-5p/GRIA4/LYPD3 axis in VSMC apoptosis and growth. Key findingsMiR-28-5p acted as an apoptosis driver while circCBFB, GRIA4 and LYPD3 exerted anti-apoptosis effects in VSMCs. Mechanically, GRIA4 and LYPD3 were suppressed by miR-28-5p. Moreover, circCBFB served as a sponge of miR-28-5p, releasing GRIA4 and LYPD3 from miR-28-5p suppression. Functionally, GRIA4, LYPD3 and miR-28-5p were required in circCBFB-mediated VSMC apoptosis. SignificanceThis work unveiled an innovative axis of circCBFB/miR-28-5p/GRIA4/LYPD3 in VSMC apoptosis, exerting its potential in providing new thoughts in AAA management.

Icariin ameliorates the cognitive function in an epilepsy neonatal rat model by blocking the GluR2/ERK I/II pathway.

The present study was performed to evaluate the protective effects of icariin on cognitive function in ahypoxia-induced neonatal epilepsy rat model. Neonatal epilepsy was induced in rat pups on postnatal day (PD) 20 by induction of hypoxia for 15 minutes. Rats were treated intraperitoneally with icariin at 75 mg/kg 1 hour before the induction of hypoxia. The effects of icariin were examined by estimating seizure stage, cognitive function and parameters of electroencephalography (EEG) in this neonatal epilepsy rat model. Parameters of oxidative stress and expression of proteins were examined in the brain tissue of the neonatal epilepsy rat model by histopathological study and Western blotting analysis, respectively. The results of this study suggest that treatment with icariin ameliorates the changes in seizure stage, number of seizures and parameters of EEG in hypoxia-induced neonatal epilepsy rats. Oxidative stress and apoptosis were decreased in the brain tissue of the icariin treatment group compared to the hypoxia group. Moreover, treatment with icariin ameliorated the altered expression of glutamate ionotropic receptor AMPA type subunit 2 (GluR2) and extracellular receptor kinase (ERK I/II) proteins in the brain tissue of hypoxia-induced epilepsy rats. Histopathological study also showed that icariin treatment improved the histopathology of brain tissue of hypoxia-induced epilepsy rats. In conclusion, the results of the present study suggest that icariin protects against neuronal injury and improves cognitive function in hypoxia-induced neonatal epilepsy rats by modulating the GluR2/ERK I/II pathway.

DRG Neurons Promote Perineural Invasion of Endometrial Cancer via GluR2.

Background: Perineural invasion (PNI) is correlated with negative prognosis in multiple cancers, but its role in endometrial cancer (EC) is still largely unknown; thus, targeted treatment for nerve infiltration is lacking as well.Methods: The interaction between nerve and EC cells were investigated by in vitro neural invasion assay and transwell coculture system. Then the nerve-related receptor gene glutamate ionotropic receptor AMPA type subunit 2 (GRIA2) was detected in EC tissues and cells using PCR array, western blotting, and immunohistochemistry. The role of GluR2 (gene name GRIA2) on EC proliferation, migration and invasion was evaluated by a GluR2 antagonist and shRNA. At the same time, the neurotransmitter effect on GluR2 (glutamate) from the cocultured conditional medium was measured using high-performance liquid chromatography (HPLC).Results: EC cell line Ishikawa (ISK) showed the ability to migrate along neurites in vitro and the numbers of migrated/invaded EC cells in the DRG neuron coculture group were significantly increased. The expression of GluR2 in EC tissue was found to be higher than that in para-carcinoma tissue. After GluR2 antagonist and GluR2 shRNA treatment, the proliferation, migration and invasion of ISK cells was markedly inhibited. Moreover, the ability of DRG neurons to promote the migration and invasion of ISK cells could also be attenuated by downregulation of GluR2, and the concentration of the neurotransmitter glutamate was notably increased in the coculture conditional medium compared to that in the DRG neuron or ISK cells alone.Conclusions: DRG neurons promote metastasis of EC cells via GluR2, which might be a risk factor for PNI in EC. Moreover, the perineural system may promote tumor invasion and metastasis under certain circumstances.

ACT-13 RESPONSE TO SEIZURE AND TUMOR-PROGRESSION BY TREATMENT WITH PERAMPANEL IN PATIENTS WITH GLIOMAS

BACKGROUNDIncreased extracellular glutamate level activates AMPA type glutamate receptors (AMPA-receptor) and induces seizures. Antagonistic activation of AMPA-receptors inhibits epilepsy and glioma progression in vivo and in vitro studies.PATIENTS AND METHODS(1) We tested perampanel (PER), an AMPA-receptor antagonist, in fifteen glioma patients with uncontrolled epilepsy. Seizure response, PER concentration, and tumor volume were assessed. (2) We tested PER in thirteen glioma patients (gr 2- 3 cases, gr 3- 4 cases, and gr 4- 4cases) after the initial treatment of surgery and RT (and CT).RESULTS(1) An objective seizure response was observed in 13 analyzed patients (100%) with 8 cases (62%) of seizure-freedom. Median plasma concentration of PER was 232 ng/ml in patients with 4mg/day PER and 518 ng/ml in patients with 8mg/day PER. High intensity lesions of MRI-FLAIR images were assessed volumetrically to analyze the tumor size. The volume reduction was detected during the 6 months period in correlation with the plasma PER levels. (2) All the 13 cases treated with PER after the initial treatment was seizure free. Two cases of gr 4 were died at 18 and 20 months after the surgical treatment. Other 11 cases are survival-free.CONCLUSIONPER treatment was effective in uncontrolled epilepsy with gliomas. MRI images showed the inhibition of tumor-progression. PER may effective for the inhibition of tumor progression.

Modified constraint-induced movement therapy alters synaptic plasticity of rat contralateral hippocampus following middle cerebral artery occlusion.

Modified constraint-induced movement therapy is an effective treatment for neurological and motor impairments in patients with stroke by increasing the use of their affected limb and limiting the contralateral limb. However, the molecular mechanism underlying its efficacy remains unclear. In this study, a middle cerebral artery occlusion (MCAO) rat model was produced by the suture method. Rats received modified constraint-induced movement therapy 1 hour a day for 14 consecutive days, starting from the 7th day after middle cerebral artery occlusion. Day 1 of treatment lasted for 10 minutes at 2 r/min, day 2 for 20 minutes at 2 r/min, and from day 3 onward for 20 minutes at 4 r/min. CatWalk gait analysis, adhesive removal test, and Y-maze test were used to investigate motor function, sensory function as well as cognitive function in rodent animals from the 1st day before MCAO to the 21st day after MCAO. On the 21st day after MCAO, the neurotransmitter receptor-related genes from both contralateral and ipsilateral hippocampi were tested by micro-array and then verified by western blot assay. The glutamate related receptor was shown by transmission electron microscopy and the glutamate content was determined by high-performance liquid chromatography. The results of behavior tests showed that modified constraint-induced movement therapy promoted motor and sensory functional recovery in the middle cerebral artery-occluded rats, but had no effect on cognitive function. The modified constraint-induced movement therapy upregulated the expression of glutamate ionotropic receptor AMPA type subunit 3 (Gria3) in the hippocampus and downregulated the expression of the beta3-adrenergic receptor gene Adrb3 and arginine vasopressin receptor 1A, Avpr1a in the middle cerebral artery-occluded rats. In the ipsilateral hippocampus, only Adra2a was downregulated, and there was no significant change in Gria3. Transmission electron microscopy revealed a denser distribution the more distribution of postsynaptic glutamate receptor 2/3, which is an α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid receptor, within 240 nm of the postsynaptic density in the contralateral cornu ammonis 3 region. The size and distribution of the synaptic vesicles within 100 nm of the presynaptic active zone were unchanged. Western blot analysis showed that modified constraint-induced movement therapy also increased the expression of glutamate receptor 2/3 and brain-derived neurotrophic factor in the hippocampus of rats with middle cerebral artery occlusion, but had no effect on Synapsin I levels. Besides, we also found modified constraint-induced movement therapy effectively reduced glutamate content in the contralateral hippocampus. This study demonstrated that modified constraint-induced movement therapy is an effective rehabilitation therapy in middle cerebral artery-occluded rats, and suggests that these positive effects occur via the upregulation of the postsynaptic membrane α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid receptor expression. This study was approved by the Institutional Animal Care and Use Committee of Fudan University, China (approval No. 201802173S) on March 3, 2018.

Prognostic Markers Identification in Glioma by Gene Expression Profile Analysis.

This study aimed to explore more gene markers associated with glioma or its prognosis. The glioma-related RNAseq data from the Gene Expression Omnibus database and The Cancer Genome Atlas dataset in UCSC Xena database were downloaded. There was a total of 971 tumor samples and 102 normal samples in the 2 datasets. The differentially expressed genes (DEGs) data between tumor and normal samples were analyzed, on which were then performed function and pathway enrichment analyses. Pearson correlation coefficient between DEGs was calculated to construct the coexpression network. Finally, prognostic genes were screened. A total of 634 upregulated and 769 downregulated DEGs were identified between tumor and control groups. These DEGs were significantly involved in 15 upregulated pathways, such as p53 signaling pathway, and 16 downregulated pathways, such as neuroactive ligand-receptor interaction, and cell adhesion molecules. In the coexpression network, pseudouridine synthase 7 (PUS7), EFR3 homolog B (EFR3B), and neuronal cell adhesion molecule (NRCAM) had the top three highest degrees. Additionally, 17 prognostic genes were selected, such as thrombospondin-1 (THBS1), caspase-8 (CASP8), glutamate ionotropic receptor AMPA type subunit 2 (GRIA2), GRIA4, and ADCYAP receptor type I (ADCYAP1R1). Pathways of p53 signaling pathway and neuroactive ligand-receptor interaction may play important roles in glioma progression. PUS7, EFR3B, and NRCAM may be potential biomarkers of glioma. THBS1, CASP8, GRIA2, GRIA4, and ADCYAP1R1 may serve as prognostic markers in glioma.

GRIK4 and GRM7 gene may be potential indicator of venlafaxine treatment reponses in Chinese of Han ethnicity.

Venlafaxine is one of commonly prescribed antidepressants for major depressive disorder (MDD). Accumulated evidence implicates the involvement of glutamatergic receptors in the pathophysiology of MDD and antidepressant treatment.By using 193 MDD patients who have been taking venlafaxine for 6 weeks, we investigated whether single nucleotide polymorphisms (SNPs) in glutamate ionotropic receptor kainate type subunit 4 (GRIK4), glutamate ionotropic receptor AMPA type subunit 1 (GRIA1) and glutamate metabotropic receptor 7 (GRM7) were associated with treatment response. 14 SNPs were selected randomly depended on association studies. Efficacy of treatment was determined by 17-item of Hamilton Rating Scale. Allele and genotype frequencies were compared between responders and non-responders.After adjusting by the false discovery rate (FDR), rs6589847 and rs56275759 in GRIK4 and rs9870680 in GRM7 showed associating with venlafaxine treatment response at week 6. (FDR: P = .018, P = .042, and P = .040, respectively).Our results indicated that genetic variants in the GRIK4 and GRM7 may associate with the treatment response in MDD patients treated by venlafaxine.

MicroRNA-186-5p controls GluA2 surface expression and synaptic scaling in hippocampal neurons

Homeostatic synaptic scaling is a negative feedback response to fluctuations in synaptic strength induced by developmental or learning-related processes, which maintains neuronal activity stable. Although several components of the synaptic scaling apparatus have been characterized, the intrinsic regulatory mechanisms promoting scaling remain largely unknown. MicroRNAs may contribute to posttranscriptional control of mRNAs implicated in different stages of synaptic scaling, but their role in these mechanisms is still undervalued. Here, we report that chronic blockade of glutamate receptors of the AMPA and NMDA types in hippocampal neurons in culture induces changes in the neuronal mRNA and miRNA transcriptomes, leading to synaptic upscaling. Specifically, we show that synaptic activity blockade persistently down-regulates miR-186-5p. Moreover, we describe a conserved miR-186-5p-binding site within the 3'UTR of the mRNA encoding the AMPA receptor GluA2 subunit, and demonstrate that GluA2 is a direct target of miR-186-5p. Overexpression of miR-186 decreased GluA2 surface levels, increased synaptic expression of GluA2-lacking AMPA receptors, and blocked synaptic scaling, whereas inhibition of miR-186-5p increased GluA2 surface levels and the amplitude and frequency of AMPA receptor-mediated currents, and mimicked excitatory synaptic scaling induced by synaptic inactivity. Our findings elucidate an activity-dependent miRNA-mediated mechanism for regulation of AMPA receptor expression.