Glutamate is a necessary excitatory neurotransmitter in human nervous system, which runs a biological function by binding with corresponding receptors. Psychiatric diseases occur when genes which encode receptors become dysfunctional. The authors have reviewed related literature and summarized the association between schizophrenia and glutamate receptor gene SNPs such as rs11146020 in GRIN1, 366C/G in GRIN2B, and rs1468412 in GRM3, etc. Due to controversial results in various studies, it is hypothesized that schizophrenia are complicated polygenic inherited diseases. Some sites such as 366C/G, 2664C/T and rs1408766 (C/T) possess with valuable genetic polymorphisms and might potentially contribute to personal identification and paternity testing. Studies in this field may have a potential significance in forensic psychiatry practice.
Forensic Psychiatry ; Humans ; Paternity ; Polymorphism, Single Nucleotide/genetics* ; Receptors, Glutamate/genetics* ; Schizophrenia/genetics*

AIMTo investigate the protective effect of mGluR1 antisense oligonucleotides and mGluR5 antisense oligonucleotides on impairment of cultured mouse cerebral cortical neurons induced by sodium glutamate (Glu).

METHODSNeuron damage induced by Glu as well as the action of mGluR1 antisense oligonucleotides and mGluR5 antisense oligonucleotides were measured by determining the leakage of lactate dehydrogenase (LDH) from neurons. Immunocytochemistry method was used to detect the expression of anti-mGluRl a and anti-mGluR5. Morphological changes of primary cortical neurons were observed by phase contrast microscope.

RESULTSFollowing the exposure of the cells to 0.1 mmol x L(-1) Glu for 15 min, LDH leakage from neurons increased. mGluR1 antisense oligonucleotides and mGluR5 antisense oligonucleotides(6 or 8 micromol x L(-1)) as well as 50 micromol x L(-1) LY367385 reduced the LDH leakage. mGluR1alpha and mGluR5 immunopositive cells showed in cultured neurons.

CONCLUSIONThe protective effects of mGluR1 antisense oligonucleotides and mGluR5 antisense oligonucleotides on neurons damaged by Glu may relate to antagonizing mGluR1a or mGItlR5.

Animals ; Cells, Cultured ; Cerebral Cortex ; cytology ; Mice ; Mice, Inbred Strains ; Neurons ; drug effects ; metabolism ; Neuroprotective Agents ; pharmacology ; Oligonucleotides, Antisense ; Receptors, Metabotropic Glutamate ; genetics ; Sodium Glutamate ; toxicity

This study investigated the effect of Suanzaoren Decoction on the expression of N-methyl-D-aspartate receptors(NMDAR) and α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid receptors(AMPAR) in the hippocampus and synaptic plasticity in rats with conditioned fear-induced anxiety. The effect of Suanzaoren Decoction on rat behaviors were evaluated through open field experiment, elevated plus maze experiment, and light/dark box experiment. Enzyme-linked immunosorbent assay(ELISA) was used to measure the levels of glutamate(Glu) and γ-aminobutyric acid(GABA) in the rat hippocampus. Real-time fluorescence quantitative PCR(qRT-PCR) and Western blot were employed to assess the gene and protein expression of ionotropic glutamate receptors in the hippocampal region. Transmission electron microscopy was utilized to observe the changes in the ultrastructure of synaptic neurons in the hippocampal region. Long-term potentiation(LTP) detection technique was employed to record the changes in population spike(PS) amplitude in the hippocampal region of mice in each group. The behavioral results showed that compared with the model group, the Suanzaoren Decoction group effectively increased the number of entries into open arms, time spent in open arms, percentage of time spent in open arms out of total movement time, number of entries into open arms out of total entries into both arms(P<0.01), and significantly increased the time spent in the light box and the number of shuttle crossings(P<0.01). There was an increasing trend in the number of grid crossings, entries into the center grid, and time spent in the center grid, indicating a significant anxiolytic effect. ELISA results showed that compared with the model group, the Suanzaoren Decoction group exhibited significantly reduced levels of Glu, Glu/GABA ratio(P<0.01), and significantly increased levels of GABA(P<0.01) in the rat hippocampus. Furthermore, Suanzaoren Decoction significantly decreased the gene and protein expression of NMDAR(GluN2B and GluN2A) and AMPAR(GluA1 and GluA2) compared with the model group. Transmission electron microscopy results demonstrated improvements in synapses, neuronal cells, and organelles in the hippocampal region of the Suanzaoren Decoction group compared with the model group. LTP detection results showed a significant increase in the PS amplitude changes in the hippocampal region of Suanzaoren Decoction group from 5 to 35 min compared with the model group(P<0.05, P<0.01). In conclusion, Suanzaoren Decoction exhibits significant anxiolytic effects, which may be attributed to the reduction in NMDAR and AMPAR expression levels and the improvement of synaptic plasticity.
Rats ; Mice ; Animals ; Receptors, Ionotropic Glutamate/metabolism* ; Hippocampus ; Neuronal Plasticity ; Receptors, N-Methyl-D-Aspartate/genetics* ; Anxiety/genetics* ; gamma-Aminobutyric Acid

OBJECTIVETo investigate whether genomic copy number variants (CNVs), within metabotropic glutamate receptors 7 (GRM 7) gene are associated with schizophrenia.

METHODSWe examined CNVs in conserved region of GRM7 using real time quantitative PCR among 180 Chinese schizophrenia cases and 33 normal controls. Products of real time quantitative PCR were sequenced bilaterally.

RESULTSReal time quantitative PCR found that a biallelic deletion existed at the 200 bps up-stream of exon 2 in a schizophrenia patient and a monoallelic deletion existed at this site in another 13 schizophrenia patients and a control subject. However, sequencing results showed a substitution of C to G at the 5bp up-stream of 3' end of reverse primer for real time PCR (GRM7-SV-1R). In addition, samples with this variant were exactly those having biallelic or monoallelic deletions, indicating that the results of the real time PCR were caused by the substitution variant at the 3' end of the primer rather than a bona fide genome deletion.

CONCLUSIONSReal-time quantitative PCR combined with sequencing can avoid false positive deletions and therefore is effective in detecting CNVs. According to our results, CNVs in GRM 7 gene is not associated with schizophrenia in the Han Chinese population. However, some potential rare CNVs may still have such relationship, and require further study.

Asian Continental Ancestry Group ; genetics ; Case-Control Studies ; DNA Copy Number Variations ; Female ; Humans ; Male ; Mutation ; Receptors, Metabotropic Glutamate ; genetics ; Schizophrenia ; genetics

To identify metabotropic glutamate receptor 4 (mGluR4) modulators by Ca2+ influx assay, we developed the functional cell-based high throughput-screening (HTS) assay. The human mGluR4 cDNA was transfected into HEK-293 stably expressing promiscuous G-protein (Ga alpha15) cells. Recombinant stable mGluR4 cell line was selected under Zeocin and validated by Ca2+ influx assay. The assay was optimized on loading time of Fluo Calcium Indicator, Dimethyl sulfoxide (DMSO) tolerance and sodium hydroxide (NaOH) tolerance using agonist (L-Glutamic acid (L-Glu)) of mGluR4. The rank order of the agonist potency for the stable human mGluR4 cell line was L-(+)-2-Amino-4-phosphonobutyric acid (L-AP4) > L-Serine-O-phosphate (L-SOP) > L-Glu, and of the antagonist potency was (RS)-alpha-Methylserine-O-phosphate (MSOP) > (RS)-alpha-Methyl-4-phosphonophenylglycine (MPPG). Z' factor value of the cell line in 96- and 384-well plate format was 0.80 and 0.65. Our data indicate a successful development of functional human mGluR4 recombinant stable cell line that was suitable for high throughput screening to identify mGluR4 agonist/antagonist.
Aminobutyrates ; pharmacology ; Cell Line ; DNA, Complementary ; genetics ; Drug Evaluation, Preclinical ; Humans ; Kidney ; cytology ; embryology ; Phosphoserine ; pharmacology ; Plasmids ; genetics ; Receptors, Metabotropic Glutamate ; agonists ; antagonists & inhibitors ; genetics ; Transfection

OBJECTIVETo investigate the expression changes of metabotropic glutamate receptor 5 (mGluR5) in neuropathic pain.

METHODSEighty-four adult male Sprague Dawley rats weighing 180-220 g were randomly divided into 7 groups (n = 12) : control group; S3, S7, and S14 groups: rats received the sham operation, the mechanical pain threshold was measured, and then the rats were decapitated and the ipsilateral lumbar spinal cord dorsal horn and dorsal root ganglion (DRG) samples were obtained on the 3rd, 7th, 14th postoperative day, respectively; C3, C7, and C14 groups: the chronic sciatic nerve constriction (CCI) model was established, the mechanical pain threshold was measured and the samples were obtained on the 3rd, 7th, 14th postoperative day, respectively. The expression level of mGluR5 mRNA and protein in the spinal cord and DRG were measured using the reverse transcriptase polymerase chain reaction and Western blot.

RESULTSIn the CCI group, the mechanical pain threshold in each observation day was significantly lower than in the sham operation group (P < 0.05). In the spinal cord, the expressions of mGluR5 mRNA and protein were significantly elevated in the C3 group than in the S3 and the control group (P < 0.05). On the 7th and the 14th postoperative day, no significant difference was found in the expression of mGluR5 mRNA and protein between CCI groups and the sham operation groups or the control group. No change was detected in DRG mRNA or protein.

CONCLUSIONmGluR5 is differentially expressed in spinal cord in response to neuropathic pain, which suggests that mGluR5 may be involved in the mechanism of neuropathic pain.

Animals ; Disease Models, Animal ; Ganglia, Spinal ; metabolism ; Male ; Neuralgia ; metabolism ; Pain Threshold ; RNA, Messenger ; genetics ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Receptor, Metabotropic Glutamate 5 ; Receptors, Metabotropic Glutamate ; genetics ; metabolism ; Sciatic Neuropathy ; metabolism ; Spinal Cord ; metabolism

OBJECTIVETo study the effects on the expression of mGluR5 mRNA and protein levels in primarily cultured hippocampal neurons after lead exposure.

METHODSPrimary embryonic rat hippocampal neuronal culture was prepared. On the 3(rd) day of incubation, lead chloride solution was added into medium to produce four different lead exposure levels: 0, 1 x 10(-8) mol/L, 1 x 10(-6) mol/L, 1 x 10(-4) mol/L Pb(2+). After 10 days of incubation, the neurons were collected to measure the alteration of mGluR5 mRNA expression by real-time fluorescent quantity PCR and the expression of mGluR5 in protein level by Western blot.

RESULTSThe studies revealed that mGluR5 mRNA expression was down-regulated after lead exposure in a dose-dependent manner. The mGluR5 mRNA expression of the lower lead-exposed neurons (Pb(2+) 10(-8) mol/L), the medium lead-exposed neurons(Pb(2+) 10(-6) mol/L), the higher lead-exposed neurons(Pb(2+) 10(-4) mol/L) were 0.724, 0.421, 0.321 times less than that of the controls, respectively. The Western blot demonstrated that mGluR5 expression in protein level should be decreased after lead exposure.

CONCLUSIONThe expression of mGluR5 in mRNA and protein levels should be down-regulated after lead exposure at different lead levels in a dose-dependent manner.

Animals ; Animals, Newborn ; Cells, Cultured ; Female ; Hippocampus ; drug effects ; metabolism ; Lead ; toxicity ; Lead Poisoning, Nervous System ; metabolism ; Neurons ; drug effects ; metabolism ; Pregnancy ; RNA, Messenger ; genetics ; Rats ; Rats, Sprague-Dawley ; Receptor, Metabotropic Glutamate 5 ; Receptors, Metabotropic Glutamate ; biosynthesis ; genetics

OBJECTIVETo investigate the expression of metabotropic glutamate receptor 4 (mGluR4) in cardiomyocytes differentiated from mouse embryonic stem cells (ES cells).

METHODSES cells were differentiated into cardiomyocytes with hanging-drop cultures. Retinoic acid (RA) and dimethyl sulfoxide (DMSO) were used as positive and negative controls, respectively. The co-expression of cardiac sarcomeric protein (α-actinin or troponin-T) and mGluR4 were verified by immunocytochemistry and flow cytometry analysis. The mRNA and protein expressions of mGluR4 were verified by RT-PCR and Western blot analysis, respectively. Meanwhile, the expression of mGluR4 in prenatal mouse heart was also examined.

RESULTSmGluR4 was expressed in both mouse ES cells and ES cell-derived cardiomyocytes. The level of mGluR4 protein expression decreased during the maturation of the cardiomyocytes. The co-expression rate of mGluR4 and Troponin T in the beating embryoid bodies (EBs) was only (3.00 ±1.00)%. On the other hand, mGluR4 gene and protein expressions showed remarkable down-regulation in the development of mouse fetal heart, which was not detected in mouse adult heart.

CONCLUSIONThe expression of mGluR4 is down-regulated in the cardiomyocyte differentiation of ES cells. The trend of expression is consistent with that in the prenatal mouse heart development.

Animals ; Cell Differentiation ; physiology ; Cell Line ; Embryonic Stem Cells ; cytology ; metabolism ; Mice ; Mice, Inbred ICR ; Myocytes, Cardiac ; cytology ; metabolism ; RNA, Messenger ; genetics ; Receptors, Metabotropic Glutamate ; genetics ; metabolism

OBJECTIVESmith-Fineman-Myers syndrome (SFMS) is an X-linked mental retardation syndrome. The authors had ascertained a large Chinese family with SFMS from Shandong and had mapped the disease locus to an interval of 19.8 Mb on Xq25 flanked by markers DXS8064 and DXS8050. Further investigation suggested that SFMS exhibited locus heterogeneity. In this study for facilitating the identification of the gene responsible for SFMS, the additional markers were analyzed to narrow down the candidate region, and four candidate genes (GPC3, MST4,GPCR2 and GLUD2) were chosen and screened for disease-causing mutation.

METHODSPCR and denaturing polyacrylamide gel electrophoresis were used to genotype 13 new polymorphic markers distributed within the candidate region. Mutation detection was accomplished by sequencing the exons and intron-exon junctions of the candidate genes.

RESULTSBy analyzing 13 additional polymorphic markers, SFMS candidate region can be reduced to an interval of 10.18 Mb bounded by XSTR3 and XSTR4, and no disease-causing mutation was identified in the coding regions of four candidate genes.

CONCLUSIONGPCR2 GPC3, MST4 and GLUD2 were excluded as pathogenic genes for SFMS. The refined SFMS locus will assist in the identification and characterization of other candidate genes for SFMS.

Abnormalities, Multiple ; genetics ; Chromosome Mapping ; Chromosomes, Human, X ; Genetic Linkage ; Glutamate Dehydrogenase ; genetics ; Glypicans ; Humans ; Intellectual Disability ; genetics ; Male ; Membrane Proteins ; genetics ; Neoplasm Proteins ; genetics ; Protein-Serine-Threonine Kinases ; genetics ; Receptors, G-Protein-Coupled ; genetics ; Syndrome

Based on the CX3C chemokine ligand 1(CX3CL1)-CX3C chemokine receptor 1(CX3CR1) axis, this study explored the potential mechanism by which Zuogui Jiangtang Jieyu Formula(ZGJTJY) improved neuroinflammation and enhanced neuroprotective effect in a rat model of diabetes mellitus complicated with depression(DD). The DD rat model was established by feeding a high-fat diet combined with streptozotocin(STZ) intraperitoneal injection for four weeks and chronic unpredictable mild stress(CUMS) combined with isolated cage rearing for five weeks. The rats were divided into a control group, a model group, a positive control group, an inhibitor group, and a ZGJTJY group. The open field test and forced swimming test were used to assess the depression-like behaviors of the rats. Enzyme-linked immunosorbent assay(ELISA) was performed to measure the expression levels of the pro-inflammatory cytokines interleukin-1β(IL-1β) and tumor necrosis factor-α(TNF-α) in plasma. Immunofluorescence staining was used to detect the expression of ionized calcium-binding adapter molecule 1(Iba1), postsynaptic density protein-95(PSD95), and synapsin-1(SYN1) in the hippocampus. Hematoxylin-eosin(HE) staining, Nissl staining, and TdT-mediated dUTP nick end labeling(TUNEL) fluorescence staining were performed to assess hippocampal neuronal damage. Western blot was used to measure the expression levels of CX3CL1, CX3CR1, A2A adenosine receptor(A2AR), glutamate receptor 2A(NR2A), glutamate receptor 2B(NR2B), and brain-derived neurotrophic factor(BDNF) in the hippocampus. Compared with the model group, the ZGJTJY group showed improved depression-like behaviors in DD rats, enhanced neuroprotective effect, increased expression of PSD95, SYN1, and BDNF(P<0.01), and decreased expression of Iba1, IL-1β, and TNF-α(P<0.01), as well as the expression of CX3CL1, CX3CR1, A2AR, NR2A, and NR2B(P<0.01). These results suggest that ZGJTJY may exert its neuroprotective effect by inhibiting the CX3CL1-CX3CR1 axis and activation of hippocampal microglia, thereby improving neuroinflammation and abnormal activation of N-methyl-D-aspartate receptor(NMDAR) subunits, and ultimately enhancing the expression of synaptic-related proteins PSD95, SYN1, and BDNF in the hippocampus.
Rats ; Animals ; Depression/drug therapy* ; Brain-Derived Neurotrophic Factor ; Neuroprotective Agents ; Tumor Necrosis Factor-alpha/metabolism* ; Neuroinflammatory Diseases ; Diabetes Mellitus ; Receptors, Glutamate ; CX3C Chemokine Receptor 1/genetics*