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Sample records for cx43 cx45 glutamate

  1. Acetylation mediates Cx43 reduction caused by electrical stimulation

    PubMed Central

    Meraviglia, Viviana; Azzimato, Valerio; Colussi, Claudia; Florio, Maria Cristina; Binda, Anna; Panariti, Alice; Qanud, Khaled; Suffredini, Silvia; Gennaccaro, Laura; Miragoli, Michele; Barbuti, Andrea; Lampe, Paul D.; Gaetano, Carlo; Pramstaller, Peter P.; Capogrossi, Maurizio C.; Recchia, Fabio A.; Pompilio, Giulio; Rivolta, Ilaria; Rossini, Alessandra

    2015-01-01

    Communication between cardiomyocytes depends upon Gap Junctions (GJ). Previous studies have demonstrated that electrical stimulation induces GJ remodeling and modifies histone acetylases (HAT) and deacetylases (HDAC) activities, although these two results have not been linked. The aim of this work was to establish whether electrical stimulation modulates GJ-mediated cardiac cell-cell communication by acetylation-dependent mechanisms. Field stimulation of HL-1 cardiomyocytes at 0.5 Hz for 24 hours significantly reduced Connexin43 (Cx43) expression and cell-cell communication. HDAC activity was down-regulated whereas HAT activity was not modified resulting in increased acetylation of Cx43. Consistent with a post-translational mechanism, we did not observe a reduction in Cx43 mRNA in electrically stimulated cells, while the proteasomal inhibitor MG132 maintained Cx43 expression. Further, the treatment of paced cells with the HAT inhibitor Anacardic Acid maintained both the levels of Cx43 and cell-cell communication. Finally, we observed increased acetylation of Cx43 in the left ventricles of dogs subjected to chronic tachypacing as a model of abnormal ventricular activation. In conclusion, our findings suggest that altered electrical activity can regulate cardiomyocyte communication by influencing the acetylation status of Cx43. PMID:26264759

  2. Involvement of connexin43 in the infrasonic noise-induced glutamate release by cultured astrocytes.

    PubMed

    Jiang, Shan; Wang, Yong-Qiang; Xu, Cheng-Feng; Li, Ya-Na; Guo, Rong; Li, Ling

    2014-05-01

    Infrasonic noise/infrasound is a type of environmental noise that threatens public health as a nonspecific biological stressor. Glutamate-related excitotoxicity is thought to be responsible for infrasound-induced impairment of learning and memory. In addition to neurons, astrocytes are also capable of releasing glutamate. In the present study, to identify the effect of infrasound on astroglial glutamate release, cultured astrocytes were exposed to infrasound at 16 Hz, 130 dB for different times. We found that infrasound exposure caused a significant increase in glutamate levels in the extracellular fluid. Moreover, blocking the connexin43 (Cx43) hemichannel or gap junction, decreasing the probability of Cx43 being open or inhibiting of Cx43 expression blocked this increase. The results suggest that glutamate release by Cx43 hemichannels/gap junctions is involved in the response of cultured astrocytes to infrasound. PMID:24634254

  3. Sulforaphane counteracts aggressiveness of pancreatic cancer driven by dysregulated Cx43-mediated gap junctional intercellular communication

    PubMed Central

    Zhang, Yiyao; Isayev, Orkhan; Heilmann, Katharina; Schoensiegel, Frank; Liu, Li; Nessling, Michelle; Richter, Karsten; Labsch, Sabrina; Nwaeburu, Clifford C.; Mattern, Juergen; Gladkich, Jury; Giese, Nathalia; Werner, Jens; Schemmer, Peter; Gross, Wolfgang; Gebhard, Martha M.; Gerhauser, Clarissa; Schaefer, Michael; Herr, Ingrid

    2014-01-01

    The extreme aggressiveness of pancreatic ductal adenocarcinoma (PDA) has been associated with blocked gap junctional intercellular communication (GJIC) and the presence of cancer stem cells (CSCs). We examined whether disturbed GJIC is responsible for a CSC phenotype in established and primary cancer cells and patient tissue of PDA using interdisciplinary methods based in physiology, cell and molecular biology, histology and epigenetics. Flux of fluorescent dyes and gemcitabine through gap junctions (GJs) was intact in less aggressive cells but not in highly malignant cells with morphological dysfunctional GJs. Among several connexins, only Cx43 was expressed on the cell surface of less aggressive and GJIC-competent cells, whereas Cx43 surface expression was absent in highly malignant, E-cadherin-negative and GJIC-incompetent cells. The levels of total Cx43 protein and Cx43 phosphorylated at Ser368 and Ser279/282 were high in normal tissue but low to absent in malignant tissue. si-RNA-mediated inhibition of Cx43 expression in GJIC-competent cells prevented GJIC and induced colony formation and the expression of stem cell-related factors. The bioactive substance sulforaphane enhanced Cx43 and E-cadherin levels, inhibited the CSC markers c-Met and CD133, improved the functional morphology of GJs and enhanced GJIC. Sulforaphane altered the phosphorylation of several kinases and their substrates and inhibition of GSK3, JNK and PKC prevented sulforaphane-induced CX43 expression. The sulforaphane-mediated expression of Cx43 was not correlated with enhanced Cx43 RNA expression, acetylated histone binding and Cx43 promoter de-methylation, suggesting that posttranslational phosphorylation is the dominant regulatory mechanism. Together, the absence of Cx43 prevents GJIC and enhances aggressiveness, whereas sulforaphane counteracts this process, and our findings highlight dietary co-treatment as a viable treatment option for PDA. PMID:24742583

  4. HPV16 E6 Controls the Gap Junction Protein Cx43 in Cervical Tumour Cells

    PubMed Central

    Sun, Peng; Dong, Li; MacDonald, Alasdair I.; Akbari, Shahrzad; Edward, Michael; Hodgins, Malcolm B.; Johnstone, Scott R.; Graham, Sheila V.

    2015-01-01

    Human papillomavirus type 16 (HPV16) causes a range of cancers including cervical and head and neck cancers. HPV E6 oncoprotein binds the cell polarity regulator hDlg (human homologue of Drosophila Discs Large). Previously we showed in vitro, and now in vivo, that hDlg also binds Connexin 43 (Cx43), a major component of gap junctions that mediate intercellular transfer of small molecules. In HPV16-positive non-tumour cervical epithelial cells (W12G) Cx43 localised to the plasma membrane, while in W12T tumour cells derived from these, it relocated with hDlg into the cytoplasm. We now provide evidence that E6 regulates this cytoplasmic pool of Cx43. E6 siRNA depletion in W12T cells resulted in restoration of Cx43 and hDlg trafficking to the cell membrane. In C33a HPV-negative cervical tumour cells expressing HPV16 or 18 E6, Cx43 was located primarily in the cytoplasm, but mutation of the 18E6 C-terminal hDlg binding motif resulted in redistribution of Cx43 to the membrane. The data indicate for the first time that increased cytoplasmic E6 levels associated with malignant progression alter Cx43 trafficking and recycling to the membrane and the E6/hDlg interaction may be involved. This suggests a novel E6-associated mechanism for changes in Cx43 trafficking in cervical tumour cells. PMID:26445057

  5. Selectivity of Cx43 channels is regulated through PKC-dependent phosphorylation

    PubMed Central

    Ek-Vitorin, Jose F.; King, Timothy J.; Heyman, Nathanael S.; Lampe, Paul D.; Burt, Janis M.

    2006-01-01

    Coordinated contractile activation of the heart and resistance to ischemic injury depend, in part, on the intercellular communication mediated by Cx43-composed gap junctions. The function of these junctions is regulated at multiple levels (assembly to degradation) through phosphorylation at specific sites in the carboxyl terminus (CT) of the Cx43 protein. We show here that the selective permeability of Cx43 junctions is regulated through PKC-dependent phosphorylation at serine 368 (S368). Selective permeability was measured in several Cx43-expressing cell lines as the rate constant for intercellular dye diffusion relative to junctional conductance. The selective permeability of Cx43 junctions under control conditions was quite variable, as was the open state behavior of the comprising channels. Co-expression of the CT of Cx43 as a distinct protein, treatment with a PKC inhibitor, or mutation of S368 to alanine all reduced (or eliminated) phosphorylation at S368, reduced the incidence of 55–70pS channels, and reduced by ten fold the selective permeability of the junctions for a small cationic dye. Since PKC activation during pre-ischemic conditioning is cardio-protective during subsequent ischemic episodes, we examined no-flow, ischemic hearts for Cx43 phosphorylated at S368 (pS368). Consistent with early activation of PKC, pS368-Cx43 was increased in ischemic hearts; despite extensive lateralization of total Cx43, pS368-Cx43 remained predominantly at intercalated disks. Our data suggest that the selectivity of gap junction channels at intercalated disks is increased early in ischemia. PMID:16709897

  6. The role of the Cx43 C-terminus in GJ plaque formation and internalization

    SciTech Connect

    Wayakanon, Praween; Bhattacharjee, Rajib; Nakahama, Ken-ichi; Morita, Ikuo

    2012-04-06

    Highlights: Black-Right-Pointing-Pointer Cx43-GFP or -DsRed fusion proteins were expressed in HeLa cells. Black-Right-Pointing-Pointer Roles of C-terminus were examined using various mutants. Black-Right-Pointing-Pointer Gap junction plaque size was dependent on the length of C-terminus. Black-Right-Pointing-Pointer C-terminus dependent gap junction plaque internalization was observed. -- Abstract: Connexin 43 (Cx43) is a major gap junction (GJ) protein found in many mammalian cell types. The C-terminal (CT) domain of Cx43 has unique characteristics in terms of amino acid (aa) sequence and its length differs from other connexins. This CT domain can be associated with protein partners to regulate GJ assembly and degradation, which results in the direct control of gap junction intercellular communication (GJIC). However, the essential roles of the CT regions involved in these mechanisms have not been fully elucidated. In this study, we aimed to investigate the specific regions of Cx43CT involved in GJ formation and internalization. Wild type Cx43{sub (382aa)} and 10 CT truncated mutants were stably expressed in HeLa cells as GFP or DsRed tagged proteins. First, we found that the deletion of 235-382aa from Cx43 resulted in failure to make GJ and establish GJIC. Second, the Cx43 with 242-382aa CT deletion could form functional GJs and be internalized as annular gap junctions (AGJs). However, the plaques consisting of Cx43 with CT deletions ({Delta}242-382aa to {Delta}271-382aa) were longer than the plaques consisting of Cx43 with CT deletions ({Delta}302-382aa). Third, co-culture experiments of cells expressing wild type Cx43{sub (382)} with cells expressing Cx43CT mutants revealed that the directions of GJ internalization were dependent on the length of the respective CT. Moreover, a specific region, 325-342aa residues of Cx43, played an important role in the direction of GJ internalization. These results showed the important roles of the Cx43 C-terminus in GJ

  7. Clathrin and Cx43 gap junction plaque endoexocytosis

    SciTech Connect

    Nickel, Beth M.; DeFranco, B. Hewa; Gay, Vernon L.; Murray, Sandra A.

    2008-10-03

    In earlier transmission electron microscopic studies, we have described pentilaminar gap junctional membrane invaginations and annular gap junction vesicles coated with short, electron-dense bristles. The similarity between these electron-dense bristles and the material surrounding clathrin-coated pits led us to suggest that the dense bristles associated with gap junction structures might be clathrin. To confirm that clathrin is indeed associated with annular gap junction vesicles and gap junction plaques, quantum dot immuno-electron microscopic techniques were used. We report here that clathrin associates with both connexin 43 (Cx43) gap junction plaques and pentilaminar gap junction vesicles. An important finding was the preferential localization of clathrin to the cytoplasmic surface of the annular or of the gap junction plaque membrane of one of the two contacting cells. This is consistent with the possibility that the direction of gap junction plaque internalization into one of two contacting cells is regulated by clathrin.

  8. Activated microglia impairs neuroglial interaction by opening Cx43 hemichannels in hippocampal astrocytes.

    PubMed

    Abudara, Verónica; Roux, Lisa; Dallérac, Glenn; Matias, Isabelle; Dulong, Jérôme; Mothet, Jean Pierre; Rouach, Nathalie; Giaume, Christian

    2015-05-01

    Glia plays an active role in neuronal functions and dysfunctions, some of which depend on the expression of astrocyte connexins, the gap junction channel and hemichannel proteins. Under neuroinflammation triggered by the endotoxin lipopolysacharide (LPS), microglia is primary stimulated and releases proinflammatory agents affecting astrocytes and neurons. Here, we investigate the effects of such microglial activation on astrocyte connexin-based channel functions and their consequences on synaptic activity in an ex vivo model. We found that LPS induces astroglial hemichannel opening in acute hippocampal slices while no change is observed in gap junctional communication. Based on pharmacological and genetic approaches we found that the LPS-induced hemichannel opening is mainly due to Cx43 hemichannel activity. This process primarily requires a microglial stimulation resulting in the release of at least two proinflammatory cytokines, IL-1β and TNF-α. Consequences of the hemichannel-mediated increase in membrane permeability are a calcium rise in astrocytes and an enhanced glutamate release associated to a reduction in excitatory synaptic activity of pyramidal neurons in response to Schaffer's collateral stimulation. As a whole our findings point out astroglial hemichannels as key determinants of the impairment of synaptic transmission during neuroinflammation.

  9. Manipulating Cx43 expression triggers gene reprogramming events in dermal fibroblasts from oculodentodigital dysplasia patients.

    PubMed

    Esseltine, Jessica L; Shao, Qing; Huang, Tao; Kelly, John J; Sampson, Jacinda; Laird, Dale W

    2015-11-15

    Oculodentodigital dysplasia (ODDD) is primarily an autosomal dominant disorder linked to over 70 GJA1 gene [connexin43 (Cx43)] mutations. For nearly a decade, our laboratory has been investigating the relationship between Cx43 and ODDD by expressing disease-linked mutants in reference cells, tissue-relevant cell lines, 3D organ cultures and by using genetically modified mouse models of human disease. Although salient features of Cx43 mutants have been revealed, these models do not necessarily reflect the complexity of the human context. To further overcome these limitations, we have acquired dermal fibroblasts from two ODDD-affected individuals harbouring D3N and V216L mutations in Cx43, along with familial controls. Using these ODDD patient dermal fibroblasts, which naturally produce less GJA1 gene product, along with RNAi and RNA activation (RNAa) approaches, we show that manipulating Cx43 expression triggers cellular gene reprogramming. Quantitative RT-PCR, Western blot and immunofluorescent analysis of ODDD patient fibroblasts show unusually high levels of extracellular matrix (ECM)-interacting proteins, including integrin α5β1, matrix metalloproteinases as well as secreted ECM proteins collagen-I and laminin. Cx43 knockdown in familial control cells produces similar effects on ECM expression, whereas Cx43 transcriptional up-regulation using RNAa decreases production of collagen-I. Interestingly, the enhanced levels of ECM-associated proteins in ODDD V216L fibroblasts is not only a consequence of increased ECM gene expression, but also due to an apparent deficit in collagen-I secretion which may further contribute to impaired collagen gel contraction in ODDD fibroblasts. These findings further illuminate the altered function of Cx43 in ODDD-affected individuals and highlight the impact of manipulating Cx43 expression in human cells. PMID:26349540

  10. Cx43-hemichannel function and regulation in physiology and pathophysiology: insights from the bovine corneal endothelial cell system and beyond

    PubMed Central

    D'hondt, Catheleyne; Iyyathurai, Jegan; Himpens, Bernard; Leybaert, Luc; Bultynck, Geert

    2014-01-01

    Intercellular communication in primary bovine corneal endothelial cells (BCECs) is mainly driven by the release of extracellular ATP through Cx43 hemichannels. Studying the characteristics of Ca2+-wave propagation in BCECs, an important form of intercellular communication, in response to physiological signaling events has led to the discovery of important insights in the functional properties and regulation of native Cx43 hemichannels. Together with ectopic expression models for Cx43 hemichannels and truncated/mutated Cx43 versions, it became very clear that loop/tail interactions play a key role in controlling the activity of Cx43 hemichannels. Interestingly, the negative regulation of Cx43 hemichannels by enhanced actin/myosin contractility seems to impinge upon loss of these loop/tail interactions essential for opening Cx43 hemichannels. Finally, these molecular insights have spurred the development of novel peptide tools that can selectively inhibit Cx43 hemichannels, but neither Cx43 gap junctions nor hemichannels formed by other Cx isoforms. These tools now set the stage to hunt for novel physiological functions for Cx43 hemichannels in primary cells and tissues and to tackle disease conditions associated with excessive, pathological Cx43-hemichannel openings. PMID:25309448

  11. Cx43-hemichannel function and regulation in physiology and pathophysiology: insights from the bovine corneal endothelial cell system and beyond.

    PubMed

    D'hondt, Catheleyne; Iyyathurai, Jegan; Himpens, Bernard; Leybaert, Luc; Bultynck, Geert

    2014-01-01

    Intercellular communication in primary bovine corneal endothelial cells (BCECs) is mainly driven by the release of extracellular ATP through Cx43 hemichannels. Studying the characteristics of Ca(2+)-wave propagation in BCECs, an important form of intercellular communication, in response to physiological signaling events has led to the discovery of important insights in the functional properties and regulation of native Cx43 hemichannels. Together with ectopic expression models for Cx43 hemichannels and truncated/mutated Cx43 versions, it became very clear that loop/tail interactions play a key role in controlling the activity of Cx43 hemichannels. Interestingly, the negative regulation of Cx43 hemichannels by enhanced actin/myosin contractility seems to impinge upon loss of these loop/tail interactions essential for opening Cx43 hemichannels. Finally, these molecular insights have spurred the development of novel peptide tools that can selectively inhibit Cx43 hemichannels, but neither Cx43 gap junctions nor hemichannels formed by other Cx isoforms. These tools now set the stage to hunt for novel physiological functions for Cx43 hemichannels in primary cells and tissues and to tackle disease conditions associated with excessive, pathological Cx43-hemichannel openings.

  12. Association of STAT3 with Cx26 and Cx43 in human uterine endometrioid adenocarcinoma

    PubMed Central

    SULKOWSKA, URSZULA; FEBP, ANDRZEJ WINCEWICZ; SULKOWSKI, STANISLAW

    2016-01-01

    Signal transducer and activator of transcription-3 (STAT3) drives endometrial carcinogenesis, while signaling via gap junctions gets weakened during cancer progression. Connexin 26 (Cx26), Cx43 and STAT3 were immunohistochemically evaluated in 78 endometrioid adenocarcinomas: Nuclear expression of STAT3 positively correlated with cytoplasmic immunoreactivity to Cx43 (P=0.004, r=0.318) and Cx26 (P=0.006, r=0.309). STAT3 correlated with Cx43 (P=0.022, r=0.411) and Cx26 (P=0.008 r=0.466) in G1 tumors. A statistically significant linkage remained in G2 cancers between STAT3 and Cx43 (P=0.061, r=0.262) and Cx26 (P=0.016, r=0.331); however, no correlations were observed in G3 tumors. STAT3 was significantly associated with Cx 43 (p=0.003, r=0.684) and Cx26 (p=0.049, r=0.500) in estrogen receptor (ER) negative adenocarcinomas. STAT3 did not correlate with Cx43 in ER positive adenocarcinomas; however, STAT3 expression remained correlated with Cx26 expression (P=0.035, r=0.268). In progesterone receptor negative tumors STAT3 was significantly associated with Cx43 (P=0.035, r=0.451) and Cx26 (P<0.0001, r=0.707). However, in PgR positive adenocarcinomas STAT3 correlated with Cx43 (P=0.03, r=0.290) but not with Cx26. Thus, it appears that hormone dependent acceleration of cancer growth breaks the association between STAT3 and Cx expression. These associations become weaker as the tumors dedifferentiate from G1 to G3 endometrioid adenocarcinomas. The present study provides evidence that the loss of correlation between STAT3 and selected Cx proteins occurs in tumors with more aggressive behavior. PMID:27313754

  13. Green Fluorescent Protein Changes the Conductance of Connexin 43 (Cx43) Hemichannels Reconstituted in Planar Lipid Bilayers*

    PubMed Central

    Carnarius, Christian; Kreir, Mohamed; Krick, Marcel; Methfessel, Christoph; Moehrle, Volker; Valerius, Oliver; Brüggemann, Andrea; Steinem, Claudia; Fertig, Niels

    2012-01-01

    In mammalian tissues, connexin 43 (Cx43) is the most prominent member of the connexin family. In a single lipid bilayer, six connexin subunits assemble into a hemichannel (connexon). Direct communication of apposing cells is realized by two adjacent hemichannels, which can form gap junction channels. Here, we established an expression system in Pichia pastoris to recombinantly produce and purify Cx43 as well as Cx43 fused to green fluorescent protein (GFP). Proteins were isolated from crude cell membrane fractions via affinity chromatography. Cx43 and Cx43-GFP hemichannels were reconstituted in giant unilamellar vesicles as proven by fluorescence microscopy, and their electrophysiological behavior was analyzed on the single channel level by planar patch clamping. Cx43 and Cx43-GFP both showed an ohmic behavior and a voltage-dependent open probability. Cx43 hemichannels exhibited one major mean conductance of 224 ± 26 picosiemens (pS). In addition, a subconductance state at 124 ± 5 pS was identified. In contrast, the analysis of Cx43-GFP single channels revealed 10 distinct conductance states in the range of 15 to 250 pS, with a larger open probability at 0 mV as compared with Cx43, which suggests that intermolecular interactions between the GFP molecules alter the electrophysiology of the protein. PMID:22139870

  14. ATP Release from Vascular Endothelia Occurs Across Cx43 Hemichannels and Is Attenuated during Hypoxia

    PubMed Central

    Zug, Stephanie; El Kasmi, Karim C.; Eltzschig, Holger K.

    2008-01-01

    Background Extracellular ATP is an important signaling molecule for vascular adaptation to limited oxygen availability (hypoxia). Here, we pursued the contribution of vascular endothelia to extracellular ATP release under hypoxic conditions. Methodology, Principal Findings We gained first insight from studying ATP release from endothelia (HMEC-1) pre-exposed to hypoxia. Surprisingly, we found that ATP release was significantly attenuated following hypoxia exposure (2% oxygen, 22±3% after 48 h). In contrast, intracellular ATP was unchanged. Similarly, lactate-dehydrogenase release into the supernatants was similar between normoxic or hypoxic endothelia, suggesting that differences in lytic ATP release between normoxia or hypoxia are minimal. Next, we used pharmacological strategies to study potential mechanisms for endothelial-dependent ATP release (eg, verapamil, dipyridamole, 18-alpha-glycyrrhetinic acid, anandamide, connexin-mimetic peptides). These studies revealed that endothelial ATP release occurs – at least in part - through connexin 43 (Cx43) hemichannels. A real-time RT-PCR screen of endothelial connexin expression showed selective repression of Cx43 transcript and additional studies confirmed time-dependent Cx43 mRNA, total and surface protein repression during hypoxia. In addition, hypoxia resulted in Cx43-serine368 phosphorylation, which is known to switch Cx43 hemi-channels from an open to a closed state. Conclusions/Significance Taken together, these studies implicate endothelial Cx43 in hypoxia-associated repression of endothelial ATP release. PMID:18665255

  15. Neurological manifestations of oculodentodigital dysplasia: a Cx43 channelopathy of the central nervous system?

    PubMed Central

    De Bock, Marijke; Kerrebrouck, Marianne; Wang, Nan; Leybaert, Luc

    2013-01-01

    The coordination of tissue function is mediated by gap junctions (GJs) that enable direct cell–cell transfer of metabolic and electric signals. GJs are formed by connexins of which Cx43 is most widespread in the human body. In the brain, Cx43 GJs are mostly found in astroglia where they coordinate the propagation of Ca2+ waves, spatial K+ buffering, and distribution of glucose. Beyond its role in direct intercellular communication, Cx43 also forms unapposed, non-junctional hemichannels in the plasma membrane of glial cells. These allow the passage of several neuro- and gliotransmitters that may, combined with downstream paracrine signaling, complement direct GJ communication among glial cells and sustain glial-neuronal signaling. Mutations in the GJA1 gene encoding Cx43 have been identified in a rare, mostly autosomal dominant syndrome called oculodentodigital dysplasia (ODDD). ODDD patients display a pleiotropic phenotype reflected by eye, hand, teeth, and foot abnormalities, as well as craniofacial and bone malformations. Remarkably, neurological symptoms such as dysarthria, neurogenic bladder (manifested as urinary incontinence), spasticity or muscle weakness, ataxia, and epilepsy are other prominent features observed in ODDD patients. Over 10 mutations detected in patients diagnosed with neurological disorders are associated with altered functionality of Cx43 GJs/hemichannels, but the link between ODDD-related abnormal channel activities and neurologic phenotype is still elusive. Here, we present an overview on the nature of the mutants conveying structural and functional changes of Cx43 channels and discuss available evidence for aberrant Cx43 GJ and hemichannel function. In a final step, we examine the possibilities of how channel dysfunction may lead to some of the neurological manifestations of ODDD. PMID:24133447

  16. Dynamic changes in protein interaction between AKAP95 and Cx43 during cell cycle progression of A549 cells

    PubMed Central

    Chen, Xiaoxuan; Kong, Xiangyu; Zhuang, Wenxin; Teng, Bogang; Yu, Xiuyi; Hua, Suhang; Wang, Su; Liang, Fengchao; Ma, Dan; Zhang, Suhui; Zou, Xuan; Dai, Yue; Yang, Wei; Zhang, Yongxing

    2016-01-01

    Here we show that A-kinase anchoring protein 95 (AKAP95) and connexin 43 (Cx43) dynamically interact during cell cycle progression of lung cancer A549 cells. Interaction between AKAP95 and Cx43 at different cell cycle phases was examined by tandem mass spectrometry(MS/MS), confocal immunofluorescence microscopy, Western blot, and co-immunoprecipitation(Co-IP). Over the course of a complete cell cycle, interaction between AKAP95 and Cx43 occurred in two stages: binding stage from late G1 to metaphase, and separating stage from anaphase to late G1. The binding stage was further subdivided into complex binding to DNA in interphase and complex separating from DNA in metaphase. In late G1, Cx43 translocated to the nucleus via AKAP95; in anaphase, Cx43 separated from AKAP95 and aggregated between two daughter nuclei. In telophase, Cx43 aggregated at the membrane of the cleavage furrow. After mitosis, Cx43 was absent from the furrow membrane and was located in the cytoplasm. Binding between AKAP95 and Cx43 was reduced by N-(2-[P-Bromocinnamylamino]-ethyl)-5-isoquinolinesulfonmide (H89) treatment and enhanced by Forskolin. dynamic interaction between AKAP95 and Cx43 varies with cell cycle progression to regulate multiple biological processes. PMID:26880274

  17. Specific functional pathologies of Cx43 mutations associated with oculodentodigital dysplasia

    PubMed Central

    Kelly, John J.; Esseltine, Jessica L.; Shao, Qing; Jabs, Ethylin Wang; Sampson, Jacinda; Auranen, Mari; Bai, Donglin; Laird, Dale W.

    2016-01-01

    Oculodentodigital dysplasia (ODDD) is a rare genetic disease that affects the development of multiple organs in the human body. More than 70 mutations in the gap junction connexin43 (Cx43) gene, GJA1, are associated with ODDD, most of which are inherited in an autosomal dominant manner. Many patients exhibit similar clinical presentations. However, there is high intrafamilial and interfamilial phenotypic variability. To better understand this variability, we established primary human dermal fibroblast cultures from several ODDD patients and unaffected controls. In the present study, we characterized three fibroblast lines expressing heterozygous p.L7V, p.G138R, and p.G143S Cx43 variants. All ODDD fibroblasts exhibited slower growth, reduced migration, and defective cell polarization, traits common to all ODDD fibroblasts studied so far. However, we found striking differences in overall expression levels, with p.L7V down-regulated at the mRNA and protein level. Although all of the Cx43 variants could traffic to the cell surface, there were stark differences in gap junction plaque formation, gap junctional intercellular communication, Cx43 phosphorylation, and hemichannel activity among Cx43 variants, as well as subtle differences in myofibroblast differentiation. Together these findings enabled us to discover mutation-specific pathologies that may help to predict future clinical outcomes. PMID:27226478

  18. Effects of ghrelin on Cx43 regulation and electrical remodeling after myocardial infarction in rats.

    PubMed

    Yuan, Ming-Jie; Huang, He; Tang, Yan-Hong; Wu, Gang; Gu, Yong-Wei; Chen, Yong-Jun; Huang, Cong-Xin

    2011-11-01

    Ghrelin is a novel growth hormone-releasing peptide, which has been shown to exert beneficial effects on ventricular remodeling. In this study, we investigated whether ghrelin could decrease vulnerability to ventricular arrhythmias in rats with myocardial infarction and the possible mechanism. Twenty-four hours after ligation of the anterior descending artery, adult male Sprague-Dawley rats were randomized to ghrelin (100 μg/kg) and saline (control group) for 4 weeks. Sham animals underwent thoracotomy and pericardiotomy, but not LAD ligation. Myocardial endothelin-1 (ET-1) levels were significantly elevated in saline-treated rats at the border zone compared with sham-operated rats. Myocardial connexin43 (Cx43) expression at the border zone was significantly decreased in saline-treated infarcted rats compared with sham-operated rats. Ghrelin significantly decreased the inducibility of ventricular tachyarrhythmias compared with control group. Arrhythmias sores during programmed stimulation in saline-treated rats were significantly higher than scores in those treated with ghrelin. The electrophysiological improvement of fatal ventricular tachyarrhythmias was accompanied with increased immunofluorescence-stained Cx43, myocardial Cx43 protein and mRNA levels in ghrelin treated rats. We also shown that ghrelin significantly decreased tissue ET-1 levels at the infarcted border zone. Thus, ghrelin showed the protective effect on ventricular arrhythmias after myocardial infarction. Although the precise mechanism by which ghrelin modulates the dephosphorylation of Cx43 remains unknown, it is most likely that the ghrelin increased expression of Cx43 through the inhibition of ET-1.

  19. Connexin-deficiency affects expression levels of glial glutamate transporters within the cerebrum.

    PubMed

    Unger, Tina; Bette, Stefanie; Zhang, Jiong; Theis, Martin; Engele, Jürgen

    2012-01-01

    The glial glutamate transporter subtypes, GLT-1/EAAT-2 and GLAST/EAAT-1 clear the bulk of extracellular glutamate and are severely dysregulated in various acute and chronic brain diseases. Despite the previous identification of several extracellular factors modulating glial glutamate transporter expression, our knowledge of the regulatory network controlling glial glutamate transport in health and disease still remains incomplete. In studies with cultured cortical astrocytes, we previously obtained evidence that glial glutamate transporter expression is also affected by gap junctions/connexins. To assess whether gap junctions would likewise control the in vivo expression of glial glutamate transporters, we have now assessed their expression levels in brains of conditional Cx43 knockout mice, total Cx30 knockouts, as well as Cx43/Cx30 double knockouts. We found that either knocking out Cx30, Cx43, or both increases GLT-1/EAAT-2 protein levels in the cerebral cortex to a similar extent. By contrast, GLAST/EAAT-1 protein levels maximally increased in cerebral cortices of Cx30/Cx43 double knockouts, implying that gap junctions differentially affect the expression of GLT-1/EAAT-2 and GLAST/EAAT-1. Quantitative PCR analysis further revealed that increases in glial glutamate transporter expression are brought about by transcriptional and translational/posttranslational processes. Moreover, GLT-1/EAAT-2- and GLAST/EAAT-1 protein levels remained unchanged in the hippocampi of Cx43/Cx30 double knockouts when compared to Cx43fl/fl controls, indicating brain region-specific effects of gap junctions on glial glutamate transport. Since astrocytic gap junction coupling is affected in various forms of brain injuries, our findings point to gap junctions/connexins as important regulators of glial glutamate turnover in the diseased cerebral cortex.

  20. Prenatal nicotine exposure enhances Cx43 and Panx1 unopposed channel activity in brain cells of adult offspring mice fed a high-fat/cholesterol diet

    PubMed Central

    Orellana, Juan A.; Busso, Dolores; Ramírez, Gigliola; Campos, Marlys; Rigotti, Attilio; Eugenín, Jaime; von Bernhardi, Rommy

    2014-01-01

    Nicotine, the most important neuroteratogen of tobacco smoke, can reproduce brain and cognitive disturbances per se when administered prenatally. However, it is still unknown if paracrine signaling among brain cells participates in prenatal nicotine-induced brain impairment of adult offspring. Paracrine signaling is partly mediated by unopposed channels formed by connexins hemichannels (HCs) and pannexins serving as aqueous pores permeable to ions and small signaling molecules, allowing exchange between the intra- and extracellular milieus. Our aim was to address whether prenatal nicotine exposure changes the activity of those channels in adult mice offspring under control conditions or subjected to a second challenge during young ages: high-fat/cholesterol (HFC) diet. To induce prenatal exposure to nicotine, osmotic minipumps were implanted in CF1 pregnant mice at gestational day 5 to deliver nicotine bitartrate or saline (control) solutions. After weaning, offspring of nicotine-treated or untreated pregnant mice were fed ad libitum with chow or HFC diets for 8 weeks. The functional state of connexin 43 (Cx43) and pannexin 1 (Panx1) unopposed channels was evaluated by dye uptake experiments in hippocampal slices from 11-week-old mice. We found that prenatal nicotine increased the opening of Cx43 HCs in astrocytes, and Panx1 channels in microglia and neurons only if offspring mice were fed with HFC diet. Blockade of inducible nitric oxide synthase (iNOS), cyclooxygenase 2 (COX2) and prostaglandin E receptor 1 (EP1), ionotropic ATP receptor type 7 (P2X7) and NMDA receptors, showed differential inhibition of prenatal nicotine-induced channel opening in glial cells and neurons. Importantly, inhibition of the above mentioned enzymes and receptors, or blockade of Cx43 and Panx1 unopposed channels greatly reduced adenosine triphosphate (ATP) and glutamate release from hippocampal slices of prenatally nicotine-exposed offspring. We propose that unregulated gliotransmitter

  1. Functional expression of Ca²⁺ dependent mammalian transmembrane gap junction protein Cx43 in slime mold Dictyostelium discoideum.

    PubMed

    Kaufmann, Stefan; Weiss, Ingrid M; Eckstein, Volker; Tanaka, Motomu

    2012-03-01

    In this paper, we expressed murine gap junction protein Cx43 in Dictyostelium discoideum by introducing the specific vector pDXA. In the first step, the successful expression of Cx43 and Cx43-eGFP was verified by (a) Western blot (anti-Cx43, anti-GFP), (b) fluorescence microscopy (eGFP-Cx43 co-expression, Cx43 immunostaining), and (c) flow cytometry analysis (eGFP-Cx43 co-expression). Although the fluorescence signals from cells expressing Cx43-eGFP detected by fluorescence microscopy seem relatively low, analysis by flow cytometry demonstrated that more than 60% of cells expressed Cx43-eGFP. In order to evaluate the function of expressed Cx43 in D. discoideum, we examined the hemi-channel function of Cx43. In this series of experiments, the passive uptake of carboxyfluorescein was monitored using flow cytometric analysis. A significant number of the transfected cells showed a prominent dye uptake in the absence of Ca(2+). The dye uptake by transfected cells in the presence of Ca(2+) was even lower than the non-specific dye uptake by non-transformed Ax3 orf+ cells, confirming that Cx43 expressed in D. discoideum retains its Ca(2+)-dependent, specific gating function. The expression of gap junction proteins expressed in slime molds opens a possibility to the biological significance of intercellular communications in development and maintenance of multicellular organisms.

  2. Functional expression of Ca²⁺ dependent mammalian transmembrane gap junction protein Cx43 in slime mold Dictyostelium discoideum.

    PubMed

    Kaufmann, Stefan; Weiss, Ingrid M; Eckstein, Volker; Tanaka, Motomu

    2012-03-01

    In this paper, we expressed murine gap junction protein Cx43 in Dictyostelium discoideum by introducing the specific vector pDXA. In the first step, the successful expression of Cx43 and Cx43-eGFP was verified by (a) Western blot (anti-Cx43, anti-GFP), (b) fluorescence microscopy (eGFP-Cx43 co-expression, Cx43 immunostaining), and (c) flow cytometry analysis (eGFP-Cx43 co-expression). Although the fluorescence signals from cells expressing Cx43-eGFP detected by fluorescence microscopy seem relatively low, analysis by flow cytometry demonstrated that more than 60% of cells expressed Cx43-eGFP. In order to evaluate the function of expressed Cx43 in D. discoideum, we examined the hemi-channel function of Cx43. In this series of experiments, the passive uptake of carboxyfluorescein was monitored using flow cytometric analysis. A significant number of the transfected cells showed a prominent dye uptake in the absence of Ca(2+). The dye uptake by transfected cells in the presence of Ca(2+) was even lower than the non-specific dye uptake by non-transformed Ax3 orf+ cells, confirming that Cx43 expressed in D. discoideum retains its Ca(2+)-dependent, specific gating function. The expression of gap junction proteins expressed in slime molds opens a possibility to the biological significance of intercellular communications in development and maintenance of multicellular organisms. PMID:22330805

  3. Novel Pharmacophores of Connexin43 Based on the “RXP” Series of Cx43-binding Peptides

    PubMed Central

    Verma, Vandana; Larsen, Bjarne Due; Coombs, Wanda; Lin, Xianming; Spagnol, Gaelle; Sorgen, Paul L; Taffet, Steven M; Delmar, Mario

    2009-01-01

    Gap junction pharmacology is a nascent field. Previous studies have identified molecules that enhance intercellular communication, and may offer potential for innovative antiarrhythmic therapy. However, their specific molecular target(s) and mechanism(s) of action remain unknown. Previously, we identified a 34-amino acid peptide (RXP-E) that binds the carboxyl terminal domain of Cx43 (Cx43CT) and prevents cardiac gap junction closure and action potential propagation block. These results supported the feasibility of a peptide-based pharmacology to Cx43, but the structure of the core active element in RXP-E, an essential step for pharmacological development, remained undefined. Here, we used a combination of molecular modeling, surface plasmon resonance, nuclear magnetic resonance and patch clamp strategies to define, for the first time, a unique ensemble of pharmacophores that bind Cx43CT and prevent closure of Cx43 channels. Two particular molecules are best representatives of this family: a cyclized heptapeptide (called CyRP-71), and a linear octapeptide of sequence RRNYRRNY. These two small compounds offer the first structural platform for the design of Cx43-interacting gap junction openers. Moreover, the structure of these compounds offers an imprint of a region of Cx43CT that is fundamental to gap junction channel function. PMID:19556520

  4. Intracellular Cleavage of the Cx43 C-Terminal Domain by Matrix-Metalloproteases: A Novel Contributor to Inflammation?

    PubMed Central

    De Bock, Marijke; Wang, Nan; Decrock, Elke; Bultynck, Geert; Leybaert, Luc

    2015-01-01

    The coordination of tissue function is mediated by gap junctions (GJs) that enable direct cell-cell transfer of metabolic and electric signals. GJs are formed by connexin (Cx) proteins of which Cx43 is most widespread in the human body. Beyond its role in direct intercellular communication, Cx43 also forms nonjunctional hemichannels (HCs) in the plasma membrane that mediate the release of paracrine signaling molecules in the extracellular environment. Both HC and GJ channel function are regulated by protein-protein interactions and posttranslational modifications that predominantly take place in the C-terminal domain of Cx43. Matrix metalloproteases (MMPs) are a major group of zinc-dependent proteases, known to regulate not only extracellular matrix remodeling, but also processing of intracellular proteins. Together with Cx43 channels, both GJs and HCs, MMPs contribute to acute inflammation and a small number of studies reports on an MMP-Cx43 link. Here, we build further on these reports and present a novel hypothesis that describes proteolytic cleavage of the Cx43 C-terminal domain by MMPs and explores possibilities of how such cleavage events may affect Cx43 channel function. Finally, we set out how aberrant channel function resulting from cleavage can contribute to the acute inflammatory response during tissue injury. PMID:26424967

  5. Effects of substitution of Cx43 by Cx32 on myocardial energy metabolism, tolerance to ischaemia and preconditioning protection

    PubMed Central

    Rodríguez-Sinovas, Antonio; Sánchez, Jose A; González-Loyola, Alejandra; Barba, Ignasi; Morente, Miriam; Aguilar, Rio; Agulló, Esperanza; Miró-Casas, Elisatet; Esquerda, Neus; Ruiz-Meana, Marisol; García-Dorado, David

    2010-01-01

    Connexin 43 (Cx43) plays an important role in cardioprotective signalling by mechanisms at least in part independent of gap junctional communication. To investigate whether this role is related to specific properties of this connexin isoform, we used a knock-in mouse model in which the coding region of Cx43 is replaced by that of Cx32. Homozygous Cx43KI32 mice showed reduced cell-to-cell Lucifer Yellow transfer (P < 0.01), but QRS duration and left ventricular fractional shortening (echocardiography) were similar to those in wild-type animals. NMR spectroscopy detected reduced ATP and increased lactate content in myocardium from homozygous Cx43KI32 animals (P < 0.05). Despite this, isolated homozygous Cx43KI32 hearts showed smaller infarcts after ischaemia–reperfusion (40 min/60 min) as compared to hearts from heterozygous and wild-type animals (13 and 31% reduction, respectively, P < 0.05). Cardiac myocytes isolated from Cx43KI32 mouse hearts also showed a reduced rate of cell death after simulated ischaemia–reperfusion. In a separate series of experiments, both ischaemic (4 cycles of 3.5 min of ischaemia and 5 min of reperfusion) and pharmacological (50 μmol l−1 diazoxide, 10 min) preconditioning reduced infarct size in hearts from wild-type mice (by 24.84 and 26.63%, respectively, P < 0.05), but only ischaemic preconditioning was effective in hearts from heterozygous animals and both preconditioning strategies failed to protect Cx43KI32 homozygous hearts. These results demonstrate that Cx43 has an important and previously unknown modulatory effect in myocardial energy metabolism and tolerance to ischaemia, and plays a critical role in preconditioning protection, by mechanisms that are specific for this connexin isoform. PMID:20156849

  6. Cx43 expression and function in the nervous system—implications for stem cell mediated regeneration

    PubMed Central

    Meier, Carola; Rosenkranz, Katja

    2014-01-01

    Pathological conditions of the brain such as ischemia cause major sensorimotor and cognitive impairments. In novel therapeutic approaches to brain injury, stem cells have been applied to ameliorate the pathological outcome. In several experimental models, including hypoxia-ischemia and trauma, transplantation of stem cells correlated with an improved functional and structural outcome. At the cellular level, brain insults also change gap junction physiology and expression, leading to altered intercellular communication. Differences in expression in response to brain injury have been detected in particular in Cx43, the major astrocytic gap junction protein, and its overexpression or deletion was associated with the pathophysiological outcome. We here focus on Cx43 changes in host tissue mediated by stem cells. Stem cell-induced changes in connexin expression, and consecutively in gap junction channel or hemichannel function, might play a part in altered cell interaction, intercellular communication, and neural cell survival, and thereby contribute to the beneficial effects of transplanted stem cells. PMID:24672489

  7. The severity of mammary gland developmental defects is linked to the overall functional status of Cx43 as revealed by genetically modified mice

    PubMed Central

    Stewart, Michael K. G.; Gong, Xiang-Qun; Barr, Kevin J.; Bai, Donglin; Fishman, Glenn I.; Laird, Dale W.

    2012-01-01

    Genetically modified mice mimicking ODDD (oculodentodigital dysplasia), a disease characterized by reduced Cx43 (connexin 43)-mediated gap junctional intercellular communication, represent an in vivo model to assess the role of Cx43 in mammary gland development and function. We previously reported that severely compromised Cx43 function delayed mammary gland development and impaired milk ejection in mice that harboured a G60S Cx43 mutant, yet there are no reports of lactation defects in ODDD patients. To address this further, we obtained a second mouse model of ODDD expressing an I130T Cx43 mutant to assess whether a mutant with partial gap junction channel activity would be sufficient to retain mammary gland development and function. The results of the present study show that virgin Cx43I130T/+ mice exhibited a temporary delay in ductal elongation at 4 weeks. In addition, Cx43I130T/+ mice develop smaller mammary glands at parturition due to reduced cell proliferation despite similar overall gland architecture. Distinct from Cx43G60S/+ mice, Cx43I130T/+ mice adequately produce and deliver milk to pups, suggesting that milk ejection is unaffected. Thus the present study suggests that a loss-of-function mutant of Cx43 with partial gap junction channel coupling conductance results in a less severe mammary gland phenotype, which may partially explain the lack of reported lactation defects associated with ODDD patients. PMID:23075222

  8. RXP-E: A CX43-BINDING PEPTIDE THAT PREVENTS ACTION POTENTIAL PROPAGATION BLOCK

    PubMed Central

    Lewandowski, Rebecca; Procida, Kristina; Vaidyanathan, Ravi; Coombs, Wanda; Jalife, Jose; Nielsen, Morten S.; Taffet, Steven M.; Delmar, Mario

    2009-01-01

    Gap junctions (GJs) provide a low-resistance pathway for cardiac electrical propagation. The role of GJ regulation in arrhythmia is unclear, partly due to limited availability of pharmacological tools. Recently, we showed that a peptide called “RXP-E” binds to the carboxyl terminal of connexin43 (Cx43) and prevents chemically-induced uncoupling in Cx43-expressing N2a cells. Here, pull-down experiments show RXP-E binding to adult cardiac Cx43. Patch-clamp studies revealed that RXP-E prevented heptanol-induced and acidification-induced uncoupling in pairs of neonatal rat ventricular myocytes (NRVM’s). Separately, RXP-E was concatenated to a cytoplasmic transduction peptide for cytoplasmic translocation (CTP-RXP-E). The effect of RXP-E on action potential (AP) propagation was assessed by high resolution optical mapping in monolayers of NRVM’s, containing ~20% of randomly distributed myofibroblasts. In contrast to control experiments, when heptanol (2 mmol/L) was added to the superfusate of monolayers loaded with CTP-RXP-E, AP propagation was maintained, albeit at a slower velocity. Similarly, intracellular acidification (pHi=6.2) caused a loss of AP propagation in control monolayers; however, propagation was maintained in CTP-RXP-E treated cells, though at a slower rate. Patch clamp experiments revealed that RXP-E did not prevent heptanol-induced block of sodium currents, nor did it alter voltage dependence or amplitude of Kir2.1/Kir2.3 currents. RXP-E is the first synthetic molecule known to: (1) bind cardiac Cx43; (2) prevent heptanol and acidification-induced uncoupling of cardiac GJ’s and 3) preserve AP propagation among cardiac myocytes. RXP-E can be used to characterize the role of GJs in the function of multicellular systems, including the heart. PMID:18669919

  9. The anti-metastatic micro-environment of the bone: Importance of osteocyte Cx43 hemichannels.

    PubMed

    Bultynck, Geert

    2016-08-01

    Bone metastases of tumor cells are a common and life-threatening feature of a variety of late-stage cancers, including breast cancers. However, until now, much less has been known about the intrinsic anti-metastatic properties of the bones and how these could be exploited to prevent or treat bone metastases. Very recently, native Cx43 hemichannels present in osteocytes have been identified as important anti-metastatic signaling complexes by establishing high local extracellular ATP levels. Moreover, bisphosphonate drugs, applied as adjuvant therapies in the treatment of breast cancer patients and bone diseases, are known to display anti-metastatic properties. Now, it became clear that these compounds exert their effects through osteocyte Cx43 hemichannels, thereby triggering their opening and promoting ATP release in the extracellular micro-environment. Hence, endogenous osteocyte Cx43 hemichannels emerge as important and promising therapeutic targets for the prevention of bone metastases and/or clinical treatment of bone-metastasized breast cancers.

  10. Anti-Arrhythmic Effect of Verapamil Is Accompanied by Preservation of Cx43 Protein in Rat Heart

    PubMed Central

    Wang, Qiu-Lin; Wu, Qi; Chen, Mai; Pei, Jian-Ming

    2013-01-01

    The present study was to test the hypothesis that anti-arrhythmic properties of verapamil may be accompanied by preserving connexin43 (Cx43) protein via calcium influx inhibition. In an in vivo study, myocardial ischemic arrhythmia was induced by occlusion of the left anterior descending (LAD) coronary artery for 45 min in Sprague-Dawley rats. Verapamil, a calcium channel antagonist, was injected i.v. into a femoral vein prior to ischemia. Effects of verapamil on arrhythmias induced by Bay K8644 (a calcium channel agonist) were also determined. In an ex vivo study, the isolated heart underwent an initial 10 min of baseline normal perfusion and was subjected to high calcium perfusion in the absence or presence of verapamil. Cardiac arrhythmia was measured by electrocardiogram (ECG) and Cx43 protein was determined by immunohistochemistry and western blotting. Administration of verapamil prior to myocardial ischemia significantly reduced the incidence of ventricular arrhythmias and total arrhythmia scores, with the reductions in heat rate, mean arterial pressure and left ventricular systolic pressure. Verapamil also inhibited arrhythmias induced by Bay K8644 and high calcium perfusion. Effect of verapamil on ischemic arrhythmia scores was abolished by heptanol, a Cx43 protein uncoupler and Gap 26, a Cx43 channels inhibitor. Immunohistochemistry data showed that ischemia-induced redistribution and reduced immunostaining of Cx43 were prevented by verapamil. In addition, diminished expression of Cx43 protein determined by western blotting was observed following myocardial ischemia in vivo or following high calcium perfusion ex vivo and was preserved after verapamil administration. Our data suggest that verapamil may confer an anti-arrhythmic effect via calcium influx inhibition, inhibition of oxygen consumption and accompanied by preservation of Cx43 protein. PMID:23951191

  11. Influence of the antiandrogen flutamide on connexin 43 (Cx43) gene and protein expression in the porcine placenta and uterus during pregnancy.

    PubMed

    Wieciech, Iwona; Grzesiak, Małgorzata; Knapczyk-Stwora, Katarzyna; Pytlik, Anna; Słomczynska, Maria

    2014-01-01

    The study focuses on the expression of connexin 43 (Cx43), a gap junctional protein in the porcine placenta and uterus. The aim was to examine Cx43 mRNA and protein expression after antiandrogen flutamide treatment. Flutamide was injected into pregnant gilts at a daily dose of 50 mg/kg body weight at different stages of pregnancy: between days 43-49 (50 dpc), 83-89 (90 dpc) and 101-107 (108 dpc) of gestation. The animals were sacrificed and tissues were collected one day after the last injection. Cx43 immunostaining was observed in epithelial and stromal cells of the fetal part of the placenta; luminal and glandular epithelial cells of maternal part of the placenta and myometrium of the uterus within placentation sites. Cx43 was also found in glandular epithelium and myometrium of non-placental uterus. Flutamide treatment caused fluctuations in Cx43 expression especially before parturition. Although significant changes in Cx43 mRNA expression were observed only in the fetal part of the placenta, Cx43 protein level was affected within the maternal part of the placenta and non-placental uterus. These results suggest the involvement of androgens in the regulation of Cx43 expression within the feto-maternal compartment in pigs. However, androgen deficiency caused pronounced changes during late pregnancy and before parturition. These results are interesting due to the functional changes in the porcine uterus during the preparturient period that is determined by Cx43 protein.

  12. Connexin targeting peptides as inhibitors of voltage- and intracellular Ca2+-triggered Cx43 hemichannel opening.

    PubMed

    Wang, Nan; De Bock, Marijke; Decrock, Elke; Bol, Mélissa; Gadicherla, Ashish; Bultynck, Geert; Leybaert, Luc

    2013-12-01

    Connexins form gap junctions that function as intercellular channels and hemichannels that form a conduit between the cytoplasm and extracellular fluid when open. Peptide inhibitors of connexin channels, especially those identical to defined connexin sequences, are interesting experimental, and possibly also therapeutic tools because they may have better selectivity than general inhibitors like carbenoxolone. Over the past ten years, several peptides have been demonstrated to block hemichannels, including Gap26, Gap27, peptide5, L2 and Gap19; some of these specifically block hemichannels but not gap junctions. Most of these peptides have only recently been investigated towards their actions at the single hemichannel level, bringing up interesting information on how they interact with the connexin protein and how they affect hemichannel gating. Hemichannels can be opened by electrical, mechanical and chemical stimuli. We here review the effect of the prototypic peptides Gap26/27 and L2/Gap19 with specific focus on their inhibition of Cx43 hemichannel opening triggered by positive membrane potentials and changes in intracellular Ca2+ concentration. Both Gap26/27 and L2/Gap19 peptide families block Cx43 hemichannel opening triggered by voltage as well as intracellular Ca2+ stimulation. Interestingly, these peptides as well as intracellular Ca2+ elevation modulate the voltage activation threshold for hemichannel opening, pointing to a common target. Moreover, L2 and Gap19 peptides are part of a sequence on the cytoplasmic loop that acts as a Ca2+/calmodulin interaction site. We here review the interesting network of interactions between Cx43 targeting peptides, voltage gating and intracellular Ca2+ as major modulators of hemichannel function. This article is part of the Special Issue Section entitled 'Current Pharmacology of Gap Junction Channels and Hemichannels'.

  13. Synergistic effects of AKAP95, Cyclin D1, Cyclin E1, and Cx43 in the development of rectal cancer

    PubMed Central

    Qi, Fengjie; Yuan, Yangyang; Zhi, Xuehong; Huang, Qian; Chen, Yuexin; Zhuang, Wenxin; Zhang, Dengcheng; Teng, Bogang; Kong, Xiangyu; Zhang, Yongxing

    2015-01-01

    Objective: To explore the expression of A-kinase anchor protein 95 (AKAP95), Cyclin D1, Cyclin E1, and Connexin43 (Cx43) in rectal cancer tissues and assess the associations between each of the proteins and pathological parameters, as well as their inter-relationships. Methods: AKAP95, Cyclin D1, Cyclin E1, and Cx43 protein expression rates were evaluated by immunohistochemistry in 50 rectal cancer specimens and 16 pericarcinoma tissues. Results: The positive rates of AKAP95, Cyclin E1, and Cyclin D1 proteins were 54.00 vs. 18.75%, 62.00 vs. 6.25%, and 72.00 vs. 31.25% in rectal cancer specimens and pericarcinoma tissues, respectively, representing statistically significant differences (P < 0.05). The positive rate of Cx43 protein expression in rectal cancer tissues was 44.00% and 62.50% in pericarcinoma tissues, and the difference between them was not significant (P > 0.05). No significant associations were found between protein expression of AKAP95, Cyclin E1, Cyclin D1, and Cx43, and the degree of differentiation, histological type, and lymph node metastasis of rectal cancer (P > 0.05). However, significant correlations were obtained between the expression rates of AKAP95 and Cyclin E1, Cyclin E1 and Cyclin D1, Cyclin E1 and Cx43 protein, and Cyclin D1 and Cx43, respectively (P < 0.05). Conclusion: AKAP95, Cyclin E1, and Cyclin D1 protein expression rates were significantly higher in rectal cancer tissues compared with pericarcinoma samples, suggesting an association between these proteins and the development and progression of rectal cancer. In addition, the significant correlations between the proteins (AKAP95 and Cyclin E1, Cyclin E1 and Cyclin D1, Cyclin E1 and Cx43 protein, and Cyclin D1 and Cx43) indicate the possible synergistic effects of these factors in the development and progression of rectal cancer. PMID:25973052

  14. Quantitative Analysis of ZO-1 Colocalization with Cx43 Gap Junction Plaques in Cultures of Rat Neonatal Cardiomyocytes

    NASA Astrophysics Data System (ADS)

    Zhu, Ching; Barker, Ralph J.; Hunter, Andrew W.; Zhang, Yuhua; Jourdan, Jane; Gourdie, Robert G.

    2005-06-01

    The gap junction (GJ) is an aggregate of intercellular channels that facilitates cytoplasmic interchange of ions, second messengers, and other molecules of less than 1000 Da between cells. In excitable organs such as heart and brain, GJs configure extended intercellular pathways for stable and long-term propagation of action potential. In a previous study in adult rat heart, we have shown that the Drosophila disks-large related protein ZO-1 shows low to moderate colocalization at myocyte borders with the GJ protein Cx43. In the present study, we detail a protocol for characterizing the pattern and level of colocalization of ZO-1 with Cx43 in cultures of neonatal myocytes at the level of individual GJ plaques. The data indicate that ZO-1 shows on average a partial 26.6% overlap (SD = 11.3%) with Cx43 GJ plaques. There is a strong positive correlation between GJ plaque size and area of ZO-1 colocalization, indicating that the level of associated ZO-1 scales with the area of the GJ plaque. Qualitatively, the most prominent colocalization occurs at the plaque perimeter. These studies may provide insight into the presently unknown biological function of ZO-1 interaction with Cx43.

  15. Expression of the gap junction gene connexin43 (Cx43) in preimplantation bovine embryos derived in vitro or in vivo.

    PubMed

    Wrenzycki, C; Herrmann, D; Carnwath, J W; Niemann, H

    1996-09-01

    In this study we have examined the presence of mRNA encoding connexin 43 (Cx43) in bovine embryos derived in vivo and in vitro. Cumulus-oocyte complexes, immature and matured oocytes liberated from cumulus cells, zygotes, 2-4-cell and 8-16-cell embryos, morulae, blastocysts and hatched blastocysts were produced in vitro from ovaries obtained from an abattoir using TCM 199 supplemented with hormones and 10% oestrous cow serum for maturation. Cumulus-oocyte complexes matured for 24 h were exposed to bull spermatozoa for 19 h and then cultured in TCM 199 supplemented with 10% oestrous cow serum to the desired developmental stage. Morulae and blastocysts derived in vivo were collected from superovulated donor cows. Total RNA was extracted from pools of 60-200 bovine oocytes or embryos using a modified phenol-chloroform extraction method and analysed by reverse transcriptase polymerase chain reaction. Before reverse transcription, aliquots of DNase-digested embryonic RNA were tested by polymerase chain reaction using bovine-specific primers to control for residual genomic DNA contamination. DNA-free, total RNA was reverse transcribed after preincubation with the Cx43 specific 3'primer. The resultant cDNA was amplified by polymerase chain reaction using Cx43 specific primers that define a 516 bp fragment of Cx43. The reverse transcriptase polymerase chain reaction product was verified by restriction enzyme analysis with Alu I and sequencing. Assays were repeated at least twice for each developmental stage and provided identical results between replicates. Cx43 transcripts were detected in bovine morulae and blastocysts grown in vivo. In contrast, whereas the early in vitro stages from cumulus-oocyte complexes to morulae expressed Cx43, blastocysts and hatched blastocysts did not have detectable concentrations of mRNA from this gene. Restriction enzyme cutting revealed three fragments of the predicted size (139, 177, 200 bp). The amplified product showed 100% identity

  16. High Glucose Alters Cx43 Expression and Gap Junction Intercellular Communication in Retinal Müller Cells: Promotes Müller Cell and Pericyte Apoptosis

    PubMed Central

    Muto, Tetsuya; Tien, Thomas; Kim, Dongjoon; Sarthy, Vijay P.; Roy, Sayon

    2014-01-01

    Purpose. To investigate whether high glucose (HG) alters connexin 43 (Cx43) expression and gap junction intercellular communication (GJIC) activity in retinal Müller cells, and promotes Müller cell and pericyte loss. Methods. Retinal Müller cells (rMC-1) and cocultures of rMC-1 and retinal pericytes were grown in normal (N) or HG (30 mM glucose) medium. Additionally, rMC-1 transfected with Cx43 small interfering RNA (siRNA) were grown as cocultures with pericytes, and rMC-1 transfected with Cx43 plasmid were grown in HG. Expression of Cx43 was determined by Western blotting and immunostaining and GJIC was assessed by scrape-loading dye transfer (SLDT) technique. Apoptosis was analyzed by TUNEL or differential staining assay, and Akt activation by assessing Akt phosphorylation. Results. In monocultures of rMC-1 and cocultures of rMC-1 and pericytes, Cx43 protein level, number of Cx43 plaques, GJIC, and Akt phosphorylation were significantly reduced in HG medium. Number of TUNEL-positive cells was also significantly increased in rMC-1 monocultures and in rMC-1 and pericyte cocultures grown in HG medium. Importantly, when rMC-1 transfected with Cx43 siRNA were grown as cocultures with pericytes, a significant decrease in GJIC, and increase in TUNEL-positive cells was observed, concomitant with decreased Akt phosphorylation. Upregulation of Cx43 rescued rMC-1 from HG-induced apoptosis. Conclusions. Gap junction communication between Müller cells and pericytes is essential for their survival. Downregulation of Cx43 that is HG induced and impairment of GJIC activity in Müller cells contributes to loss of glial and vascular cells associated with the pathogenesis of diabetic retinopathy. PMID:24938518

  17. Internal Ribosomal Entry Site (IRES) Activity Generates Endogenous Carboxyl-terminal Domains of Cx43 and Is Responsive to Hypoxic Conditions*

    PubMed Central

    Ul-Hussain, Mahboob; Olk, Stephan; Schoenebeck, Bodo; Wasielewski, Bianca; Meier, Carola; Prochnow, Nora; May, Caroline; Galozzi, Sara; Marcus, Katrin; Zoidl, Georg; Dermietzel, Rolf

    2014-01-01

    Connexin43 (Cx43) is the most abundant gap junction protein in higher vertebrate organisms and has been shown to be involved in junctional and non-junctional functions. In addition to the expression of full-length Cx43, endogenously produced carboxyl-terminal segments of Cx43 have been described and have been suggested to be involved in manifold biological functions, such as hypoxic preconditioning and neuronal migration. Molecular aspects, however, behind the separate generation of carboxyl-terminal segments of Cx43 have remained elusive. Here we report on a mechanism that may play a key role in the separate production of these domains. First, stringent evidence derived from siRNA treatment and specific knockouts revealed significant loss of the low molecular weight fragments of Cx43. By applying a dicistronic vector strategy on transfected cell lines, we were able to identify putative IRES activity (nucleotides 442–637) in the coding region of Cx43, which resides upstream from the nucleotide sequence encoding the carboxyl terminus (nucleotides 637–1149). Functional responsiveness of the endogenous expression of Cx43 fragments to hypoxic/ischemic treatment was evaluated in in vitro and in vivo models, which led to a significant increase of the fastest migrating form (20 kDa) under conditions of metabolic deprivation. By nano-MS spectrometry, we achieved stringent evidence of the identity of the 20-kDa segment as part of the carboxyl-terminal domain of full-length Cx43. Our data prove the existence of endogenously expressed carboxyl-terminal domains, which may serve as valuable tools for further translational application in ischemic disorders. PMID:24872408

  18. Diabetes Increases Cryoinjury Size with Associated Effects on Cx43 Gap Junction Function and Phosphorylation in the Mouse Heart.

    PubMed

    Palatinus, Joseph A; Gourdie, Robert G

    2016-01-01

    Diabetic patients develop larger myocardial infarctions and have an increased risk of death following a heart attack. The poor response to myocardial injury in the diabetic heart is likely related to the many metabolic derangements from diabetes that create a poor substrate in general for wound healing, response to injury and infection. Studies in rodents have implicated a role for the gap junction protein connexin 43 (Cx43) in regulating the injury response in diabetic skin wounds. In this study, we sought to determine whether diabetes alters Cx43 molecular interactions or intracellular communication in the cryoinjured STZ type I diabetic mouse heart. We found that epicardial cryoinjury size is increased in diabetic mice and this increase is prevented by preinjury insulin administration. Consistent with these findings, we found that intercellular coupling via gap junctions is decreased after insulin administration in diabetic and nondiabetic mice. This decrease in coupling is associated with a concomitant increase in phosphorylation of Cx43 at serine 368, a residue known to decrease channel conductance. Taken together, our results suggest that insulin regulates both gap junction-mediated intercellular communication and injury propagation in the mouse heart. PMID:27034963

  19. Diabetes Increases Cryoinjury Size with Associated Effects on Cx43 Gap Junction Function and Phosphorylation in the Mouse Heart

    PubMed Central

    Palatinus, Joseph A.; Gourdie, Robert G.

    2016-01-01

    Diabetic patients develop larger myocardial infarctions and have an increased risk of death following a heart attack. The poor response to myocardial injury in the diabetic heart is likely related to the many metabolic derangements from diabetes that create a poor substrate in general for wound healing, response to injury and infection. Studies in rodents have implicated a role for the gap junction protein connexin 43 (Cx43) in regulating the injury response in diabetic skin wounds. In this study, we sought to determine whether diabetes alters Cx43 molecular interactions or intracellular communication in the cryoinjured STZ type I diabetic mouse heart. We found that epicardial cryoinjury size is increased in diabetic mice and this increase is prevented by preinjury insulin administration. Consistent with these findings, we found that intercellular coupling via gap junctions is decreased after insulin administration in diabetic and nondiabetic mice. This decrease in coupling is associated with a concomitant increase in phosphorylation of Cx43 at serine 368, a residue known to decrease channel conductance. Taken together, our results suggest that insulin regulates both gap junction-mediated intercellular communication and injury propagation in the mouse heart. PMID:27034963

  20. To beat or not to beat: degradation of Cx43 imposes the heart rhythm.

    PubMed

    Martins-Marques, Tânia; Catarino, Steve; Marques, Carla; Pereira, Paulo; Girão, Henrique

    2015-06-01

    The main function of the heart is to pump blood to the different parts of the organism, a task that is efficiently accomplished through proper electric and metabolic coupling between cardiac cells, ensured by gap junctions (GJ). Cardiomyocytes are the major cell population in the heart, and as cells with low mitotic activity, are highly dependent upon mechanisms of protein degradation. In the heart, both the ubiquitin-proteasome system (UPS) and autophagy participate in the fine-tune regulation of cardiac remodelling and function, either in physiological or pathological conditions. Indeed, besides controlling cardiac signalling pathways, UPS and autophagy have been implicated in the turnover of several myocardial proteins. Degradation of Cx43, the major ventricular GJ protein, has been associated to up-regulation of autophagy at the onset of heart ischemia and ischemia/reperfusion (I/R), which can have profound implications upon cardiac function. In this review, we present recent studies devoted to the involvement of autophagy and UPS in heart homoeostasis, with a particular focus on GJ.

  1. Keratitis-ichthyosis-deafness syndrome-associated Cx26 mutants produce nonfunctional gap junctions but hyperactive hemichannels when co-expressed with wild type Cx43.

    PubMed

    García, Isaac E; Maripillán, Jaime; Jara, Oscar; Ceriani, Ricardo; Palacios-Muñoz, Angelina; Ramachandran, Jayalakshmi; Olivero, Pablo; Perez-Acle, Tomas; González, Carlos; Sáez, Juan C; Contreras, Jorge E; Martínez, Agustín D

    2015-05-01

    Mutations in Cx26 gene are found in most cases of human genetic deafness. Some mutations produce syndromic deafness associated with skin disorders, like the Keratitis-Ichthyosis-Deafness syndrome (KID). Because in the human skin connexin 26 (Cx26) is co-expressed with other connexins, like Cx43 and Cx30, and as the KID syndrome is inherited as autosomal dominant condition, it is possible that KID mutations change the way Cx26 interacts with other co-expressed connexins. Indeed, some Cx26 syndromic mutations showed gap junction dominant negative effect when co-expressed with wild-type connexins, including Cx26 and Cx43. The nature of these interactions and the consequences on hemichannels and gap junction channel (GJC) functions remain unknown. In this study, we demonstrate that syndromic mutations, at the N terminus segment of Cx26, change connexin oligomerization compatibility, allowing aberrant interactions with Cx43. Strikingly, heteromeric oligomer formed by Cx43/Cx26 (syndromic mutants) shows exacerbated hemichannel activity but nonfunctional GJCs; this also occurs for those Cx26 KID mutants that do not show functional homomeric hemichannels. Heterologous expression of these hyperactive heteromeric hemichannels increases cell membrane permeability, favoring ATP release and Ca(2+) overload. The functional paradox produced by oligomerization of Cx43 and Cx26 KID mutants could underlie the severe syndromic phenotype in human skin.

  2. Keratitis-Ichthyosis-Deafness syndrome-associated Cx26 mutants produce nonfunctional gap junctions but hyperactive hemichannels when co-expressed with wild type Cx43

    PubMed Central

    García, Isaac E.; Maripillán, Jaime; Jara, Oscar; Ceriani, Ricardo; Palacios-Muñoz, Angelina; Ramachandran, Jayalakshimi; Olivero, Pablo; Pérez-Acle, Tomás; González, Carlos; Sáez, Juan C.; Contreras, Jorge E.; Martínez, Agustín D.

    2015-01-01

    Mutations in Cx26 gene are found in most cases of human genetic deafness. Some mutations produce syndromic deafness associated with skin disorders, like Keratitis Ichthyosis Deafness syndrome (KID). Because in the human skin Cx26 is co-expressed with other connexins, like Cx43 and Cx30, and since KID syndrome is inherited as autosomal dominant condition, it is possible that KID mutations change the way Cx26 interacts with other co-expressed connexins. Indeed, some Cx26 syndromic mutations showed gap junction dominant negative effect when co-expressed with wild type connexins, including Cx26 and Cx43. The nature of these interactions and the consequences on hemichannels and gap junction channels functions remain unknown. In this study we demonstrate that syndromic mutations at the N-terminus segment of Cx26, change connexin oligomerization compatibility, allowing aberrant interactions with Cx43. Strikingly, heteromeric oligomer formed by Cx43/Cx26 (syndromic mutants) show exacerbated hemichannel activity, but nonfunctional gap junction channels; this also occurs for those Cx26 KID mutants that do not show functional homomeric hemichannels. Heterologous expression of these hyperactive heteromeric hemichannels increases cell membrane permeability, favoring ATP release and Ca2+ overload. The functional paradox produced by oligomerization of Cx43 and Cx26 KID mutants could underlie the severe syndromic phenotype in human skin. PMID:25625422

  3. Changes in Cx43 and NaV1.5 expression precede the occurrence of substantial fibrosis in calcineurin-induced murine cardiac hypertrophy.

    PubMed

    Fontes, Magda S C; Raaijmakers, Antonia J A; van Doorn, Tessa; Kok, Bart; Nieuwenhuis, Sylvia; van der Nagel, Roel; Vos, Marc A; de Boer, Teun P; van Rijen, Harold V M; Bierhuizen, Marti F A

    2014-01-01

    In mice, the calcium-dependent phosphatase calcineurin A (CnA) induces a transcriptional pathway leading to pathological cardiac hypertrophy. Interestingly, induction of CnA has been frequently noticed in human hypertrophic and failing hearts. Independently, the arrhythmia vulnerability of such hearts has been regularly associated with remodeling of parameters determining electrical conduction (expression level of connexin43 (Cx43) and NaV1.5, connective tissue architecture), for which the precise molecular basis and sequence of events is still unknown. Recently, we observed reduced Cx43 and NaV1.5 expression in 4-week old mouse hearts, overexpressing a constitutively active form of CnA (MHC-CnA model), but the order of events is still unknown. Therefore, three key parameters of conduction (Cx43, NaV1.5 and connective tissue expression) were characterized in MHC-CnA ventricles versus wild-type (WT) during postnatal development on a weekly basis. At postnatal week 1, CnA overexpression induced cardiac hypertrophy in MHC-CnA. Moreover, protein and RNA levels of both Cx43 and NaV1.5 were reduced by at least 50% as compared to WT. Cx43 immunoreactive signal was reduced at week 2 in MHC-CnA. At postnatal week 3, Cx43 was less phosphorylated and RNA level of Cx43 normalized to WT values, although the protein level was still reduced. Additionally, MHC-CnA hearts displayed substantial fibrosis relative to WT, which was accompanied by increased RNA levels for genes previously associated with fibrosis such as Col1a1, Col1a2, Col3a1, Tgfb1, Ctgf, Timp1 and microRNA miR-21. In MHC-CnA, reduction in Cx43 and NaV1.5 expression thus coincided with overexpression of CnA and hypertrophy development and preceded significant presence of fibrosis. At postnatal week 4 the alterations in conductional parameters observed in the MHC-CnA model lead to abnormal conduction and arrhythmias, similar to those observed in cardiac remodeling in heart failure patients. The MHC-CnA model, therefore

  4. Effect of FSH and LH hormones on oocyte maturation of buffalo and gene expression analysis of their receptors and Cx43 in maturing oocytes.

    PubMed

    Pandey, Alok; Gupta, S C; Gupta, Neelam

    2010-08-01

    Follicle stimulating hormone (FSH) and luteinizing hormone (LH) are commonly added to maturation media to improve cumulus expansion known to be a predictor of oocyte maturation. Therefore, effects of various concentrations of FSH (1000 ng/ml), LH (1000 ng/ml) and FSH + LH (1000 ng/ml each) in comparison with control (without FSH + LH) cultured oocytes were investigated. FSH and LH (1000 ng/ml each) induced significantly more cumulus expansion and polar body numbers, as compared with control and treatments of 1000 ng/ml FSH and 1000 ng/ml LH alone. Expression of FSH receptor (r), LHr and Cx43 mRNAs was determined by real-time PCR in cumulus-oocyte complexes (COCs) and denuded oocytes at different maturation times. Expression of all three genes was higher in COCs compared with denuded oocytes, confirming the importance of cumulus cells in oocyte maturation. FSHr and connexin 43 (Cx43) mRNA abundance in both COCs and denuded oocytes was highest at 0-6 h of maturation and decreased subsequently. However, LHr mRNA abundance increased from 6 h up to 24 h of maturation. The study concluded that FSH, LH receptors and Cx43 gene expression regulation is an index related to oocyte maturation. PMID:20128947

  5. Cx43-Dependent Skeletal Phenotypes Are Mediated by Interactions between the Hapln1a-ECM and Sema3d during Fin Regeneration

    PubMed Central

    Govindan, Jayalakshmi; Tun, Kyaw Min; Iovine, M. Kathryn

    2016-01-01

    Skeletal development is a tightly regulated process and requires proper communication between the cells for efficient exchange of information. Analysis of fin length mutants has revealed that the gap junction protein Connexin43 (Cx43) coordinates cell proliferation (growth) and joint formation (patterning) during zebrafish caudal fin regeneration. Previous studies have shown that the extra cellular matrix (ECM) protein Hyaluronan and Proteoglycan Link Protein1a (Hapln1a) is molecularly and functionally downstream of Cx43, and that hapln1a knockdown leads to reduction of the glycosaminoglycan hyaluronan. Here we find that the proteoglycan aggrecan is similarly reduced following Hapln1a knockdown. Notably, we demonstrate that both hyaluronan and aggrecan are required for growth and patterning. Moreover, we provide evidence that the Hapln1a-ECM stabilizes the secreted growth factor Semaphorin3d (Sema3d), which has been independently shown to mediate Cx43 dependent phenotypes during regeneration. Double knockdown of hapln1a and sema3d reveal synergistic interactions. Further, hapln1a knockdown phenotypes were rescued by Sema3d overexpression. Therefore, Hapln1a maintains the composition of specific components of the ECM, which appears to be required for the stabilization of at least one growth factor, Sema3d. We propose that the Hapln1a dependent ECM provides the required conditions for Sema3d stabilization and function. Interactions between the ECM and signaling molecules are complex and our study demonstrates the requirement for components of the Hapln1a-ECM for Sema3d signal transduction. PMID:26828861

  6. Phosphorylation of Ser-279/282 and Tyr-265 positions on Cx43 as possible mediators of VEGF-165 inhibition of pregnancy-adapted Ca2+ burst function in ovine uterine artery endothelial cells.

    PubMed

    Boeldt, Derek S; Grummer, Mary A; Yi, FuXian; Magness, Ronald R; Bird, Ian M

    2015-09-01

    Normal pregnancy requires increased uterine endothelial cell driven vasodilation that is related to increases in sustained Ca2+ signaling via increased connexin 43 (Cx43) gap junction function. Preeclampsia, a hypertensive disorder of pregnancy associated with endothelial dysfunction, is also linked with down regulation of Ca2+ driven vasodilator production and increased levels of vascular endothelial growth factor (VEGF). Cx43 function can be acutely down-regulated by phosphorylation of multiple inhibitory residues and VEGF is known to promote phosphorylation of Cx43. Herein, we show that VEGF-165 promotes Cx43 phosphorylation at Ser-279/282 and Tyr-265 residues and blocks pregnancy-adapted Ca2+ signaling in ovine uterine artery endothelial cells (UAEC). Pharmacological Src and ERK kinase pathway inhibitors (PP2 and U0126) reverse these phosphorylations and rescue Ca2+ signaling. We also report a nutraceutical Src inhibitor, t10,c12 conjugated linoleic acid (10,12 CLA), rescues Ca2+ signaling in UAEC and therefore may have therapeutic potential for preeclampsia.

  7. Connexin45 interacts with zonula occludens-1 in osteoblastic cells

    NASA Technical Reports Server (NTRS)

    Laing, J. G.; Manley-Markowski, R. N.; Koval, M.; Civitelli, R.; Steinberg, T. H.

    2001-01-01

    Connexin43 (Cx43) and Cx45 are co-expressed in a number of different tissues. Studies demonstrated that Cx45 transfected ROS (ROS/Cx45) cells, were less permeable to low molecular weight dyes than untransfected ROS cells, that have gap junctions made of Cx43. This suggests that there may be a functionally important interaction between Cx43 and Cx45 in these cells. One way in which these proteins may interact is by associating with the same set of proteins. In order to isolate connexin interacting proteins, we isolated Cx45 from Cx45 transfected ROS cells (ROS/Cx45 cells) under mild detergent conditions. These studies showed that Cx45 co-purified with the tight junction protein, ZO-1. Immunofluorescence studies of ROS/Cx45 cells simultaneously stained with polyclonal Cx45 antibody and a monoclonal ZO-1 antibody showed that Cx45 and ZO-1 colocalized in ROS/Cx45 cells. Furthermore we found that ZO-1 could bind to peptides derived from the carboxyl terminal of Cx45 that had been covalently bound to an agarose resin. These data suggests that Cx45 and ZO-1 directly interact in ROS/Cx45 cells.

  8. Segregated Foxc2, NFATc1 and Connexin expression at normal developing venous valves, and Connexin-specific differences in the valve phenotypes of Cx37, Cx43, and Cx47 knockout mice.

    PubMed

    Munger, Stephanie J; Geng, Xin; Srinivasan, R Sathish; Witte, Marlys H; Paul, David L; Simon, Alexander M

    2016-04-15

    Venous valves (VVs) are critical for unidirectional blood flow from superficial and deep veins towards the heart. Congenital valve aplasia or agenesis may, in some cases, be a direct cause of vascular disease, motivating an understanding of the molecular mechanisms underlying the development and maintenance of VVs. Three gap junction proteins (Connexins), Cx37, Cx43, and Cx47, are specifically expressed at VVs in a highly polarized fashion. VVs are absent from adult mice lacking Cx37; however it is not known if Cx37 is required for the initial formation of valves. In addition, the requirement of Cx43 and Cx47 for VV development has not been studied. Here, we provide a detailed description of Cx37, Cx43, and Cx47 expression during mouse vein development and show by gene knockout that each Cx is necessary for normal valve development. The valve phenotypes in the knockout lines exhibit Cx-specific differences, however, including whether peripheral or central VVs are affected by gene inactivation. In addition, we show that a Cx47 null mutation impairs peripheral VV development but does not affect lymphatic valve formation, a finding of significance for understanding how some CX47 mutations cause inherited lymphedema in humans. Finally, we demonstrate a striking segregation of Foxc2 and NFATc1 transcription factor expression between the downstream and upstream faces, respectively, of developing VV leaflets and show that this segregation is closely associated with the highly polarized expression of Cx37, Cx43, and Cx47. The partition of Foxc2 and NFATc1 expression at VV leaflets makes it unlikely that these factors directly cooperate during the leaflet elongation stage of VV development.

  9. Dexamethasone-induced muscular atrophy is mediated by functional expression of connexin-based hemichannels.

    PubMed

    Cea, Luis A; Balboa, Elisa; Puebla, Carlos; Vargas, Aníbal A; Cisterna, Bruno A; Escamilla, Rosalba; Regueira, Tomás; Sáez, Juan C

    2016-10-01

    Long-term treatment with high glucocorticoid doses induces skeletal muscle atrophy. However, the molecular mechanism of such atrophy remains unclear. We evaluated the possible involvement of connexin-based hemichannels (Cx HCs) in muscle atrophy induced by dexamethasone (DEX), a synthetic glucocorticoid, on control (Cx43(fl/fl)Cx45(fl/fl)) and Cx43/Cx45 expression-deficient (Cx43(fl/fl)Cx45(fl/fl):Myo-Cre) skeletal myofibers. Myofibers of Cx43(fl/fl)Cx45(fl/fl) mice treated with DEX (5h) expressed several proteins that form non-selective membrane channels (Cx39, Cx43, Cx45, Panx1, P2X7 receptor and TRPV2). After 5h DEX treatment in vivo, myofibers of Cx43(fl/fl)Cx45(fl/fl) mice showed Evans blue uptake, which was absent in myofibers of Cx43(fl/fl)Cx45(fl/fl):Myo-Cre mice. Similar results were obtained in vitro using ethidium as an HC permeability probe, and DEX-induced dye uptake in control myofibers was blocked by P2X7 receptor inhibitors. DEX also induced a significant increase in basal intracellular Ca(2+) signal and a reduction in resting membrane potential in Cx43(fl/fl)Cx45(fl/fl) myofibers, changes that were not elicited by myofibers deficient in Cx43/Cx45 expression. Moreover, treatment with DEX induced NFκB activation and increased mRNA levels of TNF-α in control but not in Cx43/Cx45 expression-deficient myofibers. Finally, a prolonged DEX treatment (7days) increased atrogin-1 and Murf-1 and reduced the cross sectional area of Cx43(fl/fl)Cx45(fl/fl) myofibers, but these parameters remained unaffected in Cx43(fl/fl)Cx45(fl/fl):Myo-Cre myofibers. Therefore, DEX-induced expression of Cx43 and Cx45 plays a critical role in early sarcolemma changes that lead to atrophy. Consequently, this side effect of chronic glucocorticoid treatment might be avoided by co-administration with a Cx HC blocker.

  10. Relating specific connexin co-expression ratio to connexon composition and gap junction function.

    PubMed

    Desplantez, T; Grikscheit, K; Thomas, N M; Peters, N S; Severs, N J; Dupont, E

    2015-12-01

    Cardiac connexin 43 (Cx43), Cx40 and Cx45 are co-expressed at distinct ratios in myocytes. This pattern is considered a key factor in regulating the gap junction channels composition, properties and function and remains poorly understood. This work aims to correlate gap junction function with the connexin composition of the channels at accurate ratios Cx43:Cx40 and Cx43:Cx45. Rat liver epithelial cells that endogenously express Cx43 were stably transfected to induce expression of accurate levels of Cx40 or Cx45 that may be present in various areas of the heart (e.g. atria and ventricular conduction system). Induction of Cx40 does not increase the amounts of junctional connexins (Cx43 and Cx40), whereas induction of Cx45 increases the amounts of junctional connexins (Cx43 and Cx45). Interestingly, the non-junctional fraction of Cx43 remains unaffected upon induction of Cx40 and Cx45. Co-immunoprecipitation studies show low level of Cx40/Cx43 heteromerisation and undetectable Cx45/Cx43 heteromerisation. Functional characterisation shows that induction of Cx40 and Cx45 decreases Lucifer Yellow transfer. Electrical coupling is decreased by Cx45 induction, whereas it is decreased at low induction of Cx40 and increased at high induction. These data indicate a fine regulation of the gap junction channel make-up in function of the type and the ratio of co-expressed Cxs that specifically regulates chemical and electrical coupling. This reflects specific gap junction function in regulating impulse propagation in the healthy heart, and a pro-arrhythmic potential of connexin remodelling in the diseased heart. PMID:26550940

  11. Glutamate and Neurodegenerative Disease

    NASA Astrophysics Data System (ADS)

    Schaeffer, Eric; Duplantier, Allen

    As the main excitatory neurotransmitter in the mammalian central nervous system, glutamate is critically involved in most aspects of CNS function. Given this critical role, it is not surprising that glutamatergic dysfunction is associated with many CNS disorders. In this chapter, we review the literature that links aberrant glutamate neurotransmission with CNS pathology, with a focus on neurodegenerative diseases. The biology and pharmacology of the various glutamate receptor families are discussed, along with data which links these receptors with neurodegenerative conditions. In addition, we review progress that has been made in developing small molecule modulators of glutamate receptors and transporters, and describe how these compounds have helped us understand the complex pharmacology of glutamate in normal CNS function, as well as their potential for the treatment of neurodegenerative diseases.

  12. Glutamate signalling in roots.

    PubMed

    Forde, Brian G

    2014-03-01

    As a signalling molecule, glutamate is best known for its role as a fast excitatory neurotransmitter in the mammalian nervous system, a role that requires the activity of a family of ionotropic glutamate receptors (iGluRs). The unexpected discovery in 1998 that Arabidopsis thaliana L. possesses a family of iGluR-related (GLR) genes laid the foundations for an assessment of glutamate's potential role as a signalling molecule in plants that is still in progress. Recent advances in elucidating the function of Arabidopsis GLR receptors has revealed similarities with iGluRs in their channel properties, but marked differences in their ligand specificities. The ability of plant GLR receptors to act as amino-acid-gated Ca(2+) channels with a broad agonist profile, combined with their expression throughout the plant, makes them strong candidates for a multiplicity of amino acid signalling roles. Although root growth is inhibited in the presence of a number of amino acids, only glutamate elicits a specific sequence of changes in growth, root tip morphology, and root branching. The recent finding that the MEKK1 gene is a positive regulator of glutamate sensitivity at the root tip has provided genetic evidence for the existence in plants of a glutamate signalling pathway analogous to those found in animals. This short review will discuss the most recent advances in understanding glutamate signalling in roots, considering them in the context of previous work in plants and animals.

  13. II. Glutamine and glutamate.

    PubMed

    Tapiero, H; Mathé, G; Couvreur, P; Tew, K D

    2002-11-01

    Glutamine and glutamate with proline, histidine, arginine and ornithine, comprise 25% of the dietary amino acid intake and constitute the "glutamate family" of amino acids, which are disposed of through conversion to glutamate. Although glutamine has been classified as a nonessential amino acid, in major trauma, major surgery, sepsis, bone marrow transplantation, intense chemotherapy and radiotherapy, when its consumption exceeds its synthesis, it becomes a conditionally essential amino acid. In mammals the physiological levels of glutamine is 650 micromol/l and it is one of the most important substrate for ammoniagenesis in the gut and in the kidney due to its important role in the regulation of acid-base homeostasis. In cells, glutamine is a key link between carbon metabolism of carbohydrates and proteins and plays an important role in the growth of fibroblasts, lymphocytes and enterocytes. It improves nitrogen balance and preserves the concentration of glutamine in skeletal muscle. Deamidation of glutamine via glutaminase produces glutamate a precursor of gamma-amino butyric acid, a neurotransmission inhibitor. L-Glutamic acid is a ubiquitous amino acid present in many foods either in free form or in peptides and proteins. Animal protein may contain from 11 to 22% and plants protein as much as 40% glutamate by weight. The sodium salt of glutamic acid is added to several foods to enhance flavor. L-Glutamate is the most abundant free amino acid in brain and it is the major excitatory neurotransmitter of the vertebrate central nervous system. Most free L-glutamic acid in brain is derived from local synthesis from L-glutamine and Kreb's cycle intermediates. It clearly plays an important role in neuronal differentiation, migration and survival in the developing brain via facilitated Ca++ transport. Glutamate also plays a critical role in synaptic maintenance and plasticity. It contributes to learning and memory through use-dependent changes in synaptic efficacy and

  14. Production of D-Glutamate from L-Glutamate with Glutamate Racemase and L-Glutamate Oxidase.

    PubMed

    Oikawa, T; Watanabe, M; Makiura, H; Kusakabe, H; Yamade, K; Soda, K

    1999-01-01

    We studied production of D-glutamate from L-glutamate using a bioreactor consisting of two columns of sequentially connected immobilized glutamate racemase (EC 5.1.1.3, from Bacillus subtilis IFO 3336) and L-glutamate oxidase (EC 1.4.3.11, from Streptomyces sp. X119-6): L-glutamate was racemized by the glutamate racemase column, and then L-glutamate was oxidized by the L-glutamate oxidase column. Consequently only D-glutamate remained, and was easily separated from the α-ketoglutarate formed by anion-exchange chromatography. Both enzymes were highly stabilized by immobilization. The pH and temperature optima of immobilized glutamate racemase (pH 8, 40°C) were similar to those of immobilized L-glutamate oxidase (pH 7, 50°C). Accordingly, we connected the two columns tandemly to do both enzyme reactions under the same conditions. Actually 4.5 μmol of D-glutamate was produced and isolated from 10 μmol of L-glutamate, about 90% of the theoretical yield. PMID:27373918

  15. Evidence for synergistic control of glutamate biosynthesis by glutamate dehydrogenases and glutamate in Bacillus subtilis.

    PubMed

    Stannek, Lorena; Thiele, Martin J; Ischebeck, Till; Gunka, Katrin; Hammer, Elke; Völker, Uwe; Commichau, Fabian M

    2015-09-01

    In the Gram-positive bacterium, Bacillus subtilis glutamate is synthesized by the glutamine synthetase and the glutamate synthase (GOGAT). During growth with carbon sources that exert carbon catabolite repression, the rocG glutamate dehydrogenase (GDH) gene is repressed and the transcription factor GltC activates the expression of the GOGAT encoding gltAB genes. In the presence of amino acids of the glutamate family, the GDH RocG is synthesized and the enzyme prevents GltC from binding to DNA. The dual control of glutamate biosynthesis allows the efficient utilization of the available nutrients. Here we provide genetic and biochemical evidence that, like RocG, also the paralogous GDH GudB can inhibit the transcription factor GltC, thereby controlling glutamate biosynthesis. Contradictory previous observations show that high level of GDH activity does not result in permanent inhibition of GltC. By controlling the intracellular levels of glutamate through feeding with exogenous arginine, we observed that the GDH-dependent control of GltC and thus expression of the gltAB genes inversely correlates with the glutamate pool. These results suggest that the B. subtilis GDHs RocG and GudB in fact act as glutamate sensors. In conclusion, the GDH-mediated control of glutamate biosynthesis seems to depend on the intracellular glutamate concentration. PMID:25711804

  16. Glutamic acid as anticancer agent: An overview.

    PubMed

    Dutta, Satyajit; Ray, Supratim; Nagarajan, K

    2013-10-01

    The objective of the article is to highlight various roles of glutamic acid like endogenic anticancer agent, conjugates to anticancer agents, and derivatives of glutamic acid as possible anticancer agents. Besides these emphases are given especially for two endogenous derivatives of glutamic acid such as glutamine and glutamate. Glutamine is a derivative of glutamic acid and is formed in the body from glutamic acid and ammonia in an energy requiring reaction catalyzed by glutamine synthase. It also possesses anticancer activity. So the transportation and metabolism of glutamine are also discussed for better understanding the role of glutamic acid. Glutamates are the carboxylate anions and salts of glutamic acid. Here the roles of various enzymes required for the metabolism of glutamates are also discussed.

  17. Computational Studies of Glutamate Transporters

    PubMed Central

    Setiadi, Jeffry; Heinzelmann, Germano; Kuyucak, Serdar

    2015-01-01

    Glutamate is the major excitatory neurotransmitter in the human brain whose binding to receptors on neurons excites them while excess glutamate are removed from synapses via transporter proteins. Determination of the crystal structures of bacterial aspartate transporters has paved the way for computational investigation of their function and dynamics at the molecular level. Here, we review molecular dynamics and free energy calculation methods used in these computational studies and discuss the recent applications to glutamate transporters. The focus of the review is on the insights gained on the transport mechanism through computational methods, which otherwise is not directly accessible by experimental probes. Recent efforts to model the mammalian glutamate and other amino acid transporters, whose crystal structures have not been solved yet, are included in the review. PMID:26569328

  18. Bicyclic glutamic acid derivatives.

    PubMed

    Meyer, Udo; Bisel, Philippe; Weckert, Edgar; Frahm, August Wilhelm

    2006-05-15

    For the second-generation asymmetric synthesis of the trans-tris(homoglutamic) acids via Strecker reaction of chiral ketimines, the cyanide addition as the key stereodifferentiating step produces mixtures of diastereomeric alpha-amino nitrile esters the composition of which is independent of the reaction temperature and the type of the solvent, respectively. The subsequent hydrolysis is exclusively achieved with concentrated H(2)SO(4) yielding diastereomeric mixtures of three secondary alpha-amino alpha-carbamoyl-gamma-esters and two diastereomeric cis-fused angular alpha-carbamoyl gamma-lactams as bicyclic glutamic acid derivatives, gained from in situ stereomer differentiating cyclisation of the secondary cis-alpha-amino alpha-carbamoyl-gamma-esters. Separation was achieved by CC. The pure secondary trans-alpha-amino alpha-carbamoyl-gamma-esters cyclise on heating and treatment with concentrated H(2)SO(4), respectively, to diastereomeric cis-fused angular secondary alpha-amino imides. Their hydrogenolysis led to the enantiomeric cis-fused angular primary alpha-amino imides. The configuration of all compounds was completely established by NMR methods, CD-spectra, and by X-ray analyses of the (alphaR,1R,5R)-1-carbamoyl-2-(1-phenylethyl)-2-azabicyclo[3.3.0]octan-3-one and of the trans-alphaS,1S,2R-2-ethoxycarbonylmethyl-1-(1-phenylethylamino)cyclopentanecarboxamide. PMID:16596563

  19. The Dynamics of Connexin Expression, Degradation and Localisation Are Regulated by Gonadotropins during the Early Stages of In Vitro Maturation of Swine Oocytes

    PubMed Central

    Santiquet, Nicolas; Robert, Claude; Richard, François J.

    2013-01-01

    Gap junctional communication (GJC) plays a primordial role in oocyte maturation and meiotic resumption in mammals by directing the transfer of numerous molecules between cumulus cells and the oocyte. Gap junctions are made of connexins (Cx), proteins that regulate GJC in numerous ways. Understanding the dynamic regulation of connexin arrangements during in vitro maturation (IVM) could provide a powerful tool for controlling meiotic resumption and consequently in vitro development of fully competent oocytes. However, physiological events happening during the early hours of IVM may still be elucidated. The present study reports the dynamic regulation of connexin expression, degradation and localization during this stage. Cx43, Cx45 and Cx60 were identified as the main connexins expressed in swine COC. Cx43 and Cx45 transcripts were judged too static to be a regulator of GJC, while Cx43 protein expression was highly responsive to gonadotropins, suggesting that it might be the principal regulator of GJC. In addition, the degradation of Cx43 expressed after 4.5 h of IVM in response to equine chorionic gonadotropin appeared to involve the proteasomal complex. Cx43 localisation appeared to be associated with GJC. Taken together, these results show for the first time that gonadotropins regulate Cx43 protein expression, degradation and localisation in porcine COC during the first several hours of IVM. Regulation of Cx43 may in turn, via GJC, participate in the development of fully competent oocytes. PMID:23861906

  20. Glutamate receptors at atomic resolution

    SciTech Connect

    Mayer, Mark L.

    2010-12-03

    At synapses throughout the brain and spinal cord, the amino-acid glutamate is the major excitatory neurotransmitter. During evolution, a family of glutamate-receptor ion channels seems to have been assembled from a kit consisting of discrete ligand-binding, ion-channel, modulatory and cytoplasmic domains. Crystallographic studies that exploit this unique architecture have greatly aided structural analysis of the ligand-binding core, but the results also pose a formidable challenge, namely that of resolving the allosteric mechanisms by which individual domains communicate and function in an intact receptor.

  1. 21 CFR 182.1045 - Glutamic acid.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Glutamic acid. 182.1045 Section 182.1045 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN....1045 Glutamic acid. (a) Product. Glutamic acid. (b) (c) Limitations, restrictions, or explanation....

  2. 21 CFR 182.1045 - Glutamic acid.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Glutamic acid. 182.1045 Section 182.1045 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN....1045 Glutamic acid. (a) Product. Glutamic acid. (b) (c) Limitations, restrictions, or explanation....

  3. 21 CFR 182.1045 - Glutamic acid.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Glutamic acid. 182.1045 Section 182.1045 Food and... GENERALLY RECOGNIZED AS SAFE Multiple Purpose GRAS Food Substances § 182.1045 Glutamic acid. (a) Product. Glutamic acid. (b) (c) Limitations, restrictions, or explanation. This substance is generally recognized...

  4. 21 CFR 182.1045 - Glutamic acid.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Glutamic acid. 182.1045 Section 182.1045 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN....1045 Glutamic acid. (a) Product. Glutamic acid. (b) (c) Limitations, restrictions, or explanation....

  5. 21 CFR 182.1045 - Glutamic acid.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Glutamic acid. 182.1045 Section 182.1045 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD FOR HUMAN....1045 Glutamic acid. (a) Product. Glutamic acid. (b) (c) Limitations, restrictions, or explanation....

  6. 21 CFR 582.1500 - Monoammonium glutamate.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Monoammonium glutamate. 582.1500 Section 582.1500 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Additives § 582.1500 Monoammonium glutamate. (a) Product. Monoammonium glutamate. (b) Conditions of...

  7. 21 CFR 182.1500 - Monoammonium glutamate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Monoammonium glutamate. 182.1500 Section 182.1500 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD... Substances § 182.1500 Monoammonium glutamate. (a) Product. Monoammonium glutamate. (b) Conditions of...

  8. 21 CFR 582.1500 - Monoammonium glutamate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Monoammonium glutamate. 582.1500 Section 582.1500 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Additives § 582.1500 Monoammonium glutamate. (a) Product. Monoammonium glutamate. (b) Conditions of...

  9. 21 CFR 182.1516 - Monopotassium glutamate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Monopotassium glutamate. 182.1516 Section 182.1516 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD... Substances § 182.1516 Monopotassium glutamate. (a) Product. Monopotassium glutamate. (b) Conditions of...

  10. 21 CFR 582.1500 - Monoammonium glutamate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Monoammonium glutamate. 582.1500 Section 582.1500 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Additives § 582.1500 Monoammonium glutamate. (a) Product. Monoammonium glutamate. (b) Conditions of...

  11. 21 CFR 582.1516 - Monopotassium glutamate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Monopotassium glutamate. 582.1516 Section 582.1516 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Additives § 582.1516 Monopotassium glutamate. (a) Product. Monopotassium glutamate. (b) Conditions of...

  12. 21 CFR 182.1500 - Monoammonium glutamate.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Monoammonium glutamate. 182.1500 Section 182.1500 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD... Substances § 182.1500 Monoammonium glutamate. (a) Product. Monoammonium glutamate. (b) Conditions of...

  13. 21 CFR 182.1516 - Monopotassium glutamate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 3 2013-04-01 2013-04-01 false Monopotassium glutamate. 182.1516 Section 182.1516 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD... Substances § 182.1516 Monopotassium glutamate. (a) Product. Monopotassium glutamate. (b) Conditions of...

  14. 21 CFR 582.1516 - Monopotassium glutamate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Monopotassium glutamate. 582.1516 Section 582.1516 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Additives § 582.1516 Monopotassium glutamate. (a) Product. Monopotassium glutamate. (b) Conditions of...

  15. 21 CFR 182.1500 - Monoammonium glutamate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Monoammonium glutamate. 182.1500 Section 182.1500 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD... Substances § 182.1500 Monoammonium glutamate. (a) Product. Monoammonium glutamate. (b) Conditions of...

  16. 21 CFR 582.1516 - Monopotassium glutamate.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Monopotassium glutamate. 582.1516 Section 582.1516 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Additives § 582.1516 Monopotassium glutamate. (a) Product. Monopotassium glutamate. (b) Conditions of...

  17. 21 CFR 582.1500 - Monoammonium glutamate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Monoammonium glutamate. 582.1500 Section 582.1500 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Additives § 582.1500 Monoammonium glutamate. (a) Product. Monoammonium glutamate. (b) Conditions of...

  18. 21 CFR 182.1516 - Monopotassium glutamate.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Monopotassium glutamate. 182.1516 Section 182.1516 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD... Substances § 182.1516 Monopotassium glutamate. (a) Product. Monopotassium glutamate. (b) Conditions of...

  19. 21 CFR 582.1516 - Monopotassium glutamate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Monopotassium glutamate. 582.1516 Section 582.1516 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Additives § 582.1516 Monopotassium glutamate. (a) Product. Monopotassium glutamate. (b) Conditions of...

  20. 21 CFR 182.1500 - Monoammonium glutamate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Monoammonium glutamate. 182.1500 Section 182.1500 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... glutamate. (a) Product. Monoammonium glutamate. (b) Conditions of use. This substance is...

  1. 21 CFR 182.1500 - Monoammonium glutamate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Monoammonium glutamate. 182.1500 Section 182.1500 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD... Substances § 182.1500 Monoammonium glutamate. (a) Product. Monoammonium glutamate. (b) Conditions of...

  2. 21 CFR 582.1516 - Monopotassium glutamate.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Monopotassium glutamate. 582.1516 Section 582.1516 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Additives § 582.1516 Monopotassium glutamate. (a) Product. Monopotassium glutamate. (b) Conditions of...

  3. 21 CFR 182.1516 - Monopotassium glutamate.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Monopotassium glutamate. 182.1516 Section 182.1516 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... glutamate. (a) Product. Monopotassium glutamate. (b) Conditions of use. This substance is...

  4. 21 CFR 182.1516 - Monopotassium glutamate.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Monopotassium glutamate. 182.1516 Section 182.1516 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) FOOD... Substances § 182.1516 Monopotassium glutamate. (a) Product. Monopotassium glutamate. (b) Conditions of...

  5. 21 CFR 582.1500 - Monoammonium glutamate.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Monoammonium glutamate. 582.1500 Section 582.1500 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED) ANIMAL... Additives § 582.1500 Monoammonium glutamate. (a) Product. Monoammonium glutamate. (b) Conditions of...

  6. Brain neuroprotection by scavenging blood glutamate.

    PubMed

    Zlotnik, Alexander; Gurevich, Boris; Tkachov, Sergei; Maoz, Ilana; Shapira, Yoram; Teichberg, Vivian I

    2007-01-01

    Excess glutamate in brain fluids characterizes acute brain insults such as traumatic brain injury and stroke. Its removal could prevent the glutamate excitotoxicity that causes long-lasting neurological deficits. As blood glutamate scavenging has been demonstrated to increase the efflux of excess glutamate from brain into blood, we tested the prediction that oxaloacetate-mediated blood glutamate scavenging causes neuroprotection in a pathological situation such as closed head injury (CHI), in which there is a well established deleterious increase of glutamate in brain fluids. We observed highly significant improvements of the neurological status of rats submitted to CHI following an intravenous treatment with 1 mmol oxaloacetate/100 g rat weight which decreases blood glutamate levels by 40%. No detectable therapeutic effect was obtained when rats were treated IV with 1 mmol oxaloacetate together with 1 mmol glutamate/100 g rat. The treatment with 0.005 mmol/100 g rat oxaloacetate was no more effective than saline but when it was combined with the intravenous administration of 0.14 nmol/100 g of recombinant glutamate-oxaloacetate transaminase, recovery was almost complete. Oxaloacetate provided neuroprotection when administered before CHI or at 60 min post CHI but not at 120 min post CHI. Since neurological recovery from CHI was highly correlated with the decrease of blood glutamate levels (r=0.89, P=0.001), we conclude that blood glutamate scavenging affords brain neuroprotection Blood glutamate scavenging may open now new therapeutic options.

  7. Modes of glutamate receptor gating

    PubMed Central

    Popescu, Gabriela K

    2012-01-01

    Abstract The time course of excitatory synaptic currents, the major means of fast communication between neurons of the central nervous system, is encoded in the dynamic behaviour of post-synaptic glutamate-activated channels. First-pass attempts to explain the glutamate-elicited currents with mathematical models produced reaction mechanisms that included only the most basic functionally defined states: resting vs. liganded, closed vs. open, responsive vs. desensitized. In contrast, single-molecule observations afforded by the patch-clamp technique revealed an unanticipated kinetic multiplicity of transitions: from microseconds-lasting flickers to minutes-long modes. How these kinetically defined events impact the shape of the synaptic response, how they relate to rearrangements in receptor structure, and whether and how they are physiologically controlled represent currently active research directions. Modal gating, which refers to the slowest, least frequently observed ion-channel transitions, has been demonstrated for representatives of all ion channel families. However, reaction schemes have been largely confined to the short- and medium-range time scales. For glutamate receptors as well, modal gating has only recently come under rigorous scrutiny. This article reviews the evidence for modal gating of glutamate receptors and the still developing hypotheses about the mechanism(s) by which modal shifts occur and the ways in which they may impact the time course of synaptic transmission. PMID:22106181

  8. Fluorescence imaging of glutamate release in neurons

    SciTech Connect

    Wang, Ziqiang; Yeung, Edward S.

    1999-12-01

    A noninvasive detection scheme based on glutamate dehydrogenase (GDH) enzymatic assay combined with microscopy was developed to measure the glutamate release in cultured cells from the central nervous system (CNS). The enzyme reaction is very specific and sensitive. The detection limit with charge-coupled device (CCD) imaging is down to {mu}M levels of glutamate with reasonable response time ({approx}30 s). The standard glutamate test shows a linear response over 3 orders of magnitude, from {mu}M to 0.1 mM range. The in vitro monitoring of glutamate release from cultured neuron cells demonstrated excellent spatial and temporal resolution. (c) 1999 Society for Applied Spectroscopy.

  9. Connexin-43 gap junctions are involved in multiconnexin-expressing stromal support of hemopoietic progenitors and stem cells.

    PubMed

    Cancelas, J A; Koevoet, W L; de Koning, A E; Mayen, A E; Rombouts, E J; Ploemacher, R E

    2000-07-15

    Gap junctions (GJs) provide for a unique system of intercellular communication (IC) allowing rapid transport of small molecules from cell to cell. GJs are formed by a large family of proteins named connexins (Cxs). Cx43 has been considered as the predominantly expressed Cx by hematopoietic-supporting stroma. To investigate the role of the Cx family in hemopoiesis, we analyzed the expression of 11 different Cx species in different stromal cell lines derived from murine bone marrow (BM) or fetal liver (FL). We found that up to 5 Cxs are expressed in FL stromal cells (Cx43, Cx45, Cx30.3, Cx31, and Cx31.1), whereas only Cx43, Cx45, and Cx31 were clearly detectable in BM stromal cells. In vivo, the Cx43-deficient 14.5- to 15-day FL cobblestone area-forming cells (CAFC)-week 1-4 and colony-forming unit contents were 26%-38% and 39%-47% lower than in their wild-type counterparts, respectively. The reintroduction of the Cx43 gene into Cx43-deficient FL stromal cells was able to restore their diminished IC to the level of the wild-type FL stromal cells. In addition, these Cx43-reintroduced stromal cells showed an increased support ability (3.7-fold) for CAFC-week 1 in normal mouse BM and 5-fold higher supportive ability for CAFC-week 4 in 5-fluorouracil-treated BM cells as compared with Cx43-deficient FL stromal cells. These findings suggest that stromal Cx43-mediated IC, although not responsible for all GJ-mediated IC of stromal cells, plays a role in the supportive ability for hemopoietic progenitors and stem cells. (Blood. 2000;96:498-505) PMID:10887111

  10. Ligands for Ionotropic Glutamate Receptors

    NASA Astrophysics Data System (ADS)

    Swanson, Geoffrey T.; Sakai, Ryuichi

    Marine-derived small molecules and peptides have played a central role in elaborating pharmacological specificities and neuronal functions of mammalian ionotropic glutamate receptors (iGluRs), the primary mediators of excitatory syn-aptic transmission in the central nervous system (CNS). As well, the pathological sequelae elicited by one class of compounds (the kainoids) constitute a widely-used animal model for human mesial temporal lobe epilepsy (mTLE). New and existing molecules could prove useful as lead compounds for the development of therapeutics for neuropathologies that have aberrant glutamatergic signaling as a central component. In this chapter we discuss natural source origins and pharmacological activities of those marine compounds that target ionotropic glutamate receptors.

  11. Ligands for Ionotropic Glutamate Receptors

    PubMed Central

    Swanson, Geoffrey T.; Sakai, Ryuichi

    2010-01-01

    Marine-derived small molecules and peptides have played a central role in elaborating pharmacological specificities and neuronal functions of mammalian ionotropic glutamate receptors (iGluRs), the primary mediators of excitatory synaptic transmission in the central nervous system (CNS). As well, the pathological sequelae elicited by one class of compounds (the kainoids) constitute a widely-used animal model for human mesial temporal lobe epilepsy (mTLE). New and existing molecules could prove useful as lead compounds for the development of therapeutics for neuropathologies that have aberrant glutamatergic signaling as a central component. In this chapter we discuss natural source origins and pharmacological activities of those marine compounds that target ionotropic glutamate receptors. PMID:19184587

  12. Fast skeletal myofibers of mdx mouse, model of Duchenne muscular dystrophy, express connexin hemichannels that lead to apoptosis.

    PubMed

    Cea, Luis A; Puebla, Carlos; Cisterna, Bruno A; Escamilla, Rosalba; Vargas, Aníbal A; Frank, Marina; Martínez-Montero, Paloma; Prior, Carmen; Molano, Jesús; Esteban-Rodríguez, Isabel; Pascual, Ignacio; Gallano, Pía; Lorenzo, Gustavo; Pian, Héctor; Barrio, Luis C; Willecke, Klaus; Sáez, Juan C

    2016-07-01

    Skeletal muscles of patients with Duchenne muscular dystrophy (DMD) show numerous alterations including inflammation, apoptosis, and necrosis of myofibers. However, the molecular mechanism that explains these changes remains largely unknown. Here, the involvement of hemichannels formed by connexins (Cx HCs) was evaluated in skeletal muscle of mdx mouse model of DMD. Fast myofibers of mdx mice were found to express three connexins (39, 43 and 45) and high sarcolemma permeability, which was absent in myofibers of mdx Cx43(fl/fl)Cx45(fl/fl):Myo-Cre mice (deficient in skeletal muscle Cx43/Cx45 expression). These myofibers did not show elevated basal intracellular free Ca(2+) levels, immunoreactivity to phosphorylated p65 (active NF-κB), eNOS and annexin V/active Caspase 3 (marker of apoptosis) but presented dystrophin immunoreactivity. Moreover, muscles of mdx Cx43(fl/fl)Cx45(fl/fl):Myo-Cre mice exhibited partial decrease of necrotic features (big cells and high creatine kinase levels). Accordingly, these muscles showed similar macrophage infiltration as control mdx muscles. Nonetheless, the hanging test performance of mdx Cx43(fl/fl)Cx45(fl/fl):Myo-Cre mice was significantly better than that of control mdx Cx43(fl/fl)Cx45(fl/fl) mice. All three Cxs found in skeletal muscles of mdx mice were also detected in fast myofibers of biopsy specimens from patients with muscular dystrophy. Thus, reduction of Cx expression and/or function of Cx HCs may be potential therapeutic approaches to abrogate myofiber apoptosis in DMD. PMID:26803842

  13. Fast skeletal myofibers of mdx mouse, model of Duchenne muscular dystrophy, express connexin hemichannels that lead to apoptosis.

    PubMed

    Cea, Luis A; Puebla, Carlos; Cisterna, Bruno A; Escamilla, Rosalba; Vargas, Aníbal A; Frank, Marina; Martínez-Montero, Paloma; Prior, Carmen; Molano, Jesús; Esteban-Rodríguez, Isabel; Pascual, Ignacio; Gallano, Pía; Lorenzo, Gustavo; Pian, Héctor; Barrio, Luis C; Willecke, Klaus; Sáez, Juan C

    2016-07-01

    Skeletal muscles of patients with Duchenne muscular dystrophy (DMD) show numerous alterations including inflammation, apoptosis, and necrosis of myofibers. However, the molecular mechanism that explains these changes remains largely unknown. Here, the involvement of hemichannels formed by connexins (Cx HCs) was evaluated in skeletal muscle of mdx mouse model of DMD. Fast myofibers of mdx mice were found to express three connexins (39, 43 and 45) and high sarcolemma permeability, which was absent in myofibers of mdx Cx43(fl/fl)Cx45(fl/fl):Myo-Cre mice (deficient in skeletal muscle Cx43/Cx45 expression). These myofibers did not show elevated basal intracellular free Ca(2+) levels, immunoreactivity to phosphorylated p65 (active NF-κB), eNOS and annexin V/active Caspase 3 (marker of apoptosis) but presented dystrophin immunoreactivity. Moreover, muscles of mdx Cx43(fl/fl)Cx45(fl/fl):Myo-Cre mice exhibited partial decrease of necrotic features (big cells and high creatine kinase levels). Accordingly, these muscles showed similar macrophage infiltration as control mdx muscles. Nonetheless, the hanging test performance of mdx Cx43(fl/fl)Cx45(fl/fl):Myo-Cre mice was significantly better than that of control mdx Cx43(fl/fl)Cx45(fl/fl) mice. All three Cxs found in skeletal muscles of mdx mice were also detected in fast myofibers of biopsy specimens from patients with muscular dystrophy. Thus, reduction of Cx expression and/or function of Cx HCs may be potential therapeutic approaches to abrogate myofiber apoptosis in DMD.

  14. Glutamate racemization and catabolism in Fusobacterium varium.

    PubMed

    Ramezani, Mohammad; Resmer, Kelly L; White, Robert L

    2011-07-01

    The pathways of glutamate catabolism in the anaerobic bacterium Fusobacterium varium, grown on complex, undefined medium and chemically defined, minimal medium, were investigated using specifically labelled (13)C-glutamate. The metabolic end-products acetate and butyrate were isolated from culture fluids and derivatized for analysis by nuclear magnetic resonance and mass spectrometry. On complex medium, labels from L-[1-(13)C]glutamate and L-[4-(13)C]glutamate were incorporated into C1 of acetate and equally into C1/C3 of butyrate, while label derived from L-[5-(13)C]glutamate was not incorporated. The isotopic incorporation results and the detection of glutamate mutase and 3-methylaspartate ammonia lyase in cell extracts are most consistent with the methylaspartate pathway, the best known route of glutamate catabolism in Clostridium species. When F. varium was grown on defined medium, label from L-[4-(13)C]glutamate was incorporated mainly into C4 of butyrate, demonstrating a major role for the hydroxyglutarate pathway. Upon addition of coenzyme B(12) or cobalt ion to the defined medium in replicate experiments, isotope was located equally at C1/C3 of butyrate in accord with the methylaspartate pathway. Racemization of D-glutamate and subsequent degradation of L-glutamate via the methylaspartate pathway are supported by incorporation of label into C2 of acetate and equally into C2/C4 of butyrate from D-[3-(13)C]glutamate and the detection of a cofactor-independent glutamate racemase in cell extracts. Together the results demonstrate a major role for the methylaspartate pathway of glutamate catabolism in F. varium and substantial participation of the hydroxyglutarate pathway when coenzyme B(12) is not available.

  15. The glutamate homeostasis hypothesis of addiction.

    PubMed

    Kalivas, Peter W

    2009-08-01

    Addiction is associated with neuroplasticity in the corticostriatal brain circuitry that is important for guiding adaptive behaviour. The hierarchy of corticostriatal information processing that normally permits the prefrontal cortex to regulate reinforcement-seeking behaviours is impaired by chronic drug use. A failure of the prefrontal cortex to control drug-seeking behaviours can be linked to an enduring imbalance between synaptic and non-synaptic glutamate, termed glutamate homeostasis. The imbalance in glutamate homeostasis engenders changes in neuroplasticity that impair communication between the prefrontal cortex and the nucleus accumbens. Some of these pathological changes are amenable to new glutamate- and neuroplasticity-based pharmacotherapies for treating addiction. PMID:19571793

  16. Synaptic glutamate spillover due to impaired glutamate uptake mediates heroin relapse.

    PubMed

    Shen, Hao-wei; Scofield, Michael D; Boger, Heather; Hensley, Megan; Kalivas, Peter W

    2014-04-16

    Reducing the enduring vulnerability to relapse is a therapeutic goal in treating drug addiction. Studies with animal models of drug addiction show a marked increase in extrasynaptic glutamate in the core subcompartment of the nucleus accumbens (NAcore) during reinstated drug seeking. However, the synaptic mechanisms linking drug-induced changes in extrasynaptic glutamate to relapse are poorly understood. Here, we discovered impaired glutamate elimination in rats extinguished from heroin self-administration that leads to spillover of synaptically released glutamate into the nonsynaptic extracellular space in NAcore and investigated whether restoration of glutamate transport prevented reinstated heroin seeking. Through multiple functional assays of glutamate uptake and analyzing NMDA receptor-mediated currents, we show that heroin self-administration produced long-lasting downregulation of glutamate uptake and surface expression of the transporter GLT-1. This downregulation was associated with spillover of synaptic glutamate to extrasynaptic NMDA receptors within the NAcore. Ceftriaxone restored glutamate uptake and prevented synaptic glutamate spillover and cue-induced heroin seeking. Ceftriaxone-induced inhibition of reinstated heroin seeking was blocked by morpholino-antisense targeting GLT-1 synthesis. These data reveal that the synaptic glutamate spillover in the NAcore results from reduced glutamate transport and is a critical pathophysiological mechanism underling reinstated drug seeking in rats extinguished from heroin self-administration.

  17. Scavenging of blood glutamate for enhancing brain-to-blood glutamate efflux.

    PubMed

    Li, Yunhong; Hou, Xiaolin; Qi, Qi; Wang, Le; Luo, Lan; Yang, Shaoqi; Zhang, Yumei; Miao, Zhenhua; Zhang, Yanli; Wang, Fei; Wang, Hongyan; Huang, Weidong; Wang, Zhenhai; Shen, Ying; Wang, Yin

    2014-01-01

    The presence of excess glutamate in the brain interstitial fluid characterizes several acute pathological conditions of the brain, including traumatic brain injury and stroke. It has been demonstrated that it is possible to eliminate excess glutamate in the brain by decreasing blood glutamate levels and, accordingly, accelerating the brain-to-blood glutamate efflux. It is feasible to accomplish this process by activating blood resident enzymes in the presence of the respective glutamate cosubstrates. In the present study, several glutamate cosubstrates and cofactors were studied in an attempt to identify the optimal conditions to reduce blood glutamate levels. The administration of a mixture of 1 mM pyruvate and oxaloacetate (Pyr/Oxa) for 1 h decreased blood glutamate levels by ≤50%. The addition of lipoamide to this mixture resulted in a further reduction in blood glutamate levels of >80%. In addition, in vivo experiments showed that lipoamide together with Pyr/Oxa is able to decrease blood glutamate levels to a greater extent than Pyr/Oxa alone, and accordingly, this enhances the glutamate efflux from the brain to the blood. These results may outline a novel neuroprotective strategy with increased effectiveness for the removal of excess brain glutamate in various neurodegenerative conditions.

  18. Synaptic Glutamate Spillover Due to Impaired Glutamate Uptake Mediates Heroin Relapse

    PubMed Central

    Scofield, Michael D.; Boger, Heather; Hensley, Megan; Kalivas, Peter W.

    2014-01-01

    Reducing the enduring vulnerability to relapse is a therapeutic goal in treating drug addiction. Studies with animal models of drug addiction show a marked increase in extrasynaptic glutamate in the core subcompartment of the nucleus accumbens (NAcore) during reinstated drug seeking. However, the synaptic mechanisms linking drug-induced changes in extrasynaptic glutamate to relapse are poorly understood. Here, we discovered impaired glutamate elimination in rats extinguished from heroin self-administration that leads to spillover of synaptically released glutamate into the nonsynaptic extracellular space in NAcore and investigated whether restoration of glutamate transport prevented reinstated heroin seeking. Through multiple functional assays of glutamate uptake and analyzing NMDA receptor-mediated currents, we show that heroin self-administration produced long-lasting downregulation of glutamate uptake and surface expression of the transporter GLT-1. This downregulation was associated with spillover of synaptic glutamate to extrasynaptic NMDA receptors within the NAcore. Ceftriaxone restored glutamate uptake and prevented synaptic glutamate spillover and cue-induced heroin seeking. Ceftriaxone-induced inhibition of reinstated heroin seeking was blocked by morpholino-antisense targeting GLT-1 synthesis. These data reveal that the synaptic glutamate spillover in the NAcore results from reduced glutamate transport and is a critical pathophysiological mechanism underling reinstated drug seeking in rats extinguished from heroin self-administration. PMID:24741055

  19. Effect of insulin on muscle glutamate uptake

    PubMed Central

    Aoki, T. T.; Brennan, M. F.; Müller, W. A.; Moore, F. D.; Cahill, G. F.

    1972-01-01

    For decades, investigators concerned with protein metabolism in man have performed detailed amino acid analyses of human plasma obtained under a wide range of experimental situations. A large body of information has been used to calculated rates of protein synthesis and proteolysis. During the course of an investigation of the effect of intrabrachial artery infusion of insulin (70 μU/min per kg body weight) on glutamate uptake by human forearm muscle, it was discovered that plasma arterio-deep venous glutamate difference analysis failed to document any increase in the uptake of this amino acid, suggesting that insulin had little influence on glutamate uptake by muscle. However, whole blood glutamate analyses, performed on the same blood samples, revealed that (a) the resting muscle uptake of glutamate is smaller than previously reported and (b) insulin is capable of markedly increasing glutamate uptake by muscle from whole blood. Since the hematocrit was obtained on all samples, detailed analyses of the various compartments in which glutamate could be found were performed. It was determined that circulating blood cells have a dynamic role in glutamate transport. These data underscore the need for both whole blood and plasma amino acid analysis in investigations concerned with protein synthesis and/or amino acid flux, for analysis of plasma samples alone could be misleading as illustrated in the present study. Images PMID:5080414

  20. Glutamate: Tastant and Neuromodulator in Taste Buds.

    PubMed

    Vandenbeuch, Aurelie; Kinnamon, Sue C

    2016-07-01

    In taste buds, glutamate plays a double role as a gustatory stimulus and neuromodulator. The detection of glutamate as a tastant involves several G protein-coupled receptors, including the heterodimer taste receptor type 1, member 1 and 3 as well as metabotropic glutamate receptors (mGluR1 and mGluR4). Both receptor types participate in the detection of glutamate as shown with knockout animals and selective antagonists. At the basal part of taste buds, ionotropic glutamate receptors [N-methyl-d-aspartate (NMDA) and non-NMDA] are expressed and participate in the modulation of the taste signal before its transmission to the brain. Evidence suggests that glutamate has an efferent function on taste cells and modulates the release of other neurotransmitters such as serotonin and ATP. This short article reviews the recent developments in the field with regard to glutamate receptors involved in both functions as well as the influence of glutamate on the taste signal. PMID:27422519

  1. Glutamate Transporter-Mediated Glutamate Secretion in the Mammalian Pineal Gland

    PubMed Central

    Kim, Mean-Hwan; Uehara, Shunsuke; Muroyama, Akiko; Hille, Bertil; Moriyama, Yoshinori; Koh, Duk-Su

    2008-01-01

    Glutamate transporters are expressed throughout the central nervous system where their major role is to clear released glutamate from presynaptic terminals. Here we report a novel function of the transporter in rat pinealocytes. This electrogenic transporter conducted inward current in response to L-glutamate and L- or D-aspartate and depolarized the membrane in patch clamp experiments. Ca2+ imaging demonstrated that the transporter-mediated depolarization induced a significant Ca2+ influx through voltage-gated Ca2+ channels. The Ca2+ rise finally evoked glutamate exocytosis as detected by carbon-fiber amperometry and by high-performance liquid chromatography. In pineal slices with densely packed pinealocytes, glutamate released from the cells effectively activated glutamate transporters in neighboring cells. The Ca2+ signal generated by KCl depolarization or acetylcholine propagated through several cell layers by virtue of the regenerative ‘glutamate-induced glutamate release’. Therefore we suggest that glutamate transporters mediate synchronized elevation of L-glutamate and thereby efficiently down-regulate melatonin secretion via previously identified inhibitory metabotropic glutamate receptors in the pineal gland. PMID:18945893

  2. Glutamate-based antidepressants: preclinical psychopharmacology.

    PubMed

    Pilc, Andrzej; Wierońska, Joanna M; Skolnick, Phil

    2013-06-15

    Over the past 20 years, converging lines of evidence have both linked glutamatergic dysfunction to the pathophysiology of depression and demonstrated that the glutamatergic synapse presents multiple targets for developing novel antidepressants. The robust antidepressant effects of the N-methyl-D-aspartate receptor antagonists ketamine and traxoprodil provide target validation for this family of ionotropic glutamate receptors. This article reviews the preclinical evidence that it may be possible to develop glutamate-based antidepressants by not only modulating ionotropic (N-methyl-D-aspartate and alpha-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid) and metabotropic glutamate (mGlu) receptors, including mGlu2/3, mGLu5 and mGlu7 receptors, but also by altering synaptic concentrations of glutamate via specialized transporters such as glial glutamate transporter 1 (excitatory amino-acid transporter 2).

  3. Emerging aspects of dietary glutamate metabolism in the developing gut

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Glutamate is a major constituent of dietary protein and is also consumed in many prepared foods as a flavour additive in the form of monosodium glutamate (MSG). Evidence from human and animal studies indicates that glutamate is the major oxidative fuel for the gut and that dietary glutamate is exten...

  4. Glutamate Receptor Stimulation Up-Regulates Glutamate Uptake in Human Müller Glia Cells.

    PubMed

    López-Colomé, Ana María; López, Edith; Mendez-Flores, Orquidia G; Ortega, Arturo

    2016-07-01

    Glutamate, the main excitatory amino acid in the vertebrate retina, is a well know activator of numerous signal transduction pathways, and has been critically involved in long-term synaptic changes acting through ionotropic and metabotropic glutamate receptors. However, recent findings underlining the importance of intensity and duration of glutamate stimuli for specific neuronal responses, including excitotoxicity, suggest a crucial role for Na(+)-dependent glutamate transporters, responsible for the removal of this neurotransmitter from the synaptic cleft, in the regulation of glutamate-induced signaling. Transporter proteins are expressed in neurons and glia cells, albeit most of glutamate uptake occurs in the glial compartment. Within the retina, Müller glia cells are in close proximity to glutamatergic synapses and participate in the recycling of glutamate through the glutamate/glutamine shuttle. In this context, we decided to investigate a plausible role of glutamate as a regulatory signal for its own transport in human retinal glia cells. To this end, we determined [(3)H]-D-aspartate uptake in cultures of spontaneously immortalized human Müller cells (MIO-M1) exposed to distinct glutamatergic ligands. A time and dose-dependent increase in the transporter activity was detected. This effect was dependent on the activation of the N-methyl D-aspartate subtype of glutamate receptors, due to a dual effect: an increase in affinity and an augmented expression of the transporter at the plasma membrane, as established via biotinylation experiments. Furthermore, a NMDA-dependent association of glutamate transporters with the cystoskeletal proteins ezrin and glial fibrillary acidic protein was also found. These results add a novel mediator of the glutamate transporter modulation and further strengthen the notion of the critical involvement of glia cells in synaptic function. PMID:27017513

  5. Modulation of glutamate transport and receptor binding by glutamate receptor antagonists in EAE rat brain.

    PubMed

    Sulkowski, Grzegorz; Dąbrowska-Bouta, Beata; Salińska, Elżbieta; Strużyńska, Lidia

    2014-01-01

    The etiology of multiple sclerosis (MS) is currently unknown. However, one potential mechanism involved in the disease may be excitotoxicity. The elevation of glutamate in cerebrospinal fluid, as well as changes in the expression of glutamate receptors (iGluRs and mGluRs) and excitatory amino acid transporters (EAATs), have been observed in the brains of MS patients and animals subjected to experimental autoimmune encephalomyelitis (EAE), which is the predominant animal model used to investigate the pathophysiology of MS. In the present paper, the effects of glutamatergic receptor antagonists, including amantadine, memantine, LY 367583, and MPEP, on glutamate transport, the expression of mRNA of glutamate transporters (EAATs), the kinetic parameters of ligand binding to N-methyl-D-aspartate (NMDA) receptors, and the morphology of nerve endings in EAE rat brains were investigated. The extracellular level of glutamate in the brain is primarily regulated by astrocytic glutamate transporter 1 (GLT-1) and glutamate-aspartate transporter (GLAST). Excess glutamate is taken up from the synaptic space and metabolized by astrocytes. Thus, the extracellular level of glutamate decreases, which protects neurons from excitotoxicity. Our investigations showed changes in the expression of EAAT mRNA, glutamate transport (uptake and release) by synaptosomal and glial plasmalemmal vesicle fractions, and ligand binding to NMDA receptors; these effects were partially reversed after the treatment of EAE rats with the NMDA antagonists amantadine and memantine. The antagonists of group I metabotropic glutamate receptors (mGluRs), including LY 367385 and MPEP, did not exert any effect on the examined parameters. These results suggest that disturbances in these mechanisms may play a role in the processes associated with glutamate excitotoxicity and the progressive brain damage in EAE.

  6. Regulation of Synaptic Transmission by Ambient Extracellular Glutamate

    PubMed Central

    FEATHERSTONE, DAVID E.; SHIPPY, SCOTT A.

    2008-01-01

    Many neuroscientists assume that ambient extracellular glutamate concentrations in the nervous system are biologically negligible under nonpathological conditions. This assumption is false. Hundreds of studies over several decades suggest that ambient extracellular glutamate levels in the intact mammalian brain are ~0.5 to ~5 μM. This has important implications. Glutamate receptors are desensitized by glutamate concentrations significantly lower than needed for receptor activation; 0.5 to 5 μM of glutamate is high enough to cause constitutive desensitization of most glutamate receptors. Therefore, most glutamate receptors in vivo may be constitutively desensitized, and ambient extracellular glutamate and receptor desensitization may be potent but generally unrecognized regulators of synaptic transmission. Unfortunately, the mechanisms regulating ambient extracellular glutamate and glutamate receptor desensitization remain poorly understood and understudied. PMID:17947494

  7. Construction of a potentiometric glutamate biosensor for determination of glutamate in some real samples.

    PubMed

    Y Lmaz, Demet; Karaku, Emine

    2011-12-01

    The potentiometric glutamate biosensor based on ammonium-selective poly(vinylchloride) (PVC) membrane electrode was constructed by chemically immobilizing glutamate oxidase. Ammonium ions produced after an enzymatic reaction were determined potentiometrically. We determined the optimum working conditions of the biosensor such as buffer concentration, buffer pH, lifetime, response time, linear working range, kinetic constants (K(m) and V(max)) of glutamate oxidase enzyme used for biosensor construction values, and other response characteristics. Additionally, glutamate assay in some real samples such as chicken bullion, healthy human serum, and commercial multipower amino acid mixture were also successfully carried out. The results showed good agreement with previously reported values.

  8. Brain glutamate decarboxylase and pyrroloquinoline quinone.

    PubMed

    Choi, S Y; Khemlani, L S; Churchich, J E

    1992-01-01

    Porcine brain glutamate decarboxylase was examined for the presence of covalently bound pyrroloquinoline quinone (PQQ). HPLC analysis of pure glutamate decarboxylase subjected to the hexanol extraction procedure gave negative results when monitored at 320 nm, the maximum of absorbance of 4-hydroxy-5-hexoxy-PQQ. Resolved glutamate decarboxylase exhibits a structureless absorption band at wavelengths longer than 300 nm which cannot be attributed to PQQ. The holoenzyme is not a pyridoxal-quinoprotein; its catalytic mechanism involves the participation of only one cofactor, i.e. pyridoxal-5-P. Free PQQ is a strong inhibitor of the decarboxylase (Ki = 13 microM) and the reaction with the protein results in spectral changes resembling those of polylysine treated with PQQ. If the concentration of free PQQ in some regions of the brain reaches the micromolar level, then PQQ might play a role in the regulation of glutamate decarboxylase activity.

  9. Mechanism for the activation of glutamate receptors

    Cancer.gov

    Scientists at the NIH have used a technique called cryo-electron microscopy to determine a molecular mechanism for the activation and desensitization of ionotropic glutamate receptors, a prominent class of neurotransmitter receptors in the brain and spina

  10. DNA nanopore translocation in glutamate solutions

    NASA Astrophysics Data System (ADS)

    Plesa, C.; van Loo, N.; Dekker, C.

    2015-08-01

    Nanopore experiments have traditionally been carried out with chloride-based solutions. Here we introduce silver/silver-glutamate-based electrochemistry as an alternative, and study the viscosity, conductivity, and nanopore translocation characteristics of potassium-, sodium-, and lithium-glutamate solutions. We show that it has a linear response at typical voltages and can be used to detect DNA translocations through a nanopore. The glutamate anion also acts as a redox-capable thickening agent, with high-viscosity solutions capable of slowing down the DNA translocation process by up to 11 times, with a corresponding 7 time reduction in signal. These results demonstrate that glutamate can replace chloride as the primary anion in nanopore resistive pulse sensing.

  11. [Glutamate transporter dysfunction and major mental illnesses].

    PubMed

    Tanaka, Kohichi

    2016-01-01

    Glutamate is the main excitatory neurotransmitter in the central nervous system and plays an important role in most aspects of normal brain function. In spite of its importance as a neurotransmitter, excess glutamate is toxic to neurons. Clearance of extracellular glutamate is critical for maintenance of low extracellular glutamate concentration, and occurs in large part through the activity of GLT1 (EAAT2) and GLAST (EAAT1), which are primarily expressed by astrocytes. Rare variants and down-regulation of GLT1 and GLAST, in psychiatric disorders have been reported. In this review, we demonstrate that various kinds of GLT1 and/or GLAST knockout mice replicate many aspects of the behavioral abnormalities seen in major mental illnesses including schizophrenia, depression, obsessive -compulsive disorders, autism, epilepsy and addiction. PMID:26793898

  12. Glutamic Acid Decarboxylation in Chlorella12

    PubMed Central

    Lane, T. R.; Stiller, Mary

    1970-01-01

    The decarboxylation of endogenous free glutamic acid by Chlorella pyrenoidosa, Marburg strain, was induced by a variety of metabolic poisons, by anaerobic conditions, and by freezing and thawing the cells. The rate of decarboxylation was proportional to the concentration of inhibitor present. Possible mechanisms which relate the effects of the various conditions on glutamate decarboxylation and oxygen consumption by Chlorella are discussed. Images PMID:5429350

  13. [Glutamate neurotransmission, stress and hormone secretion].

    PubMed

    Jezová, D; Juránková, E; Vigas, M

    1995-11-01

    Glutamate neurotransmission has been investigated in relation to several physiological processes (learning, memory) as well as to neurodegenerative and other disorders. Little attention has been paid to its involvement in neuroendocrine response during stress. Penetration of excitatory amino acids from blood to the brain is limited by the blood-brain barrier. As a consequence, several toxic effects but also bioavailability for therapeutic purposes are reduced. A free access to circulating glutamate is possible only in brain structures lacking the blood-brain barrier or under conditions of its increased permeability. Excitatory amino acids were shown to stimulate the pituitary hormone release, though the mechanism of their action is still not fully understood. Stress exposure in experimental animals induced specific changes in mRNA levels coding the glutamate receptor subunits in the hippocampus and hypothalamus. The results obtained with the use of glutamate receptor antagonists indicate that a number of specific receptor subtypes contribute to the stimulation of ACTH release during stress. The authors provided also data on the role of NMDA receptors in the control of catecholamine release, particularly in stress-induced secretion of epinephrine. These results were the first piece of evidence on the involvement of endogenous excitatory amino acids in neuroendocrine activation during stress. Neurotoxic effects of glutamate in animals are well described, especially after its administration in the neonatal period. In men, glutamate toxicity and its use as a food additive are a continuous subject of discussions. The authors found an increase in plasma cortisol and norepinephrine, but not epinephrine and prolactin, in response to the administration of a high dose of glutamate. It cannot be excluded that these effects might be induced even by lower doses in situations with increased vulnerability to glutamate action (age, individual variability). (Tab. 1, Fig. 6, Ref. 44

  14. Ionotropic Glutamate Receptors & CNS Disorders

    PubMed Central

    Bowie, Derek

    2008-01-01

    Disorders of the central nervous system (CNS) are complex disease states that represent a major challenge for modern medicine. Although etiology is often unknown, it is established that multiple factors such as defects in genetics and/or epigenetics, the environment as well as imbalance in neurotransmitter receptor systems are all at play in determining an individual’s susceptibility to disease. Gene therapy is currently not available and therefore, most conditions are treated with pharmacological agents that modify neurotransmitter receptor signaling. Here, I provide a review of ionotropic glutamate receptors (iGluRs) and the roles they fulfill in numerous CNS disorders. Specifically, I argue that our understanding of iGluRs has reached a critical turning point to permit, for the first time, a comprehensive re-evaluation of their role in the cause of disease. I illustrate this by highlighting how defects in AMPA receptor trafficking are important to Fragile X mental retardation and ectopic expression of kainate (KA) receptor synapses contributes to the pathology of temporal lobe epilepsy. Finally, I discuss how parallel advances in studies of other neurotransmitter systems may allow pharmacologists to work towards a cure for many CNS disorders rather than developing drugs to treat their symptoms. PMID:18537642

  15. Medroxyprogesterone acetate exacerbates glutamate excitotoxicity.

    PubMed

    Nilsen, Jon; Morales, Alison; Brinton, Roberta Diaz

    2006-07-01

    We previously demonstrated that progesterone functions as a neuroprotective agent whereas medroxyprogesterone acetate (MPA; Provera) does not. Moreover, MPA antagonized the neuroprotective and neurotrophic outcomes induced by 17beta-estradiol (E2). Towards developing effective hormone therapies for protection against neurodegeneration, we sought to determine whether formulation, chemical features or prevention versus treatment mode of exposure affected the outcome of MPA treatment in survival of primary hippocampal neurons. Results of these analyses indicated that both crystalline MPA and a pharmaceutical formulation (Depo-Provera) lacked neuroprotective efficacy, indicating that the effects were not dependent upon MPA formulation. Likewise, MPA in the prevention and treatment paradigms were equally ineffective at promoting neuronal survival, indicating that timing of MPA administration was not a factor. Further, the detrimental effects of MPA were not due to the presence of the acetate group, as medroxyprogesterone was as ineffective as MPA in promoting neuronal survival. Moreover, MPA pretreatment exacerbated neuron death induced by glutamate excitotoxicity as indicated by a 40% increase in neuron death determined by direct live/dead cell count and a commensurate increase in the number of positive cells by terminal deoxynucleotidyl transferase-mediated nick end-labeling. Collectively these results predict that the progestin formulation of hormone therapy will affect the vulnerability of the central nervous system to degenerative insults.

  16. The Degradation of 14C-Glutamic Acid by L-Glutamic Acid Decarboxylase.

    ERIC Educational Resources Information Center

    Dougherty, Charles M; Dayan, Jean

    1982-01-01

    Describes procedures and semi-micro reaction apparatus (carbon dioxide trap) to demonstrate how a particular enzyme (L-Glutamic acid decarboxylase) may be used to determine the site or sites of labeling in its substrate (carbon-14 labeled glutamic acid). Includes calculations, solutions, and reagents used. (Author/SK)

  17. Glutamate Metabolism in Major Depressive Disorder

    PubMed Central

    Abdallah, Chadi G.; Jiang, Lihong; De Feyter, Henk M.; Fasula, Madonna; Krystal, John H.; Rothman, Douglas L.; Mason, Graeme F.; Sanacora, Gerard

    2015-01-01

    Objective Emerging evidence suggests abnormalities in amino acid neurotransmitter function and impaired energy metabolism contribute to the underlying pathophysiology of Major Depressive Disorder (MDD). To test whether impairments in energetics and glutamate neurotransmitter cycling are present in MDD we used in vivo 13C magnetic resonance spectroscopy (13C MRS) to measure these fluxes in individuals diagnosed with MDD relative to non-depressed subjects. Method 1H MRS and 13C MRS data were collected on 23 medication-free MDD and 17 healthy subjects. 1H MRS provided total glutamate and GABA concentrations, and 13C MRS, coupled with intravenous infusion of [1-13C]-glucose, provided measures of the neuronal tricarboxylic acid cycle (VTCAN) for mitochondrial energy production, GABA synthesis, and glutamate/glutamine cycling, from voxels placed in the occipital cortex. Results Our main finding was that mitochondrial energy production of glutamatergic neurons was reduced by 26% in MDD subjects (t = 2.57, p = 0.01). Paradoxically we found no difference in the rate of glutamate/glutamine cycle (Vcycle). We also found a significant correlation between glutamate concentrations and Vcycle considering the total sample. Conclusions We interpret the reduction in mitochondrial energy production as being due to either mitochondrial dysfunction or a reduction in proper neuronal input or synaptic strength. Future MRS studies could help distinguish these possibilities. PMID:25073688

  18. Flavor Preferences Conditioned by Dietary Glutamate.

    PubMed

    Ackroff, Karen; Sclafani, Anthony

    2016-07-01

    Our understanding of the molecular basis of umami taste and its appetitive qualities has been greatly aided by studies in laboratory rodents. This review describes methods for testing responses to the prototypical umami substance monosodium glutamate (MSG) in rodents. Two techniques, forced exposure to MSG and 2-bottle choice tests with ascending concentrations, were used to evaluate the responses to the taste of umami itself, and 2 other methods used oral or postoral MSG to modify the responses to other flavors. Intake and preference for MSG are enhanced in mice by experience with MSG and with other nutrients with positive postoral effects. In addition, flavor preferences are enhanced in mice and rats by gastric or intestinal MSG infusions via an associative learning process. Even mice with an impaired or absent ability to taste MSG can learn to prefer a flavor added to an MSG solution, supporting the notion that glutamate acts postorally. The more complex flavor of dashi seasoning, which includes umami substances (inosinate, glutamate), is attractive to rodents, but dashi does not condition flavor preferences. Details of the postoral glutamate detection process and the nature of the signal involved in learned preferences are still uncertain but probably involve gastric or intestinal sensors or both and vagal transmission. Some findings suggest that postoral glutamate effects may enhance food preferences in humans, but this requires further study. PMID:27422522

  19. GLT-1: The elusive presynaptic glutamate transporter.

    PubMed

    Rimmele, Theresa S; Rosenberg, Paul A

    2016-09-01

    Historically, glutamate uptake in the CNS was mainly attributed to glial cells for three reasons: 1) none of the glutamate transporters were found to be located in presynaptic terminals of excitatory synapses; 2) the putative glial transporters, GLT-1 and GLAST are expressed at high levels in astrocytes; 3) studies of the constitutive GLT-1 knockout as well as pharmacological studies demonstrated that >90% of glutamate uptake into forebrain synaptosomes is mediated by the operation of GLT-1. Here we summarize the history leading up to the recognition of GLT-1a as a presynaptic glutamate transporter. A major issue now is understanding the physiological and pathophysiological significance of the expression of GLT-1 in presynaptic terminals. To elucidate the cell-type specific functions of GLT-1, a conditional knockout was generated with which to inactivate the GLT-1 gene in different cell types using Cre/lox technology. Astrocytic knockout led to an 80% reduction of GLT-1 expression, resulting in intractable seizures and early mortality as seen also in the constitutive knockout. Neuronal knockout was associated with no obvious phenotype. Surprisingly, synaptosomal uptake capacity (Vmax) was found to be significantly reduced, by 40%, in the neuronal knockout, indicating that the contribution of neuronal GLT-1 to synaptosomal uptake is disproportionate to its protein expression (5-10%). Conversely, the contribution of astrocytic GLT-1 to synaptosomal uptake was much lower than expected. In contrast, the loss of uptake into liposomes prepared from brain protein from astrocyte and neuronal knockouts was proportionate with the loss of GLT-1 protein, suggesting that a large portion of GLT-1 in astrocytic membranes in synaptosomal preparations is not functional, possibly because of a failure to reseal. These results suggest the need to reinterpret many previous studies using synaptosomal uptake to investigate glutamate transport itself as well as changes in glutamate

  20. GLT-1: The elusive presynaptic glutamate transporter.

    PubMed

    Rimmele, Theresa S; Rosenberg, Paul A

    2016-09-01

    Historically, glutamate uptake in the CNS was mainly attributed to glial cells for three reasons: 1) none of the glutamate transporters were found to be located in presynaptic terminals of excitatory synapses; 2) the putative glial transporters, GLT-1 and GLAST are expressed at high levels in astrocytes; 3) studies of the constitutive GLT-1 knockout as well as pharmacological studies demonstrated that >90% of glutamate uptake into forebrain synaptosomes is mediated by the operation of GLT-1. Here we summarize the history leading up to the recognition of GLT-1a as a presynaptic glutamate transporter. A major issue now is understanding the physiological and pathophysiological significance of the expression of GLT-1 in presynaptic terminals. To elucidate the cell-type specific functions of GLT-1, a conditional knockout was generated with which to inactivate the GLT-1 gene in different cell types using Cre/lox technology. Astrocytic knockout led to an 80% reduction of GLT-1 expression, resulting in intractable seizures and early mortality as seen also in the constitutive knockout. Neuronal knockout was associated with no obvious phenotype. Surprisingly, synaptosomal uptake capacity (Vmax) was found to be significantly reduced, by 40%, in the neuronal knockout, indicating that the contribution of neuronal GLT-1 to synaptosomal uptake is disproportionate to its protein expression (5-10%). Conversely, the contribution of astrocytic GLT-1 to synaptosomal uptake was much lower than expected. In contrast, the loss of uptake into liposomes prepared from brain protein from astrocyte and neuronal knockouts was proportionate with the loss of GLT-1 protein, suggesting that a large portion of GLT-1 in astrocytic membranes in synaptosomal preparations is not functional, possibly because of a failure to reseal. These results suggest the need to reinterpret many previous studies using synaptosomal uptake to investigate glutamate transport itself as well as changes in glutamate

  1. Mood disorders: regulation by metabotropic glutamate receptors.

    PubMed

    Pilc, Andrzej; Chaki, Shigeyuki; Nowak, Gabriel; Witkin, Jeffrey M

    2008-03-01

    Medicinal therapies for mood disorders neither fully serve the efficacy needs of patients nor are they free of side-effect issues. Although monoamine-based therapies are the primary current treatment approaches, both preclinical and clinical findings have implicated the excitatory neurotransmitter glutamate in the pathogenesis of major depressive disorders. The present commentary focuses on the metabotropic glutamate receptors and their relationship to mood disorders. Metabotropic glutamate (mGlu) receptors regulate glutamate transmission by altering the release of neurotransmitter and/or modulating the post-synaptic responses to glutamate. Convergent biochemical, pharmacological, behavioral, and clinical data will be reviewed that establish glutamatergic neurotransmission via mGlu receptors as a biologically relevant process in the regulation of mood and that these receptors may serve as novel targets for the discovery of small molecule modulators with unique antidepressant properties. Specifically, compounds that antagonize mGlu2, mGlu3, and/or mGlu5 receptors (e.g. LY341495, MGS0039, MPEP, MTEP) exhibit biochemical effects indicative of antidepressant effects as well as in vivo activity in animal models predictive of antidepressant efficacy. Both preclinical and clinical data have previously been presented to define NMDA and AMPA receptors as important targets for the modulation of major depression. In the present review, we present a model suggesting how the interplay of glutamate at the mGlu and at the ionotropic AMPA and NMDA receptors might account for the antidepressant-like effects of glutamatergic- and monoaminergic-based drugs affecting mood in patients. The current data lead to the hypothesis that mGlu-based compounds and conventional antidepressants impact a network of interactive effects that converge upon a down regulation of NMDA receptor function and an enhancement in AMPA receptor signaling. PMID:18164691

  2. Luminal l-glutamate enhances duodenal mucosal defense mechanisms via multiple glutamate receptors in rats

    PubMed Central

    Watanabe, Chikako; Mizumori, Misa; Kaunitz, Jonathan D.

    2009-01-01

    Presence of taste receptor families in the gastrointestinal mucosa suggests a physiological basis for local and early detection of a meal. We hypothesized that luminal l-glutamate, which is the primary nutrient conferring fundamental umami or proteinaceous taste, influences mucosal defense mechanisms in rat duodenum. We perfused the duodenal mucosa of anesthetized rats with l-glutamate (0.1–10 mM). Intracellular pH (pHi) of the epithelial cells, blood flow, and mucus gel thickness (MGT) were simultaneously and continuously measured in vivo. Some rats were pretreated with indomethacin or capsaicin. Duodenal bicarbonate secretion (DBS) was measured with flow-through pH and CO2 electrodes. We tested the effects of agonists or antagonists for metabotropic glutamate receptor (mGluR) 1 or 4 or calcium-sensing receptor (CaSR) on defense factors. Luminal l-glutamate dose dependently increased pHi and MGT but had no effect on blood flow in the duodenum. l-glutamate (10 mM)-induced cellular alkalinization and mucus secretion were inhibited by pretreatment with indomethacin or capsaicin. l-glutamate effects on pHi and MGT were mimicked by mGluR4 agonists and inhibited by an mGluR4 antagonist. CaSR agonists acidified cells with increased MGT and DBS, unlike l-glutamate. Perfusion of l-glutamate with inosinate (inosine 5′-monophosphate, 0.1 mM) enhanced DBS only in combination, suggesting synergistic activation of the l-glutamate receptor, typical of taste receptor type 1. l-leucine or l-aspartate had similar effects on DBS without any effect on pHi and MGT. Preperfusion of l-glutamate prevented acid-induced cellular injury, suggesting that l-glutamate protects the mucosa by enhancing mucosal defenses. Luminal l-glutamate may activate multiple receptors and afferent nerves and locally enhance mucosal defenses to prevent subsequent injury attributable to acid exposure in the duodenum. PMID:19643955

  3. Extracorporeal methods of blood glutamate scavenging: a novel therapeutic modality.

    PubMed

    Zhumadilov, Agzam; Boyko, Matthew; Gruenbaum, Shaun E; Brotfain, Evgeny; Bilotta, Federico; Zlotnik, Alexander

    2015-05-01

    Pathologically elevated glutamate concentrations in the brain's extracellular fluid are associated with several acute and chronic brain insults. Studies have demonstrated that by decreasing the concentration of glutamate in the blood, thereby increasing the concentration gradient between the brain and the blood, the rate of brain-to-blood glutamate efflux can be increased. Blood glutamate scavengers, pyruvate and oxaloacetate have shown great promise in providing neuroprotection in many animal models of acute brain insults. However, glutamate scavengers' potential systemic toxicity, side effects and pharmacokinetic properties may limit their use in clinical practice. In contrast, extracorporeal methods of blood glutamate reduction, in which glutamate is filtered from the blood and eliminated, may be an advantageous adjunct in treating acute brain insults. Here, we review the current evidence for the glutamate-lowering effects of hemodialysis, peritoneal dialysis and hemofiltration. The evidence reviewed here highlights the need for clinical trials.

  4. 21 CFR 182.1047 - Glutamic acid hydrochloride.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 3 2014-04-01 2014-04-01 false Glutamic acid hydrochloride. 182.1047 Section 182...) SUBSTANCES GENERALLY RECOGNIZED AS SAFE Multiple Purpose GRAS Food Substances § 182.1047 Glutamic acid hydrochloride. (a) Product. Glutamic acid hydrochloride. (b) (c) Limitations, restrictions, or explanation....

  5. 21 CFR 182.1047 - Glutamic acid hydrochloride.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... 21 Food and Drugs 3 2010-04-01 2009-04-01 true Glutamic acid hydrochloride. 182.1047 Section 182.1047 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... Food Substances § 182.1047 Glutamic acid hydrochloride. (a) Product. Glutamic acid hydrochloride....

  6. 21 CFR 182.1047 - Glutamic acid hydrochloride.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 3 2012-04-01 2012-04-01 false Glutamic acid hydrochloride. 182.1047 Section 182.1047 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... Food Substances § 182.1047 Glutamic acid hydrochloride. (a) Product. Glutamic acid hydrochloride....

  7. 21 CFR 182.1047 - Glutamic acid hydrochloride.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... 21 Food and Drugs 3 2011-04-01 2011-04-01 false Glutamic acid hydrochloride. 182.1047 Section 182.1047 Food and Drugs FOOD AND DRUG ADMINISTRATION, DEPARTMENT OF HEALTH AND HUMAN SERVICES (CONTINUED... Food Substances § 182.1047 Glutamic acid hydrochloride. (a) Product. Glutamic acid hydrochloride....

  8. Localization of L-glutamate and glutamate-like receptors at the squid giant synapse.

    PubMed

    Di Cosmo, A; Nardi, G; Di Cristo, C; De Santis, A; Messenger, J B

    1999-08-28

    HPLC analysis of the amino acid contents of the second- and third-order giant fibres at the giant synapse in the stellate ganglion of the squid Loligo vulgaris shows that there are significantly higher amounts of L-glutamate and L-aspartate in the second-order (presynaptic) fibre than in the third-order (postsynaptic) fibre. Immunocytochemical staining of sections of the ganglion with an antibody raised against L-glutamate produces specific positive staining in the synaptic region of the second-order fibre. In contrast, staining with antibodies raised against glutamate-receptors (mammalian GluR1 with GluR2/3) produces positive staining in the third-order fibre at the postsynaptic region. These data provide further evidence for the hypothesis that L-glutamate is an excitatory transmitter at the giant synapse.

  9. Circuit mapping by ultraviolet uncaging of glutamate.

    PubMed

    Shepherd, Gordon M G

    2012-09-01

    In laser photostimulation, small clusters of neurons in brain slices are induced to fire action potentials by focal glutamate uncaging, and synaptic connectivity between photoexcited presynaptic neurons and individual postsynaptic neurons is assessed by intracellular recording of synaptic events. With a scanner, this process can be repeated sequentially across a patterned array of stimulus locations, generating maps of neurons' local sources of synaptic inputs. Laser scanning photostimulation (LSPS) based on patterned glutamate uncaging offers an efficient, quantitative, optical-electrophysiological way to map synaptic circuits in brain slices. The efficacy of glutamate-based photostimulation for circuit mapping (in contrast to electrical stimulation) derives from the ability to stimulate neurons with high precision and speed, and without stimulating axons of passage. This protocol describes the components, assembly, and operation of a laser scanning microscope for ultraviolet (UV) uncaging, along with experimental methods for circuit mapping in brain slices. It presents a general approach and a set of guidelines for quantitative circuit mapping using "standard" LSPS methods based on single-photon glutamate uncaging using a UV laser, a pair of scanning mirror galvanometers, a patch-clamp setup, and open-source data acquisition software. PMID:22949715

  10. L-glutamate Receptor In Paramecium

    NASA Astrophysics Data System (ADS)

    Bernal-Martínez, Juan; Ortega-Soto, Arturo

    2004-09-01

    Behavioral, electrophysiological and biochemical experiments were performed in order to establish the presence of a glutamate receptor in the ciliate Paramecium. It was found that an AMPA/KA receptor is functionally expressed in Paramecium and that this receptor is immunologically and fillogenetically related to the AMPA/KA receptor present in vertebrates.

  11. Glutamate Mediated Astrocytic Filtering of Neuronal Activity

    PubMed Central

    Herzog, Nitzan; De Pittà, Maurizio; Jacob, Eshel Ben; Berry, Hugues; Hanein, Yael

    2014-01-01

    Neuron-astrocyte communication is an important regulatory mechanism in various brain functions but its complexity and role are yet to be fully understood. In particular, the temporal pattern of astrocyte response to neuronal firing has not been fully characterized. Here, we used neuron-astrocyte cultures on multi-electrode arrays coupled to Ca2+ imaging and explored the range of neuronal stimulation frequencies while keeping constant the amount of stimulation. Our results reveal that astrocytes specifically respond to the frequency of neuronal stimulation by intracellular Ca2+ transients, with a clear onset of astrocytic activation at neuron firing rates around 3-5 Hz. The cell-to-cell heterogeneity of the astrocyte Ca2+ response was however large and increasing with stimulation frequency. Astrocytic activation by neurons was abolished with antagonists of type I metabotropic glutamate receptor, validating the glutamate-dependence of this neuron-to-astrocyte pathway. Using a realistic biophysical model of glutamate-based intracellular calcium signaling in astrocytes, we suggest that the stepwise response is due to the supralinear dynamics of intracellular IP3 and that the heterogeneity of the responses may be due to the heterogeneity of the astrocyte-to-astrocyte couplings via gap junction channels. Therefore our results present astrocyte intracellular Ca2+ activity as a nonlinear integrator of glutamate-dependent neuronal activity. PMID:25521344

  12. Structural Features of the Glutamate Transporter Family

    PubMed Central

    Slotboom, Dirk Jan; Konings, Wil N.; Lolkema, Juke S.

    1999-01-01

    Neuronal and glial glutamate transporters remove the excitatory neurotransmitter glutamate from the synaptic cleft and thus prevent neurotoxicity. The proteins belong to a large and widespread family of secondary transporters, including bacterial glutamate, serine, and C4-dicarboxylate transporters; mammalian neutral-amino-acid transporters; and an increasing number of bacterial, archaeal, and eukaryotic proteins that have not yet been functionally characterized. Sixty members of the glutamate transporter family were found in the databases on the basis of sequence homology. The amino acid sequences of the carriers have diverged enormously. Homology between the members of the family is most apparent in a stretch of approximately 150 residues in the C-terminal part of the proteins. This region contains four reasonably well-conserved sequence motifs, all of which have been suggested to be part of the translocation pore or substrate binding site. Phylogenetic analysis of the C-terminal stretch revealed the presence of five subfamilies with characterized members: (i) the eukaryotic glutamate transporters, (ii) the bacterial glutamate transporters, (iii) the eukaryotic neutral-amino-acid transporters, (iv) the bacterial C4-dicarboxylate transporters, and (v) the bacterial serine transporters. A number of other subfamilies that do not contain characterized members have been defined. In contrast to their amino acid sequences, the hydropathy profiles of the members of the family are extremely well conserved. Analysis of the hydropathy profiles has suggested that the glutamate transporters have a global structure that is unique among secondary transporters. Experimentally, the unique structure of the transporters was recently confirmed by membrane topology studies. Although there is still controversy about part of the topology, the most likely model predicts the presence of eight membrane-spanning α-helices and a loop-pore structure which is unique among secondary

  13. Glutamate Receptor Agonists and Glutamate Transporter Antagonists Regulate Differentiation of Osteoblast Lineage Cells.

    PubMed

    Xie, Wenjie; Dolder, Silvia; Siegrist, Mark; Wetterwald, Antoinette; Hofstetter, Willy

    2016-08-01

    Development and function of osteoblast lineage cells are regulated by a complex microenvironment consisting of the bone extracellular matrix, cells, systemic hormones and cytokines, autocrine and paracrine factors, and mechanical load. Apart from receptors that transduce extracellular signals into the cell, molecular transporters play a crucial role in the cellular response to the microenvironment. Transporter molecules are responsible for cellular uptake of nutritional components, elimination of metabolites, ion transport, and cell-cell communication. In this report, the expression of molecular transporters in osteoblast lineage cells was investigated to assess their roles in cell development and activity. Low-density arrays, covering membrane and vesicular transport molecules, were used to assess gene expression in osteoblasts representing early and late differentiation states. Receptors and transporters for the amino acid glutamate were found to be differentially expressed during osteoblast development. Glutamate is a neurotransmitter in the central nervous system, and the mechanisms of its release, signal transduction, and cellular reabsorption in the synaptic cleft are well understood. Less clear, however, is the control of equivalent processes in peripheral tissues. In primary osteoblasts, inhibition of glutamate transporters with nonselective inhibitors leads to an increase in the concentration of extracellular glutamate. This change was accompanied by a decrease in osteoblast proliferation, stimulation of alkaline phosphatase, and the expression of transcripts encoding osteocalcin. Enzymatic removal of extracellular glutamate abolished these pro-differentiation effects, as did the inhibition of PKC- and Erk1/2-signaling pathways. These findings demonstrate that glutamate signaling promotes differentiation and activation of osteoblast lineage cells. Consequently, the glutamate system may represent a putative therapeutic target to induce an anabolic response

  14. Amperometric L-glutamate biosensor based on bacterial cell-surface displayed glutamate dehydrogenase.

    PubMed

    Liang, Bo; Zhang, Shu; Lang, Qiaolin; Song, Jianxia; Han, Lihui; Liu, Aihua

    2015-07-16

    A novel L-glutamate biosensor was fabricated using bacteria surface-displayed glutamate dehydrogenase (Gldh-bacteria). Here the cofactor NADP(+)-specific dependent Gldh was expressed on the surface of Escherichia coli using N-terminal region of ice nucleation protein (INP) as the anchoring motif. The cell fractionation assay and SDS-PAGE analysis indicated that the majority of INP-Gldh fusion proteins were located on the surface of cells. The biosensor was fabricated by successively casting polyethyleneimine (PEI)-dispersed multi-walled carbon nanotubes (MWNTs), Gldh-bacteria and Nafion onto the glassy carbon electrode (Nafion/Gldh-bacteria/PEI-MWNTs/GCE). The MWNTs could not only significantly lower the oxidation overpotential towards NAPDH, which was the product of NADP(+) involving in the oxidation of glutamate by Gldh, but also enhanced the current response. Under the optimized experimental conditions, the current-time curve of the Nafion/Gldh-bacteria/PEI-MWNTs/GCE was performed at +0.52 V (vs. SCE) by amperometry varying glutamate concentration. The current response was linear with glutamate concentration in two ranges (10 μM-1 mM and 2-10 mM). The low limit of detection was estimated to be 2 μM glutamate (S/N=3). Moreover, the proposed biosensor is stable, specific, reproducible and simple, which can be applied to real samples detection.

  15. Amperometric L-glutamate biosensor based on bacterial cell-surface displayed glutamate dehydrogenase.

    PubMed

    Liang, Bo; Zhang, Shu; Lang, Qiaolin; Song, Jianxia; Han, Lihui; Liu, Aihua

    2015-07-16

    A novel L-glutamate biosensor was fabricated using bacteria surface-displayed glutamate dehydrogenase (Gldh-bacteria). Here the cofactor NADP(+)-specific dependent Gldh was expressed on the surface of Escherichia coli using N-terminal region of ice nucleation protein (INP) as the anchoring motif. The cell fractionation assay and SDS-PAGE analysis indicated that the majority of INP-Gldh fusion proteins were located on the surface of cells. The biosensor was fabricated by successively casting polyethyleneimine (PEI)-dispersed multi-walled carbon nanotubes (MWNTs), Gldh-bacteria and Nafion onto the glassy carbon electrode (Nafion/Gldh-bacteria/PEI-MWNTs/GCE). The MWNTs could not only significantly lower the oxidation overpotential towards NAPDH, which was the product of NADP(+) involving in the oxidation of glutamate by Gldh, but also enhanced the current response. Under the optimized experimental conditions, the current-time curve of the Nafion/Gldh-bacteria/PEI-MWNTs/GCE was performed at +0.52 V (vs. SCE) by amperometry varying glutamate concentration. The current response was linear with glutamate concentration in two ranges (10 μM-1 mM and 2-10 mM). The low limit of detection was estimated to be 2 μM glutamate (S/N=3). Moreover, the proposed biosensor is stable, specific, reproducible and simple, which can be applied to real samples detection. PMID:26073813

  16. Altered Connexin 43 and Connexin 45 protein expression in the heart as a function of social and environmental stress in the prairie vole.

    PubMed

    Grippo, Angela J; Moffitt, Julia A; Henry, Matthew K; Firkins, Rachel; Senkler, Jonathan; McNeal, Neal; Wardwell, Joshua; Scotti, Melissa-Ann L; Dotson, Ashley; Schultz, Rachel

    2015-01-01

    Exposure to social and environmental stressors may influence behavior as well as autonomic and cardiovascular regulation, potentially leading to depressive disorders and cardiac dysfunction including elevated sympathetic drive, reduced parasympathetic function, and ventricular arrhythmias. The cellular mechanisms that underlie these interactions are not well understood. One mechanism may involve alterations in the expression of Connexin43 (Cx43) and Connexin45 (Cx45), gap junction proteins in the heart that play an important role in ensuring efficient cell-to-cell coupling and the maintenance of cardiac rhythmicity. The present study investigated the hypothesis that long-term social isolation, combined with mild environmental stressors, would produce both depressive behaviors and altered Cx43 and Cx45 expression in the left ventricle of prairie voles - a socially monogamous rodent model. Adult, female prairie voles were exposed to either social isolation (n = 22) or control (paired, n = 23) conditions (4 weeks), alone or in combination with chronic mild stress (CMS) (1 week). Social isolation, versus paired control conditions, produced significantly (p < 0.05) increased depressive behaviors in a 5-min forced swim test, and CMS exacerbated (p < 0.05) these behaviors. Social isolation (alone) reduced (p < 0.05) total Cx43 expression in the left ventricle; whereas CMS (but not isolation) increased (p < 0.05) total Cx45 expression and reduced (p < 0.05) the Cx43/Cx45 ratio, measured via Western blot analysis. The present findings provide insight into potential cellular mechanisms underlying altered cardiac rhythmicity associated with social and environmental stress in the prairie vole.

  17. Altered Connexin 43 and Connexin 45 Protein Expression in the Heart as a Function of Social and Environmental Stress in the Prairie Vole

    PubMed Central

    Grippo, Angela J.; Moffitt, Julia A.; Henry, Matthew K.; Firkins, Rachel; Senkler, Jonathan; McNeal, Neal; Wardwell, Joshua; Scotti, Melissa-Ann L.; Dotson, Ashley; Schultz, Rachel

    2015-01-01

    Exposure to social and environmental stressors may influence behavior as well as autonomic and cardiovascular regulation, potentially leading to depressive disorders and cardiac dysfunction including elevated sympathetic drive, reduced parasympathetic function, and ventricular arrhythmias. The cellular mechanisms that underlie these interactions are not well understood. One mechanism may involve alterations in the expression of Connexin43 (Cx43) and Connexin45 (Cx45), gap junction proteins in the heart that play an important role in ensuring efficient cell-to-cell coupling and the maintenance of cardiac rhythmicity. The present study investigated the hypothesis that long-term social isolation, combined with mild environmental stressors, would produce both depressive behaviors and altered Cx43 and Cx45 expression in the left ventricle of prairie voles – a socially monogamous rodent model. Adult, female prairie voles were exposed to either social isolation (n=22) or control (paired, n=23) conditions (4 weeks), alone or in combination with chronic mild stress (1 week). Social isolation, versus paired control conditions, produced significantly (P < 0.05) increased depressive behaviors in a 5-min forced swim test, and chronic mild stress exacerbated (P < 0.05) these behaviors. Social isolation (alone) reduced (P < 0.05) total Cx43 expression in the left ventricle; whereas chronic mild stress (but not isolation) increased (P < 0.05) total Cx45 expression and reduced (P < 0.05) the Cx43/Cx45 ratio, measured via Western blot analysis. The present findings provide insight into potential cellular mechanisms underlying altered cardiac rhythmicity associated with social and environmental stress in the prairie vole. PMID:25338193

  18. 3-Aminoglutarate is a "silent" false transmitter for glutamate neurons.

    PubMed

    Foster, Alan C; Chen, June; Runyan, Stephen; Dinh, Tim; Venadas, Steven; Ehring, George R; Li, Yong-Xin; Staubli, Ursula

    2015-10-01

    Understanding the storage and release of the excitatory neurotransmitter, L-glutamate by synaptic vesicles has lagged behind receptor characterizations due to a lack of pharmacological agents. We report that the glutamate analog, 3-aminoglutarate (3-AG) is a "silent" false transmitter for glutamate neurons that may be a useful tool to study storage and release mechanisms. Like L-glutamate itself, 3-AG is a high-affinity substrate for both the plasma membrane (EAATs) and vesicular (vGLUT) glutamate transporters. As such, EAATs facilitate 3-AG entry into neuronal cytoplasm allowing 3-AG to compete with L-glutamate for transport into vesicles thus reducing glutamate content. In a synaptosomal preparation, 3-AG inhibited calcium-dependent endogenous L-glutamate release. Unlike L-glutamate, 3-AG had low affinity for both ionotropic (NMDA and AMPA) and G-protein coupled (mGlu1-8) receptors. Consequently, 3-AG behaves as a "silent" false transmitter that may be used in physiological experiments to probe synaptic vesicle storage and release mechanisms for L-glutamate. The companion paper by Wu et al. (2015) describes initial experiments that explore the effects of 3-AG on glutamate synaptic transmission under physiological and pathophysiological conditions.

  19. Understanding safety of glutamate in food and brain.

    PubMed

    Mallick, H N

    2007-01-01

    Glutamate is ubiquitous in nature and is present in all living organisms. It is the principal excitatory neurotransmitter in central nervous system. Glutamate is being used as food additive for enhancing flavour for over last 1200 years imparting a unique taste known as "umami" in Japanese. It is being marketed for about last 100 years. The taste of umami is now recognized as the fifth basic taste. Many of the foods used in cooking for enhancing flavour contain high amount of glutamate. Breast milk has the highest concentration of glutamate amongst all amino acids. Glutamate in high doses as gavage or parenteral injection have been reported to produce neurodegeneration in infant rodents. The neurodegeneration was not produced when gluamate was given with food. The Joint FAO/WHO Expert Committee on Food Additives, based on enumerable scientific evidence, has declared that, "glutamate as an additive in food" is not an health hazard to human being. Glutamate is used as signaling molecule not only in neuronal but also in non-neuronal tissues. Excessive accumulation of glutamate in the synaptic cleft has been associated with excitotoxicty and glutamate is implicated in number of neurological disorders. Excessive accumulation could be attributed to increase release, failure of transport system for uptake mechanism, neuronal injury due to hypoxia-ischemia, trauma and associated metabolic failures. The role blood brain barrier, vesicular glutamate and sodium dependent excitatory amino acid transporters in glutamate homeostasis are emphasized in the review.

  20. Neuronal Activity and Glutamate Uptake Decrease Mitochondrial Mobility in Astrocytes and Position Mitochondria Near Glutamate Transporters

    PubMed Central

    Jackson, Joshua G.; O'Donnell, John C.; Takano, Hajime; Coulter, Douglas A.

    2014-01-01

    Within neurons, mitochondria are nonuniformly distributed and are retained at sites of high activity and metabolic demand. Glutamate transport and the concomitant activation of the Na+/K+-ATPase represent a substantial energetic demand on astrocytes. We hypothesized that mitochondrial mobility within astrocytic processes might be regulated by neuronal activity and glutamate transport. We imaged organotypic hippocampal slice cultures of rat, in which astrocytes maintain their highly branched morphologies and express glutamate transporters. Using time-lapse confocal microscopy, the mobility of mitochondria within individual astrocytic processes and neuronal dendrites was tracked. Within neurons, a greater percentage of mitochondria were mobile than in astrocytes. Furthermore, they moved faster and farther than in astrocytes. Inhibiting neuronal activity with tetrodotoxin (TTX) increased the percentage of mobile mitochondria in astrocytes. Mitochondrial movement in astrocytes was inhibited by vinblastine and cytochalasin D, demonstrating that this mobility depends on both the microtubule and actin cytoskeletons. Inhibition of glutamate transport tripled the percentage of mobile mitochondria in astrocytes. Conversely, application of the transporter substrate d-aspartate reversed the TTX-induced increase in the percentage of mobile mitochondria. Inhibition of reversed Na+/Ca2+ exchange also increased the percentage of mitochondria that were mobile. Last, we demonstrated that neuronal activity increases the probability that mitochondria appose GLT-1 particles within astrocyte processes, without changing the proximity of GLT-1 particles to VGLUT1. These results imply that neuronal activity and the resulting clearance of glutamate by astrocytes regulate the movement of astrocytic mitochondria and suggest a mechanism by which glutamate transporters might retain mitochondria at sites of glutamate uptake. PMID:24478345

  1. Three Distinct Glutamate Decarboxylase Genes in Vertebrates

    PubMed Central

    Grone, Brian P.; Maruska, Karen P.

    2016-01-01

    Gamma-aminobutyric acid (GABA) is a widely conserved signaling molecule that in animals has been adapted as a neurotransmitter. GABA is synthesized from the amino acid glutamate by the action of glutamate decarboxylases (GADs). Two vertebrate genes, GAD1 and GAD2, encode distinct GAD proteins: GAD67 and GAD65, respectively. We have identified a third vertebrate GAD gene, GAD3. This gene is conserved in fishes as well as tetrapods. We analyzed protein sequence, gene structure, synteny, and phylogenetics to identify GAD3 as a homolog of GAD1 and GAD2. Interestingly, we found that GAD3 was lost in the hominid lineage. Because of the importance of GABA as a neurotransmitter, GAD3 may play important roles in vertebrate nervous systems. PMID:27461130

  2. Concentration-Dependent Processivity of Multiple Glutamate Ligations Catalyzed by Folylpoly-γ-glutamate Synthetase

    PubMed Central

    Tomsho, John W.; Moran, Richard G.; Coward, James K.

    2010-01-01

    Folylpoly-γ-glutamate synthetase (FPGS, EC 6.3.2.17) is an ATP-dependent ligase that catalyzes formation of poly-γ-glutamate derivatives of reduced folates and anti-folates such as methotrexate and 5,10-dideaza-5,6,7,8-tetrahydrofolate (DDAH4PteGlu1). While the chemical mechanism of the reaction catalyzed by FPGS is known, it is unknown whether single or multiple glutamate residues are added following each folate binding event. A very sensitive high performance liquid chromatography method has been used to analyze the multiple ligation reactions onto radiolabeled DDAH4PteGlu1 catalyzed by FPGS in order to distinguish between distributive or processive mechanisms of catalysis. Reaction time courses, substrate trapping, and pulse-chase experiments were used to measure folate release during multiple glutamate additions. Together, the results of these experiments indicate that hFPGS can catalyze the processive addition of approximately four glutamate residues onto DDAH4PteGlu1. The degree of processivity was determined to be dependent on the concentration of the folate substrate, thus suggesting a mechanism for the regulation of folate polyglutamate synthesis in cells. PMID:18672898

  3. Ketamine and other potential glutamate antidepressants.

    PubMed

    Dutta, Arpan; McKie, Shane; Deakin, J F William

    2015-01-30

    The need for rapid acting antidepressants is widely recognised. There has been much interest in glutamate mechanisms in major depressive disorder (MDD) as a promising target for the development of new antidepressants. A single intravenous infusion of ketamine, a N-methyl-d-aspartate (NMDA) receptor antagonist anaesthetic agent, can alleviate depressive symptoms in patients within hours of administration. The mechanism of action appears to be in part through glutamate release onto non-NMDA receptors including α-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) and metabotropic receptors. However these are also reported effects on 5-HT, dopamine and intracellular effects on the mammalian target of rapamycin (mTOR) pathway. The effects of SSRI (Selective Serotonin Reuptake Inhibitor) antidepressants may also involve alterations in NMDA function. The article reviews the effect of current antidepressants on NMDA and examines the efficacy and mechanism of ketamine. Response to ketamine is also discussed and comparison with other glutamate drugs including lamotrigine, amantadine, riluzole, memantine, traxoprodil, GLYX-13, MK-0657, RO4917523, AZD2066 and Coluracetam. Future studies need to link the rapid antidepressant effects seen with ketamine to inflammatory theories in MDD.

  4. New perspectives in glutamate and anxiety.

    PubMed

    Riaza Bermudo-Soriano, Carlos; Perez-Rodriguez, M Mercedes; Vaquero-Lorenzo, Concepcion; Baca-Garcia, Enrique

    2012-02-01

    Anxiety and stress-related disorders, namely posttraumatic stress disorder (PTSD), generalized anxiety disorder (GAD), obsessive-compulsive disorder (ODC), social and specific phobias, and panic disorder, are a major public health issue. A growing body of evidence suggests that glutamatergic neurotransmission may be involved in the biological mechanisms underlying stress response and anxiety-related disorders. The glutamatergic system mediates the acquisition and extinction of fear-conditioning. Thus, new drugs targeting glutamatergic neurotransmission may be promising candidates for new pharmacological treatments. In particular, N-methyl-d-aspartate receptors (NMDAR) antagonists (AP5, AP7, CGP37849, CGP39551, LY235959, NPC17742, and MK-801), NMDAR partial agonists (DCS, ACPC), α-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate receptors (AMPARs) antagonists (topiramate), and several allosteric modulators targeting metabotropic glutamate receptors (mGluRs) mGluR1, mGluR2/3, and mGluR5, have shown anxiolytic-like effects in several animal and human studies. Several studies have suggested that polyamines (agmatine, putrescine, spermidine, and spermine) may be involved in the neurobiological mechanisms underlying stress-response and anxiety-related disorders. This could mainly be attributed to their ability to modulate ionotropic glutamate receptors, especially NR2B subunits. The aim of this review is to establish that glutamate neurotransmission and polyaminergic system play a fundamental role in the onset of anxiety-related disorders. This may open the way for new drugs that may help to treat these conditions.

  5. Glutamate involvement in calcium–dependent migration of astrocytoma cells

    PubMed Central

    2014-01-01

    Background Astrocytoma are known to have altered glutamate machinery that results in the release of large amounts of glutamate into the extracellular space but the precise role of glutamate in favoring cancer processes has not yet been fully established. Several studies suggested that glutamate might provoke active killing of neurons thereby producing space for cancer cells to proliferate and migrate. Previously, we observed that calcium promotes disassembly of integrin-containing focal adhesions in astrocytoma, thus providing a link between calcium signaling and cell migration. The aim of this study was to determine how calcium signaling and glutamate transmission cooperate to promote enhanced astrocytoma migration. Methods The wound-healing model was used to assay migration of human U87MG astrocytoma cells and allowed to monitor calcium signaling during the migration process. The effect of glutamate on calcium signaling was evaluated together with the amount of glutamate released by astrocytoma during cell migration. Results We observed that glutamate stimulates motility in serum-starved cells, whereas in the presence of serum, inhibitors of glutamate receptors reduce migration. Migration speed was also reduced in presence of an intracellular calcium chelator. During migration, cells displayed spontaneous Ca2+ transients. L-THA, an inhibitor of glutamate re-uptake increased the frequency of Ca2+ oscillations in oscillating cells and induced Ca2+ oscillations in quiescent cells. The frequency of migration-associated Ca2+ oscillations was reduced by prior incubation with glutamate receptor antagonists or with an anti-β1 integrin antibody. Application of glutamate induced increases in internal free Ca2+ concentration ([Ca2+]i). Finally we found that compounds known to increase [Ca2+]i in astrocytomas such as thapsigagin, ionomycin or the metabotropic glutamate receptor agonist t-ACPD, are able to induce glutamate release. Conclusion Our data demonstrate that

  6. Transcriptional regulation through glutamate receptors: Involvement of tyrosine kinases.

    PubMed

    López-Bayghen, Esther; Aguirre, Adán; Ortega, Arturo

    2003-12-01

    Glutamate receptors play a key role in neuronal plasticity, learning and memory, and in several neuropathologies. Short-term and long-term changes in synaptic efficacy are triggered by glutamate. Although an enhanced glutamate-dependent tyrosine phosphorylation has been described in several systems, its role in membrane-to-nuclei signaling is unclear. Taking advantage of the fact that the gene encoding the chick kainate-binding protein undergoes a glutamate-dependent transcriptional regulation via an activator protein-1 (AP-1) site, we evaluated the involvement of tyrosine kinases in this process. We describe here the participation of receptor and non-receptor tyrosine kinases in the signaling cascade triggered by glutamate. Our results suggest that in Bergmann glia cells, glutamate receptors transactivate receptor tyrosine kinases, favoring the idea of a complex network of signals activated by this excitatory neurotransmitter that results in regulation of gene expression.

  7. [Determination of glutamic acid in biological material by capillary electrophoresis].

    PubMed

    Narezhnaya, E; Krukier, I; Avrutskaya, V; Degtyareva, A; Igumnova, E A

    2015-01-01

    The conditions for the identification and determination of Glutamic acid by capillary zone electrophoresis without their preliminary derivatization have been optimized. The effect of concentration of buffer electrolyte and pH on determination of Glutamic acid has been investigated. It is shown that the 5 Mm borate buffer concentration and a pH 9.15 are optimal. Quantitative determination of glutamic acid has been carried out using a linear dependence between the concentration of the analyte and the area of the peak. The accuracy and reproducibility of the determination are confirmed by the method "introduced - found". Glutamic acid has been determined in the placenta homogenate. The duration of analysis doesn't exceed 30 minutes. The results showed a decrease in the level of glutamic acid in cases of pregnancy complicated by placental insufficiency compared with the physiological, and this fact allows to consider the level of glutamic acid as a possible marker of complicated pregnancy.

  8. Astroglial glutamate transporters coordinate excitatory signaling and brain energetics.

    PubMed

    Robinson, Michael B; Jackson, Joshua G

    2016-09-01

    In the mammalian brain, a family of sodium-dependent transporters maintains low extracellular glutamate and shapes excitatory signaling. The bulk of this activity is mediated by the astroglial glutamate transporters GLT-1 and GLAST (also called EAAT2 and EAAT1). In this review, we will discuss evidence that these transporters co-localize with, form physical (co-immunoprecipitable) interactions with, and functionally couple to various 'energy-generating' systems, including the Na(+)/K(+)-ATPase, the Na(+)/Ca(2+) exchanger, glycogen metabolizing enzymes, glycolytic enzymes, and mitochondria/mitochondrial proteins. This functional coupling is bi-directional with many of these systems both being regulated by glutamate transport and providing the 'fuel' to support glutamate uptake. Given the importance of glutamate uptake to maintaining synaptic signaling and preventing excitotoxicity, it should not be surprising that some of these systems appear to 'redundantly' support the energetic costs of glutamate uptake. Although the glutamate-glutamine cycle contributes to recycling of neurotransmitter pools of glutamate, this is an over-simplification. The ramifications of co-compartmentalization of glutamate transporters with mitochondria for glutamate metabolism are discussed. Energy consumption in the brain accounts for ∼20% of the basal metabolic rate and relies almost exclusively on glucose for the production of ATP. However, the brain does not possess substantial reserves of glucose or other fuels. To ensure adequate energetic supply, increases in neuronal activity are matched by increases in cerebral blood flow via a process known as 'neurovascular coupling'. While the mechanisms for this coupling are not completely resolved, it is generally agreed that astrocytes, with processes that extend to synapses and endfeet that surround blood vessels, mediate at least some of the signal that causes vasodilation. Several studies have shown that either genetic deletion or

  9. Exercise increases mitochondrial glutamate oxidation in the mouse cerebral cortex.

    PubMed

    Herbst, Eric A F; Holloway, Graham P

    2016-07-01

    The present study investigated the impact of acute exercise on stimulating mitochondrial respiratory function in mouse cerebral cortex. Where pyruvate-stimulated respiration was not affected by acute exercise, glutamate respiration was enhanced following the exercise bout. Additional assessment revealed that this affect was dependent on the presence of malate and did not occur when substituting glutamine for glutamate. As such, our results suggest that glutamate oxidation is enhanced with acute exercise through activation of the malate-aspartate shuttle. PMID:27184881

  10. [Autoantibodies to glutamate and GABA in opiate addiction].

    PubMed

    Vetrile, L A; Fomina, V G; Nevidimova, T I; Vetlugina, T P; Batukhtina, E I; Savochkina, D N; Zakharova, I A; Davydova, T V

    2015-01-01

    Blood serum from 129 patients with opium addiction at different stages of the disease and 63 donors (control group) was examined for the presence of autoantibodies to the exciting and inhibitory amino acids glutamate and GABA. It was shown enhanced production of autoantibodies to glutamate and GABA. Dependence of the level and frequency of detec- tion of autoantibodies to glutamate and GABA on the stage of the disease was revealed.

  11. [Autoantibodies to glutamate and GABA in opiate addiction].

    PubMed

    Vetrile, L A; Fomina, V G; Nevidimova, T I; Vetlugina, T P; Batukhtina, E I; Savochkina, D N; Zakharova, I A; Davydova, T V

    2015-01-01

    Blood serum from 129 patients with opium addiction at different stages of the disease and 63 donors (control group) was examined for the presence of autoantibodies to the exciting and inhibitory amino acids glutamate and GABA. It was shown enhanced production of autoantibodies to glutamate and GABA. Dependence of the level and frequency of detec- tion of autoantibodies to glutamate and GABA on the stage of the disease was revealed. PMID:26852594

  12. Relationship between Increase in Astrocytic GLT-1 Glutamate Transport and Late-LTP

    ERIC Educational Resources Information Center

    Pita-Almenar, Juan D.; Zou, Shengwei; Colbert, Costa M.; Eskin, Arnold

    2012-01-01

    Na[superscript +]-dependent high-affinity glutamate transporters have important roles in the maintenance of basal levels of glutamate and clearance of glutamate during synaptic transmission. Interestingly, several studies have shown that basal glutamate transport displays plasticity. Glutamate uptake increases in hippocampal slices during early…

  13. Extracellular glutamate alters mature osteoclast and osteoblast functions.

    PubMed

    Seidlitz, Eric P; Sharma, Mohit K; Singh, Gurmit

    2010-09-01

    Glutamatergic intercellular communication is involved in many aspects of metabolic homeostasis in normal bone. In bone metastasis, the balance between bone formation and degradation is disrupted. Although the responsible mechanisms are not clear, we have previously identified that cancer cell lines used in bone tumour models secrete glutamate, suggesting that tumour-derived glutamate may disrupt sensitive signalling systems in bone. This study examines the role of glutamate in mature osteoclastic bone resorption, osteoblast differentiation, and bone nodule formation. Glutamate was found to have no effect on the survival or activity of mature osteoclasts, although glutamate transporter inhibition and receptor blockade increased the number of bone resorption pits. Furthermore, transporter inhibition increased the area of resorbed bone while significantly decreasing the number of osteoclasts. Alkaline phosphatase activity and extracellular matrix mineralization were used as measurements of osteoblast differentiation. Glutamate significantly increased osteoblast differentiation and mineralization, but transport inhibitors had no effect. These studies support earlier findings suggesting that glutamate may be more important for osteoclastogenesis than for osteoclast proliferation or functions. Since glutamate is capable of changing the differentiation and activities of both osteoclast and osteoblast cell types in bone, it is reasonable to postulate that tumour-derived glutamate may impact bone homeostasis in bone metastasis.

  14. Glutamate and neurotrophic factors in neuronal plasticity and disease.

    PubMed

    Mattson, Mark P

    2008-11-01

    Glutamate's role as a neurotransmitter at synapses has been known for 40 years, but glutamate has since been shown to regulate neurogenesis, neurite outgrowth, synaptogenesis, and neuron survival in the developing and adult mammalian nervous system. Cell-surface glutamate receptors are coupled to Ca(2+) influx and release from endoplasmic reticulum stores, which causes rapid (kinase- and protease-mediated) and delayed (transcription-dependent) responses that change the structure and function of neurons. Neurotrophic factors and glutamate interact to regulate developmental and adult neuroplasticity. For example, glutamate stimulates the production of brain-derived neurotrophic factor (BDNF), which, in turn, modifies neuronal glutamate sensitivity, Ca(2+) homeostasis, and plasticity. Neurotrophic factors may modify glutamate signaling directly, by changing the expression of glutamate receptor subunits and Ca(2+)-regulating proteins, and also indirectly by inducing the production of antioxidant enzymes, energy-regulating proteins, and antiapoptotic Bcl-2 family members. Excessive activation of glutamate receptors, under conditions of oxidative and metabolic stress, may contribute to neuronal dysfunction and degeneration in diseases ranging from stroke and Alzheimer's disease to psychiatric disorders. By enhancing neurotrophic factor signaling, environmental factors such as exercise and dietary energy restriction, and chemicals such as antidepressants may optimize glutamatergic signaling and protect against neurological disorders.

  15. The Relevance of Group II Glutamate Receptors Expression to Anxiety.

    PubMed

    Ravid, Jonathan D; Mostofsky, David I

    2016-01-01

    The interface of receptor-mediated regulation of cellular signaling and neurological outputs remains an active field of investigation. The metabotropic G protein-coupled glutamate receptors, and in particular, the group II cyclic adenosine mono-phosphate (cAMP)-lowering metabotropic glutamate receptors 2 and 3 (mGlu2/3 glutamate receptors), have gained interest as therapeutic targets in different forms of neurological disorders. This review explores mGlu2/3 glutamate receptors expression, pharmacological activation, and signaling links to anxiety, as assessed in animal models and in clinical trials. PMID:27650988

  16. Emotional Learning and Glutamate: Translational Perspectives

    PubMed Central

    Gillespie, Charles F.; Ressler, Kerry J.

    2009-01-01

    Anxiety disorders are a common focus of clinical concern and certain forms of anxiety may be conceptualized as disorders of emotional learning. Behavior therapies effective in the treatment of anxiety are modeled on extinction training as a means of reducing pathological anxiety. The present understanding of human anxiety has been informed by preclinical research using rodent models to study the acquisition and extinction of fear. Glutamate appears to have a central role in both of these processes. The authors review this literature and discuss novel applications of D-cycloserine, a partial N-methyl-D-aspartate agonist, for the treatment of anxiety. PMID:16400246

  17. Connexin expression in human acute myeloid leukemia cells: Identification of patient subsets based on protein and global gene expression profiles

    PubMed Central

    REIKVAM, HÅKON; RYNINGEN, ANITA; SÆTERDAL, LARS RUNE; NEPSTAD, INA; FOSS, BRYNJAR; BRUSERUD, ØYSTEIN

    2015-01-01

    Bone marrow stromal cells support both normal and malignant hematopoiesis. Τhis support is mediated through the local cytokine network and by direct cell-cell interactions mediated via adhesion molecules and the formation of gap junctions by connexins. Previous studies on connexins in human acute myeloid leukemia (AML) have mainly focused on the investigation of leukemia cell lines. In the present study, we therefore investigated the expression of various connexins at the protein (i.e., cell surface expression) and mRNA level in primary human AML cells. The cell surface expression of the connexins, Cx26, Cx32, Cx37, Cx43 and Cx45, varied considerably between patients, and detectable levels were observed only for subsets of patients. On the whole, Cx43 and Cx45 showed the highest cell surface expression. Connexin expression was dependent on AML cell differentiation, but showed no association with cytogenetic abnormalities or mutations of the fms-related tyrosine kinase 3 (FLT3) or nucleophosmin (NPM)‑1 genes. By contrast, only Cx45 showed a significant variation between patients at the mRNA level. A high Cx45 expression was associated with the altered regulation of the mitogen-activated protein kinase (MAPK) pathway and the release of pro-inflammatory cytokines [interleukin (IL)-17, tumor necrosis factor (TNF), interferon-γ], whereas a low Cx45 expression was associated with the altered regulation of protein functions (i.e., ligase activity, protein folding and catabolism). There was no significant correlation observed between the connexin mRNA and protein levels. Thus, differences in connexin expression can be used to subclassify AML patients. Differences in connexin cell surface expression profiles are not reflected at the mRNA level and have to be directly examined, whereas variations in Cx45 mRNA expression are associated with differences in cell signaling and the regulation of protein functions. PMID:25529637

  18. Connexin expression in human acute myeloid leukemia cells: identification of patient subsets based on protein and global gene expression profiles.

    PubMed

    Reikvam, Håkon; Ryningen, Anita; Sæterdal, Lars Rune; Nepstad, Ina; Foss, Brynjar; Bruserud, Øystein

    2015-03-01

    Bone marrow stromal cells support both normal and malignant hematopoiesis. Τhis support is mediated through the local cytokine network and by direct cell‑cell interactions mediated via adhesion molecules and the formation of gap junctions by connexins. Previous studies on connexins in human acute myeloid leukemia (AML) have mainly focused on the investigation of leukemia cell lines. In the present study, we therefore investigated the expression of various connexins at the protein (i.e., cell surface expression) and mRNA level in primary human AML cells. The cell surface expression of the connexins, Cx26, Cx32, Cx37, Cx43 and Cx45, varied considerably between patients, and detectable levels were observed only for subsets of patients. On the whole, Cx43 and Cx45 showed the highest cell surface expression. Connexin expression was dependent on AML cell differentiation, but showed no association with cytogenetic abnormalities or mutations of the fms-related tyrosine kinase 3 (FLT3) or nucleophosmin (NPM)‑1 genes. By contrast, only Cx45 showed a significant variation between patients at the mRNA level. A high Cx45 expression was associated with the altered regulation of the mitogen‑activated protein kinase (MAPK) pathway and the release of pro-inflammatory cytokines [interleukin (IL)‑17, tumor necrosis factor (TNF), interferon‑γ], whereas a low Cx45 expression was associated with the altered regulation of protein functions (i.e., ligase activity, protein folding and catabolism). There was no significant correlation observed between the connexin mRNA and protein levels. Thus, differences in connexin expression can be used to subclassify AML patients. Differences in connexin cell surface expression profiles are not reflected at the mRNA level and have to be directly examined, whereas variations in Cx45 mRNA expression are associated with differences in cell signaling and the regulation of protein functions.

  19. Mesenchymal stem cells protect CNS neurons against glutamate excitotoxicity by inhibiting glutamate receptor expression and function.

    PubMed

    Voulgari-Kokota, A; Fairless, R; Karamita, M; Kyrargyri, V; Tseveleki, V; Evangelidou, M; Delorme, B; Charbord, P; Diem, R; Probert, L

    2012-07-01

    Mesenchymal stem cells (MSC) promote functional recovery in experimental models of central nervous system (CNS) pathology and are currently being tested in clinical trials for stroke, multiple sclerosis and CNS injury. Their beneficial effects are attributed to the activation of endogenous CNS protection and repair processes as well as immune regulation but their mechanisms of action are poorly understood. Here we investigated the neuroprotective effects of mouse MSC in rodent MSC-neuron co-cultures and mice using models of glutamate excitotoxicity. A 24h pre-culture of mouse primary cortical neurons with MSC protected them against glutamate (NMDA) receptor-induced death and conditioned medium from MSC (MSC CM) was sufficient for this effect. Protection by MSC CM was associated with reduced mRNA levels of genes encoding NMDA receptor subunits, and increased levels for genes associated with non-neuronal and stem cell types, as shown by RT-PCR and cDNA microarray analyses. Changes in gene expression were not associated with alterations in cell lineage representation within the cultures. Further, MSC CM-mediated neuroprotection in rat retinal ganglion cells was associated with reduced glutamate-induced calcium influx. The adoptive transfer of EGFP(+)MSC in a mouse kainic acid epilepsy model also provided neuroprotection against glutamate excitotoxicity in vivo, as shown by reduced neuron damage and glial cell activation in the hippocampus. These results show that MSC mediate direct neuroprotection by reducing neuronal sensitivity to glutamate receptor ligands and altering gene expression, and suggest a link between the therapeutic effects of MSC and the activation of cell plasticity in the damaged CNS. PMID:22561409

  20. Rat odontoblasts may use glutamate to signal dentin injury.

    PubMed

    Cho, Yi Sul; Ryu, Chang Hyun; Won, Jong Hwa; Vang, Hue; Oh, Seog Bae; Ro, Jin Young; Bae, Yong Chul

    2016-10-29

    Accumulating evidence indicates that odontoblasts act as sensor cells, capable of triggering action potentials in adjacent pulpal nociceptive axons, suggesting a paracrine signaling via a currently unknown mediator. Since glutamate can mediate signaling by non-neuronal cells, and peripheral axons may express glutamate receptors (GluR), we hypothesized that the expression of high levels of glutamate, and of sensory receptors in odontoblasts, combined with an expression of GluR in adjacent pulpal axons, is the morphological basis for odontoblastic sensory signaling. To test this hypothesis, we investigated the expression of glutamate, the thermo- and mechanosensitive ion channels transient receptor potential vanilloid 1 (TRPV1), transient receptor potential ankyrin 1 (TRPA1), and TWIK-1-related K+channel (TREK-1), and the glutamate receptor mGluR5, in a normal rat dental pulp, and following dentin injury. We also examined the glutamate release from odontoblast in cell culture. Odontoblasts were enriched with glutamate, at the level as high as in adjacent pulpal axons, and showed immunoreactivity for TRPV1, TRPA1, and TREK-1. Pulpal sensory axons adjacent to odontoblasts expressed mGluR5. Both the levels of glutamate in odontoblasts, and the expression of mGluR5 in nearby axons, were upregulated following dentin injury. The extracellular glutamate concentration was increased significantly after treating of odontoblast cell line with calcium permeable ionophore, suggesting glutamate release from odontoblasts. These findings lend morphological support to the hypothesis that odontoblasts contain glutamate as a potential neuroactive substance that may activate adjacent pulpal axons, and thus contribute to dental pain and hypersensitivity.

  1. Rat odontoblasts may use glutamate to signal dentin injury.

    PubMed

    Cho, Yi Sul; Ryu, Chang Hyun; Won, Jong Hwa; Vang, Hue; Oh, Seog Bae; Ro, Jin Young; Bae, Yong Chul

    2016-10-29

    Accumulating evidence indicates that odontoblasts act as sensor cells, capable of triggering action potentials in adjacent pulpal nociceptive axons, suggesting a paracrine signaling via a currently unknown mediator. Since glutamate can mediate signaling by non-neuronal cells, and peripheral axons may express glutamate receptors (GluR), we hypothesized that the expression of high levels of glutamate, and of sensory receptors in odontoblasts, combined with an expression of GluR in adjacent pulpal axons, is the morphological basis for odontoblastic sensory signaling. To test this hypothesis, we investigated the expression of glutamate, the thermo- and mechanosensitive ion channels transient receptor potential vanilloid 1 (TRPV1), transient receptor potential ankyrin 1 (TRPA1), and TWIK-1-related K+channel (TREK-1), and the glutamate receptor mGluR5, in a normal rat dental pulp, and following dentin injury. We also examined the glutamate release from odontoblast in cell culture. Odontoblasts were enriched with glutamate, at the level as high as in adjacent pulpal axons, and showed immunoreactivity for TRPV1, TRPA1, and TREK-1. Pulpal sensory axons adjacent to odontoblasts expressed mGluR5. Both the levels of glutamate in odontoblasts, and the expression of mGluR5 in nearby axons, were upregulated following dentin injury. The extracellular glutamate concentration was increased significantly after treating of odontoblast cell line with calcium permeable ionophore, suggesting glutamate release from odontoblasts. These findings lend morphological support to the hypothesis that odontoblasts contain glutamate as a potential neuroactive substance that may activate adjacent pulpal axons, and thus contribute to dental pain and hypersensitivity. PMID:27555550

  2. Effect of biotin on transcription levels of key enzymes and glutamate efflux in glutamate fermentation by Corynebacterium glutamicum.

    PubMed

    Cao, Yan; Duan, Zuoying; Shi, Zhongping

    2014-02-01

    Biotin is an important factor affecting the performance of glutamate fermentation by biotin auxotrophic Corynebacterium glutamicum and glutamate is over-produced only when initial biotin content is controlled at suitable levels or initial biotin is excessive but with Tween 40 addition during fermentation. The transcription levels of key enzymes at pyruvate, isocitrate and α-ketoglutarate metabolic nodes, as well as transport protein (TP) of glutamate were investigated under the conditions of varied biotin contents and Tween 40 supplementation. When biotin was insufficient, the genes encoding key enzymes and TP were down-regulated in the early production phase, in particular, the transcription level of isocitrate dehydrogenase (ICDH) which was only 2% of that of control. Although the cells' morphology transformation and TP level were not affected, low transcription level of ICDH led to lower final glutamate concentration (64 g/L). When biotin was excessive, the transcription levels of key enzymes were at comparable levels as those of control with ICDH as an exception, which was only 3-22% of control level throughout production phase. In this case, little intracellular glutamate accumulation (1.5 mg/g DCW) and impermeable membrane resulted in non glutamate secretion into broth, even though the quantity of TP was more than 10-folds of control level. Addition of Tween 40 when biotin was excessive stimulated the expression of all key enzymes and TP, intracellular glutamate content was much higher (10-12 mg/g DCW), and final glutamate concentration reached control level (75-80 g/L). Hence, the membrane alteration and TP were indispensable in glutamate secretion. Biotin and Tween 40 influenced the expression level of ICDH and glutamate efflux, thereby influencing glutamate production.

  3. Postsynaptic activation at the squid giant synapse by photolytic release of L-glutamate from a 'caged' L-glutamate.

    PubMed Central

    Corrie, J E; DeSantis, A; Katayama, Y; Khodakhah, K; Messenger, J B; Ogden, D C; Trentham, D R

    1993-01-01

    1. Pharmacological evidence suggests L-glutamate is a strong candidate as a transmitter at the giant synapse of the squid. Postsynaptic activation at the giant synapse cannot be effected by conventional application of putative neurotransmitters by iontophoresis or perfusion, apparently because the complex structure of the synapse prevents a sufficiently rapid change in concentration at the postsynaptic membrane. Flash photolytic release of L-glutamate from a pharmacologically inert 'caged' L-glutamate pre-equilibrated in the stellate ganglion of Alloteuthis or Loligo was used to determine whether L-glutamate can produce postsynaptic activation when released rapidly in the synaptic clefts. 2. The preparation, reaction mechanism and properties of the caged L-glutamate, N-1-(2-nitrophenyl)ethoxycarbonyl-L-glutamate, are described. The product quantum yield on photolysis was 0.65 (+/- 0.05). On flash photolysis glutamate release followed a single exponential time-course in the pH range 5.5-7.8. The rate constant was proportional to [H+] and was 93 s-1 at pH 5.5 and 16 degrees C in artificial sea water (ionic strength, I = 0.68 M). 3. At pH 7.8 flash photolysis of caged glutamate pre-equilibrated in the synapse caused only a slow depolarization. A second photolytic release of L-glutamate or transsynaptic activation produced no further depolarization, suggesting desensitization and inactivation of postsynaptic mechanisms by the initial pulse of L-glutamate. 4. Synaptic transmission in the giant synapse was normal at pH 5.5. Flash photolysis at pH 5.5 caused rapid production of L-glutamate within the synaptic cleft and a fast postsynaptic depolarization which generated postsynaptic action potentials.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7901400

  4. On the regulative role of the glutamate receptor in mitochondria.

    PubMed

    Selin, Alexey A; Lobysheva, Natalia V; Nesterov, Semen V; Skorobogatova, Yulia A; Byvshev, Ivan M; Pavlik, Lyubov L; Mikheeva, Irina B; Moshkov, Dmitry A; Yaguzhinsky, Lev S; Nartsissov, Yaroslav R

    2016-05-01

    The purpose of this work was to study the regulative role of the glutamate receptor found earlier in the brain mitochondria. In the present work a glutamate-dependent signaling system with similar features was detected in mitochondria of the heart. The glutamate-dependent signaling system in the heart mitochondria was shown to be suppressed by γ-aminobutyric acid (GABA). The GABA receptor presence in the heart mitochondria was shown by golding with the use of antibodies to α- and β-subunits of the receptor. The activity of glutamate receptor was assessed according to the rate of synthesis of hydrogen peroxide. The glutamate receptor in mitochondria could be activated only under conditions of hypoxic stress, which in model experiments was imitated by blocking Complex I by rotenone or fatty acids. The glutamate signal in mitochondria was shown to be calcium- and potential-dependent and the activation of the glutamate cascade was shown to be accompanied by production of hydrogen peroxide. It was discovered that H2O2 synthesis involves two complexes of the mitochondrial electron transfer system - succinate dehydrogenase (SDH) and fatty acid dehydrogenase (ETF:QO). Thus, functions of the glutamate signaling system are associated with the system of respiration-glycolysis switching (the Pasteur-Crabtree) under conditions of hypoxia. PMID:26812870

  5. Mammalian folylpoly-. gamma. -glutamate synthetase. 3. Specificity for folate analogues

    SciTech Connect

    George, S.; Cichowicz, D.J.; Shane, B.

    1987-01-27

    A variety of folate analogues were synthesized to explore the specificity of the folate binding site of hog liver folypolyglutamate synthetase and the requirements for catalysis. Modifications of the internal and terminal glutamate moieties of folate cause large drops in on rates and/or affinity for the protein. The only exceptions are glutamine, homocysteate, and ornithine analogues, indicating a less stringent specificity around the delta-carbon of glutamate. It is proposed that initial folate binding to the enzyme involves low-affinity interactions at a pterin and a glutamate site and that the first glutamate bound is the internal residue adjacent to the benzoyl group. Processive movement of the polyglutamate chain through the glutamate site and a possible conformational change in the protein when the terminal residue is bound would result in tight binding and would position the ..gamma..-carboxyl of the terminal glutamate in the correct position for catalysis. The 4-amino substitution of folate increases the on rate for monoglutamate derivatives but severely impairs catalysis with diglutamate derivatives. Pteroylornithine derivatives are the first potent and specific inhibitors of folylpolyglutamate synthetase to be identified and may act as analogues of reaction intermediates. Other folate derivatives with tetrahedral chemistry replacing the peptide bond, such as pteroyl-..gamma..-glutamyl-(psi,CH/sub 2/-NH)-glutamate, retain affinity for the protein but are considerably less effective inhibitors than the ornithine derivatives. Enzyme activity was assayed using (/sup 14/C)glutamate.

  6. 78 FR 76321 - Monosodium Glutamate From China and Indonesia

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-12-17

    ... Register of September 20, 2013 (78 FR 57881). The conference was held in Washington, DC, on October 23... COMMISSION Monosodium Glutamate From China and Indonesia Determinations On the basis of the record \\1... injured by reason of imports from China and Indonesia of monosodium glutamate, provided for in...

  7. Neuronal vs glial glutamate uptake: Resolving the conundrum.

    PubMed

    Danbolt, N C; Furness, D N; Zhou, Y

    2016-09-01

    Neither normal brain function nor the pathological processes involved in neurological diseases can be adequately understood without knowledge of the release, uptake and metabolism of glutamate. The reason for this is that glutamate (a) is the most abundant amino acid in the brain, (b) is at the cross-roads between several metabolic pathways, and (c) serves as the major excitatory neurotransmitter. In fact most brain cells express glutamate receptors and are thereby influenced by extracellular glutamate. In agreement, brain cells have powerful uptake systems that constantly remove glutamate from the extracellular fluid and thereby limit receptor activation. It has been clear since the 1970s that both astrocytes and neurons express glutamate transporters. However the relative contribution of neuronal and glial transporters to the total glutamate uptake activity, however, as well as their functional importance, has been hotly debated ever since. The present short review provides (a) an overview of what we know about neuronal glutamate uptake as well as an historical description of how we got there, and (b) a hypothesis reconciling apparently contradicting observations thereby possibly resolving the paradox. PMID:27235987

  8. 21 CFR 522.1125 - Hemoglobin glutamer-200 (bovine).

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Hemoglobin glutamer-200 (bovine). 522.1125 Section... § 522.1125 Hemoglobin glutamer-200 (bovine). (a) Specifications. Each 125 milliliter bag contains 13 grams per deciliter of polymerized hemoglobin of bovine origin in modified Lactated Ringer's...

  9. 21 CFR 522.1125 - Hemoglobin glutamer-200 (bovine).

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Hemoglobin glutamer-200 (bovine). 522.1125 Section... § 522.1125 Hemoglobin glutamer-200 (bovine). (a) Specifications. Each 125 milliliter bag contains 13 grams per deciliter of polymerized hemoglobin of bovine origin in modified Lactated Ringer's...

  10. 21 CFR 522.1125 - Hemoglobin glutamer-200 (bovine).

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Hemoglobin glutamer-200 (bovine). 522.1125 Section... § 522.1125 Hemoglobin glutamer-200 (bovine). (a) Specifications. Each 125 milliliter bag contains 13 grams per deciliter of polymerized hemoglobin of bovine origin in modified Lactated Ringer's...

  11. A review of glutamate's role in traumatic brain injury mechanisms

    NASA Astrophysics Data System (ADS)

    Good, Cameron H.

    2013-05-01

    Glutamate is the primary excitatory neurotransmitter used by the central nervous system (CNS) for synaptic communication, and its extracellular concentration is tightly regulated by glutamate transporters located on nearby astrocytes. Both animal models and human clinical studies have demonstrated elevated glutamate levels immediately following a traumatic brain event, with the duration and severity of the rise corresponding to prognosis. This rise in extracellular glutamate likely results from a combination of excessive neurotransmitter release from damaged neurons and down regulation of uptake mechanisms in local astrocytes. The immediate results of a traumatic event can lead to necrotic tissue in severely injured regions, while prolonged increases in excitatory transmission can cause secondary excitotoxic injury through activation of delayed apoptotic pathways. Initial TBI animal studies utilized a variety of broad glutamate receptor antagonists to successfully combat secondary injury mechanisms, but unfortunately this same strategy has proven inconclusive in subsequent human trials due to deleterious side effects and heterogeneity of injuries. More recent treatment strategies have utilized specific glutamate receptor subunit antagonists in an effort to minimize side effects and have shown promising results. Future challenges will be detecting the concentration and kinetics of the glutamate rise following injury, determining which patient populations could benefit from antagonist treatment based on their extracellular glutamate concentrations and when drugs should be administered to maximize efficacy.

  12. Modulation of intestinal L-glutamate transport by luminal leptin.

    PubMed

    Fanjul, Carmen; Barrenetxe, Jaione; Lostao, María Pilar; Ducroc, Robert

    2015-06-01

    Leptin is secreted into the digestive tract and contributes to the absorption of dietary molecules by regulating transporters activity. Here, we studied the effect of luminal leptin on the intestinal transport of L-glutamate, an important component of human diet. We examined the effect of leptin on L-glutamate uptake in rat intestine in vitro measuring glutamate-induced short-circuit current (Isc) in Ussing chambers and L-[(3)H (U)]-glutamate uptake in jejunal everted rings. Glutamate-induced Isc was only observed in Na(+)-free conditions. This Isc was concentration (1-60 mmol L(-1)) and pH dependent. Luminal leptin increased glutamate Isc (∼100 %). Dose-response curve showed a biphasic pattern, with maximal stimulations observed at 10(-13) and 10(-10) mmol L(-1), that were sensitive to leptin receptor antagonist. In everted rings, two glutamate transport mechanisms were distinguished: a Na(+)-dependent, H(+)-independent, that was inhibited by leptin (∼20 %), and a Na(+)-independent but H(+)-dependent, that was enhanced by leptin (∼20 %), in line with data obtained in Ussing chambers. Altogether, these data reveal original non-monotonic effect of luminal leptin in the intestine and demonstrate a new role for this hormone in the modulation of L-glutamate transport, showing that luminal active gut peptides can influence absorption of amino acids.

  13. Glial glutamate transporters: new actors in brain signaling.

    PubMed

    López-Bayghen, Esther; Ortega, Arturo

    2011-10-01

    Glutamate, the main excitatory amino acid in the vertebrate brain, is critically involved in most of the physiological functions of the central nervous system. It has traditionally been assumed that glutamate triggers a wide array of signaling cascades through the activation of specific membrane receptors. The extracellular levels are tightly regulated to prevent neurotoxic insults. Electrogenic Na(+)-dependent glial glutamate transporters remove the bulk of the neurotransmitter from the synaptic cleft. An exquisitely ordered coupling between glutamatergic neurons and surrounding glia cells is fundamental for excitatory transmission. The glutamate/glutamine and astrocyte/neuron lactate shuttles provide the biochemical framework of this compulsory association. In this context, recent advances show that glial glutamate transporters act as signal transducers that regulate the expression of proteins involved in their compartmentalization with neurons in the so-called tripartite synapse.

  14. Cortical neurons exposed to glutamate rapidly leak preloaded chromium 51

    SciTech Connect

    Maulucci-Gedde, M.; Choi, D.W.

    1987-05-01

    The acute toxic effects of excess glutamate exposure on cortical neurons in culture was followed using a novel adaptation of the /sup 51/Cr efflux assay. Although the acute, sodium-dependent phase of glutamate neurotoxicity may contribute to several acute disease settings, including sustained seizures and stroke, functional aspects of the phenomenon have not been previously studied. We report here that the earliest morphologic sign of glutamate neurotoxicity, neuronal swelling, is accompanied by a large efflux of complexed /sup 51/Cr from preloaded neurons in the first hour after exposure, and that this efflux is detectable as early as 15 min after the onset of glutamate exposure. We suggest that this pathological burst of /sup 51/Cr may result from glutamate-induced leakiness of neuronal cell membranes.

  15. A novel glutamate dehydrogenase from bovine brain: purification and characterization.

    PubMed

    Lee, J; Kim, S W; Cho, S W

    1995-08-01

    A soluble form of novel glutamate dehydrogenase has been purified from bovine brain. The preparation was homogeneous on sodium dodecyl sulfate-polyacrylamide gel electrophoresis and composed of six identical subunits having a subunit size of 57,500 Da. The biochemical properties of glutamate dehydrogenase such as N-terminal amino acids sequences, kinetic parameters, amino acids analysis, and optimum pH were examined in both reductive amination of alpha-ketoglutarate and oxidative deamination of glutamate. N-terminal amino acid sequences of the bovine brain enzyme showed the significant differences in the first 5 amino acids compared to other glutamate dehydrogenases from various sources. These results indicate that glutamate dehydrogenase isolated from bovine brain is a novel polypeptide.

  16. Influence of glutamic acid enantiomers on C-mineralization.

    PubMed

    Formánek, Pavel; Vranová, Valerie; Lojková, Lea

    2015-02-01

    Seasonal dynamics in the mineralization of glutamic acid enantiomers in soils from selected ecosystems was determined and subjected to a range of treatments: ambient x elevated CO2 level and meadow x dense x thinned forest environment. Mineralization of glutamic acid was determined by incubation of the soil with 2 mg L- or D-glutamic acid g(-1) of dry soil to induce the maximum respiration rate. Mineralization of glutamic acid enantiomers in soils fluctuates over the course of a vegetation season, following a similar trend across a range of ecosystems. Mineralization is affected by environmental changes and management practices, including elevated CO2 level and thinning intensity. L-glutamic acid metabolism is more dependent on soil type as compared to metabolism of its D-enantiomer. The results support the hypothesis that the slower rate of D- compared to L- amino acid mineralization is due to different roles in anabolism and catabolism of the soil microbial community.

  17. Protection of the glutamate pool concentration in enteric bacteria.

    PubMed

    Yan, Dalai

    2007-05-29

    The central nitrogen metabolic circuit in enteric bacteria consists of three enzymes: glutamine synthetase, glutamate synthase (GOGAT), and glutamate dehydrogenase (GDH). With the carbon skeleton provided by 2-oxoglutarate, ammonia/ammonium (NH(4)(+)) is assimilated into two central nitrogen intermediates, glutamate and glutamine. Although both serve as nitrogen donors for all biosynthetic needs, glutamate and glutamine play different roles. Internal glutamine serves as a sensor of external nitrogen availability, and its pool concentration decreases upon nitrogen limitation. A high glutamate pool concentration is required to maintain the internal K(+) pool. The configuration of high glutamate and low glutamine pools was disrupted in GOGAT(-) mutants under low NH(4)(+) conditions: the glutamate pool was low, the difference between glutamate and glutamine was diminished, and growth was defective. When a GOGAT(-) mutant was cultured in an NH(4)(+)-limited chemostat, two sequential spontaneous mutations occurred. Each resulted in a suppressor mutant that outgrew its predecessor in the chemostat. The first suppressor overexpressed GDH, and the second also had a partially impaired glutamine synthetase. The result was a triple mutant in which NH(4)(+) was assimilated by two enzymes instead of the normal three and yet glutamate and glutamine pools and growth were essentially normal. The results indicate preference for the usual ratio of glutamate and glutamine and the resilient and compensatory nature of the circuit on pool control. Analysis of other suppressor mutants selected on solid medium suggests that increased GDH expression is the key for rescue of the growth defect of GOGAT(-) mutants under low NH(4)(+) conditions.

  18. Connexins in skeletal muscle development and disease.

    PubMed

    Merrifield, Peter A; Laird, Dale W

    2016-02-01

    Gap junctions consist of clusters of intercellular channels composed of connexins that connect adjacent cells and allow the exchange of small molecules. While the 21 member multi-gene family of connexins are ubiquitously found in humans, only Cx39, Cx40, Cx43 and Cx45 have been documented in developing myoblasts and injured adult skeletal muscle while healthy adult skeletal muscle is devoid of connexins. The use of gap junctional blockers and cultured myoblast cell lines have suggested that these connexins play a critical role in myotube formation and muscle regeneration. More recent genetically-modified mouse models where Cx43 function is greatly compromized or ablated have further supported a role for Cx43 in regulating skeletal muscle development. In the last decade, we have become aware of a cohort of patients that have a development disorder known as oculodentodigital dysplasia (ODDD). These patients harbor either gain or loss of Cx43 function gene mutations that result in many organ anomalies raising questions as to whether they suffer from defects in skeletal muscle formation or regeneration upon injury. Interesting, some ODDD patients report muscle weakness and loss of limb control but it is not clear if this is neurogenic or myogenic in origin. This review will focus on the role connexins play in muscle development and repair and discuss the impact of Cx43 mutants on muscle function. PMID:26688333

  19. Serum Glutamic-Oxaloacetic Transaminase (GOT) and Glutamic-Pyruvic Transaminase (GPT) Levels in Children and Adolescents with Intellectual Disabilities

    ERIC Educational Resources Information Center

    Lin, Jin-Ding; Lin, Pei-Ying; Chen, Li-Mei; Fang, Wen-Hui; Lin, Lan-Ping; Loh, Ching-Hui

    2010-01-01

    The elevated serum glutamic-oxaloacetic transaminase (GOT) and glutamic-pyruvic transaminase (GPT) rate among people with intellectual disabilities (ID) is unknown and have not been sufficiently studies. The present paper aims to provide the profile of GOT and GPT, and their associated relationship with other biochemical levels of children or…

  20. Prefrontal changes in the glutamate-glutamine cycle and neuronal/glial glutamate transporters in depression with and without suicide.

    PubMed

    Zhao, J; Verwer, R W H; van Wamelen, D J; Qi, X-R; Gao, S-F; Lucassen, P J; Swaab, D F

    2016-11-01

    There are indications for changes in glutamate metabolism in relation to depression or suicide. The glutamate-glutamine cycle and neuronal/glial glutamate transporters mediate the uptake of the glutamate and glutamine. The expression of various components of the glutamate-glutamine cycle and the neuronal/glial glutamate transporters was determined by qPCR in postmortem prefrontal cortex. The anterior cingulate cortex (ACC) and the dorsolateral prefrontal cortex (DLPFC) were selected from young MDD patients who had committed suicide (MDD-S; n = 17), from MDD patients who died of non-suicide related causes (MDD-NS; n = 7) and from matched control subjects (n = 12). We also compared elderly depressed patients who had not committed suicide (n = 14) with matched control subjects (n = 22). We found that neuronal located components (EAAT3, EAAT4, ASCT1, SNAT1, SNAT2) of the glutamate-glutamine cycle were increased in the ACC while the astroglia located components (EAAT1, EAAT2, GLUL) were decreased in the DLPFC of MDD-S patients. In contrast, most of the components in the cycle were increased in the DLPFC of MDD-NS patients. In conclusion, the glutamate-glutamine cycle - and thus glutamine transmission - is differentially affected in depressed suicide patients and depressed non-suicide patients in an area specific way.

  1. Elevated venous glutamate levels in (pre)catabolic conditions result at least partly from a decreased glutamate transport activity.

    PubMed

    Hack, V; Stütz, O; Kinscherf, R; Schykowski, M; Kellerer, M; Holm, E; Dröge, W

    1996-06-01

    Abnormally high postabsorptive venous plasma glutamate levels have been reported for several diseases that are associated with a loss of body cell mass including cancer, human/simian immunodeficiency virus infection, and amyotrophic lateral sclerosis. Studies on exchange rates in well-nourished cancer patients now show that high venous plasma glutamate levels may serve as a bona fide indicator for a decreased uptake of glutamate by the peripheral muscle tissue in the postabsorptive period and may be indicative for a precachectic state. High glutamate levels are also moderately correlated with a decreased uptake of glucose and ketone bodies. Relatively high venous glutamate levels have also been found in non-insulin-dependent diabetes mellitus and to some extent also in the cubital vein of normal elderly subjects, i.e., in conditions commonly associated with a decreased glucose tolerance and progressive loss of body cell mass.

  2. Impact of obesity on 7,12-dimethylbenz[a]anthracene-induced altered ovarian connexin gap junction proteins in female mice

    SciTech Connect

    Ganesan, Shanthi Nteeba, Jackson Keating, Aileen F.

    2015-01-01

    The ovarian gap junction proteins alpha 4 (GJA4 or connexin 37; CX37), alpha 1 (GJA1 or connexin 43; CX43) and gamma 1 (GJC1 or connexin 45; CX45) are involved in cell communication and folliculogenesis. 7,12-dimethylbenz[a]anthracene (DMBA) alters Cx37 and Cx43 expression in cultured neonatal rat ovaries. Additionally, obesity has an additive effect on DMBA-induced ovarian cell death and follicle depletion, thus, we investigated in vivo impacts of obesity and DMBA on CX protein levels. Ovaries were collected from lean and obese mice aged 6, 12, 18, or 24 wks. A subset of 18 wk old mice (lean and obese) were dosed with sesame oil or DMBA (1 mg/kg; ip) for 14 days and ovaries collected 3 days thereafter. Cx43 and Cx45 mRNA and protein levels decreased (P < 0.05) after 18 wks while Cx37 mRNA and protein levels decreased (P < 0.05) after 24 wks in obese ovaries. Cx37 mRNA and antral follicle protein staining intensity were reduced (P < 0.05) by obesity while total CX37 protein was reduced (P < 0.05) in DMBA exposed obese ovaries. Cx43 mRNA and total protein levels were decreased (P < 0.05) by DMBA in both lean and obese ovaries while basal protein staining intensity was reduced (P < 0.05) in obese controls. Cx45 mRNA, total protein and protein staining intensity level were decreased (P < 0.05) by obesity. These data support that obesity temporally alters gap junction protein expression and that DMBA-induced ovotoxicity may involve reduced gap junction protein function. - Highlights: • Ovarian gap junction proteins are affected by ovarian aging and obesity. • DMBA exposure negatively impacts gap junction proteins. • Altered gap junction proteins may contribute to infertility.

  3. Impact of obesity on 7,12-dimethylbenz[a]anthracene-induced altered ovarian connexin gap junction proteins in female mice

    PubMed Central

    Ganesan, Shanthi; Nteeba, Jackson; Keating, Aileen F.

    2014-01-01

    The ovarian gap junction proteins alpha 4 (GJA4 or connexin 37; CX37), alpha 1 (GJA1 or connexin 43; CX43) and gamma 1 (GJC1 or connexin 45; CX45) are involved in cell communication and folliculogenesis. 7,12-dimethylbenz[a]anthracene (DMBA) alters Cx37 and Cx43 expression in cultured neonatal rat ovaries. Additionally, obesity has an additive effect on DMBA-induced ovarian cell death and follicle depletion, thus, we investigated in vivo impacts of obesity and DMBA on CX protein levels. Ovaries were collected from lean and obese mice aged 6, 12, 18, or 24 wks. A subset of 18 wk old mice (lean and obese) were dosed with sesame oil or DMBA (1mg/kg; ip) for 14 days and ovaries collected 3 days thereafter. Cx43 and Cx45 mRNA and protein levels decreased (P < 0.05) after 18 wks while Cx37 mRNA and protein levels decreased (P < 0.05) after 24 wks in obese ovaries. Cx37 mRNA and antral follicle protein staining intensity were reduced (P < 0.05) by obesity while total CX37 protein was reduced (P < 0.05) in DMBA exposed obese ovaries. Cx43 mRNA and total protein levels were decreased (P < 0.05) by DMBA in both lean and obese ovaries while basal protein staining intensity was reduced (P < 0.05) in obese controls. Cx45 mRNA, total protein and protein staining intensity level were decreased (P < 0.05) by obesity. These data support that obesity temporally alters gap junction protein expression and that DMBA-induced ovotoxicity may involve reduced gap junction protein function. PMID:25447408

  4. Translating Glutamate: From Pathophysiology to Treatment

    PubMed Central

    Javitt, Daniel C.; Schoepp, Darryle; Kalivas, Peter W.; Volkow, Nora D.; Zarate, Carlos; Merchant, Kalpana; Bear, Mark F.; Umbricht, Daniel; Hajos, Mihaly; Potter, William Z.; Lee, Chi-Ming

    2012-01-01

    The neurotransmitter glutamate is the primary excitatory neurotransmitter in mammalian brain and is responsible for most corticocortical and corticofugal neurotransmission. Disturbances in glutamatergic function have been implicated in the pathophysiology of several neuropsychiatric disorders—including schizophrenia, drug abuse and addiction, autism, and depression—that were until recently poorly understood. Nevertheless, improvements in basic information regarding these disorders have yet to translate into Food and Drug Administration–approved treatments. Barriers to translation include the need not only for improved compounds but also for improved biomarkers sensitive to both structural and functional target engagement and for improved translational models. Overcoming these barriers will require unique collaborative arrangements between pharma, government, and academia. Here, we review a recent Institute of Medicine–sponsored meeting, highlighting advances in glutamatergic theories of neuropsychiatric illness as well as remaining barriers to treatment development. PMID:21957170

  5. Untangling the glutamate dehydrogenase allosteric nightmare.

    PubMed

    Smith, Thomas J; Stanley, Charles A

    2008-11-01

    Glutamate dehydrogenase (GDH) is found in all living organisms, but only animal GDH is regulated by a large repertoire of metabolites. More than 50 years of research to better understand the mechanism and role of this allosteric network has been frustrated by its sheer complexity. However, recent studies have begun to tease out how and why this complex behavior evolved. Much of GDH regulation probably occurs by controlling a complex ballet of motion necessary for catalytic turnover and has evolved concomitantly with a long antenna-like feature of the structure of the enzyme. Ciliates, the 'missing link' in GDH evolution, might have created the antenna to accommodate changing organelle functions and was refined in humans to, at least in part, link amino acid catabolism with insulin secretion.

  6. Poly(γ-glutamic acid), coagulation? Anticoagulation?

    PubMed

    Xu, Tingting; Peng, Fang; Zhang, Tao; Chi, Bo; Xu, Hong; Mao, Chun; Feng, Shuaihui

    2016-11-01

    Poly(γ-glutamic acid) (γ-PGA) powder was usually used as hemostatic agent because of its excellent physical properties of water-absorption and water-locking. However, if γ-PGA absorbs enough water, how about its blood compatibility? Here, the other side of the coin was investigated. The anticoagulant properties of γ-PGA were characterized by in vitro coagulation tests, hemolytic assay, platelet adhesion, and platelet activation. Moreover, cytotoxicity experiments of γ-PGA were also carried out by MTT assay. Results indicated that the sufficient water-absorbed γ-PGA has good anticoagulant property and non-cytotoxicity. It means γ-PGA has good anticoagulant property, non-cytotoxicity. If γ-PGA has absorbed enough water, it can be used as an anticoagulation biomaterial. With double effects (coagulation and anticoagulation), the γ-PGA with desirable bioproperties can be readily tailored to cater to various biomedical applications. PMID:27545694

  7. The cystine/glutamate antiporter system xc− drives breast tumor cell glutamate release and cancer-induced bone pain

    PubMed Central

    Slosky, Lauren M.; BassiriRad, Neemah M.; Symons, Ashley M.; Thompson, Michelle; Doyle, Timothy; Forte, Brittany L.; Staatz, William D.; Bui, Lynn; Neumann, William L.; Mantyh, Patrick W.; Salvemini, Daniela; Largent-Milnes, Tally M.; Vanderah, Todd W.

    2016-01-01

    Abstract Bone is one of the leading sites of metastasis for frequently diagnosed malignancies, including those arising in the breast, prostate and lung. Although these cancers develop unnoticed and are painless in their primary sites, bone metastases result in debilitating pain. Deeper investigation of this pain may reveal etiology and lead to early cancer detection. Cancer-induced bone pain (CIBP) is inadequately managed with current standard-of-care analgesics and dramatically diminishes patient quality of life. While CIBP etiology is multifaceted, elevated levels of glutamate, an excitatory neurotransmitter, in the bone-tumor microenvironment may drive maladaptive nociceptive signaling. Here, we establish a relationship between the reactive nitrogen species peroxynitrite, tumor-derived glutamate, and CIBP. In vitro and in a syngeneic in vivo model of breast CIBP, murine mammary adenocarcinoma cells significantly elevated glutamate via the cystine/glutamate antiporter system xc−. The well-known system xc− inhibitor sulfasalazine significantly reduced levels of glutamate and attenuated CIBP-associated flinching and guarding behaviors. Peroxynitrite, a highly reactive species produced in tumors, significantly increased system xc− functional expression and tumor cell glutamate release. Scavenging peroxynitrite with the iron and mangano-based porphyrins, FeTMPyP and SRI10, significantly diminished tumor cell system xc− functional expression, reduced femur glutamate levels and mitigated CIBP. In sum, we demonstrate how breast cancer bone metastases upregulate a cystine/glutamate co-transporter to elevate extracellular glutamate. Pharmacological manipulation of peroxynitrite or system xc− attenuates CIBP, supporting a role for tumor-derived glutamate in CIBP and validating the targeting of system xc− as a novel therapeutic strategy for the management of metastatic bone pain. PMID:27482630

  8. Hippocampal glutamate receptors in fear memory consolidation.

    PubMed

    Cammarota, Martín; Bevilaqua, Lia R M; Bonini, Juliana S; Rossatto, Janine I; Medina, Jorge H; Izquierdo, N

    2004-01-01

    It is thought that activity-dependent changes in synaptic efficacy driven by biochemical pathways responsive to the action of the excitatory neurotransmitter glutamate are critical components of the mechanisms responsible for memory formation. In particular, the early activation of the NMDA (rNMDA) and AMPA (rAMPA) subtypes of ionotropic glutamate receptors has been demonstrated to be a necessary event for the acquisition of several types of memory. In the rat, consolidation of the long-term memory for a one-trial, step-down inhibitory avoidance task is blocked by antagonists of the rNMDA and rAMPA infused into the CA1 region of the dorsal hippocampus early after training and is associated with a rapid and reversible increase in the total number of [3H]AMPA binding sites. The learning-induced increase in [[3H]AMPA is accompanied by translocation of the GluR1 subunit of the rAMPA to the post-synaptic terminal together with its phosphorylation at Ser831. In addition, learning of the mentioned fear-motivated task induces the activation and rNMDA-dependent translocation of CaMKII to the post-synaptic density. Inhibition of this protein kinase as well as blockade of the rNMDA abolishes both the learning-induced translocation of GluR1 and its phosphorylation. Our data suggest that learning of an avoidance task enhances hippocampal rAMPA signaling through rNMDA and CaMKII-dependent mechanisms.

  9. Glutamate-dependent transcriptional regulation of GLAST: role of PKC.

    PubMed

    López-Bayghen, Esther; Ortega, Arturo

    2004-10-01

    The Na+-dependent glutamate/aspartate transporter GLAST plays a major role in the removal of glutamate from the synaptic cleft. Short-term, as well as long-term changes in transporter activity are triggered by glutamate. An important locus of regulation is the density of transporter molecules present at the plasma membrane. A substrate-dependent change in the translocation rate of the transporter molecules accounts for the short-term effect, whereas the long-term modulation apparently involves transcriptional regulation. Using cultured chick cerebellar Bergmann glial cells, we report here that glutamate receptors activation mediate a substantial reduction in the transcriptional activity of the chglast promoter through the Ca2+/diacylglicerol-dependent protein kinase (PKC) signaling cascade. Overexpression of constitutive active PKC isoforms of mimic the glutamate effect. Accordingly, increased levels of c-Jun or c-Fos, but not Jun-B, Jun-D or Fos-B, lower the chglast promoter activity. Serial deletions and electrophorectic mobility shift assays were used to define a specific region within the 5' proximal region of the chglast promoter, associated with transcriptional repression. A putative glutamate response element could be defined in the proximal promoter stretch more likely between nts -40 and -78. These results demonstrate that GLAST is under glutamate-dependent transcriptional control through PKC, and support the notion of a pivotal role of this neurotransmitter in the regulation of its own removal from the synaptic cleft, thereby modulating, mainly in the long term, glutamatergic transmission.

  10. Small molecule glutaminase inhibitors block glutamate release from stimulated microglia.

    PubMed

    Thomas, Ajit G; O'Driscoll, Cliona M; Bressler, Joseph; Kaufmann, Walter; Rojas, Camilo J; Slusher, Barbara S

    2014-01-01

    Glutaminase plays a critical role in the generation of glutamate, a key excitatory neurotransmitter in the CNS. Excess glutamate release from activated macrophages and microglia correlates with upregulated glutaminase suggesting a pathogenic role for glutaminase. Both glutaminase siRNA and small molecule inhibitors have been shown to decrease excess glutamate and provide neuroprotection in multiple models of disease, including HIV-associated dementia (HAD), multiple sclerosis and ischemia. Consequently, inhibition of glutaminase could be of interest for treatment of these diseases. Bis-2-(5-phenylacetimido-1,2,4-thiadiazol-2-yl)ethyl sulfide (BPTES) and 6-diazo-5-oxo-l-norleucine (DON), two most commonly used glutaminase inhibitors, are either poorly soluble or non-specific. Recently, several new BPTES analogs with improved physicochemical properties were reported. To evaluate these new inhibitors, we established a cell-based microglial activation assay measuring glutamate release. Microglia-mediated glutamate levels were significantly augmented by tumor necrosis factor (TNF)-α, phorbol 12-myristate 13-acetate (PMA) and Toll-like receptor (TLR) ligands coincident with increased glutaminase activity. While several potent glutaminase inhibitors abrogated the increase in glutamate, a structurally related analog devoid of glutaminase activity was unable to block the increase. In the absence of glutamine, glutamate levels were significantly attenuated. These data suggest that the in vitro microglia assay may be a useful tool in developing glutaminase inhibitors of therapeutic interest. PMID:24269238

  11. Evidence for Glutamate as a Neuroglial Transmitter within Sensory Ganglia

    PubMed Central

    Kung, Ling-Hsuan; Gong, Kerui; Adedoyin, Mary; Ng, Johnson; Bhargava, Aditi; Ohara, Peter T.; Jasmin, Luc

    2013-01-01

    This study examines key elements of glutamatergic transmission within sensory ganglia of the rat. We show that the soma of primary sensory neurons release glutamate when depolarized. Using acute dissociated mixed neuronal/glia cultures of dorsal root ganglia (DRG) or trigeminal ganglia and a colorimetric assay, we show that when glutamate uptake by satellite glial cells (SGCs) is inhibited, KCl stimulation leads to simultaneous increase of glutamate in the culture medium. With calcium imaging we see that the soma of primary sensory neurons and SGCs respond to AMPA, NMDA, kainate and mGluR agonists, and selective antagonists block this response. Using whole cell patch-clamp technique, inward currents were recorded from small diameter (<30 µm) DRG neurons from intact DRGs (ex-vivo whole ganglion preparation) in response to local application of the above glutamate receptor agonists. Following a chronic constriction injury (CCI) of either the inferior orbital nerve or the sciatic nerve, glutamate expression increases in the trigeminal ganglia and DRG respectively. This increase occurs in neurons of all diameters and is present in the somata of neurons with injured axons as well as in somata of neighboring uninjured neurons. These data provides additional evidence that glutamate can be released within the sensory ganglion, and that the somata of primary sensory neurons as well as SGCs express functional glutamate receptors at their surface. These findings, together with our previous gene knockdown data, suggest that glutamatergic transmission within the ganglion could impact nociceptive threshold. PMID:23844184

  12. Dysfunctional TCA-Cycle Metabolism in Glutamate Dehydrogenase Deficient Astrocytes.

    PubMed

    Nissen, Jakob D; Pajęcka, Kamilla; Stridh, Malin H; Skytt, Dorte M; Waagepetersen, Helle S

    2015-12-01

    Astrocytes take up glutamate in the synaptic area subsequent to glutamatergic transmission by the aid of high affinity glutamate transporters. Glutamate is converted to glutamine or metabolized to support intermediary metabolism and energy production. Glutamate dehydrogenase (GDH) and aspartate aminotransferase (AAT) catalyze the reversible reaction between glutamate and α-ketoglutarate, which is the initial step for glutamate to enter TCA cycle metabolism. In contrast to GDH, AAT requires a concomitant interconversion of oxaloacetate and aspartate. We have investigated the role of GDH in astrocyte glutamate and glucose metabolism employing siRNA mediated knock down (KD) of GDH in cultured astrocytes using stable and radioactive isotopes for metabolic mapping. An increased level of aspartate was observed upon exposure to [U-(13) C]glutamate in astrocytes exhibiting reduced GDH activity. (13) C Labeling of aspartate and TCA cycle intermediates confirmed that the increased amount of aspartate is associated with elevated TCA cycle flux from α-ketoglutarate to oxaloacetate, i.e. truncated TCA cycle. (13) C Glucose metabolism was elevated in GDH deficient astrocytes as observed by increased de novo synthesis of aspartate via pyruvate carboxylation. In the absence of glucose, lactate production from glutamate via malic enzyme was lower in GDH deficient astrocytes. In conclusions, our studies reveal that metabolism via GDH serves an important anaplerotic role by adding net carbon to the TCA cycle. A reduction in GDH activity seems to cause the astrocytes to up-regulate activity in pathways involved in maintaining the amount of TCA cycle intermediates such as pyruvate carboxylation as well as utilization of alternate substrates such as branched chain amino acids.

  13. Relationship between increase in astrocytic GLT-1 glutamate transport and late-LTP

    PubMed Central

    Pita-Almenar, Juan D.; Zou, Shengwei; Colbert, Costa M.; Eskin, Arnold

    2012-01-01

    Na+-dependent high-affinity glutamate transporters have important roles in the maintenance of basal levels of glutamate and clearance of glutamate during synaptic transmission. Interestingly, several studies have shown that basal glutamate transport displays plasticity. Glutamate uptake increases in hippocampal slices during early long-term potentiation (E-LTP) and late long-term potentiation (L-LTP). Four issues were addressed in this research: Which glutamate transporter is responsible for the increase in glutamate uptake during L-LTP? In what cell type in the hippocampus does the increase in glutamate uptake occur? Does a single type of cell contain all the mechanisms to respond to an induction stimulus with a change in glutamate uptake? What role does the increase in glutamate uptake play during L-LTP? We have confirmed that GLT-1 is responsible for the increase in glutamate uptake during L-LTP. Also, we found that astrocytes were responsible for much, if not all, of the increase in glutamate uptake in hippocampal slices during L-LTP. Additionally, we found that cultured astrocytes alone were able to respond to an induction stimulus with an increase in glutamate uptake. Inhibition of basal glutamate uptake did not affect the induction of L-LTP, but inhibition of the increase in glutamate uptake did inhibit both the expression of L-LTP and induction of additional LTP. It seems likely that heightened glutamate transport plays an ongoing role in the ability of hippocampal circuitry to code and store information. PMID:23166293

  14. Efficient synthesis of novel glutamate homologues and investigation of their affinity and selectivity profile at ionotropic glutamate receptors.

    PubMed

    Pinto, Andrea; Tamborini, Lucia; Mastronardi, Federica; Ettari, Roberta; Romano, Diego; Nielsen, Birgitte; De Micheli, Carlo; Conti, Paola

    2014-04-15

    A convenient synthesis of four new enantiomerically pure acidic amino acids is reported and their affinity at ionotropic glutamate receptors was determined. The new compounds are higher homologues of glutamic acid in which the molecular complexity has been increased by introducing an aromatic/heteroaromatic ring, that is a phenyl or a thiophene ring, that could give additional electronic interactions with the receptors. The results of the present investigation indicate that the insertion of an aromatic/heteroaromatic ring into the amino acid skeleton of glutamate higher homologues is well tolerated and this modification could be exploited to generate a new class of NMDA antagonists. PMID:24630559

  15. Glutamate neurons within the midbrain dopamine regions.

    PubMed

    Morales, M; Root, D H

    2014-12-12

    Midbrain dopamine systems play important roles in Parkinson's disease, schizophrenia, addiction, and depression. The participation of midbrain dopamine systems in diverse clinical contexts suggests these systems are highly complex. Midbrain dopamine regions contain at least three neuronal phenotypes: dopaminergic, GABAergic, and glutamatergic. Here, we review the locations, subtypes, and functions of glutamatergic neurons within midbrain dopamine regions. Vesicular glutamate transporter 2 (VGluT2) mRNA-expressing neurons are observed within each midbrain dopamine system. Within rat retrorubral field (RRF), large populations of VGluT2 neurons are observed throughout its anteroposterior extent. Within rat substantia nigra pars compacta (SNC), VGluT2 neurons are observed centrally and caudally, and are most dense within the laterodorsal subdivision. RRF and SNC rat VGluT2 neurons lack tyrosine hydroxylase (TH), making them an entirely distinct population of neurons from dopaminergic neurons. The rat ventral tegmental area (VTA) contains the most heterogeneous populations of VGluT2 neurons. VGluT2 neurons are found in each VTA subnucleus but are most dense within the anterior midline subnuclei. Some subpopulations of rat VGluT2 neurons co-express TH or glutamic acid decarboxylase (GAD), but most of the VGluT2 neurons lack TH or GAD. Different subsets of rat VGluT2-TH neurons exist based on the presence or absence of vesicular monoamine transporter 2, dopamine transporter, or D2 dopamine receptor. Thus, the capacity by which VGluT2-TH neurons may release dopamine will differ based on their capacity to accumulate vesicular dopamine, uptake extracellular dopamine, or be autoregulated by dopamine. Rat VTA VGluT2 neurons exhibit intrinsic VTA projections and extrinsic projections to the accumbens and to the prefrontal cortex. Mouse VTA VGluT2 neurons project to accumbens shell, prefrontal cortex, ventral pallidum, amygdala, and lateral habenula. Given their molecular

  16. Neuroprotective Effects of Glutamate Antagonists and Extracellular Acidity

    NASA Astrophysics Data System (ADS)

    Kaku, David A.; Giffard, Rona G.; Choi, Dennis W.

    1993-06-01

    Glutamate antagonists protect neurons from hypoxic injury both in vivo and in vitro, but in vitro studies have not been done under the acidic conditions typical of hypoxia-ischemia in vivo. Consistent with glutamate receptor antagonism, extracellular acidity reduced neuronal death in murine cortical cultures that were deprived of oxygen and glucose. Under these acid conditions, N-methyl-D-aspartate and α-amino-3-hydroxy-5-methyl-4-isox-azolepropionate-kainate antagonists further reduced neuronal death, such that some neurons tolerated prolonged oxygen and glucose deprivation almost as well as did astrocytes. Neuroprotection induced by this combination exceeded that induced by glutamate antagonists alone, suggesting that extracellular acidity has beneficial effects beyond the attenuation of ionotropic glutamate receptor activation.

  17. Bidirectional Control of Synaptic GABAAR Clustering by Glutamate and Calcium

    PubMed Central

    Bannai, Hiroko; Niwa, Fumihiro; Sherwood, Mark W.; Shrivastava, Amulya Nidhi; Arizono, Misa; Miyamoto, Akitoshi; Sugiura, Kotomi; Lévi, Sabine; Triller, Antoine; Mikoshiba, Katsuhiko

    2015-01-01

    Summary GABAergic synaptic transmission regulates brain function by establishing the appropriate excitation-inhibition (E/I) balance in neural circuits. The structure and function of GABAergic synapses are sensitive to destabilization by impinging neurotransmitters. However, signaling mechanisms that promote the restorative homeostatic stabilization of GABAergic synapses remain unknown. Here, by quantum dot single-particle tracking, we characterize a signaling pathway that promotes the stability of GABAA receptor (GABAAR) postsynaptic organization. Slow metabotropic glutamate receptor signaling activates IP3 receptor-dependent calcium release and protein kinase C to promote GABAAR clustering and GABAergic transmission. This GABAAR stabilization pathway counteracts the rapid cluster dispersion caused by glutamate-driven NMDA receptor-dependent calcium influx and calcineurin dephosphorylation, including in conditions of pathological glutamate toxicity. These findings show that glutamate activates distinct receptors and spatiotemporal patterns of calcium signaling for opposing control of GABAergic synapses. PMID:26711343

  18. A further experimental study of the antisilicotic effect of glutamate.

    PubMed Central

    Morosova, K I; Katsnelson, B A; Rotenberg YuS; Belobragina, G V

    1984-01-01

    Two groups of rats were exposed to quartz dust for six months and in addition one group was given drinking water containing 1.5% sodium glutamate while the second received only water. In the rats receiving glutamate we observed (a) evidence for a considerably reduced cytotoxic effect of the quartz on cells obtained by bronchopulmonary lavage, (b) a reduction in dust retention in the lungs, especially in the tracheobronchial lymph nodes, (c) a considerable reduction in the weight gain in the lungs and in their hydroxyproline and lipid contents, and (d) the inhibition of the formation of silicotic nodules. Polarographic studies of the oxygen consumption of peritoneal macrophages from rats receiving glutamate showed that glutamate prevents the adverse effects of quartz on mitochondrial oxidative processes. PMID:6093851

  19. Transport Mechanism of a Bacterial Homologue of Glutamate Transporters

    SciTech Connect

    Reyes, N.; Ginter, C; Boudker, O

    2009-01-01

    Glutamate transporters are integral membrane proteins that catalyse a thermodynamically uphill uptake of the neurotransmitter glutamate from the synaptic cleft into the cytoplasm of glia and neuronal cells by harnessing the energy of pre-existing electrochemical gradients of ions. Crucial to the reaction is the conformational transition of the transporters between outward and inward facing states, in which the substrate binding sites are accessible from the extracellular space and the cytoplasm, respectively. Here we describe the crystal structure of a double cysteine mutant of a glutamate transporter homologue from Pyrococcus horikoshii, GltPh, which is trapped in the inward facing state by cysteine crosslinking. Together with the previously determined crystal structures of Glt{sub Ph} in the outward facing state, the structure of the crosslinked mutant allows us to propose a molecular mechanism by which Glt{sub Ph} and, by analogy, mammalian glutamate transporters mediate sodium-coupled substrate uptake.

  20. Metabolic Control of Vesicular Glutamate Transport and Release

    PubMed Central

    Juge, Narinobu; Gray, John A.; Omote, Hiroshi; Miyaji, Takaaki; Inoue, Tsuyoshi; Hara, Chiaki; Uneyama, Hisayuki; Edwards, Robert H.; Nicoll, Roger A.; Moriyama, Yoshinori

    2010-01-01

    Fasting has been used to control epilepsy since antiquity, but the mechanism of coupling between metabolic state and excitatory neurotransmission remains unknown. Previous work has shown that the vesicular glutamate transporters (VGLUTs) required for exocytotic release of glutamate undergo an unusual form of regulation by Cl−. Using functional reconstitution of the purified VGLUTs into proteoliposomes, we now show that Cl− acts as an allosteric activator, and the ketone bodies that increase with fasting inhibit glutamate release by competing with Cl− at the site of allosteric regulation. Consistent with these observations, acetoacetate reduced quantal size at hippocampal synapses, and suppresses glutamate release and seizures evoked with 4-aminopyridine in the brain. The results indicate an unsuspected link between metabolic state and excitatory neurotransmission through anion-dependent regulation of VGLUT activity. PMID:20920794

  1. Vesicular release of glutamate from unmyelinated axons in white matter

    PubMed Central

    Ziskin, Jennifer L; Nishiyama, Akiko; Rubio, Maria; Fukaya, Masahiro; Bergles, Dwight E

    2007-01-01

    Directed fusion of transmitter-laden vesicles enables rapid intercellular signaling in the central nervous system and occurs at synapses within gray matter. Here we show that action potentials also induce the release of glutamate from axons in the corpus callosum, a white matter region responsible for interhemispheric communication. Callosal axons release glutamate by vesicular fusion, which induces quantal AMPA receptor–mediated currents in NG2+ glial progenitors at anatomically distinct axo–glial synaptic junctions. Glutamate release from axons was facilitated by repetitive stimulation and could be inhibited through activation of metabotropic autoreceptors. Although NG2+ cells form associations with nodes of Ranvier in white matter, measurements of conduction velocity indicated that unmyelinated fibers are responsible for glutamatergic signaling with NG2+ glia. This activity-dependent secretion of glutamate was prevalent in the developing and mature mouse corpus callosum, indicating that axons within white matter both conduct action potentials and engage in rapid neuron-glia communication. PMID:17293857

  2. Glutamic Acid Selective Chemical Cleavage of Peptide Bonds.

    PubMed

    Nalbone, Joseph M; Lahankar, Neelam; Buissereth, Lyssa; Raj, Monika

    2016-03-01

    Site-specific hydrolysis of peptide bonds at glutamic acid under neutral aqueous conditions is reported. The method relies on the activation of the backbone amide chain at glutamic acid by the formation of a pyroglutamyl (pGlu) imide moiety. This activation increases the susceptibility of a peptide bond toward hydrolysis. The method is highly specific and demonstrates broad substrate scope including cleavage of various bioactive peptides with unnatural amino acid residues, which are unsuitable substrates for enzymatic hydrolysis.

  3. Transport mechanism of a glutamate transporter homologue GltPh

    PubMed Central

    Ji, Yurui; Postis, Vincent L.G.; Wang, Yingying; Bartlam, Mark; Goldman, Adrian

    2016-01-01

    Glutamate transporters are responsible for uptake of the neurotransmitter glutamate in mammalian central nervous systems. Their archaeal homologue GltPh, an aspartate transporter isolated from Pyrococcus horikoshii, has been the focus of extensive studies through crystallography, MD simulations and single-molecule FRET (smFRET). Here, we summarize the recent research progress on GltPh, in the hope of gaining some insights into the transport mechanism of this aspartate transporter. PMID:27284058

  4. Functional architecture of olfactory ionotropic glutamate receptors

    PubMed Central

    Abuin, Liliane; Bargeton, Benoîte; Ulbrich, Maximilian H.; Isacoff, Ehud Y.; Kellenberger, Stephan; Benton, Richard

    2010-01-01

    Ionotropic glutamate receptors (iGluRs) are ligand-gated ion channels that mediate chemical communication between neurons at synapses. A variant iGluR subfamily, the Ionotropic Receptors (IRs), was recently proposed to detect environmental volatile chemicals in olfactory cilia. Here we elucidate how these peripheral chemosensors have evolved mechanistically from their iGluR ancestors. Using a Drosophila model, we demonstrate that IRs act in combinations of up to three subunits, comprising individual odor-specific receptors and one or two broadly expressed co-receptors. Heteromeric IR complex formation is necessary and sufficient for trafficking to cilia and mediating odor-evoked electrophysiological responses in vivo and in vitro. IRs display heterogeneous ion conduction specificities related to their variable pore sequences, and divergent ligand-binding domains function in odor recognition and cilia localization. Our results provide insights into the conserved and distinct architecture of these olfactory and synaptic ion channels and offer perspectives into use of IRs as genetically encoded chemical sensors. PMID:21220098

  5. [Glutamate Metabotropic Receptors: Structure, Localisation, Functions].

    PubMed

    Perfilova, V N; Tyurenkov, I N

    2016-01-01

    The data on the structure, location and functions of the metabotropic glutamate receptor is shown. The family consists of 8 mGluRs subtypes and is divided into three groups: I group--mGluRs1/mGluRs5, II group--mGluRs2/mGluRs3, III group--mGluRs4/mGluRs6/mGluRs7/mGluRs8. They are associated with G-protein; signaling in the cells is carried out by IP3 or adenylate cyclase signaling pathways, in the result of which, mGluRs modify glial and neuronal excitability. Receptors are localized in the CNS and periphery in non-neuronal tissues: bone, heart, kidney, pancreas pod and platelets, the gastrointestinal tract, immune system. Their participation in the mechanisms of neurodegenerative diseases, mental and cognitive disorders, autoimmune processes, etc. is displayed. Agonists, antagonists, allosteric modulators of mGluRs are considered as potential medicines for treatment of mental diseases, including depression, fragile X syndrome, anxiety, obsessive-compulsive disorders, Parkinson's disease, etc. PMID:27530046

  6. Reduced hippocampal glutamate in Alzheimer disease.

    PubMed

    Rupsingh, R; Borrie, M; Smith, M; Wells, J L; Bartha, R

    2011-05-01

    Altered neurometabolic profiles have been detected in Alzheimer disease (AD) using (1)H magnetic resonance spectroscopy (MRS), but no definitive biomarker of mild cognitive impairment (MCI) or AD has been established. This study used MRS to compare hippocampal metabolite levels between normal elderly controls (NEC) and subjects with MCI and AD. Short echo-time (TE=46 ms) (1)H spectra were acquired at 4T from the right hippocampus of 23 subjects with AD, 12 subjects with MCI and 15 NEC. Absolute metabolite levels and metabolite ratios were compared between groups using a multivariate analysis of covariance (covariates: age, sex) followed by post hoc Tukey's test (p<0.05 significant). Subjects with AD had decreased glutamate (Glu) as well as decreased Glu/creatine (Cr), Glu/myo-inositol (mI), Glu/N-acetylaspartate (NAA), and NAA/Cr ratios compared to NEC. Subjects with AD also had decreased Glu/mI ratio compared to MCI. There were no differences between subjects with MCI and NEC. Therefore, in addition to NAA/Cr, decreased hippocampal Glu may be an indicator of AD.

  7. [Glutamic acid as a universal extracellular signal].

    PubMed

    Yoneda, Yukio

    2015-08-01

    The prevailing view is that both glutamic (Glu) and gamma-aminobutyric (GABA) acids play a role as an amino acid neurotransmitter released from neurons. However, little attention has been paid to the possible expression and functionality of signaling machineries required for amino acidergic neurotransmission in cells other than central neurons. In line with our first demonstration of the presence of Glu receptors outside the brain, in this review I will outline our recent findings accumulated since then on the physiological and pathological significance of neuronal amino acids as an extracellular signal essential for homeostasis in a variety of phenotypic cells. In undifferentiated neural progenitor cells, for instance, functional expression is seen with different signaling machineries used for glutamatergic and GABAergic neurotransmission in neurons. Moreover, Glu plays a role in mechanisms underlying suppression of proliferation for self-replication in undifferentiated mesenchymal stem cells. There is more accumulating evidence for neuronal amino acids playing a role as an extracellular autocrine or paracrine signal commonly used in different phenotypic cells. Evaluation of drugs currently used could be thus beneficial for the efficient prophylaxis and/or the therapy of a variety of diseases relevant to disturbance of amino acid signaling in diverse organs.

  8. Podocytes: a new player for glutamate signaling.

    PubMed

    Armelloni, S; Li, M; Messa, P; Rastaldi, M P

    2012-12-01

    In the renal glomerulus, podocytes envelop the external side of the capillary basement membrane with their intertwining ramifications, and ensure elimination of metabolic waste within the urine, while proteins and important blood components are retained into the circulation. To preserve the integrity of the glomerular filter, which is constantly exposed to a high variety of stimuli, podocytes need to communicate by rapid and precise signaling, likely similar to that used by neuronal cells. In the last years, we and others have shown that podocytes are indeed molecularly equipped for communicating in a synaptic-like way, where glutamate and its receptors seem to have a pivotal role, because altering glutamatergic communication leads to podocyte damage and increased filter permeability. Major components of glutamatergic signaling are organized at foot process junctions by adhesion molecules, chiefly by nephrin, and are connected to the actin cytoskeleton, that governs the health of podocytes. Further advances in understanding podocyte physiological behavior and signaling properties have the potential to improve the knowledge of podocyte diseases, first among them idiopathic focal segmental glomerulosclerosis that still needs more precise molecular-based diagnosis and targeted treatment. PMID:23018105

  9. Nitrogen isotope effects on glutamate decarboxylase from Escherichia coli

    SciTech Connect

    Abell, L.M.; O'Leary, M.H.

    1988-05-03

    The nitrogen isotope effect on the decarboxylation of glutamic acid by glutamate decarboxylase from Escherichia coli has been measured by comparison of the isotopic composition of the amino nitrogen of the product ..gamma..-aminobutyric acid isolated after 10-20% reaction with that of the starting glutamic acid. At pH 4.7, 37 /sup 0/C, the isotope effect is k/sup 14//k/sup 15/ = 0.9855 +/- 0.0006 when compared to unprotonated glutamic acid. Interpretation of this result requires knowledge of the equilibrium nitrogen isotope effect for Schiff base formation. This equilibrium isotope effect is K/sup 14//K/sup 15/ - 0.9824 for the formation of the unprotonated Schiff base between unprotonated valine and salicylaldehyde. Analysis of the nitrogen isotope effect on decarboxylation of glutamic acid and of the previously measured carbon isotope effect on this same reaction shows that decarboxylation and Schiff base formation are jointly rate limiting. The enzyme-bound Schiff base between glutamate and pyridoxal 5'-phosphate partitions approximately 2:1 between decarboxylation and return to the starting state. The nitrogen isotope effect also reveals that the Schiff base nitrogen is protonated in this intermediate.

  10. Effect of dexamethasone on fetal hepatic glutamine-glutamate exchange.

    PubMed

    Timmerman, M; Teng, C; Wilkening, R B; Fennessey, P; Battaglia, F C; Meschia, G

    2000-05-01

    Intravenous infusion of dexamethasone (Dex) in the fetal lamb causes a two- to threefold increase in plasma glutamine and other glucogenic amino acids and a decrease of plasma glutamate to approximately one-third of normal. To explore the underlying mechanisms, hepatic amino acid uptake and conversion of L-[1-(13)C]glutamine to L-[1-(13)C]glutamate and (13)CO(2) were measured in six sheep fetuses before and in the last 2 h of a 26-h Dex infusion. Dex decreased hepatic glutamine and alanine uptakes (P < 0.01) and hepatic glutamate output (P < 0.001). Hepatic outputs of the glutamate (R(Glu,Gln)) and CO(2) formed from plasma glutamine decreased to 21 (P < 0.001) and 53% (P = 0.009) of control, respectively. R(Glu,Gln), expressed as a fraction of both outputs, decreased (P < 0.001) from 0.36 +/- 0.02 to 0.18 +/- 0.04. Hepatic glucose output remained virtually zero throughout the experiment. We conclude that Dex decreases fetal hepatic glutamate output by increasing the routing of glutamate carbon into the citric acid cycle and by decreasing the hepatic uptake of glucogenic amino acids. PMID:10780940

  11. Glutamate biosensors based on diamond and graphene platforms.

    PubMed

    Hu, Jingping; Wisetsuwannaphum, Sirikarn; Foord, John S

    2014-01-01

    l-Glutamate is one of the most important neurotransmitters in the mammalian central nervous system, playing a vital role in many physiological processes and implicated in several neurological disorders, for which monitoring of dynamic levels of extracellular glutamate in the living brain tissues may contribute to medical understanding and treatments. Electrochemical sensing of glutamate has been developed recently mainly using platinum, carbon fibre and carbon nanotube electrodes. In the present work, we explore the fabrication and properties of electrochemical glutamate sensors fabricated on doped chemical vapour deposition diamond electrodes and graphene nanoplatelet structures. The sensors incorporate platinum nanoparticles to catalyse the electrooxidation of hydrogen peroxide, glutamate oxidase to oxidise glutamate, and a layer of poly-phenylenediamine to impart selectivity. The performance of the devices was compared to a similar sensor fabricated on glassy carbon. Both the diamond and the graphene sensor showed very competitive performance compared to the majority of existing electrochemical sensors. The graphene based sensor showed the best performance of the three investigated in terms of sensitivity, linear dynamic range and long term stability, whereas it was found that the diamond device showed the best limit of detection.

  12. Glutamate Receptor Ion Channels: Structure, Regulation, and Function

    PubMed Central

    Wollmuth, Lonnie P.; McBain, Chris J.; Menniti, Frank S.; Vance, Katie M.; Ogden, Kevin K.; Hansen, Kasper B.; Yuan, Hongjie; Myers, Scott J.; Dingledine, Ray

    2010-01-01

    The mammalian ionotropic glutamate receptor family encodes 18 gene products that coassemble to form ligand-gated ion channels containing an agonist recognition site, a transmembrane ion permeation pathway, and gating elements that couple agonist-induced conformational changes to the opening or closing of the permeation pore. Glutamate receptors mediate fast excitatory synaptic transmission in the central nervous system and are localized on neuronal and non-neuronal cells. These receptors regulate a broad spectrum of processes in the brain, spinal cord, retina, and peripheral nervous system. Glutamate receptors are postulated to play important roles in numerous neurological diseases and have attracted intense scrutiny. The description of glutamate receptor structure, including its transmembrane elements, reveals a complex assembly of multiple semiautonomous extracellular domains linked to a pore-forming element with striking resemblance to an inverted potassium channel. In this review we discuss International Union of Basic and Clinical Pharmacology glutamate receptor nomenclature, structure, assembly, accessory subunits, interacting proteins, gene expression and translation, post-translational modifications, agonist and antagonist pharmacology, allosteric modulation, mechanisms of gating and permeation, roles in normal physiological function, as well as the potential therapeutic use of pharmacological agents acting at glutamate receptors. PMID:20716669

  13. Glutamate biosensors based on diamond and graphene platforms.

    PubMed

    Hu, Jingping; Wisetsuwannaphum, Sirikarn; Foord, John S

    2014-01-01

    l-Glutamate is one of the most important neurotransmitters in the mammalian central nervous system, playing a vital role in many physiological processes and implicated in several neurological disorders, for which monitoring of dynamic levels of extracellular glutamate in the living brain tissues may contribute to medical understanding and treatments. Electrochemical sensing of glutamate has been developed recently mainly using platinum, carbon fibre and carbon nanotube electrodes. In the present work, we explore the fabrication and properties of electrochemical glutamate sensors fabricated on doped chemical vapour deposition diamond electrodes and graphene nanoplatelet structures. The sensors incorporate platinum nanoparticles to catalyse the electrooxidation of hydrogen peroxide, glutamate oxidase to oxidise glutamate, and a layer of poly-phenylenediamine to impart selectivity. The performance of the devices was compared to a similar sensor fabricated on glassy carbon. Both the diamond and the graphene sensor showed very competitive performance compared to the majority of existing electrochemical sensors. The graphene based sensor showed the best performance of the three investigated in terms of sensitivity, linear dynamic range and long term stability, whereas it was found that the diamond device showed the best limit of detection. PMID:25427169

  14. Comparative evaluation of glutamate-sensitive radiopharmaceuticals: Technetium-99m-glutamic acid and technetium-99m-diethylenetriaminepentaacetic acid-bis(glutamate) conjugate for tumor imaging.

    PubMed

    Kakkar, Dipti; Tiwari, Anjani K; Chuttani, Krishna; Kaul, Ankur; Singh, Harpal; Mishra, Anil K

    2010-12-01

    Single-photon emission computed tomography has become a significant imaging modality with huge potential to visualize and provide information of anatomic dysfunctions that are predictive of future diseases. This imaging tool is complimented by radiopharmaceuticals/radiosubstrates that help in imaging specific physiological aspects of the human body. The present study was undertaken to explore the utility of technetium-99m (⁹⁹(m)Tc)-labeled glutamate conjugates for tumor scintigraphy. As part of our efforts to further utilize the application of chelating agents, glutamic acid was conjugated with a multidentate ligand, diethylenetriaminepentaacetic acid (DTPA). The DTPA-glutamate conjugate [DTPA-bis(Glu)] was well characterized by IR, NMR, and mass spectroscopy. The biological activity of glutamic acid was compared with its DTPA conjugate by radiocomplexation with ⁹⁹(m)Tc (labeling efficiency ≥98%). In vivo studies of both the radiolabeled complexes ⁹⁹(m)Tc-Glu and ⁹⁹(m)Tc-DTPA-bis(Glu) were then carried out, followed by gamma scintigraphy in New Zealand albino rabbits. Improved serum stability of ⁹⁹(m)Tc-labeled DTPA conjugate indicated that ⁹⁹(m)Tc remained bound to the conjugate up to 24 hours. Blood clearance showed a relatively slow washout of the DTPA conjugate when compared with the labeled glutamate. Biodistribution characteristics of the conjugate in Balb/c mice revealed that DTPA conjugation of glutamic acid favors less accumulation in the liver and bone and rapid renal clearance. Tumor scintigraphy in mice showed increasing tumor accumulation, stable up to 4 hours. These preliminary studies show that ⁹⁹(m)Tc-DTPA-bis(Glu) can be a useful radiopharmaceutical for diagnostic applications in single-photon emission computed tomography imaging.

  15. Type 1 metabotropic glutamate receptors (mGlu1) trigger the gating of GluD2 delta glutamate receptors

    PubMed Central

    Ady, Visou; Perroy, Julie; Tricoire, Ludovic; Piochon, Claire; Dadak, Selma; Chen, Xiaoru; Dusart, Isabelle; Fagni, Laurent; Lambolez, Bertrand; Levenes, Carole

    2014-01-01

    The orphan GluD2 receptor belongs to the ionotropic glutamate receptor family but does not bind glutamate. Ligand-gated GluD2 currents have never been evidenced, and whether GluD2 operates as an ion channel has been a long-standing question. Here, we show that GluD2 gating is triggered by type 1 metabotropic glutamate receptors, both in a heterologous expression system and in Purkinje cells. Thus, GluD2 is not only an adhesion molecule at synapses but also works as a channel. This gating mechanism reveals new properties of glutamate receptors that emerge from their interaction and opens unexpected perspectives regarding synaptic transmission and plasticity. PMID:24357660

  16. Bacterial cell-surface displaying of thermo-tolerant glutamate dehydrogenase and its application in L-glutamate assay.

    PubMed

    Song, Jianxia; Liang, Bo; Han, Dongfei; Tang, Xiangjiang; Lang, Qiaolin; Feng, Ruirui; Han, Lihui; Liu, Aihua

    2015-03-01

    In this paper, glutamate dehydrogenase (Gldh) is reported to efficiently display on Escherichia coli cell surface by using N-terminal region of ice the nucleation protein as an anchoring motif. The presence of Gldh was confirmed by SDS-PAGE and enzyme activity assay. Gldh was detected mainly in the outer membrane fraction, suggesting that the Gldh was displayed on the bacterial cell surface. The optimal temperature and pH for the bacteria cell-surface displayed Gldh (bacteria-Gldh) were 70°C and 9.0, respectively. Additionally, the fusion protein retained almost 100% of its initial enzymatic activity after 1 month incubation at 4°C. Transition metal ions could inhibit the enzyme activity to different extents, while common anions had little adverse effect on enzyme activity. Importantly, the displayed Gldh is most specific to l-glutamate reported so far. The bacterial Gldh was enabled to catalyze oxidization of l-glutamate with NADP(+) as cofactor, and the resultant NADPH can be detected spectrometrically at 340nm. The bacterial-Gldh based l-glutamate assay was established, where the absorbance at 340nm increased linearly with the increasing l-glutamate concentration within the range of 10-400μM. Further, the proposed approach was successfully applied to measure l-glutamate in real samples. PMID:25659635

  17. Among the twenty classical L-amino acids, only glutamate directly activates metabotropic glutamate receptors.

    PubMed

    Frauli, Mélanie; Neuville, Pascal; Vol, Claire; Pin, Jean-Philippe; Prézeau, Laurent

    2006-02-01

    Under pathophysiological conditions, cellular amino acids can be profusely released from cells into the cerebral interstitial space. Because several class-C G protein coupled receptors (GPCRs) display a broad natural ligand spectrum, being sensitive to more than one endogenous ligand, we wondered whether the related metabotropic glutamate (mGlu) receptors could be modulated by various types of L-amino acids, allowing them to sense large increase in extracellular amino acid concentration. Here, the agonist, antagonist and allosteric effects of the twenty classical L-amino acids were evaluated on the eight mGlu receptor subtypes. We show that, in addition to glutamate (Glu), cysteine, aspartate and asparagine also lead to the activation of mGlu3, 4 and 5. Interestingly, our data demonstrate that the effect of these three amino acids did not result from a direct activation of the receptors, but from an indirect action involving Glu-transporters/exchangers. These data first demonstrate that mGlu receptors, unlike other class-C GPCRs, display an extremely high selectivity towards one ligand. Moreover, our results also show that Glu transport systems allow mGlu receptors to sense large increase in the extracellular concentration of some amino acids. Such a system will certainly lead to a large increase in some mGlu receptor activity under pathological conditions, such as seizure, ischemia or other brain injuries. PMID:16310227

  18. Human Articular Chondrocytes Express Multiple Gap Junction Proteins

    PubMed Central

    Mayan, Maria D.; Carpintero-Fernandez, Paula; Gago-Fuentes, Raquel; Martinez-de-Ilarduya, Oskar; Wang, Hong-Zhang; Valiunas, Virginijus; Brink, Peter; Blanco, Francisco J.

    2014-01-01

    Osteoarthritis (OA) is the most common joint disease and involves progressive degeneration of articular cartilage. The aim of this study was to investigate if chondrocytes from human articular cartilage express gap junction proteins called connexins (Cxs). We show that human chondrocytes in tissue express Cx43, Cx45, Cx32, and Cx46. We also find that primary chondrocytes from adults retain the capacity to form functional voltage-dependent gap junctions. Immunohistochemistry experiments in cartilage from OA patients revealed significantly elevated levels of Cx43 and Cx45 in the superficial zone and down through the next approximately 1000 μm of tissue. These zones corresponded with regions damaged in OA that also had high levels of proliferative cell nuclear antigen. An increased number of Cxs may help explain the increased proliferation of cells in clusters that finally lead to tissue homeostasis loss. Conversely, high levels of Cxs in OA cartilage reflect the increased number of adjacent cells in clusters that are able to interact directly by gap junctions as compared with hemichannels on single cells in normal cartilage. Our data provide strong evidence that OA patients have a loss of the usual ordered distribution of Cxs in the damaged zones and that the reductions in Cx43 levels are accompanied by the loss of correct Cx localization in the nondamaged areas. PMID:23416160

  19. Metabotropic glutamate receptor ligands as potential therapeutics for addiction

    PubMed Central

    Olive, M. F.

    2009-01-01

    There is now compelling evidence that the excitatory amino acid neurotransmitter glutamate plays a pivotal role in drug addiction and alcoholism. As a result, there has been increasing interest in developing glutamate-based therapies for the treatment of addictive disorders. Receptors for glutamate are primarily divided into two classes: ionotropic glutamate receptors (iGluRs) that mediate fast excitatory glutamate transmission, and metabotropic glutamate receptors (mGluRs), which are G-protein coupled receptors that mediate slower, modulatory glutamate transmission. Most iGluR antagonists, while showing some efficacy in animal models of addiction, exhibit serious side effects when tested in humans. mGluR ligands, on the other hand, which have been advanced to testing in clinical trials for various medical conditions, have demonstrated the ability to reduce drug reward, reinforcement, and relapse-like behaviors in animal studies. mGluR ligands that have been shown to be primarily effective are Group I (mGluR1 and mGluR5) negative allosteric modulators and Group II (mGluR2 and mGluR3) orthosteric presynaptic autoreceptor agonists. In this review, we will summarize findings from animal studies suggesting that these mGluR ligands may be of potential benefit in reducing on-going drug self-administration and may aid in the prevention of relapse. The neuroanatomical distribution of mGluR1, mGluR2/3, and mGluR5 receptors and the pharmacological properties of Group I negative allosteric modulators and Group II agonists will also be overviewed. Finally, we will discuss the current status of mGluR ligands in human clinical trials. PMID:19630739

  20. Current approaches to enhance glutamate transporter function and expression.

    PubMed

    Fontana, Andréia C K

    2015-09-01

    L-glutamate is the predominant excitatory neurotransmitter in the CNS and has a central role in a variety of brain functions. The termination of glutamate neurotransmission by excitatory amino acid transporters (EAATs) is essential to maintain glutamate concentration low in extracellular space and avoid excitotoxicity. EAAT2/GLT-1, being the most abundant subtype of glutamate transporter in the CNS, plays a key role in regulation of glutamate transmission. Dysfunction of EAAT2 has been correlated with various pathologies such as traumatic brain injury, stroke, amyotrophic lateral sclerosis, Alzheimer's disease, among others. Therefore, activators of the function or enhancers of the expression of EAAT2/GLT-1 could serve as a potential therapy for these conditions. Translational activators of EAAT2/GLT-1, such as ceftriaxone and LDN/OSU-0212320, have been described to have significant protective effects in animal models of amyotrophic lateral sclerosis and epilepsy. In addition, pharmacological activators of the activity of EAAT2/GLT-1 have been explored for decades and are currently emerging as promising tools for neuroprotection, having potential advantages over expression activators. This review describes the current status of the search for EAAT2/GLT-1 activators and addresses challenges and limitations that this approach might encounter. Termination of glutamate neurotransmission by glutamate transporter EAAT2 is essential to maintain homeostasis in the brain and to avoid excitotoxicity. Dysfunction of EAAT2 has been correlated with various neurological pathologies. Therefore, activators of the function or enhancers of the expression of EAAT2 (green arrows) could serve as a potential therapy for these conditions. This review describes the current status of the search for EAAT2 activators and addresses challenges and limitations of this approach. PMID:26096891

  1. Therapeutic effects of glutamic acid in piglets challenged with deoxynivalenol.

    PubMed

    Wu, Miaomiao; Xiao, Hao; Ren, Wenkai; Yin, Jie; Tan, Bie; Liu, Gang; Li, Lili; Nyachoti, Charles Martin; Xiong, Xia; Wu, Guoyao

    2014-01-01

    The mycotoxin deoxynivalenol (DON), one of the most common food contaminants, primarily targets the gastrointestinal tract to affect animal and human health. This study was conducted to examine the protective function of glutamic acid on intestinal injury and oxidative stress caused by DON in piglets. Twenty-eight piglets were assigned randomly into 4 dietary treatments (7 pigs/treatment): 1) uncontaminated control diet (NC), 2) NC+DON at 4 mg/kg (DON), 3) NC+2% glutamic acid (GLU), and 4) NC+2% glutamic acid + DON at 4 mg/kg (DG). At day 15, 30 and 37, blood samples were collected to determine serum concentrations of CAT (catalase), T-AOC (total antioxidant capacity), H2O2 (hydrogen peroxide), NO (nitric oxide), MDA (maleic dialdehyde), DAO (diamine oxidase) and D-lactate. Intestinal morphology, and the activation of Akt/mTOR/4EBP1 signal pathway, as well as the concentrations of H2O2, MDA, and DAO in kidney, liver and small intestine, were analyzed at day 37. Results showed that DON significantly (P<0.05) induced oxidative stress in piglets, while this stress was remarkably reduced with glutamic acid supplementation according to the change of oxidative parameters in blood and tissues. Meanwhile, DON caused obvious intestinal injury from microscopic observations and permeability indicators, which was alleviated by glutamic acid supplementation. Moreover, the inhibition of DON on Akt/mTOR/4EBP1 signal pathway was reduced by glutamic acid supplementation. Collectively, these data suggest that glutamic acid may be a useful nutritional regulator for DON-induced damage manifested as oxidative stress, intestinal injury and signaling inhibition.

  2. Therapeutic Effects of Glutamic Acid in Piglets Challenged with Deoxynivalenol

    PubMed Central

    Ren, Wenkai; Yin, Jie; Tan, Bie; Liu, Gang; Li, Lili; Nyachoti, Charles Martin; Xiong, Xia; Wu, Guoyao

    2014-01-01

    The mycotoxin deoxynivalenol (DON), one of the most common food contaminants, primarily targets the gastrointestinal tract to affect animal and human health. This study was conducted to examine the protective function of glutamic acid on intestinal injury and oxidative stress caused by DON in piglets. Twenty-eight piglets were assigned randomly into 4 dietary treatments (7 pigs/treatment): 1) uncontaminated control diet (NC), 2) NC+DON at 4 mg/kg (DON), 3) NC+2% glutamic acid (GLU), and 4) NC+2% glutamic acid + DON at 4 mg/kg (DG). At day 15, 30 and 37, blood samples were collected to determine serum concentrations of CAT (catalase), T-AOC (total antioxidant capacity), H2O2 (hydrogen peroxide), NO (nitric oxide), MDA (maleic dialdehyde), DAO (diamine oxidase) and D-lactate. Intestinal morphology, and the activation of Akt/mTOR/4EBP1 signal pathway, as well as the concentrations of H2O2, MDA, and DAO in kidney, liver and small intestine, were analyzed at day 37. Results showed that DON significantly (P<0.05) induced oxidative stress in piglets, while this stress was remarkably reduced with glutamic acid supplementation according to the change of oxidative parameters in blood and tissues. Meanwhile, DON caused obvious intestinal injury from microscopic observations and permeability indicators, which was alleviated by glutamic acid supplementation. Moreover, the inhibition of DON on Akt/mTOR/4EBP1 signal pathway was reduced by glutamic acid supplementation. Collectively, these data suggest that glutamic acid may be a useful nutritional regulator for DON-induced damage manifested as oxidative stress, intestinal injury and signaling inhibition. PMID:24984001

  3. HIV-1, Methamphetamine and Astrocyte Glutamate Regulation: Combined Excitotoxic Implications for Neuro-AIDS

    PubMed Central

    Cisneros, Irma E; Ghorpade, Anuja

    2012-01-01

    Glutamate, the most abundant excitatory transmitter in the brain can lead to neurotoxicity when not properly regulated. Excitotoxicity is a direct result of abnormal regulation of glutamate concentrations in the synapse, and is a common neurotoxic mediator associated with neurodegenerative disorders. It is well accepted that methamphetamine (METH), a potent central nervous stimulant with high abuse potential, and human immunodeficiency virus (HIV)-1 are implicated in the progression of neurocognitive malfunction. Both have been shown to induce common neurodegenerative effects such as astrogliosis, compromised blood brain barrier integrity, and excitotoxicity in the brain. Reduced glutamate uptake from neuronal synapses likely leads to the accumulation of glutamate in the extracellular spaces. Astrocytes express the glutamate transporters responsible for majority of the glutamate uptake from the synapse, as well as for vesicular glutamate release. However, the cellular and molecular mechanisms of astrocyte-mediated excitotoxicity in the context of METH and HIV-1 are undefined. Topics reviewed include dysregulation of the glutamate transporters, specifically excitatory amino acid transporter-2, metabotropic glutamate receptor(s) expression and the release of glutamate by vesicular exocytosis. We also discuss glutamate concentration dysregulation through astrocytic expression of enzymes for glutamate synthesis and metabolism. Lastly, we discuss recent evidence of various astrocyte and neuron crosstalk mechanisms implicated in glutamate regulation. Astrocytes play an essential role in the neuropathologies associated with METH/HIV-1-induced excitotoxicity. We hope to shed light on common cellular and molecular pathways astrocytes share in glutamate regulation during drug abuse and HIV-1 infection. PMID:22591363

  4. HIV-1, methamphetamine and astrocyte glutamate regulation: combined excitotoxic implications for neuro-AIDS.

    PubMed

    Cisneros, Irma E; Ghorpade, Anuja

    2012-07-01

    Glutamate, the most abundant excitatory transmitter in the brain can lead to neurotoxicity when not properly regulated. Excitotoxicity is a direct result of abnormal regulation of glutamate concentrations in the synapse, and is a common neurotoxic mediator associated with neurodegenerative disorders. It is well accepted that methamphetamine (METH), a potent central nervous stimulant with high abuse potential, and human immunodeficiency virus (HIV)-1 are implicated in the progression of neurocognitive malfunction. Both have been shown to induce common neurodegenerative effects such as astrogliosis, compromised blood brain barrier integrity, and excitotoxicity in the brain. Reduced glutamate uptake from neuronal synapses likely leads to the accumulation of glutamate in the extracellular spaces. Astrocytes express the glutamate transporters responsible for majority of the glutamate uptake from the synapse, as well as for vesicular glutamate release. However, the cellular and molecular mechanisms of astrocyte-mediated excitotoxicity in the context of METH and HIV-1 are undefined. Topics reviewed include dysregulation of the glutamate transporters, specifically excitatory amino acid transporter-2, metabotropic glutamate receptor(s) expression and the release of glutamate by vesicular exocytosis. We also discuss glutamate concentration dysregulation through astrocytic expression of enzymes for glutamate synthesis and metabolism. Lastly, we discuss recent evidence of various astrocyte and neuron crosstalk mechanisms implicated in glutamate regulation. Astrocytes play an essential role in the neuropathologies associated with METH/HIV-1-induced excitotoxicity. We hope to shed light on common cellular and molecular pathways astrocytes share in glutamate regulation during drug abuse and HIV-1 infection.

  5. Connexin expression in renin-producing cells.

    PubMed

    Kurtz, Lisa; Janssen-Bienhold, Ulrike; Kurtz, Armin; Wagner, Charlotte

    2009-03-01

    Absence of connexin 40 (Cx40) leads to ectopic juxtaglomerular renin expression and abrogates recruitment of renin-expressing cells in the adult kidney but does not disturb renin expression during kidney development. To find an explanation for these observations, we aimed to analyze the expression pattern of major vascular Cxs in normal juxtaglomerular epithelioid cells, in recruited renin-expressing cells, and in fetal renin-expressing cells. We found that during kidney development, the appearance of renin-producing cells paralleled the expression of Cx40 and, to a lesser extent, Cx45 but not other Cxs. In the adult kidney, juxtaglomerular epithelioid cells expressed Cx40 and lesser amounts of Cx37 and Cx43 but not Cx45, which localized to arteriolar smooth muscle cells. Recruitment of renin-producing cells in adult kidneys in response to long-term salt deprivation of mice correlated with the reappearance of only Cx40. Cx40-null renin-producing cells did not express Cx37, Cx43, or Cx45. These findings suggest that Cx40 expression is a characteristic of renin-producing cells in the kidney, and it seems to be essential in the recruitment of renin-producing cells in the adult but not the fetal kidney. PMID:19073828

  6. Electrogenic Steps Associated with Substrate Binding to the Neuronal Glutamate Transporter EAAC1.

    PubMed

    Tanui, Rose; Tao, Zhen; Silverstein, Nechama; Kanner, Baruch; Grewer, Christof

    2016-05-27

    Glutamate transporters actively take up glutamate into the cell, driven by the co-transport of sodium ions down their transmembrane concentration gradient. It was proposed that glutamate binds to its binding site and is subsequently transported across the membrane in the negatively charged form. With the glutamate binding site being located partially within the membrane domain, the possibility has to be considered that glutamate binding is dependent on the transmembrane potential and, thus, is electrogenic. Experiments presented in this report test this possibility. Rapid application of glutamate to the wild-type glutamate transporter subtype EAAC1 (excitatory amino acid carrier 1) through photo-release from caged glutamate generated a transient inward current, as expected for the electrogenic inward movement of co-transported Na(+) In contrast, glutamate application to a transporter with the mutation A334E induced transient outward current, consistent with movement of negatively charged glutamate into its binding site within the dielectric of the membrane. These results are in agreement with electrostatic calculations, predicting a valence for glutamate binding of -0.27. Control experiments further validate and rule out other possible explanations for the transient outward current. Electrogenic glutamate binding can be isolated in the mutant glutamate transporter because reactions, such as glutamate translocation and/or Na(+) binding to the glutamate-bound state, are inhibited by the A334E substitution. Electrogenic glutamate binding has to be considered together with other voltage-dependent partial reactions to cooperatively determine the voltage dependence of steady-state glutamate uptake and glutamate buffering at the synapse. PMID:27044739

  7. Mediator-less highly sensitive voltammetric detection of glutamate using glutamate dehydrogenase/vertically aligned CNTs grown on silicon substrate.

    PubMed

    Gholizadeh, Azam; Shahrokhian, Saeed; zad, Azam Iraji; Mohajerzadeh, Shamsoddin; Vosoughi, Manouchehr; Darbari, Sara; Sanaee, Zeinab

    2012-01-15

    A sensitive glutamate biosensor is prepared based on glutamate dehydrogenase/vertically aligned carbon nanotubes (GLDH, VACNTs). Vertically aligned carbon nanotubes were grown on a silicon substrate by direct current plasma enhanced chemical vapor deposition (DC-PECVD) method. The electrochemical behavior of the synthesized VACNTs was investigated by cyclic voltammetry and electrochemical impedance spectroscopic methods. Glutamate dehydrogenase covalently attached on tip of VACNTs. The electrochemical performance of the electrode for detection of glutamate was investigated by cyclic and differential pulse voltammetry. Differential pulse voltammetric determinations of glutamate are performed in mediator-less condition and also, in the presence of 1 and 5 μM thionine as electron mediator. The linear calibration curve of the concentration of glutamate versus peak current is investigated in a wide range of 0.1-500 μM. The mediator-less biosensor has a low detection limit of 57 nM and two linear ranges of 0.1-20 μM with a sensitivity of 0.976 mA mM(-1) cm(-2) and 20-300 μM with a sensitivity of 0.182 mA mM(-1) cm(-2). In the presence of 1 μM thionine as an electron mediator, the prepared biosensor shows a low detection limit of 68 nM and two linear ranges of 0.1-20 with a calibration sensitivity of 1.17 mA mM(-1) cm(-2) and 20-500 μM with a sensitivity of 0.153 mA mM(-1) cm(-2). The effects of the other biological compounds on the voltammetric behavior of the prepared biosensor and its response stability are investigated. The results are demonstrated that the GLDH/VACNTs electrode even without electron mediator is a suitable basic electrode for detection of glutamate. PMID:22040749

  8. Methylphenidate Increases Glutamate Uptake in Bergmann Glial Cells.

    PubMed

    Guillem, Alain M; Martínez-Lozada, Zila; Hernández-Kelly, Luisa C; López-Bayghen, Esther; López-Bayghen, Bruno; Calleros, Oscar A; Campuzano, Marco R; Ortega, Arturo

    2015-11-01

    Glutamate, the main excitatory transmitter in the vertebrate brain, exerts its actions through the activation of specific membrane receptors present in neurons and glial cells. Over-stimulation of glutamate receptors results in neuronal death, phenomena known as excitotoxicity. A family of glutamate uptake systems, mainly expressed in glial cells, removes the amino acid from the synaptic cleft preventing an excessive glutamatergic stimulation and thus neuronal damage. Autism spectrum disorders comprise a group of syndromes characterized by impaired social interactions and anxiety. One or the most common drugs prescribed to treat these disorders is Methylphenidate, known to increase dopamine extracellular levels, although it is not clear if its sedative effects are related to a plausible regulation of the glutamatergic tone via the regulation of the glial glutamate uptake systems. To gain insight into this possibility, we used the well-established model system of cultured chick cerebellum Bergmann glia cells. A time and dose-dependent increase in the activity and protein levels of glutamate transporters was detected upon Methylphenidate exposure. Interestingly, this increase is the result of an augmentation of both the synthesis as well as the insertion of these protein complexes in the plasma membrane. These results favour the notion that glial cells are Methylphenidate targets, and that by these means could regulate dopamine turnover.

  9. Ghrelin Regulates Glucose and Glutamate Transporters in Hypothalamic Astrocytes

    PubMed Central

    Fuente-Martín, Esther; García-Cáceres, Cristina; Argente-Arizón, Pilar; Díaz, Francisca; Granado, Miriam; Freire-Regatillo, Alejandra; Castro-González, David; Ceballos, María L.; Frago, Laura M.; Dickson, Suzanne L.; Argente, Jesús; Chowen, Julie A.

    2016-01-01

    Hypothalamic astrocytes can respond to metabolic signals, such as leptin and insulin, to modulate adjacent neuronal circuits and systemic metabolism. Ghrelin regulates appetite, adiposity and glucose metabolism, but little is known regarding the response of astrocytes to this orexigenic hormone. We have used both in vivo and in vitro approaches to demonstrate that acylated ghrelin (acyl-ghrelin) rapidly stimulates glutamate transporter expression and glutamate uptake by astrocytes. Moreover, acyl-ghrelin rapidly reduces glucose transporter (GLUT) 2 levels and glucose uptake by these glial cells. Glutamine synthetase and lactate dehydrogenase decrease, while glycogen phosphorylase and lactate transporters increase in response to acyl-ghrelin, suggesting a change in glutamate and glucose metabolism, as well as glycogen storage by astrocytes. These effects are partially mediated through ghrelin receptor 1A (GHSR-1A) as astrocytes do not respond equally to desacyl-ghrelin, an isoform that does not activate GHSR-1A. Moreover, primary astrocyte cultures from GHSR-1A knock-out mice do not change glutamate transporter or GLUT2 levels in response to acyl-ghrelin. Our results indicate that acyl-ghrelin may mediate part of its metabolic actions through modulation of hypothalamic astrocytes and that this effect could involve astrocyte mediated changes in local glucose and glutamate metabolism that alter the signals/nutrients reaching neighboring neurons. PMID:27026049

  10. 4-Hydroxyhexenal (HHE) impairs glutamate transport in astrocyte cultures.

    PubMed

    Lovell, Mark A; Bradley, Melissa A; Fister, Shuling X

    2012-01-01

    Multiple studies show elevations of α,β-unsaturated aldehydic by-products of lipid peroxidation including 4-hydroxynonenal and acrolein in vulnerable brain regions of subjects throughout the progression of Alzheimer's disease (AD). More recently 4-hydroxyhexenal (HHE), a diffusible α,β-unsaturated aldehyde resulting from peroxidation of ω-3 polyunsaturated fatty acids, was shown to be elevated in the hippocampus/parahippocampal gyrus (HPG) of subjects with preclinical AD (PCAD) and in late stage AD (LAD). HHE treatment of primary rat cortical neuron cultures led to a time- and concentration-dependent decrease in survival and glucose uptake. To determine if HHE also impairs glutamate uptake, primary rat astrocyte cultures were exposed to HHE for 4 hours and glutamate transport measured. Results show subtoxic (2.5 μM) HHE concentrations significantly (p < 0.05) impair glutamate uptake in primary astrocytes. Immunoprecipitation of excitatory amino acid transporter-2 (EAAT-2), the primary glutamate transporter in brain, from normal control, mild cognitive impairment (MCI), PCAD, and LAD HPG followed by quantification of HHE immunolabeling showed a significant increase in HHE positive EAAT-2 in MCI and LAD HPG. Together these data suggest HHE can significantly impair glutamate uptake and may play a role in the pathogenesis of AD. PMID:22766736

  11. Ghrelin Regulates Glucose and Glutamate Transporters in Hypothalamic Astrocytes.

    PubMed

    Fuente-Martín, Esther; García-Cáceres, Cristina; Argente-Arizón, Pilar; Díaz, Francisca; Granado, Miriam; Freire-Regatillo, Alejandra; Castro-González, David; Ceballos, María L; Frago, Laura M; Dickson, Suzanne L; Argente, Jesús; Chowen, Julie A

    2016-03-30

    Hypothalamic astrocytes can respond to metabolic signals, such as leptin and insulin, to modulate adjacent neuronal circuits and systemic metabolism. Ghrelin regulates appetite, adiposity and glucose metabolism, but little is known regarding the response of astrocytes to this orexigenic hormone. We have used both in vivo and in vitro approaches to demonstrate that acylated ghrelin (acyl-ghrelin) rapidly stimulates glutamate transporter expression and glutamate uptake by astrocytes. Moreover, acyl-ghrelin rapidly reduces glucose transporter (GLUT) 2 levels and glucose uptake by these glial cells. Glutamine synthetase and lactate dehydrogenase decrease, while glycogen phosphorylase and lactate transporters increase in response to acyl-ghrelin, suggesting a change in glutamate and glucose metabolism, as well as glycogen storage by astrocytes. These effects are partially mediated through ghrelin receptor 1A (GHSR-1A) as astrocytes do not respond equally to desacyl-ghrelin, an isoform that does not activate GHSR-1A. Moreover, primary astrocyte cultures from GHSR-1A knock-out mice do not change glutamate transporter or GLUT2 levels in response to acyl-ghrelin. Our results indicate that acyl-ghrelin may mediate part of its metabolic actions through modulation of hypothalamic astrocytes and that this effect could involve astrocyte mediated changes in local glucose and glutamate metabolism that alter the signals/nutrients reaching neighboring neurons.

  12. [Glutamate receptor-mediated retinal neuronal injury in experimental glaucoma].

    PubMed

    Wang, Zhong-Feng; Yang, Xiong-Li

    2016-08-25

    Glaucoma, the second leading cause of blindness, is a neurodegenerative disease characterized by optic nerve degeneration related to apoptotic death of retinal ganglion cells (RGCs). In the pathogenesis of RGC death following the onset of glaucoma, functional changes of glutamate receptors are commonly regarded as important risk factors. During the past several years, we have explored the mechanisms underlying RGC apoptosis and retinal Müller cell reactivation (gliosis) in a rat chronic ocular hypertension (COH) model. We demonstrated that elevated intraocular pressure in COH rats may induce changes of various signaling pathways, which are involved in RGC apoptosis by modulating glutamate NMDA and AMPA receptors. Moreover, we also demonstrated that over-activation of group I metabotropic glutamate receptors (mGluR I) by excessive extracellular glutamate in COH rats could contribute to Müller cell gliosis by suppressing Kir4.1 channels. In this review, incorporating our results, we discuss glutamate receptor- mediated RGC apoptosis and Müller cell gliosis in experimental glaucoma. PMID:27546508

  13. Ionotropic glutamate receptor expression in human white matter.

    PubMed

    Christensen, Pia Crone; Samadi-Bahrami, Zahra; Pavlov, Vlady; Stys, Peter K; Moore, G R Wayne

    2016-09-01

    Glutamate is the key excitatory neurotransmitter of the central nervous system (CNS). Its role in human grey matter transmission is well understood, but this is less clear in white matter (WM). Ionotropic glutamate receptors (iGluR) are found on both neuronal cell bodies and glia as well as on myelinated axons in rodents, and rodent WM tissue is capable of glutamate release. Thus, rodent WM expresses many of the components of the traditional grey matter neuron-to-neuron synapse, but to date this has not been shown for human WM. We demonstrate the presence of iGluRs in human WM by immunofluorescence employing high-resolution spectral confocal imaging. We found that the obligatory N-methyl-d-aspartic acid (NMDA) receptor subunit GluN1 and the α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptor subunit GluA4 co-localized with myelin, oligodendroglial cell bodies and processes. Additionally, GluA4 colocalized with axons, often in distinct clusters. These findings may explain why human WM is vulnerable to excitotoxic events following acute insults such as stroke and traumatic brain injury and in more chronic inflammatory conditions such as multiple sclerosis (MS). Further exploration of human WM glutamate signalling could pave the way for developing future therapies modulating the glutamate-mediated damage in these and other CNS disorders. PMID:27443784

  14. Single rodent mesohabenular axons release glutamate and GABA

    PubMed Central

    Root, David H.; Mejias-Aponte, Carlos; Zhang, Shiliang; Wang, Huiling; Hoffman, Alexander F.; Lupica, Carl R.; Morales, Marisela

    2016-01-01

    The lateral habenula (LHb) is involved in reward, aversion, addiction, and depression, through descending interactions with several brain structures, including the ventral tegmental area (VTA). VTA provides reciprocal inputs to LHb, but their actions are unclear. Here we show that the majority of rat and mouse VTA neurons innervating LHb co-express markers for both glutamate-signaling (vesicular glutamate transporter 2, VGluT2) and GABA-signaling (glutamate decarboxylase, GAD; and vesicular GABA transporter, VGaT). A single axon from these mesohabenular neurons co-expresses VGluT2-protein and VGaT-protein, and surprisingly establishes symmetric and asymmetric synapses on LHb neurons. In LHb slices, light activation of mesohabenular fibers expressing channelrhodopsin-2 (ChR2) driven by VGluT2 or VGaT promoters elicits release of both glutamate and GABA onto single LHb neurons. In vivo light-activation of mesohabenular terminals inhibits or excites LHb neurons. Our findings reveal an unanticipated type of VTA neuron that co-transmits glutamate and GABA, and provides the majority of mesohabenular inputs. PMID:25242304

  15. Frontal glutamate and reward processing in adolescence and adulthood.

    PubMed

    Gleich, Tobias; Lorenz, Robert C; Pöhland, Lydia; Raufelder, Diana; Deserno, Lorenz; Beck, Anne; Heinz, Andreas; Kühn, Simone; Gallinat, Jürgen

    2015-11-01

    The fronto-limbic network interaction, driven by glutamatergic and dopaminergic neurotransmission, represents a core mechanism of motivated behavior and personality traits. Reward seeking behavior undergoes tremendous changes in adolescence paralleled by neurobiological changes of this network including the prefrontal cortex, striatum and amygdala. Since fronto-limbic dysfunctions also underlie major psychiatric diseases beginning in adolescence, this investigation focuses on network characteristics separating adolescents from adults. To investigate differences in network interactions, the brain reward system activity (slot machine task) together with frontal glutamate concentration (anterior cingulate cortex, ACC) was measured in 28 adolescents and 26 adults employing functional magnetic resonance imaging and magnetic resonance spectroscopy, respectively. An inverse coupling of glutamate concentrations in the ACC and activation of the ventral striatum was observed in adolescents. Further, amygdala response in adolescents was negatively correlated with the personality trait impulsivity. For adults, no significant associations of network components or correlations with impulsivity were found. The inverse association between frontal glutamate concentration and striatal activation in adolescents is in line with the triadic model of motivated behavior stressing the important role of frontal top-down inhibition on limbic structures. Our data identified glutamate as the mediating neurotransmitter of this inhibitory process and demonstrates the relevance of glutamate on the reward system and related behavioral traits like impulsivity. This fronto-limbic coupling may represent a vulnerability factor for psychiatric disorders starting in adolescence but not in adulthood.

  16. The role of glutamate on the action of antidepressants.

    PubMed

    Hashimoto, Kenji

    2011-08-15

    Major depressive disorder (MDD) is a common, chronic, recurrent mental illness that affects millions of individuals worldwide. Currently available antidepressants are known to affect the monoaminergic (e.g., serotonin, norepinephrine, and dopamine) systems in the brain. Accumulating evidence suggests that the glutamatergic neurotransmission via the excitatory amino acid glutamate also plays an important role in the neurobiology and treatment of this disease. Clinical studies have demonstrated that the non-competitive N-methyl-d-aspartate (NMDA) receptor antagonist ketamine has rapid antidepressant effects in treatment-resistant patients with MDD, suggesting the role of glutamate in the pathophysiology of treatment-resistant MDD. Furthermore, a number of preclinical studies demonstrated that the agents which act at glutamate receptors such as NMDA receptors, α-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) receptors and metabotropic glutamate receptors (mGluRs) might have antidepressant-like activities in animal models of depression. In this article, the author reviews the role of glutamate in the neuron-glia communication induced by potential antidepressants.

  17. Central Role of Glutamate Metabolism in the Maintenance of Nitrogen Homeostasis in Normal and Hyperammonemic Brain

    PubMed Central

    Cooper, Arthur J. L.; Jeitner, Thomas M.

    2016-01-01

    Glutamate is present in the brain at an average concentration—typically 10–12 mM—far in excess of those of other amino acids. In glutamate-containing vesicles in the brain, the concentration of glutamate may even exceed 100 mM. Yet because glutamate is a major excitatory neurotransmitter, the concentration of this amino acid in the cerebral extracellular fluid must be kept low—typically µM. The remarkable gradient of glutamate in the different cerebral compartments: vesicles > cytosol/mitochondria > extracellular fluid attests to the extraordinary effectiveness of glutamate transporters and the strict control of enzymes of glutamate catabolism and synthesis in well-defined cellular and subcellular compartments in the brain. A major route for glutamate and ammonia removal is via the glutamine synthetase (glutamate ammonia ligase) reaction. Glutamate is also removed by conversion to the inhibitory neurotransmitter γ-aminobutyrate (GABA) via the action of glutamate decarboxylase. On the other hand, cerebral glutamate levels are maintained by the action of glutaminase and by various α-ketoglutarate-linked aminotransferases (especially aspartate aminotransferase and the mitochondrial and cytosolic forms of the branched-chain aminotransferases). Although the glutamate dehydrogenase reaction is freely reversible, owing to rapid removal of ammonia as glutamine amide, the direction of the glutamate dehydrogenase reaction in the brain in vivo is mainly toward glutamate catabolism rather than toward the net synthesis of glutamate, even under hyperammonemia conditions. During hyperammonemia, there is a large increase in cerebral glutamine content, but only small changes in the levels of glutamate and α-ketoglutarate. Thus, the channeling of glutamate toward glutamine during hyperammonemia results in the net synthesis of 5-carbon units. This increase in 5-carbon units is accomplished in part by the ammonia-induced stimulation of the anaplerotic enzyme pyruvate

  18. Central Role of Glutamate Metabolism in the Maintenance of Nitrogen Homeostasis in Normal and Hyperammonemic Brain.

    PubMed

    Cooper, Arthur J L; Jeitner, Thomas M

    2016-01-01

    Glutamate is present in the brain at an average concentration-typically 10-12 mM-far in excess of those of other amino acids. In glutamate-containing vesicles in the brain, the concentration of glutamate may even exceed 100 mM. Yet because glutamate is a major excitatory neurotransmitter, the concentration of this amino acid in the cerebral extracellular fluid must be kept low-typically µM. The remarkable gradient of glutamate in the different cerebral compartments: vesicles > cytosol/mitochondria > extracellular fluid attests to the extraordinary effectiveness of glutamate transporters and the strict control of enzymes of glutamate catabolism and synthesis in well-defined cellular and subcellular compartments in the brain. A major route for glutamate and ammonia removal is via the glutamine synthetase (glutamate ammonia ligase) reaction. Glutamate is also removed by conversion to the inhibitory neurotransmitter γ-aminobutyrate (GABA) via the action of glutamate decarboxylase. On the other hand, cerebral glutamate levels are maintained by the action of glutaminase and by various α-ketoglutarate-linked aminotransferases (especially aspartate aminotransferase and the mitochondrial and cytosolic forms of the branched-chain aminotransferases). Although the glutamate dehydrogenase reaction is freely reversible, owing to rapid removal of ammonia as glutamine amide, the direction of the glutamate dehydrogenase reaction in the brain in vivo is mainly toward glutamate catabolism rather than toward the net synthesis of glutamate, even under hyperammonemia conditions. During hyperammonemia, there is a large increase in cerebral glutamine content, but only small changes in the levels of glutamate and α-ketoglutarate. Thus, the channeling of glutamate toward glutamine during hyperammonemia results in the net synthesis of 5-carbon units. This increase in 5-carbon units is accomplished in part by the ammonia-induced stimulation of the anaplerotic enzyme pyruvate carboxylase

  19. [Features of glutamate dehydrogenase in fetal and adult rumen tissue].

    PubMed

    Kalachniuk, H I; Fomenko, I S; Kalachniuk, L H; Kavai, Sh; Marounek, M; Savka, O H

    2001-01-01

    Glutamate dehydrogenase (GDH) from rumen mucosa of cow fetus, liver and two forms from mucosa (bacterial and tissue) of the adult animal were partly purified and characterized. The activity of the bacterial glutamate dehydrogenase was shown to depend on qualities of a biomass of microbes, adhered on surface of rumen mucosa. All enzymes from tissues (GDHTRF, TRC, TLC), revealed the hypersensibility to increase in the concentration medium of Zn2+, guanosine triphosphate (GTP), acting here in a role of negative modulators, and also adenosine monophosphate (AMP) and leucine, which acted as activators. However, in the same concentrations these effectors do not influence the activity of the bacterial glutamate dehydrogenase. And if all tissues enzymes are highly specific to coenzyme NADH, the bacterial ones almost in 3 times is more active at NADPH use. PMID:11642036

  20. Aminotransferase and glutamate dehydrogenase activities in lactobacilli and streptococci.

    PubMed

    Peralta, Guillermo Hugo; Bergamini, Carina Viviana; Hynes, Erica Rut

    2016-01-01

    Aminotransferases and glutamate dehydrogenase are two main types of enzymes involved in the initial steps of amino acid catabolism, which plays a key role in the cheese flavor development. In the present work, glutamate dehydrogenase and aminotransferase activities were screened in twenty one strains of lactic acid bacteria of dairy interest, either cheese-isolated or commercial starters, including fifteen mesophilic lactobacilli, four thermophilic lactobacilli, and two streptococci. The strains of Streptococcus thermophilus showed the highest glutamate dehydrogenase activity, which was significantly elevated compared with the lactobacilli. Aspartate aminotransferase prevailed in most strains tested, while the levels and specificity of other aminotransferases were highly strain- and species-dependent. The knowledge of enzymatic profiles of these starter and cheese-isolated cultures is helpful in proposing appropriate combinations of strains for improved or increased cheese flavor. PMID:27266631

  1. [PECULIARITIES OF THE CEREBROVASCULAR EFFECTS OF GLUTAMIC ACID].

    PubMed

    Gan'shina, T S; Kurza, E V; Kurdyumov, I N; Maslennikov, D V; Mirzoyan, R S

    2016-01-01

    Experiments on nonlinear rats subjected to global transient cerebral ischemia revealed the ability of glutamic acid to improve cerebral circulation. Consequently, the excitatory amino acid can produce adverse (neurotoxic) and positive (anti-ischemic) effects in cerebral ischemia. The cerebrovascular effect of glutamic acid in cerebral ischemia is attenuated on the background action of the MNDA receptor blocker MK-801 (0.5 mg/kg intravenously) and eliminated by bicuculline. When glutamic acid is combined with the non-competitive MNDA receptor antagonist MK-801, neither one nor another drug shows its vasodilator effect. The results are indicative of the interaction between excitatory and inhibitory systems on the level of cerebral vessels and once again confirm our previous conclusion about the decisive role of GABA(A) receptors in brain vessels in the implementation of anti-ischemic activity of endogenous compounds (melatonin) and well-known pharmacological substances (mexidol, afobazole), and new chemical compounds based on GABA-containing lipid derivatives.

  2. Glutamate transporters in brain ischemia: to modulate or not?

    PubMed Central

    Krzyżanowska, Weronika; Pomierny, Bartosz; Filip, Małgorzata; Pera, Joanna

    2014-01-01

    In this review, we briefly describe glutamate (Glu) metabolism and its specific transports and receptors in the central nervous system (CNS). Thereafter, we focus on excitatory amino acid transporters, cystine/glutamate antiporters (system xc-) and vesicular glutamate transporters, specifically addressing their location and roles in CNS and the molecular mechanisms underlying the regulation of Glu transporters. We provide evidence from in vitro or in vivo studies concerning alterations in Glu transporter expression in response to hypoxia or ischemia, including limited human data that supports the role of Glu transporters in stroke patients. Moreover, the potential to induce brain tolerance to ischemia through modulation of the expression and/or activities of Glu transporters is also discussed. Finally we present strategies involving the application of ischemic preconditioning and pharmacological agents, eg β-lactam antibiotics, amitriptyline, riluzole and N-acetylcysteine, which result in the significant protection of nervous tissues against ischemia. PMID:24681894

  3. Aminotransferase and glutamate dehydrogenase activities in lactobacilli and streptococci.

    PubMed

    Peralta, Guillermo Hugo; Bergamini, Carina Viviana; Hynes, Erica Rut

    2016-01-01

    Aminotransferases and glutamate dehydrogenase are two main types of enzymes involved in the initial steps of amino acid catabolism, which plays a key role in the cheese flavor development. In the present work, glutamate dehydrogenase and aminotransferase activities were screened in twenty one strains of lactic acid bacteria of dairy interest, either cheese-isolated or commercial starters, including fifteen mesophilic lactobacilli, four thermophilic lactobacilli, and two streptococci. The strains of Streptococcus thermophilus showed the highest glutamate dehydrogenase activity, which was significantly elevated compared with the lactobacilli. Aspartate aminotransferase prevailed in most strains tested, while the levels and specificity of other aminotransferases were highly strain- and species-dependent. The knowledge of enzymatic profiles of these starter and cheese-isolated cultures is helpful in proposing appropriate combinations of strains for improved or increased cheese flavor.

  4. Inflammation, glutamate, and glia in depression: a literature review.

    PubMed

    McNally, Leah; Bhagwagar, Zubin; Hannestad, Jonas

    2008-06-01

    Multiple lines of evidence suggest that inflammation and glutamate dysfunction contribute to the pathophysiology of depression. In this review we provide an overview of how these two systems may interact. Excess levels of inflammatory mediators occur in a subgroup of depressed patients. Studies of acute experimental activation of the immune system with endotoxin and of chronic activation during interferon-alpha treatment show that inflammation can cause depression. Peripheral inflammation leads to microglial activation which could interfere with excitatory amino acid metabolism leading to inappropriate glutamate receptor activation. Loss of astroglia, a feature of depression, upsets the balance of anti- and pro-inflammatory mediators and further impairs the removal of excitatory amino acids. Microglia activated by excess inflammation, astroglial loss, and inappropriate glutamate receptor activation ultimately disrupt the delicate balance of neuroprotective versus neurotoxic effects in the brain, potentially leading to depression. PMID:18567974

  5. Enzymatic production of α-ketoglutaric acid from l-glutamic acid via l-glutamate oxidase.

    PubMed

    Niu, Panqing; Dong, Xiaoxiang; Wang, Yuancai; Liu, Liming

    2014-06-10

    In this study, a novel strategy for α-ketoglutaric acid (α-KG) production from l-glutamic acid using recombinant l-glutamate oxidase (LGOX) was developed. First, by analyzing the molecular structure characteristics of l-glutamic acid and α-KG, LGOX was found to be the best catalyst for oxidizing the amino group of l-glutamic acid to a ketonic group without the need for exogenous cofactor. Then the LGOX gene was expressed in Escherichia coli BL21 (DE3) in a soluble and active form, and the recombinant LGOX activity reached to a maximum value of 0.59U/mL at pH 6.5, 30°C. Finally, the maximum α-KG concentration reached 104.7g/L from 110g/L l-glutamic acid in 24h, under the following optimum conditions: 1.5U/mL LGOX, 250U/mL catalase, 3mM MnCl2, 30°C, and pH 6.5.

  6. Glutamate Signaling in Benign and Malignant Disorders: Current Status, Future Perspectives, and Therapeutic Implications

    PubMed Central

    Willard, Stacey S.; Koochekpour, Shahriar

    2013-01-01

    Glutamate, a nonessential amino acid, is the major excitatory neurotransmitter in the central nervous system. As such, glutamate has been shown to play a role in not only neural processes, such as learning and memory, but also in bioenergetics, biosynthetic and metabolic oncogenic pathways. Glutamate has been the target of intense investigation for its involvement not only in the pathogenesis of benign neurodegenerative diseases (NDDs) such as Parkinson's disease, Alzheimer's disease, schizophrenia, multiple sclerosis, and amyotropic lateral sclerosis (ALS), but also in carcinogenesis and progression of malignant diseases. In addition to its intracellular activities, glutamate in secreted form is a phylogenetically conserved cell signaling molecule. Glutamate binding activates multiple major receptor families including the metabotropic glutamate receptors (mGluRs) and ionotropic glutamate receptors (iGluRs), both of which have been implicated in various signaling pathways in cancer. Inhibition of extracellular glutamate release or glutamate receptor activation via competitive or non-competitive antagonists decreases growth, migration and invasion and induces apoptosis in breast cancer, melanoma, glioma and prostate cancer cells. In this review, we discuss the current state of glutamate signaling research as it relates to benign and malignant diseases. In addition, we provide a synopsis of clinical trials using glutamate antagonists for the treatment of NDD and malignant diseases. We conclude that in addition to its potential role as a metabolic biomarker, glutamate receptors and glutamate-initiated signaling pathways may provide novel therapeutic opportunities for cancer. PMID:23983606

  7. Electrogenic glutamate uptake in glial cells is activated by intracellular potassium

    NASA Astrophysics Data System (ADS)

    Barbour, Boris; Brew, Helen; Attwell, David

    1988-09-01

    Uptake of glutamate into glial cells in the CNS maintains the extracellular glutamate concentration below neurotoxic levels and helps terminate its action as a neurotransmitter 1. The co-transport of two sodium ions on the glutamate carrier is thought to provide the energy needed to transport glutamate into cells2,3. We have shown recently that glutamate uptake can be detected electrically because the excess of Na+ ions transported with each glutamate anion results in a net current flow into the cell4. We took advantage of the control of the environment, both inside and outside the cell, provided by whole-cell patch-clamping and now report that glutamate uptake is activated by intracellular potassium and inhibited by extracellular potassium. Our results indicate that one K+ ion is transported out of the cell each time a glutamate anion and three Na+ ions are transported in. A carrier with this stoichiometry can accumulate glutamate against a much greater concentration gradient than a carrier co-transporting one glutamate anion and two Na+ ions. Pathological rises in extracellular potassium concentration will inhibit glutamate uptake by depolarizing glial cells and by preventing the loss of K+ from the glutamate carrier. This will facilitate a rise in the extracellular glutamate concentration to neurotoxic levels and contribute to the neuronal death occurring in brain anoxia and ischaemia.

  8. Chronic Glutamate Toxicity in Neurodegenerative Diseases—What is the Evidence?

    PubMed Central

    Lewerenz, Jan; Maher, Pamela

    2015-01-01

    Together with aspartate, glutamate is the major excitatory neurotransmitter in the brain. Glutamate binds and activates both ligand-gated ion channels (ionotropic glutamate receptors) and a class of G-protein coupled receptors (metabotropic glutamate receptors). Although the intracellular glutamate concentration in the brain is in the millimolar range, the extracellular glutamate concentration is kept in the low micromolar range by the action of excitatory amino acid transporters that import glutamate and aspartate into astrocytes and neurons. Excess extracellular glutamate may lead to excitotoxicity in vitro and in vivo in acute insults like ischemic stroke via the overactivation of ionotropic glutamate receptors. In addition, chronic excitotoxicity has been hypothesized to play a role in numerous neurodegenerative diseases including amyotrophic lateral sclerosis, Alzheimer's disease and Huntington's disease. Based on this hypothesis, a good deal of effort has been devoted to develop and test drugs that either inhibit glutamate receptors or decrease extracellular glutamate. In this review, we provide an overview of the different pathways that are thought to lead to an over-activation of the glutamatergic system and glutamate toxicity in neurodegeneration. In addition, we summarize the available experimental evidence for glutamate toxicity in animal models of neurodegenerative diseases. PMID:26733784

  9. Glutamate Excitotoxicity Mediates Neuronal Apoptosis After Hypothermic Circulatory Arrest

    PubMed Central

    Tseng, Elaine E.; Brock, Malcolm V.; Lange, Mary S.; Troncoso, Juan C.; Blue, Mary E.; Lowenstein, Charles J.; Johnston, Michael V.; Baumgartner, William A.

    2011-01-01

    Background Prolonged hypothermic circulatory arrest results in neuronal cell death and neurologic injury. We have previously shown that hypothermic circulatory arrest causes both neuronal apoptosis and necrosis in a canine model. Inhibition of neuronal nitric oxide synthase reduced neuronal apoptosis, while glutamate receptor antagonism reduced necrosis in our model. This study was undertaken to determine whether glutamate receptor antagonism reduces nitric oxide formation and neuronal apoptosis after hypothermic circulatory arrest. Methods Sixteen hound dogs underwent 2 hours of circulatory arrest at 18°C and were sacrificed after 8 hours. Group 1 (n=8) was treated with MK-801, 0.75 mg/kg IV prior to arrest followed by 75 μg/kg/hr infusion. Group 2 dogs (n=8) received vehicle only. Intracerebral levels of excitatory amino acids and citrulline, an equal co-product of nitric oxide, were measured. Apoptosis, identified by H&E staining and confirmed by electron microscopy, was blindly scored from 0 (normal) to 100 (severe injury), while nick-end labeling demonstrated DNA fragmentation. Results Group 1 and 2 dogs had similar intracerebral levels of glutamate. However, MK-801 significantly reduced intracerebral glycine and citrulline levels as compared to HCA controls. MK-801 significantly inhibited apoptosis (7.92 ± 7.85 vs. 62.08 ± 6.28, Group 1 vs. 2, p<0.001). Conclusions Our results showed that glutamate receptor antagonism significantly reduced nitric oxide formation and neuronal apoptosis. We provide evidence that glutamate excitotoxicity mediates neuronal apoptosis in addition to necrosis after hypothermic circulatory arrest. Clinical glutamate receptor antagonists may have therapeutic benefit in ameliorating both types of neurologic injury after hypothermic circulatory arrest. PMID:20103318

  10. Glutamate-dopamine-GABA interactions in the aging basal ganglia.

    PubMed

    Mora, Francisco; Segovia, Gregorio; Del Arco, Alberto

    2008-08-01

    The study of neurotransmitter interactions gives a better understanding of the physiology of specific circuits in the brain. In this review we focus mostly on our own results on the interaction of the neurotransmitters glutamate, dopamine and GABA in the basal ganglia during the normal process of aging. We review first the studies on the action of endogenous glutamate on the extracellular concentrations of dopamine and GABA in the neostriatum and nucleus accumbens during aging. It was found that there exists an age-related change in the interaction of glutamate, dopamine and GABA and that these effects of aging exhibit a dorsal-to-ventral pattern of effects with no changes in the dorsal parts (dorsal striatum) and changes in the most ventral parts (nucleus accumbens). Second we reviewed the data on the effects of different ionotropic and metabotropic glutamate receptor agonists on the extracellular concentrations of dopamine and GABA in the nucleus accumbens. The results obtained clearly show the different contribution of each glutamate receptor subtype in the age-related changes produced on the interaction of glutamate, dopamine and GABA in this area of the brain. Third the effects of an enriched environment on the action of AMPA and NMDA-receptor agonists in the nucleus accumbens of rats during aging are also evaluated. Finally, and since the nucleus accumbens has been suggested to play a role in emotion and motivation and also motor behaviour, we speculated on the possibility of a specific contribution for the different glutamatergic pathways terminating in the nucleus accumbens and their interaction with a decreased dopamine playing a relevant role in motor behaviour during aging.

  11. [Enzymatic production of α-ketoglutaric acid by L-glutamate oxidase from L-glutamic acid].

    PubMed

    Niu, Panqing; Zhang, Zhenyu; Liu, Liming

    2014-08-01

    We produced α-ketoglutaric acid (α-KG) from L-glutamic acid, using enzymatic transformation approach with L-glutamate oxidase (LGOX). First, wild strain Streptomyces sp. FMME066 was mutated with NTG, a genetically stable mutant Streptomyces sp. FMME067 was obtained. Under the optimal nutrition conditions with fructose 10 g/L, peptone 7.5 g/L, KH2PO4 1 g/L and CaCl2 0.05 g/L, the maximum LGOX activity reached 0.14 U/mL. The LGOX was stable to pH and temperature, and Mn2+ had a stimulating effect. Finally, after 24 h enzymatic conversion under the optimal conditions, the maximum titer of α-KG reached 38.1 g/L from 47 g/L L-glutamic acid. Enzymatic transformation by LGOX is a potential approach for α-KG production.

  12. Cerebellar Ataxia and Glutamic Acid Decarboxylase Antibodies

    PubMed Central

    Ariño, Helena; Gresa-Arribas, Nuria; Blanco, Yolanda; Martínez-Hernández, Eugenia; Sabater, Lidia; Petit-Pedrol, Mar; Rouco, Idoia; Bataller, Luis; Dalmau, Josep O.; Saiz, Albert; Graus, Francesc

    2016-01-01

    IMPORTANCE Current clinical and immunologic knowledge on cerebellar ataxia (CA) with glutamic acid decarboxylase 65 antibodies (GAD65-Abs) is based on case reports and small series with short-term follow-up data. OBJECTIVE To report the symptoms, additional antibodies, prognostic factors, and long-term outcomes in a cohort of patients with CA and GAD65-Abs. DESIGN, SETTING, AND PARTICIPANTS Retrospective cohort study and laboratory investigations at a center for autoimmune neurologic disorders among 34 patients with CA and GAD65-Abs, including 25 with long-term follow-up data (median, 5.4 years; interquartile range, 3.1-10.3 years). MAIN OUTCOMES AND MEASURES Analysis of clinicoimmunologic features and predictors of response to immunotherapy. Immunochemistry on rat brain, cultured neurons, and human embryonic kidney cells expressing GAD65, GAD67, α1-subunit of the glycine receptor, and a repertoire of known cell surface autoantigens were used to identify additional antibodies. Twenty-eight patients with stiff person syndrome and GAD65-Abs served as controls. RESULTS The median age of patients was 58 years (range, 33-80 years); 28 of 34 patients (82%) were women. Nine patients (26%) reported episodes of brainstem and cerebellar dysfunction or persistent vertigo several months before developing CA. The clinical presentation was subacute during a period of weeks in 13 patients (38%). Nine patients (26%) had coexisting stiff person syndrome symptoms. Systemic organ-specific autoimmunities (type 1 diabetes mellitus and others) were present in 29 patients (85%). Twenty of 25 patients with long-term follow-up data received immunotherapy (intravenous immunoglobulin in 10 and corticosteroids and intravenous immunoglobulin or other immunosuppressors in 10), and 7 of them (35%) improved. Predictors of clinical response included subacute onset of CA (odds ratio [OR], 0.50; 95% CI, 0.25-0.99; P = .047) and prompt immunotherapy (OR, 0.98; 95% CI, 0.96-0.99; P = .01). Similar

  13. Rapid Microelectrode Measurements and the Origin and Regulation of Extracellular Glutamate in Rat Prefrontal Cortex

    PubMed Central

    Hascup, E.R.; Hascup, K.N.; Stephens, M.; Pomerleau, F.; Huettl, P.; Gratton, A.; Gerhardt, G.A.

    2010-01-01

    Glutamate in the prefrontal cortex (PFC) plays a significant role in several mental illnesses, including schizophrenia, addiction and anxiety. Previous studies on PFC glutamate-mediated function have used techniques that raise questions on the neuronal vs. astrocytic origin of glutamate. The present studies used enzyme-based microelectrode arrays (MEAs) to monitor second-by-second resting glutamate levels in the PFC of awake rats. Locally-applied drugs were employed in an attempt to discriminate between the neuronal or glial components of the resting glutamate signal. Local application of tetrodotoxin (TTX; sodium channel blocker), produced a significant (~40%) decline in resting glutamate levels. In addition significant reductions in extracellular glutamate were seen with locally-applied ω-conotoxin (MVIIC; ~50%; calcium channel blocker), and the mGluR⅔ agonist, LY379268 (~20%), and a significant increase with the mGluR⅔ antagonist LY341495 (~40%), effects all consistent with a large neuronal contribution to the resting glutamate levels. Local administration of D,L-threo-β-benzyloxyaspartate (TBOA; glutamate transporter inhibitor) produced an ~120% increase in extracellular glutamate levels, supporting that excitatory amino acid transporters, which are largely located on glia, modulate clearance of extracellular glutamate. Interestingly, local application of (S)-4-carboxyphenylglycine (CPG; cystine/glutamate antiporter inhibitor), produced small, non-significant bi-phasic changes in extracellular glutamate versus vehicle control. Finally, pre-administration of TTX completely blocked the glutamate response to tail pinch stress. Taken together, these results support that PFC resting glutamate levels in rats as measured by the MEA technology are at least 40-50% derived from neurons. Furthermore, these data support that the impulse flow-dependent glutamate release from a physiologically-evoked event is entirely neuronally derived. PMID:20969570

  14. Zinc-positive boutons in the cerebral cortex of lizards show glutamate immunoreactivity.

    PubMed

    Martinez-Guijarro, F J; Soriano, E; Del Rio, J A; Lopez-Garcia, C

    1991-10-01

    Zinc-positive boutons, originating in the medial cortex of lizards, exhibit glutamate immunoreactivity. This finding supports the presumed homology between lizard zinc-positive boutons and the hippocampal mossy fibres of mammals, which are also glutamate-immunoreactive and zinc-positive. Zinc-positive boutons of lizards contain a chelatable pool of zinc located in the hippocampal mossy fibres of mammals. These synaptic systems also contain glutamate, which indicates a possible simultaneous action of zinc and glutamate during synaptic transmission.

  15. Glutamate secretion and metabotropic glutamate receptor 1 expression during Kaposi's sarcoma-associated herpesvirus infection promotes cell proliferation.

    PubMed

    Valiya Veettil, Mohanan; Dutta, Dipanjan; Bottero, Virginie; Bandyopadhyay, Chirosree; Gjyshi, Olsi; Sharma-Walia, Neelam; Dutta, Sujoy; Chandran, Bala

    2014-10-01

    Kaposi's sarcoma associated herpesvirus (KSHV) is etiologically associated with endothelial Kaposi's sarcoma (KS) and B-cell proliferative primary effusion lymphoma (PEL), common malignancies seen in immunocompromised HIV-1 infected patients. The progression of these cancers occurs by the proliferation of cells latently infected with KSHV, which is highly dependent on autocrine and paracrine factors secreted from the infected cells. Glutamate and glutamate receptors have emerged as key regulators of intracellular signaling pathways and cell proliferation. However, whether they play any role in the pathological changes associated with virus induced oncogenesis is not known. Here, we report the first systematic study of the role of glutamate and its metabotropic glutamate receptor 1 (mGluR1) in KSHV infected cell proliferation. Our studies show increased glutamate secretion and glutaminase expression during de novo KSHV infection of endothelial cells as well as in KSHV latently infected endothelial and B-cells. Increased mGluR1 expression was detected in KSHV infected KS and PEL tissue sections. Increased c-Myc and glutaminase expression in the infected cells was mediated by KSHV latency associated nuclear antigen 1 (LANA-1). In addition, mGluR1 expression regulating host RE-1 silencing transcription factor/neuron restrictive silencer factor (REST/NRSF) was retained in the cytoplasm of infected cells. KSHV latent protein Kaposin A was also involved in the over expression of mGluR1 by interacting with REST in the cytoplasm of infected cells and by regulating the phosphorylation of REST and interaction with β-TRCP for ubiquitination. Colocalization of Kaposin A with REST was also observed in KS and PEL tissue samples. KSHV infected cell proliferation was significantly inhibited by glutamate release inhibitor and mGluR1 antagonists. These studies demonstrated that elevated glutamate secretion and mGluR1 expression play a role in KSHV induced cell proliferation and

  16. 40 CFR 721.3821 - L-Glutamic acid, N-(1-oxododecyl)-.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 30 2010-07-01 2010-07-01 false L-Glutamic acid, N-(1-oxododecyl... Substances § 721.3821 L-Glutamic acid, N-(1-oxododecyl)-. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as L-Glutamic acid, N-(1-oxododecyl)- (PMN...

  17. Unveiling the secret lives of glutamate transporters: VGLUTs engage in multiple transport modes.

    PubMed

    Accardi, Alessio

    2014-12-17

    Accumulation of glutamate in synaptic vesicles is mediated by vesicular glutamate transporters called VGLUTs. In the current issue of Neuron, Preobraschenski et al. (2014) show that the VGLUTs, in addition to transporting glutamate, also provide the conductances necessary to maintain the appropriate voltage and pH inside these vesicles. PMID:25521371

  18. 40 CFR 721.3821 - L-Glutamic acid, N-(1-oxododecyl)-.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 40 Protection of Environment 32 2012-07-01 2012-07-01 false L-Glutamic acid, N-(1-oxododecyl... Substances § 721.3821 L-Glutamic acid, N-(1-oxododecyl)-. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as L-Glutamic acid, N-(1-oxododecyl)- (PMN...

  19. 40 CFR 721.3821 - L-Glutamic acid, N-(1-oxododecyl)-.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 40 Protection of Environment 32 2013-07-01 2013-07-01 false L-Glutamic acid, N-(1-oxododecyl... Substances § 721.3821 L-Glutamic acid, N-(1-oxododecyl)-. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as L-Glutamic acid, N-(1-oxododecyl)- (PMN...

  20. 40 CFR 721.3821 - L-Glutamic acid, N-(1-oxododecyl)-.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 31 2011-07-01 2011-07-01 false L-Glutamic acid, N-(1-oxododecyl... Substances § 721.3821 L-Glutamic acid, N-(1-oxododecyl)-. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as L-Glutamic acid, N-(1-oxododecyl)- (PMN...

  1. 40 CFR 721.3821 - L-Glutamic acid, N-(1-oxododecyl)-.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 40 Protection of Environment 31 2014-07-01 2014-07-01 false L-Glutamic acid, N-(1-oxododecyl... Substances § 721.3821 L-Glutamic acid, N-(1-oxododecyl)-. (a) Chemical substance and significant new uses subject to reporting. (1) The chemical substance identified as L-Glutamic acid, N-(1-oxododecyl)- (PMN...

  2. Control of cortical neuronal migration by glutamate and GABA

    PubMed Central

    Luhmann, Heiko J.; Fukuda, A.; Kilb, W.

    2015-01-01

    Neuronal migration in the cortex is controlled by the paracrine action of the classical neurotransmitters glutamate and GABA. Glutamate controls radial migration of pyramidal neurons by acting primarily on NMDA receptors and regulates tangential migration of inhibitory interneurons by activating non-NMDA and NMDA receptors. GABA, acting on ionotropic GABAA-rho and GABAA receptors, has a dichotomic action on radially migrating neurons by acting as a GO signal in lower layers and as a STOP signal in upper cortical plate (CP), respectively. Metabotropic GABAB receptors promote radial migration into the CP and tangential migration of interneurons. Besides GABA, the endogenous GABAergic agonist taurine is a relevant agonist controlling radial migration. To a smaller extent glycine receptor activation can also influence radial and tangential migration. Activation of glutamate and GABA receptors causes increases in intracellular Ca2+ transients, which promote neuronal migration by acting on the cytoskeleton. Pharmacological or genetic manipulation of glutamate or GABA receptors during early corticogenesis induce heterotopic cell clusters in upper layers and loss of cortical lamination, i.e., neuronal migration disorders which can be associated with neurological or neuropsychiatric diseases. The pivotal role of NMDA and ionotropic GABA receptors in cortical neuronal migration is of major clinical relevance, since a number of drugs acting on these receptors (e.g., anti-epileptics, anesthetics, alcohol) may disturb the normal migration pattern when present during early corticogenesis. PMID:25688185

  3. Control of cortical neuronal migration by glutamate and GABA.

    PubMed

    Luhmann, Heiko J; Fukuda, A; Kilb, W

    2015-01-01

    Neuronal migration in the cortex is controlled by the paracrine action of the classical neurotransmitters glutamate and GABA. Glutamate controls radial migration of pyramidal neurons by acting primarily on NMDA receptors and regulates tangential migration of inhibitory interneurons by activating non-NMDA and NMDA receptors. GABA, acting on ionotropic GABAA-rho and GABAA receptors, has a dichotomic action on radially migrating neurons by acting as a GO signal in lower layers and as a STOP signal in upper cortical plate (CP), respectively. Metabotropic GABAB receptors promote radial migration into the CP and tangential migration of interneurons. Besides GABA, the endogenous GABAergic agonist taurine is a relevant agonist controlling radial migration. To a smaller extent glycine receptor activation can also influence radial and tangential migration. Activation of glutamate and GABA receptors causes increases in intracellular Ca(2+) transients, which promote neuronal migration by acting on the cytoskeleton. Pharmacological or genetic manipulation of glutamate or GABA receptors during early corticogenesis induce heterotopic cell clusters in upper layers and loss of cortical lamination, i.e., neuronal migration disorders which can be associated with neurological or neuropsychiatric diseases. The pivotal role of NMDA and ionotropic GABA receptors in cortical neuronal migration is of major clinical relevance, since a number of drugs acting on these receptors (e.g., anti-epileptics, anesthetics, alcohol) may disturb the normal migration pattern when present during early corticogenesis.

  4. Delineation of an in vivo inhibitor for Aspergillus glutamate dehydrogenase.

    PubMed

    Choudhury, Rajarshi; Noor, Shahid; Varadarajalu, Lakshmi Prabha; Punekar, Narayan S

    2008-01-01

    NADP-glutamate dehydrogenase (NADP-GDH) along with glutamine synthetase plays a pivotal role in ammonium assimilation. Specific inhibitors were valuable in defining the importance of glutamine synthetase in nitrogen metabolism. Selective in vivo inhibition of NADP-GDH has so far been an elusive desideratum. Isophthalate, a potent in vitro inhibitor of Aspergillus niger NADP-GDH [Noor S, Punekar NS. Allosteric NADP-glutamate dehydrogenase from aspergilli: purification, characterization and implications for metabolic regulation at the carbon-nitrogen interface. Microbiology 2005;151:1409-19], was evaluated for its efficacy in vivo. Dimethyl ester of isophthalate (DMIP), but not isophthalate, inhibited A. niger growth on agar as well as in liquid culture. This was ascribed to the inability of isophthalate to enter fungal mycelia. Subsequent to DMIP addition however, intracellular isophthalate could be demonstrated. Apart from NAD-GDH, no other enzyme including NAD-glutamate synthase was inhibited by isophthalate. A cross-over at NADP-GDH step of metabolism was observed as a direct consequence of isophthalate (formed in vivo from DMIP) inhibiting this enzyme. Addition of ammonium to DMIP-treated A. niger mycelia resulted in intensive vacuolation, retraction of cytoplasm and autolysis. Taken together, these results implicate glutamate dehydrogenase and NADP-GDH in particular, as a key target of in vivo isophthalate inhibition during ammonium assimilation. PMID:22578865

  5. Caffeine promotes glutamate and histamine release in the posterior hypothalamus

    PubMed Central

    Kodama, Tohru; Siegel, Jerome M.

    2014-01-01

    Histamine neurons are active during waking and largely inactive during sleep, with minimal activity during rapid-eye movement (REM) sleep. Caffeine, the most widely used stimulant, causes a significant increase of sleep onset latency in rats and humans. We hypothesized that caffeine increases glutamate release in the posterior hypothalamus (PH) and produces increased activity of wake-active histamine neurons. Using in vivo microdialysis, we collected samples from the PH after caffeine administration in freely behaving rats. HPLC analysis and biosensor measurements showed a significant increase in glutamate levels beginning 30 min after caffeine administration. Glutamate levels remained elevated for at least 140 min. GABA levels did not significantly change over the same time period. Histamine level significantly increased beginning 30 min after caffeine administration and remained elevated for at least 140 min. Immunostaining showed a significantly elevated number of c-Fos-labeled histamine neurons in caffeine-treated rats compared with saline-treated animals. We conclude that increased glutamate levels in the PH activate histamine neurons and contribute to caffeine-induced waking and alertness. PMID:25031227

  6. Microbial production and chemical transformation of poly-γ-glutamate.

    PubMed

    Ashiuchi, Makoto

    2013-11-01

    Poly-γ-glutamate (PGA), a novel polyamide material with industrial applications, possesses a nylon-like backbone, is structurally similar to polyacrylic acid, is biodegradable and is safe for human consumption. PGA is frequently found in the mucilage of natto, a Japanese traditional fermented food. To date, three different types of PGA, namely a homo polymer of D-glutamate (D-PGA), a homo polymer of L-glutamate (L-PGA), and a random copolymer consisting of D- and L-glutamate (DL-PGA), are known. This review will detail the occurrence and physiology of PGA. The proposed reaction mechanism of PGA synthesis including its localization and the structure of the involved enzyme, PGA synthetase, are described. The occurrence of multiple carboxyl residues in PGA likely plays a role in its relative unsuitability for the development of bio-nylon plastics and thus, establishment of an efficient PGA-reforming strategy is of great importance. Aside from the potential applications of PGA proposed to date, a new technique for chemical transformation of PGA is also discussed. Finally, some techniques for PGA and its derivatives in advanced material technology are presented.

  7. Metabotropic glutamate receptors: From the workbench to the bedside

    PubMed Central

    Nicoletti, F.; Bockaert, J.; Collingridge, G.L.; Conn, P.J.; Ferraguti, F.; Schoepp, D.D.; Wroblewski, J.T.; Pin, J.P.

    2013-01-01

    Metabotropic glutamate (mGlu) receptors were discovered in the mid 1980s and originally described as glutamate receptors coupled to polyphosphoinositide hydrolysis. Almost 6500 articles have been published since then, and subtype-selective mGlu receptor ligands are now under clinical development for the treatment of a variety of disorders such as Fragile-X syndrome, schizophrenia, Parkinson’s disease and L-DOPA-induced dyskinesias, generalized anxiety disorder, chronic pain, and gastroesophageal reflux disorder. Prof. Erminio Costa was linked to the early times of the mGlu receptor history, when a few research groups challenged the general belief that glutamate could only activate ionotropic receptors and all metabolic responses to glutamate were secondary to calcium entry. This review moves from those nostalgic times to the most recent advances in the physiology and pharmacology of mGlu receptors, and highlights the role of individual mGlu receptor subtypes in the pathophysiology of human disorders. This article is part of a Special Issue entitled ‘Trends in Neuropharmacology: In Memory of Erminio Costa’. PMID:21036182

  8. Continuous glutamate production using an immobilized whole-cell system

    SciTech Connect

    Kim, H.S.; Ryu, D.D.Y.

    1982-10-01

    For the purpose of saving the energy and raw materials required in a glutamate fermentation, an immobilized whole-cell system was prepared and its performance in a continuous reactor system was evaluated. Corynebacterium glutamicum (a mutant strain of ATCC 13058) whole cell was immobilized in k-carrageenan matrix and the gel structure was strengthened by treatment with a hardening agent. The effective diffusivities of carrageenan gel for glucose and oxygen were formed to decrease significantly with an increase in carrageenan concentration, while the gel strength showed an increasing trend. Based on the physical and chemical properties of carrageenan gel, the immobilized method was improved and the operation of the continuous reactor system was partially optimized. In an air-stirred fermentor, the continuous production of glutamate was carried out. The effect of the dilution rate of glutamate production and operation stability was investigated. The performance of the continuous wbole-cell reactor system was evaluated by measuring glutamate productivity for a period of 30 days; it was found to be far superior to the performance of convention batch reactor systems using free cells.

  9. Microbial production and chemical transformation of poly-γ-glutamate

    PubMed Central

    Ashiuchi, Makoto

    2013-01-01

    Poly-γ-glutamate (PGA), a novel polyamide material with industrial applications, possesses a nylon-like backbone, is structurally similar to polyacrylic acid, is biodegradable and is safe for human consumption. PGA is frequently found in the mucilage of natto, a Japanese traditional fermented food. To date, three different types of PGA, namely a homo polymer of d-glutamate (D-PGA), a homo polymer of l-glutamate (L-PGA), and a random copolymer consisting of d- and l-glutamate (DL-PGA), are known. This review will detail the occurrence and physiology of PGA. The proposed reaction mechanism of PGA synthesis including its localization and the structure of the involved enzyme, PGA synthetase, are described. The occurrence of multiple carboxyl residues in PGA likely plays a role in its relative unsuitability for the development of bio-nylon plastics and thus, establishment of an efficient PGA-reforming strategy is of great importance. Aside from the potential applications of PGA proposed to date, a new technique for chemical transformation of PGA is also discussed. Finally, some techniques for PGA and its derivatives in advanced material technology are presented. PMID:23855427

  10. Paraventricular Stimulation with Glutamate Elicits Bradycardia and Pituitary Responses

    NASA Technical Reports Server (NTRS)

    Darlington, Daniel N.; Miyamoto, Michael; Keil, Lanny C.; Dallman, Mary F.

    1989-01-01

    The excitatory neurotransmitter, L-glutamate (0.5 M, pH 7.4), or the organic acid, acetate (0.5 M, pH 7.4), was microinjected (50 nl over 2 min) directly into the paraventricular nuclei (PVN) of pentobarbital sodium-anesthetized rats while arterial blood pressure and heart rate and plasma adrenocorticotropic hormone (ACTH), vasopressin, and oxytocin were measured. Activation of PVN neurons with L-glutamate led to increases in plasma ACTH, vasopressin, and oxytocin and a profound bradycardia (-80 beats/min) with little change in arterial blood pressure. Microinjection of acetate had no effect on the above variables. The decrease in heart rate was shown to be dependent on the concentration of glutamate injected and the volume of injectate. The bradycardia was mediated through the autonomic nervous system because ganglionic blockade (pentolinium tartrate) eliminated the response; atropine and propranolol severely attenuated the bradycardia. The bradycardia was greatest when L-glutamate was microinjected into the caudal PVN. Injections into the rostral PVN or into nuclei surrounding the PVN led to small or nonsignificant decreases in heart rate. Focal electric stimulation (2-50 pA) of the PVN also led to decreases in heart rate and arterial blood pressure. These data suggest that activation of PVN neurons leads to the release of ACTH, vasopressin, and oxytocin from the pituitary and a bradycardia that is mediated by the autonomic nervous system.

  11. Does formate reduce alpha-ketoglutarate and ammonia to glutamate?

    NASA Technical Reports Server (NTRS)

    Maughan, Q.; Miller, S. L.; Bada, J. L. (Principal Investigator)

    1999-01-01

    The reported reduction of alpha-ketoglutarate and ammonia by formate is much slower than described (Morowitz et al., 1995). The formate reduction if any is small under these conditions. Glutamate is produced from a reduction by a second molecule of alpha-ketoglutarate involving an oxidative decarboxylation.

  12. Microbial production and chemical transformation of poly-γ-glutamate.

    PubMed

    Ashiuchi, Makoto

    2013-11-01

    Poly-γ-glutamate (PGA), a novel polyamide material with industrial applications, possesses a nylon-like backbone, is structurally similar to polyacrylic acid, is biodegradable and is safe for human consumption. PGA is frequently found in the mucilage of natto, a Japanese traditional fermented food. To date, three different types of PGA, namely a homo polymer of D-glutamate (D-PGA), a homo polymer of L-glutamate (L-PGA), and a random copolymer consisting of D- and L-glutamate (DL-PGA), are known. This review will detail the occurrence and physiology of PGA. The proposed reaction mechanism of PGA synthesis including its localization and the structure of the involved enzyme, PGA synthetase, are described. The occurrence of multiple carboxyl residues in PGA likely plays a role in its relative unsuitability for the development of bio-nylon plastics and thus, establishment of an efficient PGA-reforming strategy is of great importance. Aside from the potential applications of PGA proposed to date, a new technique for chemical transformation of PGA is also discussed. Finally, some techniques for PGA and its derivatives in advanced material technology are presented. PMID:23855427

  13. Deeper Insights into the Allosteric Modulation of Ionotropic Glutamate Receptors.

    PubMed

    Regan, Michael C; Furukawa, Hiro

    2016-09-21

    Two articles in this issue of Neuron (Yelshanskaya et al., 2016; Yi et al., 2016) explore the structural basis of allosteric inhibition in ionotropic glutamate receptors, providing key insights into how iGluRs function in the brain as well as how they might be pharmacologically modulated in neurological disorders and disease. PMID:27657445

  14. Blood and Brain Glutamate Levels in Children with Autistic Disorder

    ERIC Educational Resources Information Center

    Hassan, Tamer H.; Abdelrahman, Hadeel M.; Fattah, Nelly R. Abdel; El-Masry, Nagda M.; Hashim, Haitham M.; El-Gerby, Khaled M.; Fattah, Nermin R. Abdel

    2013-01-01

    Despite of the great efforts that move forward to clarify the pathophysiologic mechanisms in autism, the cause of this disorder, however, remains largely unknown. There is an increasing body of literature concerning neurochemical contributions to the pathophysiology of autism. We aimed to determine blood and brain levels of glutamate in children…

  15. Glutamate and GABA in Vestibulo-Sympathetic Pathway Neurons

    PubMed Central

    Holstein, Gay R.; Friedrich, Victor L. Jr.; Martinelli, Giorgio P.

    2016-01-01

    The vestibulo-sympathetic reflex (VSR) actively modulates blood pressure during changes in posture. This reflex allows humans to stand up and quadrupeds to rear or climb without a precipitous decline in cerebral perfusion. The VSR pathway conveys signals from the vestibular end organs to the caudal vestibular nuclei. These cells, in turn, project to pre-sympathetic neurons in the rostral and caudal ventrolateral medulla (RVLM and CVLM, respectively). The present study assessed glutamate- and GABA-related immunofluorescence associated with central vestibular neurons of the VSR pathway in rats. Retrograde FluoroGold tract tracing was used to label vestibular neurons with projections to RVLM or CVLM, and sinusoidal galvanic vestibular stimulation (GVS) was employed to activate these pathways. Central vestibular neurons of the VSR were identified by co-localization of FluoroGold and cFos protein, which accumulates in some vestibular neurons following galvanic stimulation. Triple-label immunofluorescence was used to co-localize glutamate- or GABA- labeling in the identified VSR pathway neurons. Most activated projection neurons displayed intense glutamate immunofluorescence, suggestive of glutamatergic neurotransmission. To support this, anterograde tracer was injected into the caudal vestibular nuclei. Vestibular axons and terminals in RVLM and CVLM co-localized the anterograde tracer and vesicular glutamate transporter-2 signals. Other retrogradely-labeled cFos-positive neurons displayed intense GABA immunofluorescence. VSR pathway neurons of both phenotypes were present in the caudal medial and spinal vestibular nuclei, and projected to both RVLM and CVLM. As a group, however, triple-labeled vestibular cells with intense glutamate immunofluorescence were located more rostrally in the vestibular nuclei than the GABAergic neurons. Only the GABAergic VSR pathway neurons showed a target preference, projecting predominantly to CVLM. These data provide the first

  16. Glutamate and GABA in Vestibulo-Sympathetic Pathway Neurons.

    PubMed

    Holstein, Gay R; Friedrich, Victor L; Martinelli, Giorgio P

    2016-01-01

    The vestibulo-sympathetic reflex (VSR) actively modulates blood pressure during changes in posture. This reflex allows humans to stand up and quadrupeds to rear or climb without a precipitous decline in cerebral perfusion. The VSR pathway conveys signals from the vestibular end organs to the caudal vestibular nuclei. These cells, in turn, project to pre-sympathetic neurons in the rostral and caudal ventrolateral medulla (RVLM and CVLM, respectively). The present study assessed glutamate- and GABA-related immunofluorescence associated with central vestibular neurons of the VSR pathway in rats. Retrograde FluoroGold tract tracing was used to label vestibular neurons with projections to RVLM or CVLM, and sinusoidal galvanic vestibular stimulation (GVS) was employed to activate these pathways. Central vestibular neurons of the VSR were identified by co-localization of FluoroGold and cFos protein, which accumulates in some vestibular neurons following galvanic stimulation. Triple-label immunofluorescence was used to co-localize glutamate- or GABA- labeling in the identified VSR pathway neurons. Most activated projection neurons displayed intense glutamate immunofluorescence, suggestive of glutamatergic neurotransmission. To support this, anterograde tracer was injected into the caudal vestibular nuclei. Vestibular axons and terminals in RVLM and CVLM co-localized the anterograde tracer and vesicular glutamate transporter-2 signals. Other retrogradely-labeled cFos-positive neurons displayed intense GABA immunofluorescence. VSR pathway neurons of both phenotypes were present in the caudal medial and spinal vestibular nuclei, and projected to both RVLM and CVLM. As a group, however, triple-labeled vestibular cells with intense glutamate immunofluorescence were located more rostrally in the vestibular nuclei than the GABAergic neurons. Only the GABAergic VSR pathway neurons showed a target preference, projecting predominantly to CVLM. These data provide the first

  17. Glutamate-mediated excitotoxicity in schizophrenia: a review.

    PubMed

    Plitman, Eric; Nakajima, Shinichiro; de la Fuente-Sandoval, Camilo; Gerretsen, Philip; Chakravarty, M Mallar; Kobylianskii, Jane; Chung, Jun Ku; Caravaggio, Fernando; Iwata, Yusuke; Remington, Gary; Graff-Guerrero, Ariel

    2014-10-01

    Findings from neuroimaging studies in patients with schizophrenia suggest widespread structural changes although the mechanisms through which these changes occur are currently unknown. Glutamatergic activity appears to be increased in the early phases of schizophrenia and may contribute to these structural alterations through an excitotoxic effect. The primary aim of this review was to describe the possible role of glutamate-mediated excitotoxicity in explaining the presence of neuroanatomical changes within schizophrenia. A Medline(®) literature search was conducted, identifying English language studies on the topic of glutamate-mediated excitotoxicity in schizophrenia, using the terms "schizophreni" and "glutam" and (("MRS" or "MRI" or "magnetic resonance") or ("computed tomography" or "CT")). Studies concomitantly investigating glutamatergic activity and brain structure in patients with schizophrenia were included. Results are discussed in the context of findings from preclinical studies. Seven studies were identified that met the inclusion criteria. These studies provide inconclusive support for the role of glutamate-mediated excitotoxicity in the occurrence of structural changes within schizophrenia, with the caveat that there is a paucity of human studies investigating this topic. Preclinical data suggest that an excitotoxic effect may occur as a result of a paradoxical increase in glutamatergic activity following N-methyl-D-aspartate receptor hypofunction. Based on animal literature, glutamate-mediated excitotoxicity may account for certain structural changes present in schizophrenia, but additional human studies are required to substantiate these findings. Future studies should adopt a longitudinal design and employ magnetic resonance imaging techniques to investigate whether an association between glutamatergic activity and structural changes exists in patients with schizophrenia.

  18. Glutamate, GABA, and glutamine are synchronously upregulated in the mouse lateral septum during the postpartum period

    PubMed Central

    Zhao, Changjiu; Gammie, Stephen C.

    2014-01-01

    Dramatic structural and functional remodeling occurs in the postpartum brain for the establishment of maternal care, which is essential for the growth and development of young offspring. Glutamate and GABA signaling are critically important in modulating multiple behavioral performances. Large scale signaling changes occur in the postpartum brain, but it is still not clear to what extent the neurotransmitters glutamate and GABA change and whether the ratio of glutamate/GABA remains balanced. In this study, we examined the glutamate/GABA-glutamine cycle in the lateral septum (LS) of postpartum female mice. In postpartum females (relative to virgins), tissue levels of glutamate and GABA were elevated in LS and increased mRNA was found for the respective enzymes producing glutamate and GABA, glutaminase (Gls) and glutamate decarboxylase 1 and 2 (Gad1 and Gad2). The common precursor, glutamine, was elevated as was the enzyme that produces it, glutamate-ammonia ligase (Glul). Additionally, glutamate, GABA, and glutamine were positively correlated and the glutamate/GABA ratio was almost identical in the postpartum and virgin females. Collectively, these findings indicate that glutamate and GABA signaling are increased and that the ratio of glutamate/GABA is well balanced in the maternal LS. The postpartum brain may provide a useful model system for understanding how glutamate and GABA are linked despite large signaling changes. Given that some mental health disorders, including depression and schizophrenia display dysregulated glutamate/GABA ratio, and there is increased vulnerability to mental disorders in mothers, it is possible that these postpartum disorders emerge when glutamate and GABA changes are not properly coordinated. PMID:25451092

  19. Glutamate, GABA, and glutamine are synchronously upregulated in the mouse lateral septum during the postpartum period.

    PubMed

    Zhao, Changjiu; Gammie, Stephen C

    2014-12-01

    Dramatic structural and functional remodeling occurs in the postpartum brain for the establishment of maternal care, which is essential for the growth and development of young offspring. Glutamate and GABA signaling are critically important in modulating multiple behavioral performances. Large scale signaling changes occur in the postpartum brain, but it is still not clear to what extent the neurotransmitters glutamate and GABA change and whether the ratio of glutamate/GABA remains balanced. In this study, we examined the glutamate/GABA-glutamine cycle in the lateral septum (LS) of postpartum female mice. In postpartum females (relative to virgins), tissue levels of glutamate and GABA were elevated in LS and increased mRNA was found for the respective enzymes producing glutamate and GABA, glutaminase (Gls) and glutamate decarboxylase 1 and 2 (Gad1 and Gad2). The common precursor, glutamine, was elevated as was the enzyme that produces it, glutamate-ammonia ligase (Glul). Additionally, glutamate, GABA, and glutamine were positively correlated and the glutamate/GABA ratio was almost identical in the postpartum and virgin females. Collectively, these findings indicate that glutamate and GABA signaling are increased and that the ratio of glutamate/GABA is well balanced in the maternal LS. The postpartum brain may provide a useful model system for understanding how glutamate and GABA are linked despite large signaling changes. Given that some mental health disorders, including depression and schizophrenia display dysregulated glutamate/GABA ratio, and there is increased vulnerability to mental disorders in mothers, it is possible that these postpartum disorders emerge when glutamate and GABA changes are not properly coordinated.

  20. Calmodulin binding to glutamate decarboxylase is required for regulation of glutamate and GABA metabolism and normal development in plants.

    PubMed Central

    Baum, G; Lev-Yadun, S; Fridmann, Y; Arazi, T; Katsnelson, H; Zik, M; Fromm, H

    1996-01-01

    Glutamate decarboxylase (GAD) catalyzes the decarboxylation of glutamate to CO2 and gamma-aminobutyrate (GABA). GAD is ubiquitous in prokaryotes and eukaryotes, but only plant GAD has been shown to bind calmodulin (CaM). Here, we assess the role of the GAD CaM-binding domain in vivo. Transgenic tobacco plants expressing a mutant petunia GAD lacking the CaM-binding domain (GADdeltaC plants) exhibit severe morphological abnormalities, such as short stems, in which cortex parenchyma cells fail to elongate, associated with extremely high GABA and low glutamate levels. The morphology of transgenic plants expressing the full-length GAD (GAD plants) is indistinguishable from that of wild-type (WT) plants. In WT and GAD plant extracts, GAD activity is inhibited by EGTA and by the CaM antagonist trifluoperazine, and is associated with a CaM-containing protein complex of approximately 500 kDa. In contrast, GADdeltaC plants lack normal GAD complexes, and GAD activity in their extracts is not affected by EGTA and trifluoperazine. We conclude that CaM binding to GAD is essential for the regulation of GABA and glutamate metabolism, and that regulation of GAD activity is necessary for normal plant development. This study is the first to demonstrate an in vivo function for CaM binding to a target protein in plants. Images PMID:8670800

  1. Calmodulin binding to glutamate decarboxylase is required for regulation of glutamate and GABA metabolism and normal development in plants.

    PubMed

    Baum, G; Lev-Yadun, S; Fridmann, Y; Arazi, T; Katsnelson, H; Zik, M; Fromm, H

    1996-06-17

    Glutamate decarboxylase (GAD) catalyzes the decarboxylation of glutamate to CO2 and gamma-aminobutyrate (GABA). GAD is ubiquitous in prokaryotes and eukaryotes, but only plant GAD has been shown to bind calmodulin (CaM). Here, we assess the role of the GAD CaM-binding domain in vivo. Transgenic tobacco plants expressing a mutant petunia GAD lacking the CaM-binding domain (GADdeltaC plants) exhibit severe morphological abnormalities, such as short stems, in which cortex parenchyma cells fail to elongate, associated with extremely high GABA and low glutamate levels. The morphology of transgenic plants expressing the full-length GAD (GAD plants) is indistinguishable from that of wild-type (WT) plants. In WT and GAD plant extracts, GAD activity is inhibited by EGTA and by the CaM antagonist trifluoperazine, and is associated with a CaM-containing protein complex of approximately 500 kDa. In contrast, GADdeltaC plants lack normal GAD complexes, and GAD activity in their extracts is not affected by EGTA and trifluoperazine. We conclude that CaM binding to GAD is essential for the regulation of GABA and glutamate metabolism, and that regulation of GAD activity is necessary for normal plant development. This study is the first to demonstrate an in vivo function for CaM binding to a target protein in plants.

  2. Deletion of genes involved in glutamate metabolism to improve poly-gamma-glutamic acid production in B. amyloliquefaciens LL3.

    PubMed

    Zhang, Wei; He, Yulian; Gao, Weixia; Feng, Jun; Cao, Mingfeng; Yang, Chao; Song, Cunjiang; Wang, Shufang

    2015-02-01

    Here, we attempted to elevate poly-gamma-glutamic acid (γ-PGA) production by modifying genes involved in glutamate metabolism in Bacillus amyloliquefaciens LL3. Products of rocR, rocG and gudB facilitate the conversion from glutamate to 2-oxoglutarate in Bacillus subtillis. The gene odhA is responsible for the synthesis of a component of the 2-oxoglutarate dehydrogenase complex that catalyzes the oxidative decarboxylation of 2-oxoglutarate to succinyl coenzyme A. In-frame deletions of these four genes were performed. In shake flask experiments the gudB/rocG double mutant presented enhanced production of γ-PGA, a 38 % increase compared with wild type. When fermented in a 5-L fermenter with pH control, the γ-PGA yield of the rocR mutant was increased to 5.83 g/L from 4.55 g/L for shake flask experiments. The gudB/rocG double mutant produced 5.68 g/L γ-PGA compared with that of 4.03 g/L for the wild type, a 40 % increase. Those results indicated the possibility of improving γ-PGA production by modifying glutamate metabolism, and identified potential genetic targets to improve γ-PGA production.

  3. Construction of glutamate biosensor based on covalent immobilization of glutamate oxidase on polypyrrole nanoparticles/polyaniline modified gold electrode.

    PubMed

    Batra, Bhawna; Kumari, Seema; Pundir, Chandra Shekhar

    2014-04-10

    A method is described for construction of a highly sensitive electrochemical biosensor for detection of glutamate. The biosensor is based on covalent immobilization of glutamate oxidase (GluOx) onto polypyrrole nanoparticles and polyaniline composite film (PPyNPs/PANI) electrodeposited onto Au electrode. The enzyme electrode was characterized by cyclic voltammetry (CV), scanning electron microscopy (SEM), X-ray diffraction (XRD), transmission electron microscopy (TEM), Fourier transform infra-red spectroscopy (FTIR) and electrochemical impedance spectroscopy (EIS). The biosensor showed optimum response within 3s at pH 7.5 (0.1 M sodium phosphate) and 35 °C, when operated at 50 mV s⁻¹. It exhibited excellent sensitivity (detection limit as 0.1 nM), fast response time and wider linear range (from 0.02 to 400 μM). Analytical recovery of added glutamate (5 mM and 10 mM) was 95.56 and 97%, while within batch and between batch coefficients of variation were 3.2% and 3.35% respectively. The enzyme electrode was used 100 times over a period of 60 days, when stored at 4 °C. The biosensor measured glutamate level in food stuff, which correlated well with a standard colorimetric method (r=0.99). PMID:24629270

  4. The cystine/glutamate antiporter: when too much of a good thing goes bad.

    PubMed

    Reissner, Kathryn J

    2014-08-01

    Glutamate excitotoxicity represents a major cellular component of ischemic brain injury. In this issue of the JCI, Soria and colleagues reveal that the cystine/glutamate exchanger is an important source of excitotoxic glutamate in response to ischemia induced by oxygen and glucose deprivation. As the exchanger is a primary determinant of both extracellular glutamate and intracellular glutathione, the findings from this study not only provide important insight into the mechanisms of brain ischemia but also demonstrate the complexity of the yin and yang of glutamate homeostasis and cellular redox balance.

  5. Electrogenic glutamate uptake is a major current carrier in the membrane of axolotl retinal glial cells

    NASA Astrophysics Data System (ADS)

    Brew, Helen; Attwell, David

    1987-06-01

    Glutamate is taken up avidly by glial cells in the central nervous system1. Glutamate uptake may terminate the transmitter action of glutamate released from neurons1, and keep extracellular glutamate at concentrations below those which are neurotoxic. We report here that glutamate evokes a large inward current in retinal glial cells which have their membrane potential and intracellular ion concentrations controlled by the whole-cell patch-clamp technique2. This current seems to be due to an electrogenic glutamate uptake carrier, which transports at least two sodium ions with every glutamate anion carried into the cell. Glutamate uptake is strongly voltage-dependent, decreasing at depolarized potentials: when fully activated, it contributes almost half of the conductance in the part of the glial cell membrane facing the retinal neurons. The spatial localization, glutamate affinity and magnitude of the uptake are appropriate for terminating the synaptic action of glutamate released from photoreceptors and bipolar cells. These data challenge present explanations of how the b-wave of the electroretinogram is generated, and suggest a mechanism for non-vesicular voltage-dependent release of glutamate from neurons.

  6. Hypoxia regulates glutamate metabolism and membrane transport in rat PC12 cells.

    PubMed

    Kobayashi, S; Millhorn, D E

    2001-03-01

    We investigated the effect of hypoxia on glutamate metabolism and uptake in rat pheochromocytoma (PC12) cells. Various key enzymes relevant to glutamate production, metabolism and transport were coordinately regulated by hypoxia. PC12 cells express two glutamate-metabolizing enzymes, glutamine synthetase (GS) and glutamate decarboxylase (GAD), as well as the glutamate-producing enzyme, phosphate-activated glutaminase (PAG). Exposure to hypoxia (1% O(2)) for 6 h or longer increased expression of GS mRNA and protein and enhanced GS enzymatic activity. In contrast, hypoxia caused a significant decrease in expression of PAG mRNA and protein, and also decreased PAG activity. In addition, hypoxia led to an increase in GAD65 and GAD67 protein levels and GAD enzymatic activity. PC12 cells express three Na(+)-dependent glutamate transporters; EAAC1, GLT-1 and GLAST. Hypoxia increased EAAC1 and GLT-1 protein levels, but had no effect on GLAST. Chronic hypoxia significantly enhanced the Na(+)-dependent component of glutamate transport. Furthermore, chronic hypoxia decreased cellular content of glutamate, but increased that of glutamine. Taken together, the hypoxia-induced changes in enzymes related to glutamate metabolism and transport are consistent with a decrease in the extracellular concentration of glutamate. This may have a role in protecting PC12 cells from the cytotoxic effects of glutamate during chronic hypoxia. PMID:11259512

  7. An examination of aspartate decarboxylase and glutamate decarboxylase activity in mosquitoes

    PubMed Central

    Richardson, Graham; Ding, Haizhen; Rocheleau, Tom; Mayhew, George; Reddy, Erin; Han, Qian; Christensen, Bruce M.; Li, Jianyong

    2010-01-01

    A major pathway of beta-alanine synthesis in insects is through the alpha-decarboxylation of aspartate, but the enzyme involved in the decarboxylation of aspartate has not been clearly defined in mosquitoes and characterized in any insect species. In this study, we expressed two putative mosquito glutamate decarboxylase-like enzymes of mosquitoes and critically analyzed their substrate specificity and biochemical properties. Our results provide clear biochemical evidence establishing that one of them is an aspartate decarboxylase and the other is a glutamate decarboxylase. The mosquito aspartate decarboxylase functions exclusively on the production of beta-alanine with no activity with glutamate. Likewise the mosquito glutamate decarboxylase is highly specific to glutamate with essentially no activity with aspartate. Although insect aspartate decarboxylase shares high sequence identity with glutamate decarboxylase, we are able to closely predict aspartate decarboxylase from glutamate decarboxylase based on the difference of their active site residues. PMID:19842059

  8. Effect of insulin on the compartmentation of glutamate for protein synthesis

    SciTech Connect

    Brown, A.B.; Mohan, C.; Bessman, S.P.

    1986-03-05

    The effect of insulin on the formation of CO/sub 2/ and incorporation of 1-/sup 14/C glutamine and U-/sup 14/C acetate into protein was studied in isolated rat hepatocytes. Insulin caused an 18% increase in /sup 14/CO/sub 2/ production from U-/sup 14/C acetate in comparison to a 10% increase from 1-/sup 14/C glutamate. Insulin caused a greater increase in the incorporation of tracer acetate carbons into hepatocyte protein. Hydrolysis of labeled protein and subsequent determination of glutamate specific activity revealed that incorporation of acetate carbons into protein as glutamate was about 52% greater in the presence of insulin. These results demonstrate the existence of two compartments of glutamate for protein synthesis: (i) glutamate generated in the Krebs cycle through transamination of a-ketoglutarate; (ii) cytosolic glutamate. Insulin had a greater stimulatory effect on the incorporation of glutamate generated in the Krebs cycle.

  9. Metabolic pathways and activity-dependent modulation of glutamate concentration in the human brain.

    PubMed

    Mangia, Silvia; Giove, Federico; Dinuzzo, Mauro

    2012-11-01

    Glutamate is one of the most versatile molecules present in the human brain, involved in protein synthesis, energy production, ammonia detoxification, and transport of reducing equivalents. Aside from these critical metabolic roles, glutamate plays a major part in brain function, being not only the most abundant excitatory neurotransmitter, but also the precursor for γ-aminobutyric acid, the predominant inhibitory neurotransmitter. Regulation of glutamate levels is pivotal for normal brain function, as abnormal extracellular concentration of glutamate can lead to impaired neurotransmission, neurodegeneration and even neuronal death. Understanding how the neuron-astrocyte functional and metabolic interactions modulate glutamate concentration during different activation status and under physiological and pathological conditions is a challenging task, and can only be tentatively estimated from current literature. In this paper, we focus on describing the various metabolic pathways which potentially affect glutamate concentration in the brain, and emphasize which ones are likely to produce the variations in glutamate concentration observed during enhanced neuronal activity in human studies.

  10. Genes involved in Drosophila glutamate receptor expression and localization

    PubMed Central

    Liebl, Faith LW; Featherstone, David E

    2005-01-01

    Background A clear picture of the mechanisms controlling glutamate receptor expression, localization, and stability remains elusive, possibly due to an incomplete understanding of the proteins involved. We screened transposon mutants generated by the ongoing Drosophila Gene Disruption Project in an effort to identify the different types of genes required for glutamate receptor cluster development. Results To enrich for non-silent insertions with severe disruptions in glutamate receptor clustering, we identified and focused on homozygous lethal mutants in a collection of 2185 BG and KG transposon mutants generated by the BDGP Gene Disruption Project. 202 lethal mutant lines were individually dissected to expose glutamatergic neuromuscular junctions, stained using antibodies that recognize neuronal membrane and the glutamate receptor subunit GluRIIA, and viewed using laser-scanning confocal microscopy. We identified 57 mutants with qualitative differences in GluRIIA expression and/or localization. 84% of mutants showed loss of receptors and/or clusters; 16% of mutants showed an increase in receptors. Insertion loci encode a variety of protein types, including cytoskeleton proteins and regulators, kinases, phosphatases, ubiquitin ligases, mucins, cell adhesion proteins, transporters, proteins controlling gene expression and protein translation, and proteins of unknown/novel function. Expression pattern analyses and complementation tests, however, suggest that any single mutant – even if a mutant gene is uniquely tagged – must be interpreted with caution until the mutation is validated genetically and phenotypically. Conclusion Our study identified 57 transposon mutants with qualitative differences in glutamate receptor expression and localization. Despite transposon tagging of every insertion locus, extensive validation is needed before one can have confidence in the role of any individual gene. Alternatively, one can focus on the types of genes identified, rather

  11. Metabotropic glutamate receptors in the control of mood disorders.

    PubMed

    Witkin, Jeffrey M; Marek, Gerard J; Johnson, Bryan G; Schoepp, Darryle D

    2007-04-01

    Current treatments for depression are less than optimal in terms of onset of action, response and remission rates, and side-effect profiles. Glutamate is the major excitatory neurotransmitter controlling synaptic excitability and plasticity in most brain circuits, including limbic pathways involved in depression. Thus, drugs that target glutamate neuronal transmission offer novel approaches to treat depression. Recently, the NMDA receptor antagonist ketamine has demonstrated clinical efficacy in a randomized clinical trial of depressed patients. Metabotropic glutamate (mGlu) receptors function to regulate glutamate neuronal transmission by altering the release of neurotransmitter or modulating the post-synaptic responses to glutamate. Accumulating evidence from biochemical and behavioral studies support the idea that the regulation of glutamatergic neurotransmission via mGlu receptors is linked to mood disorders and that these receptors may serve as novel targets for the discovery of small molecule modulators with unique antidepressant properties. For example, mGlu receptor modulation can facilitate neuronal stem cell proliferation (neurogenesis) and the release of neurotransmitters that are associated with treatment response to depression in humans (serotonin, norepinephrine, dopamine). In particular, compounds that antagonize mGlu2, mGlu3 and/or mGlu5 receptors (e.g. LY341495, MSG0039, MPEP) have been linked to the above pharmacology and have also shown in vivo activity in animal models predictive of antidepressant efficacy such as the forced-swim test. The in vivo actions of these agents can be antagonized by compounds that block AMPA receptors, suggesting that their actions are direct downstream consequences of the enhancement of glutamate neuronal transmission in brain regions involved in depression. These data provide new approaches to finding mechanistically distinct drugs for depression that may have advantages over current therapies for some patients

  12. Glutamic acid decarboxylase and glutamate receptor changes during tolerance and dependence to benzodiazepines

    PubMed Central

    Izzo, Emanuela; Auta, James; Impagnatiello, Francesco; Pesold, Christine; Guidotti, Alessandro; Costa, Erminio

    2001-01-01

    Protracted administration of diazepam elicits tolerance, whereas discontinuation of treatment results in signs of dependence. Tolerance to the anticonvulsant action of diazepam is present in an early phase (6, 24, and 36 h) but disappears in a late phase (72–96 h) of withdrawal. In contrast, signs of dependence such as decrease in open-arm entries on an elevated plus-maze and increased susceptibility to pentylenetetrazol-induced seizures were apparent 96 h (but not 12, 24, or 48 h) after diazepam withdrawal. During the first 72 h of withdrawal, tolerance is associated with changes in the expression of GABAA (γ-aminobutyric acid type A) receptor subunits (decrease in γ2 and α1; increase in α5) and with an increase of mRNA expression of the most abundant form of glutamic acid decarboxylase (GAD), GAD67. In contrast, dl-α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor GluR1 subunit mRNA and cognate protein, which are normal during the early phase of diazepam withdrawal, increase by approximately 30% in cortex and hippocampus in association with the appearance of signs of dependence 96 h after diazepam withdrawal. Immunohistochemical studies of GluR1 subunit expression with gold-immunolabeling technique reveal that the increase of GluR1 subunit protein is localized to layer V pyramidal neurons and their apical dendrites in the cortex, and to pyramidal neurons and in their dendritic fields in hippocampus. The results suggest an involvement of GABA-mediated processes in the development and maintenance of tolerance to diazepam, whereas excitatory amino acid-related processes (presumably via AMPA receptors) may be involved in the expression of signs of dependence after withdrawal. PMID:11248104

  13. Glutamic acid decarboxylase and glutamate receptor changes during tolerance and dependence to benzodiazepines.

    PubMed

    Izzo, E; Auta, J; Impagnatiello, F; Pesold, C; Guidotti, A; Costa, E

    2001-03-13

    Protracted administration of diazepam elicits tolerance, whereas discontinuation of treatment results in signs of dependence. Tolerance to the anticonvulsant action of diazepam is present in an early phase (6, 24, and 36 h) but disappears in a late phase (72-96 h) of withdrawal. In contrast, signs of dependence such as decrease in open-arm entries on an elevated plus-maze and increased susceptibility to pentylenetetrazol-induced seizures were apparent 96 h (but not 12, 24, or 48 h) after diazepam withdrawal. During the first 72 h of withdrawal, tolerance is associated with changes in the expression of GABA(A) (gamma-aminobutyric acid type A) receptor subunits (decrease in gamma(2) and alpha(1); increase in alpha(5)) and with an increase of mRNA expression of the most abundant form of glutamic acid decarboxylase (GAD), GAD(67). In contrast, dl-alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA) receptor GluR1 subunit mRNA and cognate protein, which are normal during the early phase of diazepam withdrawal, increase by approximately 30% in cortex and hippocampus in association with the appearance of signs of dependence 96 h after diazepam withdrawal. Immunohistochemical studies of GluR1 subunit expression with gold-immunolabeling technique reveal that the increase of GluR1 subunit protein is localized to layer V pyramidal neurons and their apical dendrites in the cortex, and to pyramidal neurons and in their dendritic fields in hippocampus. The results suggest an involvement of GABA-mediated processes in the development and maintenance of tolerance to diazepam, whereas excitatory amino acid-related processes (presumably via AMPA receptors) may be involved in the expression of signs of dependence after withdrawal.

  14. Sodium-Dependent Glutamate Uptake by an Alkaliphilic, Thermophilic Bacillus Strain, TA2.A1

    PubMed Central

    Peddie, Catherine J.; Cook, Gregory M.; Morgan, Hugh W.

    1999-01-01

    A strain of Bacillus designated TA2.A1, isolated from a thermal spring in Te Aroha, New Zealand, grew optimally at pH 9.2 and 70°C. Bacillus strain TA2.A1 utilized glutamate as a sole carbon and energy source for growth, and sodium chloride (>5 mM) was an obligate requirement for growth. Growth on glutamate was inhibited by monensin and amiloride, both inhibitors that collapse the sodium gradient (ΔpNa) across the cell membrane. N,N-Dicyclohexylcarbodiimide inhibited the growth of Bacillus strain TA2.A1, suggesting that an F1F0-ATPase (H type) was being used to generate cellular ATP needed for anabolic reactions. Vanadate, an inhibitor of V-type ATPases, did not affect the growth of Bacillus strain TA2.A1. Glutamate transport by Bacillus strain TA2.A1 could be driven by an artificial membrane potential (ΔΨ), but only when sodium was present. In the absence of sodium, the rate of ΔΨ-driven glutamate uptake was fourfold lower. No glutamate transport was observed in the presence of ΔpNa alone (i.e., no ΔΨ). Glutamate uptake was specifically inhibited by monensin, and the Km for sodium was 5.6 mM. The Hill plot had a slope of approximately 1, suggesting that sodium binding was noncooperative and that the glutamate transporter had a single binding site for sodium. Glutamate transport was not affected by the protonophore carbonyl cyanide m-chlorophenylhydrazone, suggesting that the transmembrane pH gradient was not required for glutamate transport. The rate of glutamate transport increased with increasing glutamate concentration; the Km for glutamate was 2.90 μM, and the Vmax was 0.7 nmol · min−1 mg of protein. Glutamate transport was specifically inhibited by glutamate analogues. PMID:10322019

  15. Vitamin C neuroprotection against dose-dependent glutamate-induced neurodegeneration in the postnatal brain.

    PubMed

    Shah, Shahid Ali; Yoon, Gwang Ho; Kim, Hyun-Ok; Kim, Myeong Ok

    2015-05-01

    Glutamate-induced excitotoxicity due to over-activation of glutamate receptors and associated energy depletion (phosphorylation and activation of AMPK) results in neuronal cell death in various neurological disorders. Restoration of energy balance during an excitotoxic insult is critical for neuronal survival. Ascorbic acid (vitamin C), an essential nutrient with well-known antioxidant potential, protects the brain from oxidative damage in various models of neurodegeneration. In this study, we reported the therapeutic efficacy of vitamin C in response to glutamate-induced excitation, resulting in energy depletion and apoptosis in the hippocampus of the developing rat brain. A single subcutaneous injection of glutamate at two different concentrations (5 and 10 mg/kg) in postnatal day 7 rat pups increased brain glutamate levels and increased the protein expression of neuronal apoptotic markers. Both doses of glutamate upregulated the ratio of pro-apoptotic Bax to anti-apoptotic Bcl-2, cytochrome-c release, caspase-3 activation and the expression of PARP-1. However, co-treatment of vitamin C (250 mg/kg) with glutamate decreased brain glutamate levels and reversed the changes induced by glutamate in the developing hippocampus. Interestingly, only a high dose of glutamate caused the phosphorylation and activation of AMPK and induced neuronal cell death, whereas a low dose of glutamate failed to mediate these effects. Vitamin C supplementation reduced the glutamate-induced phosphorylation of AMPK and attenuated neuronal cell death, as assessed morphologically by Fluoro Jade B in the hippocampal CA1 region of the developing brain. Taken together, our results indicated that glutamate in both concentrations is toxic to the immature rat brain, whereas vitamin C is pharmacologically effective against glutamate-induced neurodegeneration. PMID:25701025

  16. Repeated exposure to moderate doses of ethanol augments hippocampal glutamate neurotransmission by increasing release

    PubMed Central

    Chefer, Vladimir; Meis, Jennifer; Wang, Grace; Kuzmin, Alexander; Bakalkin, Georgy; Shippenberg, Toni

    2013-01-01

    The present study used conventional and quantitative microdialysis to assess glutamatergic and GABAergic neurotransmission in the hippocampal CA3 area of the rat following a moderate-dose ethanol treatment regimen. Male Wistar rats received 3.4 g/kg of ethanol or water for 6 days via gastric gavage. Microdialysis experiments commenced 2 days later. Basal and depolarization-induced glutamate overflow were significantly elevated in ethanol-treated animals. Basal and depolarization-induced gamma-aminobutyric acid (GABA) overflow were unaltered. Quantitative no-net-flux microdialysis was used to determine if changes in dialysate glutamate levels following ethanol administration are due to an increase in release or a decrease in uptake.To confirm the validity of this method for quantifying basal glutamate dynamics, extracellular concentrations of glutamate and the extraction fraction, which reflects changes in analyte clearance, were quantified in response to retro-dialysis of the glutamate uptake blocker trans-pyrrolidine-2,4-dicarboxylic acid (tPDC). tPDC significantly decreased the extraction fraction for glutamate, resulting in augmented extracellular glutamate concentrations. Repeated ethanol administration did not alter the glutamate extraction fraction. However, extracellular glutamate concentrations were significantly elevated, indicating that glutamate release is increased as a consequence of repeated ethanol administration. These data demonstrate that repeated bouts of moderate ethanol consumption alter basal glutamate dynamics in the CA3 region of the dorsal hippocampus. Basal glutamate release is augmented, whereas glutamate uptake is unchanged. Furthermore, they suggest that dysregulation of glutamate transmission in this region may contribute to the previously documented deficits in cognitive function associated with moderate dose ethanol use. PMID:21182572

  17. Vitamin C neuroprotection against dose-dependent glutamate-induced neurodegeneration in the postnatal brain.

    PubMed

    Shah, Shahid Ali; Yoon, Gwang Ho; Kim, Hyun-Ok; Kim, Myeong Ok

    2015-05-01

    Glutamate-induced excitotoxicity due to over-activation of glutamate receptors and associated energy depletion (phosphorylation and activation of AMPK) results in neuronal cell death in various neurological disorders. Restoration of energy balance during an excitotoxic insult is critical for neuronal survival. Ascorbic acid (vitamin C), an essential nutrient with well-known antioxidant potential, protects the brain from oxidative damage in various models of neurodegeneration. In this study, we reported the therapeutic efficacy of vitamin C in response to glutamate-induced excitation, resulting in energy depletion and apoptosis in the hippocampus of the developing rat brain. A single subcutaneous injection of glutamate at two different concentrations (5 and 10 mg/kg) in postnatal day 7 rat pups increased brain glutamate levels and increased the protein expression of neuronal apoptotic markers. Both doses of glutamate upregulated the ratio of pro-apoptotic Bax to anti-apoptotic Bcl-2, cytochrome-c release, caspase-3 activation and the expression of PARP-1. However, co-treatment of vitamin C (250 mg/kg) with glutamate decreased brain glutamate levels and reversed the changes induced by glutamate in the developing hippocampus. Interestingly, only a high dose of glutamate caused the phosphorylation and activation of AMPK and induced neuronal cell death, whereas a low dose of glutamate failed to mediate these effects. Vitamin C supplementation reduced the glutamate-induced phosphorylation of AMPK and attenuated neuronal cell death, as assessed morphologically by Fluoro Jade B in the hippocampal CA1 region of the developing brain. Taken together, our results indicated that glutamate in both concentrations is toxic to the immature rat brain, whereas vitamin C is pharmacologically effective against glutamate-induced neurodegeneration.

  18. High-Throughput Assay Development for Cystine-Glutamate Antiporter (xc-) Highlights Faster Cystine Uptake than Glutamate Release in Glioma Cells

    PubMed Central

    Thomas, Ajit G.; Sattler, Rita; Tendyke, Karen; Loiacono, Kara A.; Hansen, Hans; Sahni, Vishal; Hashizume, Yutaka; Rojas, Camilo; Slusher, Barbara S.

    2015-01-01

    The cystine-glutamate antiporter (system xc-) is a Na+-independent amino acid transporter that exchanges extracellular cystine for intracellular glutamate. It is thought to play a critical role in cellular redox processes through regulation of intracellular glutathione synthesis via cystine uptake. In gliomas, system xc- expression is universally up-regulated while that of glutamate transporters down-regulated, leading to a progressive accumulation of extracellular glutamate and excitotoxic cell death of the surrounding non-tumorous tissue. Additionally, up-regulation of system xc- in activated microglia has been implicated in the pathogenesis of several neurodegenerative disorders mediated by excess glutamate. Consequently, system xc- is a new drug target for brain cancer and neuroinflammatory diseases associated with excess extracellular glutamate. Unfortunately no potent and selective small molecule system xc- inhibitors exist and to our knowledge, no high throughput screening (HTS) assay has been developed to identify new scaffolds for inhibitor design. To develop such an assay, various neuronal and non-neuronal human cells were evaluated as sources of system xc-. Human glioma cells were chosen based on their high system xc- activity. Using these cells, [14C]-cystine uptake and cystine-induced glutamate release assays were characterized and optimized with respect to cystine and protein concentrations and time of incubation. A pilot screen of the LOPAC/NINDS libraries using glutamate release demonstrated that the logistics of the assay were in place but unfortunately, did not yield meaningful pharmacophores. A larger, HTS campaign using the 384-well cystine-induced glutamate release as primary assay and the 96-well 14C-cystine uptake as confirmatory assay is currently underway. Unexpectedly, we observed that the rate of cystine uptake was significantly faster than the rate of glutamate release in human glioma cells. This was in contrast to the same rates of

  19. High-Throughput Assay Development for Cystine-Glutamate Antiporter (xc-) Highlights Faster Cystine Uptake than Glutamate Release in Glioma Cells.

    PubMed

    Thomas, Ajit G; Sattler, Rita; Tendyke, Karen; Loiacono, Kara A; Hansen, Hans; Sahni, Vishal; Hashizume, Yutaka; Rojas, Camilo; Slusher, Barbara S

    2015-01-01

    The cystine-glutamate antiporter (system xc-) is a Na+-independent amino acid transporter that exchanges extracellular cystine for intracellular glutamate. It is thought to play a critical role in cellular redox processes through regulation of intracellular glutathione synthesis via cystine uptake. In gliomas, system xc- expression is universally up-regulated while that of glutamate transporters down-regulated, leading to a progressive accumulation of extracellular glutamate and excitotoxic cell death of the surrounding non-tumorous tissue. Additionally, up-regulation of system xc- in activated microglia has been implicated in the pathogenesis of several neurodegenerative disorders mediated by excess glutamate. Consequently, system xc- is a new drug target for brain cancer and neuroinflammatory diseases associated with excess extracellular glutamate. Unfortunately no potent and selective small molecule system xc- inhibitors exist and to our knowledge, no high throughput screening (HTS) assay has been developed to identify new scaffolds for inhibitor design. To develop such an assay, various neuronal and non-neuronal human cells were evaluated as sources of system xc-. Human glioma cells were chosen based on their high system xc- activity. Using these cells, [14C]-cystine uptake and cystine-induced glutamate release assays were characterized and optimized with respect to cystine and protein concentrations and time of incubation. A pilot screen of the LOPAC/NINDS libraries using glutamate release demonstrated that the logistics of the assay were in place but unfortunately, did not yield meaningful pharmacophores. A larger, HTS campaign using the 384-well cystine-induced glutamate release as primary assay and the 96-well 14C-cystine uptake as confirmatory assay is currently underway. Unexpectedly, we observed that the rate of cystine uptake was significantly faster than the rate of glutamate release in human glioma cells. This was in contrast to the same rates of

  20. Alcohol consumption increases basal extracellular glutamate in the nucleus accumbens core of Sprague-Dawley rats without increasing spontaneous glutamate release.

    PubMed

    Pati, Dipanwita; Kelly, Kyle; Stennett, Bethany; Frazier, Charles J; Knackstedt, Lori A

    2016-07-01

    Glutamate neurotransmission in the nucleus accumbens core (NAc) mediates ethanol consumption. Previous studies using non-contingent and voluntary alcohol administration in inbred rodents have reported increased basal extracellular glutamate levels in the NAc. Here, we assessed basal glutamate levels in the NAc following intermittent alcohol consumption in male Sprague-Dawley rats that had access to ethanol for 7 weeks on alternating days. We found increased basal NAc glutamate at 24 h withdrawal from ethanol and thus sought to identify the source of this glutamate. To do so, we employed a combination of microdialysis, slice electrophysiology and western blotting. Reverse dialysis of the voltage-gated sodium channel blocker tetrodotoxin did not affect glutamate levels in either group. Electrophysiological recordings in slices made after 24 h withdrawal revealed a decrease in spontaneous excitatory postsynaptic current (sEPSC) frequency relative to controls, with no change in sEPSC amplitude. No change in metabotropic glutamate receptor 2/3 (mGlu2/3) function was detected as bath application of the mGlu2/3 agonist LY379268 decreased spontaneous and miniature EPSC frequency in slices from both control and ethanol-consuming rats. The increase in basal glutamate was not associated with changes in the surface expression of GLT-1, however, a decrease in slope of the no-net-flux dialysis function was observed following ethanol consumption, indicating a potential decrease in glutamate reuptake. Taken together, these findings indicate that the increase in basal extracellular glutamate occurring after chronic ethanol consumption is not mediated by an increase in action potential-dependent glutamate release or a failure of mGlu2/3 autoreceptors to regulate such release. PMID:27207718

  1. Theoretical analysis of the glutamate dehydrogenase kinetics under physiological conditions.

    PubMed

    Popova, S V; Reich, J G

    1983-01-01

    A kinetic model of the glutamate dehydrogenase reaction has been formulated for the reversible reaction including all seven reactants (substrates and cofactors NAD(H) and NADP(H)). The model parameters have been evaluated from published initial-rate data. Analysis of the model at cofactor concentration near to that in the intact mitochondrion has shown that the competition for active sites between cofactors and substrates simultaneously present in mitochondria diminishes the steady-state rate of the reaction by a factor of 10 to 100 as compared to the maximal reaction rate. The model predicts near-equilibrium of the reaction substrates with NAD+/NADH cofactor pair and off-equilibrium with NADP+/NADPH. Substrate cycling with futile transfer of hydrogen from NADP+-system to NAD+-system has been found to account under in vivo conditions for no more than 2% of the maximal glutamate dehydrogenase activity in the mitochondria.

  2. The Role of Metabotropic Glutamate Receptor Genes in Schizophrenia.

    PubMed

    Maj, Carlo; Minelli, Alessandra; Giacopuzzi, Edoardo; Sacchetti, Emilio; Gennarelli, Massimo

    2016-01-01

    Genomic studies revealed two main components in the genetic architecture of schizophrenia, one constituted by common variants determining a distributed polygenic effect and one represented by a large number of heterogeneous rare and highly disruptive mutations. These gene modifications often affect neural transmission and different studies proved an involvement of metabotropic glutamate receptors in schizophrenia phenotype. Through the combination of literature information with genomic data from public repositories, we analyzed the current knowledge on the involvement of genetic variations of the human metabotropic glutamate receptors in schizophrenia and related endophenotypes. Despite the analysis did not reveal a definitive connection, different suggestive associations have been identified and in particular a relevant role has emerged for GRM3 in affecting specific schizophrenia endophenotypes. This supports the hypothesis that these receptors are directly involved in schizophrenia disorder. PMID:27296644

  3. Synthesis of biobased succinonitrile from glutamic acid and glutamine.

    PubMed

    Lammens, Tijs M; Le Nôtre, Jérôme; Franssen, Maurice C R; Scott, Elinor L; Sanders, Johan P M

    2011-06-20

    Succinonitrile is the precursor of 1,4-diaminobutane, which is used for the industrial production of polyamides. This paper describes the synthesis of biobased succinonitrile from glutamic acid and glutamine, amino acids that are abundantly present in many plant proteins. Synthesis of the intermediate 3-cyanopropanoic amide was achieved from glutamic acid 5-methyl ester in an 86 mol% yield and from glutamine in a 56 mol % yield. 3-Cyanopropanoic acid can be converted into succinonitrile, with a selectivity close to 100% and a 62% conversion, by making use of a palladium(II)-catalyzed equilibrium reaction with acetonitrile. Thus, a new route to produce biobased 1,4-diaminobutane has been discovered. PMID:21557494

  4. The Role of Metabotropic Glutamate Receptor Genes in Schizophrenia.

    PubMed

    Maj, Carlo; Minelli, Alessandra; Giacopuzzi, Edoardo; Sacchetti, Emilio; Gennarelli, Massimo

    2016-01-01

    Genomic studies revealed two main components in the genetic architecture of schizophrenia, one constituted by common variants determining a distributed polygenic effect and one represented by a large number of heterogeneous rare and highly disruptive mutations. These gene modifications often affect neural transmission and different studies proved an involvement of metabotropic glutamate receptors in schizophrenia phenotype. Through the combination of literature information with genomic data from public repositories, we analyzed the current knowledge on the involvement of genetic variations of the human metabotropic glutamate receptors in schizophrenia and related endophenotypes. Despite the analysis did not reveal a definitive connection, different suggestive associations have been identified and in particular a relevant role has emerged for GRM3 in affecting specific schizophrenia endophenotypes. This supports the hypothesis that these receptors are directly involved in schizophrenia disorder.

  5. Leptin regulates glutamate and glucose transporters in hypothalamic astrocytes

    PubMed Central

    Fuente-Martín, Esther; García-Cáceres, Cristina; Granado, Miriam; de Ceballos, María L.; Sánchez-Garrido, Miguel Ángel; Sarman, Beatrix; Liu, Zhong-Wu; Dietrich, Marcelo O.; Tena-Sempere, Manuel; Argente-Arizón, Pilar; Díaz, Francisca; Argente, Jesús; Horvath, Tamas L.; Chowen, Julie A.

    2012-01-01

    Glial cells perform critical functions that alter the metabolism and activity of neurons, and there is increasing interest in their role in appetite and energy balance. Leptin, a key regulator of appetite and metabolism, has previously been reported to influence glial structural proteins and morphology. Here, we demonstrate that metabolic status and leptin also modify astrocyte-specific glutamate and glucose transporters, indicating that metabolic signals influence synaptic efficacy and glucose uptake and, ultimately, neuronal function. We found that basal and glucose-stimulated electrical activity of hypothalamic proopiomelanocortin (POMC) neurons in mice were altered in the offspring of mothers fed a high-fat diet. In adulthood, increased body weight and fasting also altered the expression of glucose and glutamate transporters. These results demonstrate that whole-organism metabolism alters hypothalamic glial cell activity and suggest that these cells play an important role in the pathology of obesity. PMID:23064363

  6. Glutamate modulators in the treatment of obsessive-compulsive disorder

    PubMed Central

    Pittenger, Christopher

    2015-01-01

    Established treatments for obsessive-compulsive disorder (OCD) are of benefit in approximately 3 of every 4 patients, but refractory disease remains distressingly common, and many treatment responders continue to experience considerable morbidity. This motivates a search for new insights into pathophysiology that may inform novel treatment strategies. Much recent work has focused on the neurotransmitter glutamate. Several lines of neurochemical and genetic evidence suggests that glutamate dysregulation may contribute to OCD, although much remains unclear. The off-label use of a number of pharmacological agents approved for other indications has been investigated in refractory OCD. We summarize investigations of memantine, riluzole, ketamine, D-cycloserine, glycine, N-acetylserine, topiramate, and lamotrigine. Evidence exists for benefit from each of these in some patients; though none has been proven effective with sufficient clarity to be considered part of standard care, these agents are options in individuals whose symptoms are refractory to better-established therapeutic strategies. PMID:26236057

  7. [Cardioprotective properties of new glutamic acid derivative under stress conditions].

    PubMed

    Perfilova, V N; Sadikova, N V; Berestovitskaia, V M; Vasil'eva, O S

    2014-01-01

    The effect of new glutamic acid derivative on the cardiac ino- and chronotropic functions has been studied in experiments on rats exposed to 24-hour immobilization-and-pain stress. It is established that glutamic acid derivative RGPU-238 (glufimet) at a dose of 28.7 mg/kg increases the increment of myocardial contractility and relaxation rates and left ventricular pressure in stress-tested animals by 13 1,1, 72.4, and 118.6%, respectively, as compared to the control group during the test for adrenoreactivity. Compound RGPU-238 increases the increment of the maximum intensity of myocardium functioning by 196.5 % at 30 sec of isometric workload as compared to the control group. The cardioprotective effect of compound RGPU-238 is 1.5 - 2 times higher than that of the reference drug phenibut.

  8. Metabotropic glutamate receptors: beyond the regulation of synaptic transmission.

    PubMed

    Nicoletti, Ferdinando; Battaglia, Giuseppe; Storto, Marianna; Ngomba, Richard T; Iacovelli, Luisa; Arcella, Antonietta; Gradini, Roberto; Sale, Patrizio; Rampello, Liborio; De Vita, Teresa; Di Marco, Roberto; Melchiorri, Daniela; Bruno, Valeria

    2007-08-01

    Metabotropic glutamate (mGlu) receptors are G-protein coupled receptors activated by glutamate, the major excitatory neurotransmitter of the CNS. A growing body of evidence suggests that the function of mGlu receptors is not restricted to the regulation of synaptic transmission. mGlu receptors are expressed in a variety of peripheral cells, including inter alia hepatocytes, pancreatic cells, osteoblasts and immune cells. Within the immunological synapses, mGlu receptors expressed by T cells might contribute to the vast array of signals generated by the antigen-presenting cells. mGlu receptors are also found in embryonic and neural stem cells. This suggests their involvement in the pathophysiology of brain tumors, which likely originates from cancer stem cells similar to neural stem cells. Ligands of mGlu3 and mGlu4 receptors are potential candidates for the experimental treatment of malignant gliomas and medulloblastomas, respectively. PMID:17651904

  9. Role of astrocytic glutamate transporter in alcohol use disorder

    PubMed Central

    Ayers-Ringler, Jennifer R; Jia, Yun-Fang; Qiu, Yan-Yan; Choi, Doo-Sup

    2016-01-01

    Alcohol use disorder (AUD) is one of the most widespread neuropsychiatric conditions, having a significant health and socioeconomic impact. According to the 2014 World Health Organization global status report on alcohol and health, the harmful use of alcohol is responsible for 5.9% of all deaths worldwide. Additionally, 5.1% of the global burden of disease and injury is ascribed to alcohol (measured in disability adjusted life years, or disability adjusted life years). Although the neurobiological basis of AUD is highly complex, the corticostriatal circuit contributes significantly to the development of addictive behaviors. In-depth investigation into the changes of the neurotransmitters in this circuit, dopamine, gamma-aminobutyricacid, and glutamate, and their corresponding neuronal receptors in AUD and other addictions enable us to understand the molecular basis of AUD. However, these discoveries have also revealed a dearth of knowledge regarding contributions from non-neuronal sources. Astrocytes, though intimately involved in synaptic function, had until recently been noticeably overlooked in their potential role in AUD. One major function of the astrocyte is protecting neurons from excitotoxicity by removing glutamate from the synapse via excitatory amino acid transporter type 2. The importance of this key transporter in addiction, as well as ethanol withdrawal, has recently become evident, though its regulation is still under investigation. Historically, pharmacotherapy for AUD has been focused on altering the activity of neuronal glutamate receptors. However, recent clinical evidence has supported the animal-based findings, showing that regulating glutamate homeostasis contributes to successful management of recovery from AUD. PMID:27014596

  10. Possible significance of adverse reactions to glutamate in humans.

    PubMed

    Reif-Lehrer, L

    1976-09-01

    Of those exposed to Chinese restaurant food, our studies indicate that 25% report adverse reactions (Chinese restaurant syndrome (CRS)), presumably to the mono-sodium glutamate (MSG) content. The possible significance of the symptoms is discussed in the light of the known neuroexcitatory activity of MSG. It is suggested that CRS may result from a "benign" inborn "error" of metabolism that is deserving of further study, particularly in individuals with certain other metabolic abnormalities or who are on certain types of drug therapy.

  11. [PECULIARITIES OF THE CEREBROVASCULAR EFFECTS OF GLUTAMIC ACID].

    PubMed

    Gan'shina, T S; Kurza, E V; Kurdyumov, I N; Maslennikov, D V; Mirzoyan, R S

    2016-01-01

    Experiments on nonlinear rats subjected to global transient cerebral ischemia revealed the ability of glutamic acid to improve cerebral circulation. Consequently, the excitatory amino acid can produce adverse (neurotoxic) and positive (anti-ischemic) effects in cerebral ischemia. The cerebrovascular effect of glutamic acid in cerebral ischemia is attenuated on the background action of the MNDA receptor blocker MK-801 (0.5 mg/kg intravenously) and eliminated by bicuculline. When glutamic acid is combined with the non-competitive MNDA receptor antagonist MK-801, neither one nor another drug shows its vasodilator effect. The results are indicative of the interaction between excitatory and inhibitory systems on the level of cerebral vessels and once again confirm our previous conclusion about the decisive role of GABA(A) receptors in brain vessels in the implementation of anti-ischemic activity of endogenous compounds (melatonin) and well-known pharmacological substances (mexidol, afobazole), and new chemical compounds based on GABA-containing lipid derivatives. PMID:27455572

  12. Group III metabotropic glutamate receptors and drug addiction

    PubMed Central

    Mao, Limin; Guo, Minglei; Jin, Daozhong; Xue, Bing; Wang, John Q.

    2014-01-01

    Neuroadaptations of glutamatergic transmission in the limbic reward circuitry are linked to persistent drug addiction. Accumulating data have demonstrated roles of ionotropic glutamate receptors and group I and II metabotropic glutamate receptors (mGluRs) in this event. Emerging evidence also identifies Gαi/o-coupled group III mGluRs (mGluR4/7/8 subtypes enriched in the limbic system) as direct substrates of drugs of abuse and active regulators of drug action. Auto- and heteroreceptors of mGluR4/7/8 reside predominantly on nerve terminals of glutamatergic corticostriatal and GABAergic striatopallidal pathways, respectively. These presynaptic receptors regulate basal and/or phasic release of respective transmitters to maintain basal ganglia homeostasis. In response to operant administration of common addictive drugs, such as psychostimulants (cocaine and amphetamine), alcohol and opiates, limbic group III mGluRs undergo drastic adaptations to contribute to the enduring remodeling of excitatory synapses and to usually suppress drug seeking behavior. As a result, a loss-of-function mutation (knockout) of individual group III receptor subtypes often promotes drug seeking. This review summarizes the data from recent studies on three group III receptor subtypes (mGluR4/7/8) expressed in the basal ganglia and analyzes their roles in the regulation of dopamine and glutamate signaling in the striatum and their participation in the addictive properties of three major classes of drugs (psychostimulants, alcohol, and opiates). PMID:24078068

  13. TAAR1 Modulates Cortical Glutamate NMDA Receptor Function

    PubMed Central

    Espinoza, Stefano; Lignani, Gabriele; Caffino, Lucia; Maggi, Silvia; Sukhanov, Ilya; Leo, Damiana; Mus, Liudmila; Emanuele, Marco; Ronzitti, Giuseppe; Harmeier, Anja; Medrihan, Lucian; Sotnikova, Tatyana D; Chieregatti, Evelina; Hoener, Marius C; Benfenati, Fabio; Tucci, Valter; Fumagalli, Fabio; Gainetdinov, Raul R

    2015-01-01

    Trace Amine-Associated Receptor 1 (TAAR1) is a G protein-coupled receptor expressed in the mammalian brain and known to influence subcortical monoaminergic transmission. Monoamines, such as dopamine, also play an important role within the prefrontal cortex (PFC) circuitry, which is critically involved in high-o5rder cognitive processes. TAAR1-selective ligands have shown potential antipsychotic, antidepressant, and pro-cognitive effects in experimental animal models; however, it remains unclear whether TAAR1 can affect PFC-related processes and functions. In this study, we document a distinct pattern of expression of TAAR1 in the PFC, as well as altered subunit composition and deficient functionality of the glutamate N-methyl-D-aspartate (NMDA) receptors in the pyramidal neurons of layer V of PFC in mice lacking TAAR1. The dysregulated cortical glutamate transmission in TAAR1-KO mice was associated with aberrant behaviors in several tests, indicating a perseverative and impulsive phenotype of mutants. Conversely, pharmacological activation of TAAR1 with selective agonists reduced premature impulsive responses observed in the fixed-interval conditioning schedule in normal mice. Our study indicates that TAAR1 plays an important role in the modulation of NMDA receptor-mediated glutamate transmission in the PFC and related functions. Furthermore, these data suggest that the development of TAAR1-based drugs could provide a novel therapeutic approach for the treatment of disorders related to aberrant cortical functions. PMID:25749299

  14. Dysfunction of Glutamate Receptors in Microglia May Cause Neurodegeneration.

    PubMed

    Noda, Mami

    2016-01-01

    Dysregulation of glutamate signalling is important in Alzheimer's disease and other pathologies. There has been a focus on changes in neuronal glutamate signalling, but microglia also express glutamate receptors (GluRs), which are known to modulate their responses to neuropathology. Microglia express both metabotropic and ionotropic GluRs. Among ionotropic GluRs, microglial AMPA (α-amino-hydroxy-5-methyl-isoxazole-4-propionate)-type of GluRs (AMPA-Rs) are Ca2+ impermeable due to the expression of subunit GluA2. Upon activation of microglia, expression level of surface GluA2 subunits significantly increase, while expression of GluA1, A3 and A4 subunits on membrane surface significantly decrease. Owing to the GluA2 subunits-dominant composition, AMPA-Rs in activated microglia show little response to Glu. On the other hand, microglia lacking GluA2 show higher Ca(2+)-permeability, consequently inducing a significant increase in the release of the pro-inflammatory cytokine, such as TNF-α. It is suggested that membrane translocation of GluA2-containing AMPA-Rs in activated microglia has functional importance. Thus, dysfunction or decreased expression of GluA2 reported in patients with neurodegenerative diseases such as Alzheimer's and Creutzfeldt-Jakob disease may accelerate Glu neurotoxicity via excess release of proinflammatory cytokines from microglia, causing more neuronal death. PMID:26567741

  15. MicroRNA-223 is neuroprotective by targeting glutamate receptors.

    PubMed

    Harraz, Maged M; Eacker, Stephen M; Wang, Xueqing; Dawson, Ted M; Dawson, Valina L

    2012-11-13

    Stroke is a major cause of mortality and morbidity worldwide. Extracellular glutamate accumulation leading to overstimulation of the ionotropic glutamate receptors mediates neuronal injury in stroke and in neurodegenerative disorders. Here we show that miR-223 controls the response to neuronal injury by regulating the functional expression of the glutamate receptor subunits GluR2 and NR2B in brain. Overexpression of miR-223 lowers the levels of GluR2 and NR2B by targeting 3'-UTR target sites (TSs) in GluR2 and NR2B, inhibits NMDA-induced calcium influx in hippocampal neurons, and protects the brain from neuronal cell death following transient global ischemia and excitotoxic injury. MiR-223 deficiency results in higher levels of NR2B and GluR2, enhanced NMDA-induced calcium influx, and increased miniature excitatory postsynaptic currents in hippocampal neurons. In addition, the absence of MiR-223 leads to contextual, but not cued memory deficits and increased neuronal cell death following transient global ischemia and excitotoxicity. These data identify miR-223 as a major regulator of the expression of GluR2 and NR2B, and suggest a therapeutic role for miR-223 in stroke and other excitotoxic neuronal disorders.

  16. Targeting glutamate uptake to treat alcohol use disorders

    PubMed Central

    Rao, P.S.S.; Bell, Richard L.; Engleman, Eric A.; Sari, Youssef

    2015-01-01

    Alcoholism is a serious public health concern that is characterized by the development of tolerance to alcohol's effects, increased consumption, loss of control over drinking and the development of physical dependence. This cycle is often times punctuated by periods of abstinence, craving and relapse. The development of tolerance and the expression of withdrawal effects, which manifest as dependence, have been to a great extent attributed to neuroadaptations within the mesocorticolimbic and extended amygdala systems. Alcohol affects various neurotransmitter systems in the brain including the adrenergic, cholinergic, dopaminergic, GABAergic, glutamatergic, peptidergic, and serotonergic systems. Due to the myriad of neurotransmitter and neuromodulator systems affected by alcohol, the efficacies of current pharmacotherapies targeting alcohol dependence are limited. Importantly, research findings of changes in glutamatergic neurotransmission induced by alcohol self- or experimenter-administration have resulted in a focus on therapies targeting glutamatergic receptors and normalization of glutamatergic neurotransmission. Glutamatergic receptors implicated in the effects of ethanol include the ionotropic glutamate receptors (AMPA, Kainate, and NMDA) and some metabotropic glutamate receptors. Regarding glutamatergic homeostasis, ceftriaxone, MS-153, and GPI-1046, which upregulate glutamate transporter 1 (GLT1) expression in mesocorticolimbic brain regions, reduce alcohol intake in genetic animal models of alcoholism. Given the hyperglutamatergic/hyperexcitable state of the central nervous system induced by chronic alcohol abuse and withdrawal, the evidence thus far indicates that a restoration of glutamatergic concentrations and activity within the mesocorticolimbic system and extended amygdala as well as multiple memory systems holds great promise for the treatment of alcohol dependence. PMID:25954150

  17. Glutamate dehydrogenase from pumpkin cotyledons: characterization and isoenzymes.

    PubMed

    Chou, K H; Splittstoesser, W E

    1972-04-01

    Glutamate dehydrogenase from pumpkin (Cucurbita moschata Pior. cultivar Dickinson Field) cotyledons was found in both soluble and particulate fractions with the bulk of the activity in the soluble fraction. Both enzymes used NAD(H) and NADP(H) but NAD(H) was favored. The enzymes were classified as glutamate-NAD oxidoreductase, deaminating (EC 1.4.1.3). Both enzymes were heat stable, had a pH optimum for reductive amination of 8.0, and were inhibited by high concentrations of NH(4) (+) or alpha-ketoglutarate. The soluble enzyme was more sensitive to NH(4) (+) inhibition and was activated by metal ions after ammonium sulfate fractionation while the solubilized particulate enzyme was not. Inhibition by ethylenediaminetetraacetate was restored by several divalent ions and inhibition by p-hydroxymercuribenzoate was reversed by glutathione. Particulate glutamate dehydrogenase showed a greater activity with NADP. The molecular weights of the enzymes are 250,000. Separation of the enzymes by disc gel electrophoresis showed that during germination the soluble isoenzymes increased from 1 to 7 in number, while only one particulate isoenzyme was found at any time. This particulate isoenzyme was identical with one of the soluble isoenzymes. A number of methods indicated that the soluble isoenzymes were not simply removed from the particulate fraction and that true isoenzymes were found.

  18. Glutamate Receptor Homologs in Plants: Functions and Evolutionary Origins

    PubMed Central

    Price, Michelle Beth; Jelesko, John; Okumoto, Sakiko

    2012-01-01

    The plant glutamate-like receptor homologs (GLRs) are homologs of mammalian ionotropic glutamate receptors (iGluRs) which were discovered more than 10 years ago, and are hypothesized to be potential amino acid sensors in plants. Although initial progress on this gene family has been hampered by gene redundancy and technical issues such as gene toxicity; genetic, pharmacological, and electrophysiological approaches are starting to uncover the functions of this protein family. In parallel, there has been tremendous progress in elucidating the structure of animal glutamate receptors (iGluRs), which in turn will help understanding of the molecular mechanisms of plant GLR functions. In this review, we will summarize recent progress on the plant GLRs. Emerging evidence implicates plant GLRs in various biological processes in and beyond N sensing, and implies that there is some overlap in the signaling mechanisms of amino acids between plants and animals. Phylogenetic analysis using iGluRs from metazoans, plants, and bacteria showed that the plant GLRs are no more closely related to metazoan iGluRs as they are to bacterial iGluRs, indicating the separation of plant, other eukaryotic, and bacterial GLRs might have happened as early on as the last universal common ancestor. Structural similarities and differences with animal iGluRs, and the implication thereof, are also discussed. PMID:23115559

  19. Crystal structure of a chimaeric bacterial glutamate dehydrogenase.

    PubMed

    Oliveira, Tânia; Sharkey, Michael A; Engel, Paul C; Khan, Amir R

    2016-06-01

    Glutamate dehydrogenases (EC 1.4.1.2-4) catalyse the oxidative deamination of L-glutamate to α-ketoglutarate using NAD(P)(+) as a cofactor. The bacterial enzymes are hexameric, arranged with 32 symmetry, and each polypeptide consists of an N-terminal substrate-binding segment (domain I) followed by a C-terminal cofactor-binding segment (domain II). The catalytic reaction takes place in the cleft formed at the junction of the two domains. Distinct signature sequences in the nucleotide-binding domain have been linked to the binding of NAD(+) versus NADP(+), but they are not unambiguous predictors of cofactor preference. In the absence of substrate, the two domains move apart as rigid bodies, as shown by the apo structure of glutamate dehydrogenase from Clostridium symbiosum. Here, the crystal structure of a chimaeric clostridial/Escherichia coli enzyme has been determined in the apo state. The enzyme is fully functional and reveals possible determinants of interdomain flexibility at a hinge region following the pivot helix. The enzyme retains the preference for NADP(+) cofactor from the parent E. coli domain II, although there are subtle differences in catalytic activity. PMID:27303899

  20. An Optimized Glutamate Receptor Photoswitch with Sensitized Azobenzene Isomerization.

    PubMed

    Gascón-Moya, Marta; Pejoan, Arnau; Izquierdo-Serra, Mercè; Pittolo, Silvia; Cabré, Gisela; Hernando, Jordi; Alibés, Ramon; Gorostiza, Pau; Busqué, Félix

    2015-10-16

    A new azobenzene-based photoswitch, 2, has been designed to enable optical control of ionotropic glutamate receptors in neurons via sensitized two-photon excitation with NIR light. In order to develop an efficient and versatile synthetic route for this molecule, a modular strategy is described which relies on the use of a new linear fully protected glutamate derivative stable in basic media. The resulting compound undergoes one-photon trans-cis photoisomerization via two different mechanisms: direct excitation of its azoaromatic unit and irradiation of the pyrene sensitizer, a well-known two-photon sensitive chromophore. Moreover, 2 presents large thermal stability of its cis isomer, in contrast to other two-photon responsive switches relying on the intrinsic nonlinear optical properties of push-pull substituted azobenzenes. As a result, the molecular system developed herein is a very promising candidate for evoking large photoinduced biological responses during the multiphoton operation of neuronal glutamate receptors with NIR light, which require accumulation of the protein-bound cis state of the switch upon repeated illumination.

  1. Pharmacology of Glutamate Transport in the CNS: Substrates and Inhibitors of Excitatory Amino Acid Transporters (EAATs) and the Glutamate/Cystine Exchanger System x c -

    NASA Astrophysics Data System (ADS)

    Bridges, Richard J.; Patel, Sarjubhai A.

    As the primary excitatory neurotransmitter in the mammalian CNS, l-glutamate participates not only in standard fast synaptic communication, but also contributes to higher order signal processing, as well as neuropathology. Given this variety of functional roles, interest has been growing as to how the extracellular concentrations of l-glutamate surrounding neurons are regulated by cellular transporter proteins. This review focuses on two prominent systems, each of which appears capable of influencing both the signaling and pathological actions of l-glutamate within the CNS: the sodium-dependent excitatory amino acid transporters (EAATs) and the glutamate/cystine exchanger, system x c - (Sx c -). While the family of EAAT subtypes limit access to glutamate receptors by rapidly and efficiently sequestering l-glutamate in neurons and glia, Sxc - provides a route for the export of glutamate from cells into the extracellular environment. The primary intent of this work is to provide an overview of the inhibitors and substrates that have been developed to delineate the pharmacological specificity of these transport systems, as well as be exploited as probes with which to selectively investigate function. Particular attention is paid to the development of small molecule templates that mimic the structural properties of the endogenous substrates, l-glutamate, l-aspartate and l-cystine and how strategic control of functional group position and/or the introduction of lipophilic R-groups can impact multiple aspects of the transport process, including: subtype selectivity, inhibitory potency, and substrate activity.

  2. Effect of carnitine on muscular glutamate uptake and intramuscular glutathione in malignant diseases.

    PubMed

    Breitkreutz, R; Babylon, A; Hack, V; Schuster, K; Tokus, M; Böhles, H; Hagmüller, E; Edler, L; Holm, E; Dröge, W

    2000-01-01

    Abnormally low intramuscular glutamate and glutathione (GSH) levels and/or a decreased muscular uptake of glutamate by the skeletal muscle tissue have previously been found in malignant diseases and simian immunodeficiency virus (SIV) infection and may contribute to the development of cachexia. We tested the hypothesis that an impaired mitochondrial energy metabolism may compromise the Na+-dependent glutamate transport. A randomized double-blind clinical trial was designed to study the effects of L-carnitine, i.e. an agent known to enhance mitochondrial integrity and function, on the glutamate transport and plasma glutamate level of cancer patients. The effect of carnitine on the intramuscular glutamate and GSH levels was examined in complementary experiments with tumour-bearing mice. In the mice, L-carnitine treatment ameliorated indeed the tumour-induced decrease in muscular glutamate and GSH levels and the increase in plasma glutamate levels. The carnitine-treated group in the randomized clinical study showed also a significant decrease in the plasma glutamate levels but only a moderate and statistically not significant increase in the relative glutamate uptake in the lower extremities. Further studies may be warranted to determine the effect of L-carnitine on the intramuscular GSH levels in cancer patients.

  3. Morphine Protects Spinal Cord Astrocytes from Glutamate-Induced Apoptosis via Reducing Endoplasmic Reticulum Stress

    PubMed Central

    Zhang, Chao; Wang, Chendan; Ren, Jianbo; Guo, Xiangjie; Yun, Keming

    2016-01-01

    Glutamate is not only a neurotransmitter but also an important neurotoxin in central nervous system (CNS). Chronic elevation of glutamate induces both neuronal and glial cell apoptosis. However, its effect on astrocytes is complex and still remains unclear. In this study, we investigated whether morphine, a common opioid ligand, could affect glutamate-induced apoptosis in astrocytes. Primary cultured astrocytes were incubated with glutamate in the presence/absence of morphine. It was found that morphine could reduce glutamate-induced apoptosis of astrocytes. Furthermore, glutamate activated Ca2+ release, thereby inducing endoplasmic reticulum (ER) stress in astrocytes, while morphine attenuated this deleterious effect. Using siRNA to reduce the expression of κ-opioid receptor, morphine could not effectively inhibit glutamate-stimulated Ca2+ release in astrocytes, the protective effect of morphine on glutamate-injured astrocytes was also suppressed. These results suggested that morphine could protect astrocytes from glutamate-induced apoptosis via reducing Ca2+ overload and ER stress pathways. In conclusion, this study indicated that excitotoxicity participated in the glutamate mediated apoptosis in astrocytes, while morphine attenuated this deleterious effect via regulating Ca2+ release and ER stress. PMID:27783050

  4. Glutamate-mediated protection of crayfish glial cells from PDT-induced apoptosis

    NASA Astrophysics Data System (ADS)

    Rudkovskii, M. V.; Romanenko, N. P.; Berezhnaya, E. V.; Kovaleva, V. D.; Uzdensky, A. B.

    2010-10-01

    Photodynamic treatment that causes intense oxidative stress and kills cells is currently used in neurooncology. However, along with tumor it damages surrounding healthy neurons and glial cells. In order to study the possible role of glutamate-related signaling pathways in photodynamic injury of neurons and glia, we investigated photodynamic effect of alumophthalocyanine Photosens on isolated crayfish stretch receptor that consists of a single neuron surrounded by glial cells. The laser diode (670 nm, 0.4 W/cm2) was used for dye photoexcitation. Application of glutamate increased photodynamically induced necrosis of neurons and glial cells but significantly decreased glial apoptosis. The natural neuroglial mediator N-acetylaspartylglutamate, which releases glutamate after cleavage in the extracellular space by glutamate carboxypeptidase II, also inhibited photoinduced apoptosis. Inhibition of glutamate carboxypeptidase II, oppositely, enhanced apoptosis of glial cells. These data confirm the anti-apoptotic activity of glutamate. Application of NMDA or inhibition of NMDA receptors by MK801 did not influence photodynamic death of neurons and glial cells that indicated nonparticipation of NMDA receptors in these processes. Inhibition of metabotropic glutamate receptors by AP-3 decreased PDT-induced apoptosis. One can suggest that crayfish neurons naturally secrete NAAG, which being cleaved by GCOP produces glutamate. Glutamate prevents photoinduced apoptosis of glial cells possibly through metabotropic but not ionotropic glutamate receptors.

  5. Glutamate-mediated protection of crayfish glial cells from PDT-induced apoptosis

    NASA Astrophysics Data System (ADS)

    Rudkovskii, M. V.; Romanenko, N. P.; Berezhnaya, E. V.; Kovaleva, V. D.; Uzdensky, A. B.

    2011-03-01

    Photodynamic treatment that causes intense oxidative stress and kills cells is currently used in neurooncology. However, along with tumor it damages surrounding healthy neurons and glial cells. In order to study the possible role of glutamate-related signaling pathways in photodynamic injury of neurons and glia, we investigated photodynamic effect of alumophthalocyanine Photosens on isolated crayfish stretch receptor that consists of a single neuron surrounded by glial cells. The laser diode (670 nm, 0.4 W/cm2) was used for dye photoexcitation. Application of glutamate increased photodynamically induced necrosis of neurons and glial cells but significantly decreased glial apoptosis. The natural neuroglial mediator N-acetylaspartylglutamate, which releases glutamate after cleavage in the extracellular space by glutamate carboxypeptidase II, also inhibited photoinduced apoptosis. Inhibition of glutamate carboxypeptidase II, oppositely, enhanced apoptosis of glial cells. These data confirm the anti-apoptotic activity of glutamate. Application of NMDA or inhibition of NMDA receptors by MK801 did not influence photodynamic death of neurons and glial cells that indicated nonparticipation of NMDA receptors in these processes. Inhibition of metabotropic glutamate receptors by AP-3 decreased PDT-induced apoptosis. One can suggest that crayfish neurons naturally secrete NAAG, which being cleaved by GCOP produces glutamate. Glutamate prevents photoinduced apoptosis of glial cells possibly through metabotropic but not ionotropic glutamate receptors.

  6. A Glutamic Acid-Producing Lactic Acid Bacteria Isolated from Malaysian Fermented Foods

    PubMed Central

    Zareian, Mohsen; Ebrahimpour, Afshin; Bakar, Fatimah Abu; Mohamed, Abdul Karim Sabo; Forghani, Bita; Ab-Kadir, Mohd Safuan B.; Saari, Nazamid

    2012-01-01

    l-glutamaic acid is the principal excitatory neurotransmitter in the brain and an important intermediate in metabolism. In the present study, lactic acid bacteria (218) were isolated from six different fermented foods as potent sources of glutamic acid producers. The presumptive bacteria were tested for their ability to synthesize glutamic acid. Out of the 35 strains showing this capability, strain MNZ was determined as the highest glutamic-acid producer. Identification tests including 16S rRNA gene sequencing and sugar assimilation ability identified the strain MNZ as Lactobacillus plantarum. The characteristics of this microorganism related to its glutamic acid-producing ability, growth rate, glucose consumption and pH profile were studied. Results revealed that glutamic acid was formed inside the cell and excreted into the extracellular medium. Glutamic acid production was found to be growth-associated and glucose significantly enhanced glutamic acid production (1.032 mmol/L) compared to other carbon sources. A concentration of 0.7% ammonium nitrate as a nitrogen source effectively enhanced glutamic acid production. To the best of our knowledge this is the first report of glutamic acid production by lactic acid bacteria. The results of this study can be further applied for developing functional foods enriched in glutamic acid and subsequently γ-amino butyric acid (GABA) as a bioactive compound. PMID:22754309

  7. Effect of carnitine on muscular glutamate uptake and intramuscular glutathione in malignant diseases

    PubMed Central

    Breitkreutz, R; Babylon, A; Hack, V; Schuster, K; Tokus, M; Böhles, H; Hagmüller, E; Edler, L; Holm, E; Dröge, W

    2000-01-01

    Abnormally low intramuscular glutamate and glutathione (GSH) levels and/or a decreased muscular uptake of glutamate by the skeletal muscle tissue have previously been found in malignant diseases and simian immunodeficiency virus (SIV) infection and may contribute to the development of cachexia. We tested the hypothesis that an impaired mitochondrial energy metabolism may compromise the Na+-dependent glutamate transport. A randomized double-blind clinical trial was designed to study the effects of L -carnitine, i.e. an agent known to enhance mitochondrial integrity and function, on the glutamate transport and plasma glutamate level of cancer patients. The effect of carnitine on the intramuscular glutamate and GSH levels was examined in complementary experiments with tumour-bearing mice. In the mice, L -carnitine treatment ameliorated indeed the tumour-induced decrease in muscular glutamate and GSH levels and the increase in plasma glutamate levels. The carnitine-treated group in the randomized clinical study showed also a significant decrease in the plasma glutamate levels but only a moderate and statistically not significant increase in the relative glutamate uptake in the lower extremities. Further studies may be warranted to determine the effect of L -carnitine on the intramuscular GSH levels in cancer patients. © 2000 Cancer Research Campaign PMID:10646895

  8. Delayed post-conditioning reduces post-ischemic glutamate level and improves protein synthesis in brain.

    PubMed

    Bonova, Petra; Burda, Jozef; Danielisova, Viera; Nemethova, Miroslava; Gottlieb, Miroslav

    2013-05-01

    In the clinic delayed post-conditioning would represent an attractive strategy for the survival of vulnerable neurons after an ischemic event. In this paper we studied the impact of ischemia and delayed post-conditioning on blood and brain tissue concentrations of glutamate and protein synthesis. We designed two groups of animals for analysis of brain tissues and blood after global ischemia and post-conditioning, and one for analysis of blood glutamate after transient focal ischemia. Our results showed elevated blood glutamate in two models of transient brain ischemia and decreases in blood glutamate to control in the first 20min of post-conditioning recirculation followed by a consecutive drop of about 20.5% on the first day. Similarly, we recorded reduced protein synthesis in hippocampus and cortex 2 and 3days after ischemia. However, increased glutamate was registered only in the hippocampus. Post-conditioning improves protein synthesis in CA1 and dentate gyrus and, surprisingly, leads to 50% reduction in glutamate in whole hippocampus and cortex. In conclusion, ischemia leads to meaningful elevation of blood and tissue glutamate. Post-conditioning activates mechanisms resulting in rapid elimination of glutamate from brain tissue and/or in the circulatory system that could otherwise impede brain-to-blood glutamate efflux mechanisms. Moreover, post-conditioning induces protein synthesis renewing in ischemia affected tissues that could also contribute to elimination of excitotoxicity. In addition, the potential of glutamate for monitoring the progress of ischemia and efficacy of therapy was shown.

  9. Glutamate-related gene expression changes with age in the mouse auditory midbrain.

    PubMed

    Tadros, Sherif F; D'Souza, Mary; Zettel, Martha L; Zhu, Xiaoxia; Waxmonsky, Nicole C; Frisina, Robert D

    2007-01-01

    Glutamate is the main excitatory neurotransmitter in both the peripheral and central auditory systems. Changes of glutamate and glutamate-related genes with age may be an important factor in the pathogenesis of age-related hearing loss-presbycusis. In this study, changes in glutamate-related mRNA gene expression in the CBA mouse inferior colliculus with age and hearing loss were examined and correlations were sought between these changes and functional hearing measures, such as the auditory brainstem response (ABR) and distortion product otoacoustic emissions (DPOAEs). Gene expression of 68 glutamate-related genes was investigated using both genechip microarray and real-time PCR (qPCR) molecular techniques for four different age/hearing loss CBA mouse subject groups. Two genes showed consistent differences between groups for both the genechip and qPCR. Pyrroline-5-carboxylate synthetase enzyme (Pycs) showed down-regulation with age and a high-affinity glutamate transporter (Slc1a3) showed up-regulation with age and hearing loss. Since Pycs plays a role in converting glutamate to proline, its deficiency in old age may lead to both glutamate increases and proline deficiencies in the auditory midbrain, playing a role in the subsequent inducement of glutamate toxicity and loss of proline neuroprotective effects. The up-regulation of Slc1a3 gene expression may reflect a cellular compensatory mechanism to protect against age-related glutamate or calcium excitoxicity.

  10. Protein kinase C -dependent regulation of synaptosomal glutamate uptake under conditions of hypergravity

    NASA Astrophysics Data System (ADS)

    Borisova, Tatiana; Krisanova, Natalia; Borisov, Arseniy; Sivko, Roman

    Glutamate is not only the main excitatory neurotransmitter in the mammalian CNS, but also a potent neurotoxin. Excessive concentration of ambient glutamate over activates glutamate receptors and causes neurotoxicity. Uptake of glutamate from the extracellular space into nerve cells was mediated by sodium-dependent glutamate transporters located in the plasma membrane. It was shown that the activity of glutamate transporters in rat brain nerve terminals was decreased after hypergravity (centrifugation of rats in special containers at 10 G for 1 hour). This decrease may result from the reduction in the number of glutamate transporters expressed in the plasma membrane of nerve terminals after hypergravity that was regulated by protein kinase C. The possibility of the involvement of protein kinase C in the regulation of the activity of glutamate transporters was assessed under conditions of hypergravity. The effect of protein kinase C inhibitor GF 109 203X on synaptosomal L-[14C]glutamate uptake was analysed. It was shown that the inhibitor decreased L-[14C]glutamate uptake by 15 % in control but did not influence it after hypergravity. In control, the initial velocity of L-[14C]glutamate uptake in the presence of the inhibitor decreased from 2.5 ± 0.2 nmol x min-1 x mg-1 of proteins to 2.17 ± 0.1 nmol x min-1 x mg-1 of proteins, whereas after hypergravity this value lowered from 2.05 ± 0.1 nmol x min-1 x mg-1 of proteins to 2.04 ± 0.1 nmol x min-1 x mg-1 of proteins. Thus, protein kinase C -dependent alteration in the cell surface expression of glutamate transporters may be one of the causes of a decrease in the activity of glutamate transporters after hypergravity.

  11. The glutamate and neutral amino acid transporter family: physiological and pharmacological implications.

    PubMed

    Kanai, Yoshikatsu; Hediger, Matthias A

    2003-10-31

    The solute carrier family 1 (SLC1) is composed of five high affinity glutamate transporters, which exhibit the properties of the previously described system XAG-, as well as two Na+-dependent neutral amino acid transporters with characteristics of the so-called "ASC" (alanine, serine and cysteine). The SLC1 family members are structurally similar, with almost identical hydropathy profiles and predicted membrane topologies. The transporters have eight transmembrane domains and a structure reminiscent of a pore loop between the seventh and eighth domains [Neuron 21 (1998) 623]. However, each of these transporters exhibits distinct functional properties. Glutamate transporters mediate transport of L-Glu, L-Asp and D-Asp, accompanied by the cotransport of 3 Na+ and one 1 H+, and the countertransport of 1 K+, whereas ASC transporters mediate Na+-dependent exchange of small neutral amino acids such as Ala, Ser, Cys and Thr. Given the high concentrating capacity provided by the unique ion coupling pattern of glutamate transporters, they play crucial roles in protecting neurons against glutamate excitotoxicity in the central nervous system (CNS). The regulation and manipulation of their function is a critical issue in the pathogenesis and treatment of CNS disorders involving glutamate excitotoxicity. Loss of function of the glial glutamate transporter GLT1 (SLC1A2) has been implicated in the pathogenesis of amyotrophic lateral sclerosis (ALS), resulting in damage of adjacent motor neurons. The importance of glial glutamate transporters in protecting neurons from extracellular glutamate was further demonstrated in studies of the slc1A2 glutamate transporter knockout mouse. The findings suggest that therapeutic upregulation of GLT1 may be beneficial in a variety of pathological conditions. Selective inhibition of the neuronal glutamate transporter EAAC1 (SLC1A1) but not the glial glutamate transporters may be of therapeutic interest, allowing blockage of glutamate exit from

  12. A high throughput assay of diffusion through Cx43 gap junction channels with a microfluidic chip

    PubMed Central

    Bathany, Cédric; Beahm, Derek; Felske, James D.; Sachs, Frederick; Hua, Susan Z.

    2011-01-01

    This paper describes a microfluidic-based assay capable of measuring gap-junction mediated dye diffusion in cultured cells. The technique exploits multi-stream laminar flow to selectively expose cells to different environments, enabling continuous loading of cells in one compartment while monitoring, in real time, dye diffusion into cells of a neighboring compartment. A simple one dimensional diffusion model fit to the data extracted the diffusion coefficient of four different dyes, 5-(6)-carboxyfluorescein (CFDA), 5-chloromethylfluorescein (CMFDA), Oregon green 488 carboxylic acid and calcein. Different inhibitors were assayed for their ability to reduce dye coupling. The chip can screen multiple inhibitors in parallel in the same cell preparation, demonstrating its potential for high throughput. The technique provides a convenient method to measure gap junction mediated diffusion and a screen for drugs that affect gap junction communication. PMID:21182279

  13. Disease mutations in desmoplakin inhibit Cx43 membrane targeting mediated by desmoplakin–EB1 interactions

    PubMed Central

    Patel, Dipal M.; Dubash, Adi D.; Kreitzer, Geri

    2014-01-01

    Mechanisms by which microtubule plus ends interact with regions of cell–cell contact during tissue development and morphogenesis are not fully understood. We characterize a previously unreported interaction between the microtubule binding protein end-binding 1 (EB1) and the desmosomal protein desmoplakin (DP), and demonstrate that DP–EB1 interactions enable DP to modify microtubule organization and dynamics near sites of cell–cell contact. EB1 interacts with a region of the DP N terminus containing a hotspot for pathogenic mutations associated with arrhythmogenic cardiomyopathy (AC). We show that a subset of AC mutations, in addition to a mutation associated with skin fragility/woolly hair syndrome, impair gap junction localization and function by misregulating DP–EB1 interactions and altering microtubule dynamics. This work identifies a novel function for a desmosomal protein in regulating microtubules that affect membrane targeting of gap junction components, and elucidates a mechanism by which DP mutations may contribute to the development of cardiac and cutaneous diseases. PMID:25225338

  14. Glutamate and GABA-metabolizing enzymes in post-mortem cerebellum in Alzheimer's disease: phosphate-activated glutaminase and glutamic acid decarboxylase.

    PubMed

    Burbaeva, G Sh; Boksha, I S; Tereshkina, E B; Savushkina, O K; Prokhorova, T A; Vorobyeva, E A

    2014-10-01

    Enzymes of glutamate and GABA metabolism in postmortem cerebellum from patients with Alzheimer's disease (AD) have not been comprehensively studied. The present work reports results of original comparative study on levels of phosphate-activated glutaminase (PAG) and glutamic acid decarboxylase isoenzymes (GAD65/67) in autopsied cerebellum samples from AD patients and matched controls (13 cases in each group) as well as summarizes published evidence for altered levels of PAG and GAD65/67 in AD brain. Altered (decreased) levels of these enzymes and changes in links between amounts of these enzymes and other glutamate-metabolizing enzymes (such as glutamate dehydrogenase and glutamine synthetase-like protein) in AD cerebella suggest significantly impaired glutamate and GABA metabolism in this brain region, which was previously regarded as not substantially involved in AD pathogenesis.

  15. Decreased glutamate, glutamine and citrulline concentrations in plasma and muscle in endotoxemia cannot be reversed by glutamate or glutamine supplementation: a primary intestinal defect?

    PubMed

    Boutry, Claire; Matsumoto, Hideki; Bos, Cécile; Moinard, Christophe; Cynober, Luc; Yin, Yulong; Tomé, Daniel; Blachier, François

    2012-10-01

    Endotoxemia affects intestinal physiology. A decrease of circulating citrulline concentration is considered as a reflection of the intestinal function. Citrulline can be produced in enterocytes notably from glutamate and glutamine. The aim of this work was to determine if glutamate, glutamine and citrulline concentrations in blood, intestine and muscle are decreased by endotoxemia, and if supplementation with glutamate or glutamine can restore normal concentrations. We induced endotoxemia in rats by an intraperitoneal injection of 0.3 mg kg(-1) lipopolysaccharide (LPS). This led to a rapid anorexia, negative nitrogen balance and a transient increase of the circulating level of IL-6 and TNF-α. When compared with the values measured in pair fed (PF) animals, almost all circulating amino acids (AA) including citrulline decreased, suggesting a decrease of intestinal function. However, at D2 after LPS injection, most circulating AA concentrations were closed to the values recorded in the PF group. At that time, among AA, only glutamate, glutamine and citrulline were decreased in gastrocnemius muscle without change in intestinal mucosa. A supplementation with 4% monosodium glutamate (MSG) or an isomolar amount of glutamine failed to restore glutamate, glutamine and citrulline concentrations in plasma and muscle. However, MSG supplementation led to an accumulation of glutamate in the intestinal mucosa. In conclusion, endotoxemia rapidly but transiently decreased the circulating concentrations of almost all AA and more durably of glutamate, glutamine and citrulline in muscle. Supplementation with glutamate or glutamine failed to restore glutamate, glutamine and citrulline concentrations in plasma and muscles. The implication of a loss of the intestinal capacity for AA absorption and/or metabolism in endotoxemia (as judged from decreased citrulline plasma concentration) for explaining such results are discussed.

  16. Glutamate and tumor-associated epilepsy: glial cell dysfunction in the peritumoral environment

    PubMed Central

    Buckingham, Susan C.; Robel, Stefanie

    2013-01-01

    Seizures are a serious and debilitating co-morbidity of primary brain tumors that affect most patients, yet their etiology is poorly understood. In many CNS pathologies, including epilepsy and brain injury, high levels of extracellular glutamate have been implicated in seizure generation. It has been shown that gliomas release neurotoxic levels of glutamate through their high expression of system xc-. More recently it was shown that the surrounding peritumoral cortex is spontaneously hyperexcitable. In this review, we discuss how gliomas induce changes in the surrounding environment that may further contribute to elevated extracellular glutamate and tumor-associated seizures. Peritumoral astrocytes become reactive and lose their ability to remove glutamate, while microglia, in response to signals from glioma cells, may release glutamate. In addition, gliomas increase blood brain barrier permeability, allowing seizure-inducing serum components, including glutamate, into the peritumoral region. These factors, working together or alone, may influence the frequency and severity of tumor-associated epilepsy. PMID:23385090

  17. Recent progress in the synthesis and characterization of group II metabotropic glutamate receptor allosteric modulators.

    PubMed

    Sheffler, Douglas J; Pinkerton, Anthony B; Dahl, Russell; Markou, Athina; Cosford, Nicholas D P

    2011-08-17

    Group II metabotropic glutamate (mGlu) receptors consist of the metabotropic glutamate 2 (mGlu(2)) and metabotropic glutamate 3 (mGlu(3)) receptor subtypes which modulate glutamate transmission by second messenger activation to negatively regulate the activity of adenylyl cyclase. Excessive accumulation of glutamate in the perisynaptic extracellular region triggers mGlu(2) and mGlu(3) receptors to inhibit further release of glutamate. There is growing evidence that the modulation of glutamatergic neurotransmission by small molecule modulators of Group II mGlu receptors has significant potential for the treatment of several neuropsychiatric and neurodegenerative diseases. This review provides an overview of recent progress on the synthesis and pharmacological characterization of positive and negative allosteric modulators of the Group II mGlu receptors. PMID:22860167

  18. Recent Progress in the Synthesis and Characterization of Group II Metabotropic Glutamate Receptor Allosteric Modulators

    PubMed Central

    2011-01-01

    Group II metabotropic glutamate (mGlu) receptors consist of the metabotropic glutamate 2 (mGlu2) and metabotropic glutamate 3 (mGlu3) receptor subtypes which modulate glutamate transmission by second messenger activation to negatively regulate the activity of adenylyl cyclase. Excessive accumulation of glutamate in the perisynaptic extracellular region triggers mGlu2 and mGlu3 receptors to inhibit further release of glutamate. There is growing evidence that the modulation of glutamatergic neurotransmission by small molecule modulators of Group II mGlu receptors has significant potential for the treatment of several neuropsychiatric and neurodegenerative diseases. This review provides an overview of recent progress on the synthesis and pharmacological characterization of positive and negative allosteric modulators of the Group II mGlu receptors. PMID:22860167

  19. Enzyme-based flow injection analysis system for glutamine and glutamate in mammalian cell culture media.

    PubMed

    Mayer, C; Frauer, A; Schalkhammer, T; Pittner, F

    1999-03-01

    We present the setup of a flow injection analysis system designed for on-line monitoring of glutamate and glutamine. These amino acids represent a major energy source in mammalian cell culture. A cycling assay consisting of glutamate dehydrogenase and aspartate aminotransferase produces NADH proportional to the glutamate concentration in the sample. NADH is then measured spectrophotometrically. Glutamine is determined by conversion to glutamate which is fed into the cycling assay. The conversion of glutamine to glutamate is catalyzed by asparaginase. Asparaginase was used in place of glutaminase due to its relatively high reactivity with glutamine and a pH optimum similar to that of glutamate dehydrogenase. The enzymes were immobilized covalently to activated controlled pore glass beads and integrated into the flow injection analysis system. The application of the immobilized enzymes and the technical setup are presented in this paper.

  20. [The effect of cerebral glutamate enhanced level on the respiratory system of anesthetized rats].

    PubMed

    Aleksandrov, V G; Buĭ Tkhi, Kh; Aleksandrova, N P

    2012-07-01

    A cerebral level of glutamate is one of the determinants of the central mechanisms of respiratory control. It had been hypothesized that endogenous glutamate could have a modulating effect on the functioning of mechanisms for neural control of respiratory function. Acute experiments on spontaneuosly breathing, urethane-anesthetized rats had been performed to study the respiratory effects of cerebroventricular microinjection of glutamate. It has been shown that a higher level of cerebral glutamate increases breathing rate and electrical activity of the diaphragm, and strengthen the Hering-Breuer reflex. These effects had a clear character of the phase. The results confirm the hypothesis suggested and prove that the increase in cerebral levels of glutamate leads to the activation of glutamate receptors of various types.

  1. Magnesium Sulfate Protects Against the Bioenergetic Consequences of Chronic Glutamate Receptor Stimulation

    PubMed Central

    Clerc, Pascaline; Young, Christina A.; Bordt, Evan A.; Grigore, Alina M.; Fiskum, Gary; Polster, Brian M.

    2013-01-01

    Extracellular glutamate is elevated following brain ischemia or trauma and contributes to neuronal injury. We tested the hypothesis that magnesium sulfate (MgSO4, 3 mM) protects against metabolic failure caused by excitotoxic glutamate exposure. Rat cortical neuron preparations treated in medium already containing a physiological concentration of Mg2+ (1 mM) could be segregated based on their response to glutamate (100 µM). Type I preparations responded with a decrease or small transient increase in oxygen consumption rate (OCR). Type II neurons responded with >50% stimulation in OCR, indicating a robust response to increased energy demand without immediate toxicity. Pre-treatment with MgSO4 improved the initial bioenergetic response to glutamate and ameliorated subsequent loss of spare respiratory capacity, measured following addition of the uncoupler FCCP, in Type I but not Type II neurons. Spare respiratory capacity in Type I neurons was also improved by incubation with MgSO4 or NMDA receptor antagonist MK801 in the absence of glutamate treatment. This finding indicates that the major difference between Type I and Type II preparations is the amount of endogenous glutamate receptor activity. Incubation of Type II neurons with 5 µM glutamate prior to excitotoxic (100 µM) glutamate exposure recapitulated a Type I phenotype. MgSO4 protected against an excitotoxic glutamate-induced drop in neuronal ATP both with and without prior 5 µM glutamate exposure. Results indicate that MgSO4 protects against chronic moderate glutamate receptor stimulation and preserves cellular ATP following treatment with excitotoxic glutamate. PMID:24236167

  2. Detection of glutamate release from neurons by genetically encoded surface-displayed FRET nanosensors

    NASA Astrophysics Data System (ADS)

    Okumoto, Sakiko; Looger, Loren L.; Micheva, Kristina D.; Reimer, Richard J.; Smith, Stephen J.; Frommer, Wolf B.

    2005-06-01

    Glutamate is the predominant excitatory neurotransmitter in the mammalian brain. Once released, its rapid removal from the synaptic cleft is critical for preventing excitotoxicity and spillover to neighboring synapses. Despite consensus on the role of glutamate in normal and disease physiology, technical issues limit our understanding of its metabolism in intact cells. To monitor glutamate levels inside and at the surface of living cells, genetically encoded nanosensors were developed. The fluorescent indicator protein for glutamate (FLIPE) consists of the glutamate/aspartate binding protein ybeJ from Escherichia coli fused to two variants of the green fluorescent protein. Three sensors with lower affinities for glutamate were created by mutation of residues peristeric to the ybeJ binding pocket. In the presence of ligands, FLIPEs show a concentration-dependent decrease in FRET efficiency. When expressed on the surface of rat hippocampal neurons or PC12 cells, the sensors respond to extracellular glutamate with a reversible concentration-dependent decrease in FRET efficiency. Depolarization of neurons leads to a reduction in FRET efficiency corresponding to 300 nM glutamate at the cell surface. No change in FRET was observed when cells expressing sensors in the cytosol were superfused with up to 20 mM glutamate, consistent with a minimal contribution of glutamate uptake to cytosolic glutamate levels. The results demonstrate that FLIPE sensors can be used for real-time monitoring of glutamate metabolism in living cells, in tissues, or in intact organisms, providing tools for studying metabolism or for drug discovery. aspartate | hippocampal neuron | neurotransmitter | secretion | transport

  3. Astrocytic energetics during excitatory neurotransmission: What are contributions of glutamate oxidation and glycolysis?

    PubMed

    Dienel, Gerald A

    2013-10-01

    Astrocytic energetics of excitatory neurotransmission is controversial due to discrepant findings in different experimental systems in vitro and in vivo. The energy requirements of glutamate uptake are believed by some researchers to be satisfied by glycolysis coupled with shuttling of lactate to neurons for oxidation. However, astrocytes increase glycogenolysis and oxidative metabolism during sensory stimulation in vivo, indicating that other sources of energy are used by astrocytes during brain activation. Furthermore, glutamate uptake into cultured astrocytes stimulates glutamate oxidation and oxygen consumption, and glutamate maintains respiration as well as glucose. The neurotransmitter pool of glutamate is associated with the faster component of total glutamate turnover in vivo, and use of neurotransmitter glutamate to fuel its own uptake by oxidation-competent perisynaptic processes has two advantages, substrate is supplied concomitant with demand, and glutamate spares glucose for use by neurons and astrocytes. Some, but not all, perisynaptic processes of astrocytes in adult rodent brain contain mitochondria, and oxidation of only a small fraction of the neurotransmitter glutamate taken up into these structures would be sufficient to supply the ATP required for sodium extrusion and conversion of glutamate to glutamine. Glycolysis would, however, be required in perisynaptic processes lacking oxidative capacity. Three lines of evidence indicate that critical cornerstones of the astrocyte-to-neuron lactate shuttle model are not established and normal brain does not need lactate as supplemental fuel: (i) rapid onset of hemodynamic responses to activation delivers oxygen and glucose in excess of demand, (ii) total glucose utilization greatly exceeds glucose oxidation in awake rodents during activation, indicating that the lactate generated is released, not locally oxidized, and (iii) glutamate-induced glycolysis is not a robust phenotype of all astrocyte cultures

  4. Coupling of glutamate and glucose uptake in cultured Bergmann glial cells.

    PubMed

    Mendez-Flores, Orquidia G; Hernández-Kelly, Luisa C; Suárez-Pozos, Edna; Najimi, Mustapha; Ortega, Arturo

    2016-09-01

    Glutamate, the main excitatory neurotransmitter in the vertebrate brain, exerts its actions through specific membrane receptors present in neurons and glial cells. Over-stimulation of glutamate receptors results in neuronal death, phenomena known as excitotoxicity. A family of sodium-dependent, glutamate uptake transporters mainly expressed in glial cells, removes the amino acid from the synaptic cleft preventing neuronal death. The sustained sodium influx associated to glutamate removal in glial cells, activates the sodium/potassium ATPase restoring the ionic balance, additionally, glutamate entrance activates glutamine synthetase, both events are energy demanding, therefore glia cells increase their ATP expenditure favouring glucose uptake, and triggering several signal transduction pathways linked to proper neuronal glutamate availability, via the glutamate/glutamine shuttle. To further characterize these complex transporters interactions, we used the well-established model system of cultured chick cerebellum Bergmann glia cells. A time and dose-dependent increase in the activity, plasma membrane localization and protein levels of glucose transporters was detected upon d-aspartate exposure. Interestingly, this increase is the result of a protein kinase C-dependent signaling cascade. Furthermore, a glutamate-dependent glucose and glutamate transporters co-immunoprecipitation was detected. These results favour the notion that glial cells are involved in glutamatergic neuronal physiology. PMID:27184733

  5. Kinetic studies to determine the mechanism of regulation of bovine liver glutamate dehydrogenase by nucleotide effectors.

    PubMed

    Cook, P F

    1982-01-01

    A combination of kinetic and isotope effect studies in the presence and absence of the effectors ADP and GTP was used to elucidate the mechanism of regulation of bovine liver glutamate dehydrogenase. ADP at low concentrations of glutamate competes with TPN for free enzyme. GTP exhibits a similar effect at high concentrations (100 microM and above). When ADP binds at its allosteric site, it increases the off rates of both alpha-ketoglutarate and TPNH from their product complexes. This results in a decrease in V/K for both substrates, an increase in V, and an increase in the deuterium isotope effects for all three parameters so that they are all about 1.3. The rate of release of glutamate from E-TPNH-glutamate is also apparently enhanced since no substrate inhibition by glutamate is observed in the presence of ADP. The effect of GTP is in opposition to that of ADP in that GTP decreases the off rates for both TPN and glutamate from E-TPN-glutamate as well as the off rates for alpha-ketoglutarate and TPNH. This results in an increase in the V/K's for both substrates, a decrease in V, and a decrease in the deuterium isotope effects for all three parameters to a value of 1. Substrate inhibition by glutamate is also eliminated by GTP probably by preventing any significant accumulation of E-TPNH to which glutamate binds as an inhibitor.

  6. Differential effects of diltiazem on glutamate potentials and excitatory junctional potentials at the crayfish neuromuscular junction.

    PubMed Central

    Ishida, M; Shinozaki, H

    1980-01-01

    1. The effects of diltiazem on glutamate potentials and excitatory junctional potentials (e.j.p.s) were investigated in the crayfish neuromuscular junction. 2. When diltiazem (0.3 mM) was added to the perfusion fluid, the ionophoretic glutamate potential was reduced to about half, whereas the peak amplitude of successive e.j.p.s elicited by a train of pulses of 100/sec increased by about 2 times. 3. It was suggested that diltiazem was a non-competitive inhibitor of L-glutamate. The reduction of the response to applied glutamate was not due to the acceleration of desensitization of the glutamate receptor. The rate of recovery from desensitization was delayzed by diltiazem. 4. The increase in amplitude of e.j.p.s caused by diltiazem was due to the increase in membrane resistance. The quantum content and size of extracellular e.j.p.s were not affected by diltiazem. 5. It was substantiated using the micro-electrode technique that the glutamate sensitive area coincided with the neuromuscular junctional area. 6. The pharmacological difference between glutamate potentials and e.j.p.s revealed in the present study is difficult to explain on the glutamate transmitter hypothesis. One explanation worthy to be considered is that there are two pharmacologically different kinds of receptors sensitive to L-glutamate. PMID:7359406

  7. On the mechanism of enhanced ATP formation in hypoxic myocardium caused by glutamic acid.

    PubMed

    Pisarenko, O I; Solomatina, E S; Ivanov, V E; Studneva, I M; Kapelko, V I; Smirnov, V N

    1985-01-01

    The effect of glutamic acid on the cardiac contractile function and sources of anaerobic ATP formation in hypoxic myocardium was studied in isovolumic rat hearts. The presence of glutamic acid (5 mM) in the perfusate significantly diminished an increment in diastolic pressure caused by 60 min hypoxia, and facilitated its complete recovery during 30 min reoxygenation. This effect was combined with the maintenance of a higher ATP level during hypoxia and reoxygenation. The total content of lactate in the heart-perfusate system rose exactly as during hypoxia without glutamic acid, while pyruvate content decreased due to increased alanine formation. Restoration of tissue content of glutamate and aspartate in the presence of exogenous glutamic acid was accompanied by a more than 2-fold increase in succinate formation, the end-product of the Krebs' cycle under anaerobic conditions. The products of glutamic acid transamination with oxaloacetic acid, aspartic and alpha-ketoglutaric acids (5mM each), induced the same functional and metabolic alterations as glutamic acid. Amino-oxyacetic acid, a tramsaminase inhibitor, eliminated the effects caused by glutamic acid. Moreover, the inhibition of transamination was accompanied by a decreased succinate and alanine synthesis as well as insignificantly increased lactate formation compared to hypoxia without additives. The results suggest that the beneficial effect of glutamic acid is due to the activation of anaerobic ATP formation in the mitochondria rather than stimulation of glycolysis.

  8. On the defensive action of glutamate against the cytotoxicity and fibrogenicity of quartz dust.

    PubMed Central

    Morosova, K I; Aronova, G V; Katsnelson, B A; Velichkovski, B T; Genkin, A M; Elnichnykh, L N; Privalova, L I

    1982-01-01

    The cytotoxic action of quartz (DQ12) particles on cultures of rat peritoneal macrophages, as estimated by the inhibition of the TTC-reductase activity, is considerably reduced by preincubation with glutamic acid and by adding sodium glutamate (15 mg/ml) to the drinking water of the rats donating the macrophages. This increase in macrophage resistance under the influence of glutamate is the most probable cause of the delay in the development of silicotic fibrosis shown in several experiments on rats intratracheally injected with quartz and then treated by prolonged administration of glutamate. This effect is probably connected with the influence of glutamate on the stability of the macrophage membranes, which can in its turn be explained by different mechanisms, including the influence on the synthesis and phosphorylation of adenosine nucleotides. Such an influence was shown in rats receiving glutamate by the change of the ATP/ADP ratio in macrophages, but not in erythrocytes. The resistance of rat erythrocytes to the haemolytic action of quartz is also not influenced by the action of glutamate neither in vitro nor in vivo. Such differences in the influences of glutamate on two types of cells, equally susceptible to quartz cytotoxicity but considerably differing in the character of energy metabolism, is an indirect proof of the role of the latter in the realisation of the anticytotoxic, and thereby antifibrogenic, effect of glutamate. PMID:6124270

  9. Local glutamate release in the rat ventral lateral thalamus evoked by high-frequency stimulation

    NASA Astrophysics Data System (ADS)

    Agnesi, Filippo; Blaha, Charles D.; Lin, Jessica; Lee, Kendall H.

    2010-04-01

    Thalamic deep brain stimulation (DBS) is proven therapy for essential tremor, Parkinson's disease and Tourette's syndrome. We tested the hypothesis that high-frequency electrical stimulation results in local thalamic glutamate release. Enzyme-linked glutamate amperometric biosensors were implanted in anesthetized rat thalamus adjacent to the stimulating electrode. Electrical stimulation was delivered to investigate the effect of frequency, pulse width, voltage-controlled or current-controlled stimulation, and charge balancing. Monophasic electrical stimulation-induced glutamate release was linearly dependent on stimulation frequency, intensity and pulse width. Prolonged stimulation evoked glutamate release to a plateau that subsequently decayed back to baseline after stimulation. Glutamate release was less pronounced with voltage-controlled stimulation and not present with charge balanced current-controlled stimulation. Using fixed potential amperometry in combination with a glutamate bioprobe and adjacent microstimulating electrode, the present study has shown that monophasic current-controlled stimulation of the thalamus in the anesthetized rat evoked linear increases in local extracellular glutamate concentrations that were dependent on stimulation duration, frequency, intensity and pulse width. However, the efficacy of monophasic voltage-controlled stimulation, in terms of evoking glutamate release in the thalamus, was substantially lower compared to monophasic current-controlled stimulation and entirely absent with biphasic (charge balanced) current-controlled stimulation. It remains to be determined whether similar glutamate release occurs with human DBS electrodes and similar charge balanced stimulation. As such, the present results indicate the importance of evaluating local neurotransmitter dynamics in studying the mechanism of action of DBS.

  10. Resting Glutamate Levels and Rapid Glutamate Transients in the Prefrontal Cortex of the Flinders Sensitive Line Rat: A Genetic Rodent Model of Depression

    PubMed Central

    Hascup, Kevin N; Hascup, Erin R; Stephens, Michelle L; Glaser, Paul EA; Yoshitake, Takashi; Mathé, Aleksander A; Gerhardt, Greg A; Kehr, Jan

    2011-01-01

    Despite the numerous drugs targeting biogenic amines for major depressive disorder (depression), the search for novel therapeutics continues because of their poor response rates (∼30%) and slow onset of action (2–4 weeks). To better understand role of glutamate in depression, we used an enzyme-based microelectrode array (MEA) that was selective for glutamate measures with fast temporal (2 Hz) and high spatial (15 × 333 μm) resolution. These MEAs were chronically implanted into the prefrontal cortex of 3- to 6-month-old and 12- to 15-month-old Flinders Sensitive Line (FSL) and control Flinders Resistant Line (FRL) rats, a validated genetic rodent model of depression. Although no changes in glutamate dynamics were observed between 3 and 6 months FRL and FSL rats, a significant increase in resting glutamate levels was observed in the 12- to 15-month-old FSL rats compared with the 3- to 6-month-old FSL and age-matched FRL rats on days 3–5 post-implantation. Our MEA also recorded, for the first time, a unique phenomenon in all the four rat groups of fluctuations in resting glutamate, which we have termed glutamate transients. Although these events lasted only for seconds, they did occur throughout the testing paradigm. The average concentration of these glutamate-burst events was significantly increased in the 12- to 15-month-old FSL rats compared with 3- to 6-month-old FSL and age-matched FRL rats. These studies lay the foundation for future studies of both tonic and phasic glutamate signaling in rat models of depression to better understand the potential role of glutamate signaling in depression. PMID:21525860

  11. The dissolution of natural and artificial dusts in glutamic acid

    NASA Astrophysics Data System (ADS)

    Ling, Zhang; Faqin, Dong; Xiaochun, He

    2015-06-01

    This article describes the characteristics of natural dusts, industrial dusts, and artificial dusts, such as mineral phases, chemical components, morphological observation and size. Quartz and calcite are the main phases of natural dusts and industrial dusts with high SiO2 and CaO and low K2O and Na2O in the chemical composition. The dissolution and electrochemical action of dusts in glutamic acid liquor at the simulated human body temperature (37 °C) in 32 h was investigated. The potential harm that the dust could lead to in body glutamic acid acidic environment, namely biological activity, is of great importance for revealing the human toxicological mechanism. The changes of pH values and electric conductivity of suspension of those dusts were similar, increased slowly in the first 8 h, and then the pH values increased rapidly. The total amount of dissolved ions of K, Ca, Na, and Mg was 35.4 to 429 mg/kg, particularly Ca was maximal of 20 to 334 mg/kg. The total amount of dissolved ions of Fe, Zn, Mn, Pb, and Ba was 0.18 to 5.59 mg/kg and in Al and Si was 3.0 to 21.7 mg/kg. The relative solubility order of dusts in glutamic acid is wollastonite > serpentine > sepiolite, the cement plant industrial dusts > natural dusts > power plant industrial dusts. The wollastonite and cement plant industrial dusts have the highest solubility, which also have high content of CaO; this shows that there are a poorer corrosion-resisting ability and lower bio-resistibility. Sepiolite and power plant industrial dusts have lowest solubility, which also have high content of SiO2; this shows that there are a higher corrosion-resisting ability and stronger bio-resistibility.

  12. Monosodium glutamate and aspartame in perceived pain in fibromyalgia.

    PubMed

    Vellisca, María Y; Latorre, José I

    2014-07-01

    Our aim was to assess the effect of dietary elimination of monosodium glutamate (MSG) and aspartame on perceived pain in fibromyalgia. A total of 72 female patients with fibromyalgia were randomized to discontinuation of dietary MSG and aspartame (n = 36) or waiting list (n = 36). Patients were requested to rate their pain using a seven-point scale. Comparisons between both groups showed no significant differences on pain referred during the baseline or after the elimination of dietary MSG and aspartame. The discontinuation of dietary MSG and aspartame did not improve the symptoms of fibromyalgia.

  13. Glutamate and GABA imbalance following traumatic brain injury.

    PubMed

    Guerriero, Réjean M; Giza, Christopher C; Rotenberg, Alexander

    2015-05-01

    Traumatic brain injury (TBI) leads to multiple short- and long-term changes in neuronal circuits that ultimately conclude with an imbalance of cortical excitation and inhibition. Changes in neurotransmitter concentrations, receptor populations, and specific cell survival are important contributing factors. Many of these changes occur gradually, which may explain the vulnerability of the brain to multiple mild impacts, alterations in neuroplasticity, and delays in the presentation of posttraumatic epilepsy. In this review, we provide an overview of normal glutamate and GABA homeostasis and describe acute, subacute, and chronic changes that follow injury. We conclude by highlighting opportunities for therapeutic interventions in this paradigm. PMID:25796572

  14. Glutamate and GABA imbalance following traumatic brain injury

    PubMed Central

    Guerriero, Réjean M.; Giza, Christopher C.; Rotenberg, Alexander

    2015-01-01

    Traumatic brain injury (TBI) leads to multiple short and long term changes in neuronal circuits that ultimately conclude with an imbalance of cortical excitation and inhibition. Changes in neurotransmitter concentrations, receptor populations and specific cell survival are important contributing factors. Many of these changes occur gradually, which may explain the vulnerability of the brain to multiple mild impacts, alterations in neuroplasticity, and delays in the presentation of post-traumatic epilepsy. In this review we provide an overview of normal glutamate and GABA homeostasis, and describe acute, subacute and chronic changes that follow injury. We conclude by highlighting opportunities for therapeutic interventions in this paradigm. PMID:25796572

  15. Metabotropic Glutamate Receptors and Interacting Proteins in Epileptogenesis.

    PubMed

    Qian, Feng; Tang, Feng-Ru

    2016-01-01

    Neurotransmitter and receptor systems are involved in different neurological and neuropsychological disorders such as Parkinson's disease, depression, Alzheimer's disease and epilepsy. Recent advances in studies of signal transduction pathways or interacting proteins of neurotransmitter receptor systems suggest that different receptor systems may share the common signal transduction pathways or interacting proteins which may be better therapeutic targets for development of drugs to effectively control brain diseases. In this paper, we reviewed metabotropic glutamate receptors (mGluRs) and their related signal transduction pathways or interacting proteins in status epilepticus and temporal lobe epilepsy, and proposed some novel therapeutical drug targets for controlling epilepsy and epileptogenesis. PMID:27030135

  16. Glutamate Metabolism and HIV-Associated Neurocognitive Disorders

    PubMed Central

    Vázquez-Santiago, Fabián J.; Noel, Richard J.; Porter, James T.; Rivera-Amill, Vanessa

    2014-01-01

    HIV-1 infection can lead to neurocognitive impairment collectively known as HIV-Associated Neurocognitive Disorders (HAND). Although combined antiretroviral treatment (cART) has significantly ameliorated HIV’s morbidity and mortality, persistent neuroinflammation and neurocognitive dysfunction continue. This review focuses on the current clinical and molecular evidence of the viral and host factors that influence glutamate-mediated neurotoxicity and neuropathogenesis as an important underlying mechanism during the course of HAND development. In addition, discusses potential pharmacological strategies targeting the glutamatergic system that may help prevent and improve neurological outcomes in HIV-1 infected subjects. PMID:24867611

  17. Genetic insights into migraine and glutamate: a protagonist driving the headache.

    PubMed

    Gasparini, Claudia F; Smith, Robert A; Griffiths, Lyn R

    2016-08-15

    Migraine is a complex polygenic disorder that continues to be a great source of morbidity in the developed world with a prevalence of 12% in the Caucasian population. Genetic and pharmacological studies have implicated the glutamate pathway in migraine pathophysiology. Glutamate profoundly impacts brain circuits that regulate core symptom domains in a range of neuropsychiatric conditions and thus remains a "hot" target for drug discovery. Glutamate has been implicated in cortical spreading depression (CSD), the phenomenon responsible for migraine with aura and in animal models carrying FHM mutations. Genotyping case-control studies have shown an association between glutamate receptor genes, namely, GRIA1 and GRIA3 with migraine with indirect supporting evidence from GWAS. New evidence localizes PRRT2 at glutamatergic synapses and shows it affects glutamate signalling and glutamate receptor activity via interactions with GRIA1. Glutamate-system defects have also been recently implicated in a novel FHM2 ATP1A2 disease-mutation mouse model. Adding to the growing evidence neurophysiological findings support a role for glutamate in cortical excitability. In addition to the existence of multiple genes to choreograph the functions of fast-signalling glutamatergic neurons, glutamate receptor diversity and regulation is further increased by the post-translational mechanisms of RNA editing and miRNAs. Ongoing genetic studies, GWAS and meta-analysis implicate neurogenic mechanisms in migraine pathology and the first genome-wide associated locus for migraine on chromosome X. Finally, in addition to glutamate modulating therapies, the kynurenine pathway has emerged as a candidate for involvement in migraine pathophysiology. In this review we discuss recent genetic evidence and glutamate modulating therapies that bear on the hypothesis that a glutamatergic mechanism may be involved in migraine susceptibility. PMID:27423601

  18. Peripherally restricted viral challenge elevates extracellular glutamate and enhances synaptic transmission in the hippocampus.

    PubMed

    Hunsberger, Holly C; Wang, Desheng; Petrisko, Tiffany J; Alhowail, Ahmad; Setti, Sharay E; Suppiramaniam, Vishnu; Konat, Gregory W; Reed, Miranda N

    2016-07-01

    Peripheral infections increase the propensity and severity of seizures in susceptible populations. We have previously shown that intraperitoneal injection of a viral mimic, polyinosinic-polycytidylic acid (PIC), elicits hypersusceptibility of mice to kainic acid (KA)-induced seizures. This study was undertaken to determine whether this seizure hypersusceptibility entails alterations in glutamate signaling. Female C57BL/6 mice were intraperitoneally injected with PIC, and after 24 h, glutamate homeostasis in the hippocampus was monitored using the enzyme-based microelectrode arrays. PIC challenge robustly increased the level of resting extracellular glutamate. While pre-synaptic potassium-evoked glutamate release was not affected, glutamate uptake was profoundly impaired and non-vesicular glutamate release was augmented, indicating functional alterations of astrocytes. Electrophysiological examination of hippocampal slices from PIC-challenged mice revealed a several fold increase in the basal synaptic transmission as compared to control slices. PIC challenge also increased the probability of pre-synaptic glutamate release as seen from a reduction of paired-pulse facilitation and synaptic plasticity as seen from an enhancement of long-term potentiation. Altogether, our results implicate a dysregulation of astrocytic glutamate metabolism and an alteration of excitatory synaptic transmission as the underlying mechanism for the development of hippocampal hyperexcitability, and consequently seizure hypersusceptibility following peripheral PIC challenge. Peripheral infections/inflammations enhance seizure susceptibility. Here, we explored the effect of peritoneal inflammation induced by a viral mimic on glutamate homeostasis and glutamatergic neurotransmission in the mouse hippocampus. We found that peritoneal inflammation elevated extracellular glutamate concentration and enhanced the probability of pre-synaptic glutamate release resulting in hyperexcitability of

  19. Researching glutamate – induced cytotoxicity in different cell lines: a comparative/collective analysis/study

    PubMed Central

    Kritis, Aristeidis A.; Stamoula, Eleni G.; Paniskaki, Krystallenia A.; Vavilis, Theofanis D.

    2015-01-01

    Although glutamate is one of the most important excitatory neurotransmitters of the central nervous system, its excessive extracellular concentration leads to uncontrolled continuous depolarization of neurons, a toxic process called, excitotoxicity. In excitotoxicity glutamate triggers the rise of intracellular Ca2+ levels, followed by up regulation of nNOS, dysfunction of mitochondria, ROS production, ER stress, and release of lysosomal enzymes. Excessive calcium concentration is the key mediator of glutamate toxicity through over activation of ionotropic and metabotropic receptors. In addition, glutamate accumulation can also inhibit cystine (CySS) uptake by reversing the action of the CySS/glutamate antiporter. Reversal of the antiporter action reinforces the aforementioned events by depleting neurons of cysteine and eventually glutathione’s reducing potential. Various cell lines have been employed in the pursuit to understand the mechanism(s) by which excitotoxicity affects the cells leading them ultimately to their demise. In some cell lines glutamate toxicity is exerted mainly through over activation of NMDA, AMPA, or kainate receptors whereas in other cell lines lacking such receptors, the toxicity is due to glutamate induced oxidative stress. However, in the greatest majority of the cell lines ionotropic glutamate receptors are present, co-existing to CySS/glutamate antiporters and metabotropic glutamate receptors, supporting the assumption that excitotoxicity effect in these cells is accumulative. Different cell lines differ in their responses when exposed to glutamate. In this review article the responses of PC12, SH-SY5Y, HT-22, NT-2, OLCs, C6, primary rat cortical neurons, RGC-5, and SCN2.2 cell systems are systematically collected and analyzed. PMID:25852482

  20. Glutamate cycling may drive organic anion transport on the basal membrane of human placental syncytiotrophoblast.

    PubMed

    Lofthouse, Emma M; Brooks, Suzanne; Cleal, Jane K; Hanson, Mark A; Poore, Kirsten R; O'Kelly, Ita M; Lewis, Rohan M

    2015-10-15

    The organic anion transporter OAT4 (SLC22A11) and organic anion transporting polypeptide OATP2B1 (SLCO2B1) are expressed in the basal membrane of the placental syncytiotrophoblast. These transporters mediate exchange whereby uptake of one organic anion is coupled to efflux of a counter-ion. In placenta, these exchangers mediate placental uptake of substrates for oestrogen synthesis as well as clearing waste products and xenobiotics from the fetal circulation. However, the identity of the counter-ion driving this transport in the placenta, and in other tissues, is unclear. While glutamate is not a known OAT4 or OATP2B1 substrate, we propose that its high intracellular concentration has the potential to drive accumulation of substrates from the fetal circulation. In the isolated perfused placenta, glutamate exchange was observed between the placenta and the fetal circulation. This exchange could not be explained by known glutamate exchangers. However, glutamate efflux was trans-stimulated by an OAT4 and OATP2B1 substrate (bromosulphothalein). Exchange of glutamate for bromosulphothalein was only observed when glutamate reuptake was inhibited (by addition of aspartate). To determine if OAT4 and/or OATP2B1 mediate glutamate exchange, uptake and efflux of glutamate were investigated in Xenopus laevis oocytes. Our data demonstrate that in Xenopus oocytes expressing either OAT4 or OATP2B1 efflux of intracellular [(14)C]glutamate could be stimulated by conditions including extracellular glutamate (OAT4), estrone-sulphate and bromosulphothalein (both OAT4 and OATP2B1) or pravastatin (OATP2B1). Cycling of glutamate across the placenta involving efflux via OAT4 and OATP2B1 and subsequent reuptake will drive placental uptake of organic anions from the fetal circulation.

  1. Three-dimensional models of non-NMDA glutamate receptors.

    PubMed Central

    Sutcliffe, M J; Wo, Z G; Oswald, R E

    1996-01-01

    Structural models have been produced for three types of non-NMDA inotropic glutamate receptors: an AMPA receptor, GluR1, a kainate receptor, GluR6; and a low-molecular-weight kainate receptor from goldfish, GFKAR alpha. Modeling was restricted to the domains of the proteins that bind the neurotransmitter glutamate and that form the ion channel. Model building combined homology modeling, distance geometry, molecular mechanics, interactive modeling, and known constraints. The models indicate new potential interactions in the extracellular domain between protein and agonists, and suggest that the transition from the "closed" to the "open" state involves the movement of a conserved positive residue away from, and two conserved negative residues into, the extracellular entrance to the pore upon binding. As a first approximation, the ion channel domain was modeled with a structure comprising a central antiparallel beta-barrel that partially crosses the membrane, and against which alpha-helices from each subunit are packed; a third alpha-helix packs against these two helices in each subunit. Much, but not all, of the available data were consistent with this structure. Modifying the beta-barrel to a loop-like topology produced a model consistent with available data. Images FIGURE 1 FIGURE 2 FIGURE 3 FIGURE 4 FIGURE 5 FIGURE 7 PMID:8785317

  2. Metabotropic Glutamate Receptor Dependent Cortical Plasticity in Chronic Pain.

    PubMed

    Koga, Kohei; Li, Shermaine; Zhuo, Min

    2016-01-01

    Many cortical areas play crucial roles in higher order brain functions such as pain and emotion-processing, decision-making, and cognition. Among them, anterior cingulate cortex (ACC) and insular cortex (IC) are two key areas. Glutamate mediates major excitatory transmission during long-term plasticity in both physiological and pathological conditions. Specifically related to nociceptive or pain behaviors, metabotropic glutamate subtype receptors (mGluRs) have been involved in different types of synaptic modulation and plasticity from periphery to the spinal cord. However, less is known about their functional roles in plasticity related to pain and its related behaviors within cortical regions. In this review, we first summarized previous studies of synaptic plasticity in both the ACC and IC, and discussed how mGluRs may be involved in both cortical long-term potentiation (LTP) and long-term depression (LTD)-especially in LTD. The activation of mGluRs contributes to the induction of LTD in both ACC and IC areas. The loss of LTD caused by peripheral amputation or nerve injury can be rescued by priming ACC or IC with activations of mGluR1 receptors. We also discussed the potential functional roles of mGluRs for pain-related behaviors. We propose that targeting mGluRs in the cortical areas including the ACC and IC may provide a new therapeutic strategy for the treatment of chronic pain, phantom pain or anxiety. PMID:27296638

  3. Activity-Dependent Plasticity of Astroglial Potassium and Glutamate Clearance

    PubMed Central

    Cheung, Giselle; Sibille, Jérémie; Zapata, Jonathan; Rouach, Nathalie

    2015-01-01

    Recent evidence has shown that astrocytes play essential roles in synaptic transmission and plasticity. Nevertheless, how neuronal activity alters astroglial functional properties and whether such properties also display specific forms of plasticity still remain elusive. Here, we review research findings supporting this aspect of astrocytes, focusing on their roles in the clearance of extracellular potassium and glutamate, two neuroactive substances promptly released during excitatory synaptic transmission. Their subsequent removal, which is primarily carried out by glial potassium channels and glutamate transporters, is essential for proper functioning of the brain. Similar to neurons, different forms of short- and long-term plasticity in astroglial uptake have been reported. In addition, we also present novel findings showing robust potentiation of astrocytic inward currents in response to repetitive stimulations at mild frequencies, as low as 0.75 Hz, in acute hippocampal slices. Interestingly, neurotransmission was hardly affected at this frequency range, suggesting that astrocytes may be more sensitive to low frequency stimulation and may exhibit stronger plasticity than neurons to prevent hyperexcitability. Taken together, these important findings strongly indicate that astrocytes display both short- and long-term plasticity in their clearance of excess neuroactive substances from the extracellular space, thereby regulating neuronal activity and brain homeostasis. PMID:26346563

  4. Glutamate dehydrogenase: structure, allosteric regulation, and role in insulin homeostasis.

    PubMed

    Li, Ming; Li, Changhong; Allen, Aron; Stanley, Charles A; Smith, Thomas J

    2014-01-01

    Glutamate dehydrogenase (GDH) is a homohexameric enzyme that catalyzes the reversible oxidative deamination of L-glutamate to 2-oxoglutarate. Only in the animal kingdom is this enzyme heavily allosterically regulated by a wide array of metabolites. The major activators are ADP and leucine and inhibitors include GTP, palmitoyl CoA, and ATP. Spontaneous mutations in the GTP inhibitory site that lead to the hyperinsulinism/hyperammonemia (HHS) syndrome have shed light as to why mammalian GDH is so tightly regulated. Patients with HHS exhibit hypersecretion of insulin upon consumption of protein and concomitantly extremely high levels of ammonium in the serum. The atomic structures of four new inhibitors complexed with GDH complexes have identified three different allosteric binding sites. Using a transgenic mouse model expressing the human HHS form of GDH, at least three of these compounds blocked the dysregulated form of GDH in pancreatic tissue. EGCG from green tea prevented the hyper-response to amino acids in whole animals and improved basal serum glucose levels. The atomic structure of the ECG-GDH complex and mutagenesis studies is directing structure-based drug design using these polyphenols as a base scaffold. In addition, all of these allosteric inhibitors are elucidating the atomic mechanisms of allostery in this complex enzyme.

  5. The structure and allosteric regulation of mammalian glutamate dehydrogenase.

    PubMed

    Li, Ming; Li, Changhong; Allen, Aron; Stanley, Charles A; Smith, Thomas J

    2012-03-15

    Glutamate dehydrogenase (GDH) is a homohexameric enzyme that catalyzes the reversible oxidative deamination of l-glutamate to 2-oxoglutarate. Only in the animal kingdom is this enzyme heavily allosterically regulated by a wide array of metabolites. The major activators are ADP and leucine, while the most important inhibitors include GTP, palmitoyl CoA, and ATP. Recently, spontaneous mutations in the GTP inhibitory site that lead to the hyperinsulinism/hyperammonemia (HHS) syndrome have shed light as to why mammalian GDH is so tightly regulated. Patients with HHS exhibit hypersecretion of insulin upon consumption of protein and concomitantly extremely high levels of ammonium in the serum. The atomic structures of four new inhibitors complexed with GDH complexes have identified three different allosteric binding sites. Using a transgenic mouse model expressing the human HHS form of GDH, at least three of these compounds were found to block the dysregulated form of GDH in pancreatic tissue. EGCG from green tea prevented the hyper-response to amino acids in whole animals and improved basal serum glucose levels. The atomic structure of the ECG-GDH complex and mutagenesis studies is directing structure-based drug design using these polyphenols as a base scaffold. In addition, all of these allosteric inhibitors are elucidating the atomic mechanisms of allostery in this complex enzyme.

  6. Conformation of poly(γ-glutamic acid) in aqueous solution.

    PubMed

    Muroga, Yoshio; Nakaya, Asami; Inoue, Atsuki; Itoh, Daiki; Abiru, Masaya; Wada, Kaori; Takada, Masako; Ikake, Hiroki; Shimizu, Shigeru

    2016-04-01

    Local conformation and overall conformation of poly(γ-DL-glutamic acid) (PγDLGA) and poly(γ-L-glutamic acid) (PγLGA) in aqueous solution was studied as a function of degree of ionization ε by (1) H-NMR, circular dichroism, and potentiometric titration. It was clarified that their local conformation is represented by random coil over an entire ε range and their overall conformation is represented by expanded random-coil in a range of ε > ε(*) , where ε(*) is about 0.3, 0.35, 0.45, and 0.5 for added-salt concentration of 0.02M, 0.05M, 0.1M, and 0.2M, respectively. In a range of ε < ε(*) , however, ε dependence of their overall conformation is significantly differentiated from each other. PγDLGA tends to aggregate intramolecularly and/or intermolecularly with decreasing ε, but PγLGA still behaves as expanded random-coil. It is speculated that spatial arrangement of adjacent carboxyl groups along the backbone chain essentially affects the overall conformation of PγGA in acidic media.

  7. Targeting of metabotropic glutamate receptors for the treatment of schizophrenia.

    PubMed

    Chaki, Shigeyuki; Hikichi, Hirohiko

    2011-01-01

    The glutamatergic system is involved in a wide range of physiological processes in the brain, and its dysfunction plays an important role in the etiology and pathophysiology of psychiatric disorders, including schizophrenia. Among the glutamate receptors, metabotropic receptors (mGlu receptors) have emerged as attractive therapeutic targets for the development of novel interventions for psychiatric disorders. Among them, group II mGlu receptors, such as mGlu2 and mGlu3 receptors, are of particular interest because of their unique distribution and the regulatory roles they have in neurotransmission. Recently, potent agonists for mGlu2/3 receptor have been synthesized, and their pharmacological roles have been intensively investigated in animal models. The efficacy for the treatment of schizophrenia has also been proven in a clinical trial. Recently, much attention has been paid to mGlu2 receptor potentiators, which potentiate the glutamate response without affecting the actual activity of the mGlu2 receptor. In addition, mGlu1 receptor antagonists have recently been proposed as an attractive approach to developing novel antipsychotics in animal models. This review describes the potential of both mGlu2/3 receptor agonists/potentiators and mGlu1 receptor antagonists for the treatment of schizophrenia. PMID:21355835

  8. Metabotropic glutamate receptors: their therapeutic potential in anxiety.

    PubMed

    Spooren, Will; Lesage, Anne; Lavreysen, Hilde; Gasparini, Fabrizio; Steckler, Thomas

    2010-01-01

    Psychiatric and neurological disorders are linked to changes in synaptic excitatory processes with a key role for glutamate, that is, the most abundant excitatory amino-acid. Molecular cloning of the metabotropic glutamate (mGlu) receptors has led to the identification of eight mGlu receptors, which, in contrast to ligand-gated ion channels (responsible for fast excitatory transmission), modulate and fine-tune the efficacy of synaptic transmission. mGlu receptors are G protein-coupled and constitute a new group of "drugable" targets for the treatment of various CNS disorders. The recent discovery of small molecules that selectively bind to receptors of Groups I (mGlu1 and mGlu5) and II (mGlu2 and mGlu3) allowed significant advances in our understanding of the roles of these receptors in brain function and dysfunction including anxiety. Although investigation of the role of the Group III (mGlu4, 6, 7, and 8) receptors is less advanced, the generation of genetically manipulated animals and recent advances in the identification of subtype-selective compounds have revealed some first insights into the therapeutic potential of this group of receptors. PMID:21309118

  9. Metabotropic glutamate receptors: their therapeutic potential in anxiety.

    PubMed

    Spooren, Will; Lesage, Anne; Lavreysen, Hilde; Gasparini, Fabrizio; Steckler, Thomas

    2010-01-01

    Psychiatric and neurological disorders are linked to changes in synaptic excitatory processes with a key role for glutamate, that is, the most abundant excitatory amino-acid. Molecular cloning of the metabotropic glutamate (mGlu) receptors has led to the identification of eight mGlu receptors, which, in contrast to ligand-gated ion channels (responsible for fast excitatory transmission), modulate and fine-tune the efficacy of synaptic transmission. mGlu receptors are G protein-coupled and constitute a new group of "drugable" targets for the treatment of various CNS disorders. The recent discovery of small molecules that selectively bind to receptors of Groups I (mGlu1 and mGlu5) and II (mGlu2 and mGlu3) allowed significant advances in our understanding of the roles of these receptors in brain function and dysfunction including anxiety. Although investigation of the role of the Group III (mGlu4, 6, 7, and 8) receptors is less advanced, the generation of genetically manipulated animals and recent advances in the identification of subtype-selective compounds have revealed some first insights into the therapeutic potential of this group of receptors.

  10. Abnormal glutamate release in aged BTBR mouse model of autism.

    PubMed

    Wei, Hongen; Ding, Caiyun; Jin, Guorong; Yin, Haizhen; Liu, Jianrong; Hu, Fengyun

    2015-01-01

    Autism is a neurodevelopmental disorder characterized by abnormal reciprocal social interactions, communication deficits, and repetitive behaviors with restricted interests. Most of the available research on autism is focused on children and young adults and little is known about the pathological alternation of autism in older adults. In order to investigate the neurobiological alternation of autism in old age stage, we compared the morphology and synaptic function of excitatory synapses between the BTBR mice with low level sociability and B6 mice with high level sociability. The results revealed that the number of excitatory synapse colocalized with pre- and post-synaptic marker was not different between aged BTBR and B6 mice. The aged BTBR mice had a normal structure of dendritic spine and the expression of Shank3 protein in the brain as well as that in B6 mice. The baseline and KCl-evoked glutamate release from the cortical synaptoneurosome in aged BTBR mice was lower than that in aged B6 mice. Overall, the data indicate that there is a link between disturbances of the glutamate transmission and autism. These findings provide new evidences for the hypothesis of excitation/inhibition imbalance in autism. Further work is required to determine the cause of this putative abnormality.

  11. Memantine: targeting glutamate excitotoxicity in Alzheimer's disease and other dementias.

    PubMed

    Molinuevo, José L; Lladó, Albert; Rami, Lorena

    2005-01-01

    The management of dementia has changed since the development of new antidementia drugs. The benefits observed in Alzheimer's disease (AD) with selective cholinergic transmission treatments are mainly symptomatic, without clear evidence of neuroprotection. The hypothesis that glutamate-mediated neurotoxicity is involved in the pathogenesis of AD is finding increasingly more acceptance in the scientific community. Glutamate receptors are overactive, and N-methyl-D-aspartate (NMDA) receptor antagonists have therapeutic potential for the treatment of AD and other neurological disorders. Memantine is a noncompetitive NMDA antagonist that is considered a neuroprotective drug. Memantine's capacity has been demonstrated in preclinical studies, and it is considered a useful symptomatic treatment for AD. Memantine has been shown to benefit cognition, function, and global outcome in patients with moderate to severe AD, and it is currently approved by the US Food and Drug Administration (FDA) for the treatment of moderate to severe AD. Recently, memantine has also demonstrated efficacy in the initial stages of AD, although FDA authorization is pending. This review highlights the important pharmacological and clinical aspects of memantine, as well as some basic mechanisms mediating glutamatergic neurodegeneration.

  12. Yokukansan, a kampo medicine, protects PC12 cells from glutamate-induced death by augmenting gene expression of cystine/glutamate antiporter system Xc-.

    PubMed

    Kanno, Hitomi; Kawakami, Zenji; Mizoguchi, Kazushige; Ikarashi, Yasushi; Kase, Yoshio

    2014-01-01

    Effects of the kampo medicine yokukansan on gene expression of the cystine/glutamate antiporter system Xc-, which protects against glutamate-induced cytotoxicity, were examined in Pheochromocytoma cells (PC12 cells). Yokukansan inhibited glutamate-induced PC12 cell death. Similar cytoprotective effects were found in Uncaria hook. Experiments to clarify the active compounds revealed that geissoschizine methyl ether, hirsuteine, hirsutine, and procyanidin B1 in Uncaria hook, had cytoprotective effects. These components enhanced gene expressions of system Xc- subunits xCT and 4F2hc, and also ameliorated the glutamate-induced decrease in glutathione levels. These results suggest that the cytoprotective effect of yokukansan may be attributed to geissoschizine methyl ether, hirsuteine, hirsutine, and procyanidin B1 in Uncaria hook.

  13. Ethanol-Associated Changes in Glutamate Reward Neurocircuitry: A Minireview of Clinical and Preclinical Genetic Findings.

    PubMed

    Bell, Richard L; Hauser, Sheketha R; McClintick, Jeanette; Rahman, Shafiqur; Edenberg, Howard J; Szumlinski, Karen K; McBride, William J

    2016-01-01

    Herein, we have reviewed the role of glutamate, the major excitatory neurotransmitter in the brain, in a number of neurochemical, -physiological, and -behavioral processes mediating the development of alcohol dependence. The findings discussed include results from both preclinical as well as neuroimaging and postmortem clinical studies. Expression levels for a number of glutamate-associated genes and/or proteins are modulated by alcohol abuse and dependence. These changes in expression include metabotropic receptors and ionotropic receptor subunits as well as different glutamate transporters. Moreover, these changes in gene expression parallel the pharmacologic manipulation of these same receptors and transporters. Some of these gene expression changes may have predated alcohol abuse and dependence because a number of glutamate-associated polymorphisms are related to a genetic predisposition to develop alcohol dependence. Other glutamate-associated polymorphisms are linked to age at the onset of alcohol-dependence and initial level of response/sensitivity to alcohol. Finally, findings of innate and/or ethanol-induced glutamate-associated gene expression differences/changes observed in a genetic animal model of alcoholism, the P rat, are summarized. Overall, the existing literature indicates that changes in glutamate receptors, transporters, enzymes, and scaffolding proteins are crucial for the development of alcohol dependence and there is a substantial genetic component to these effects. This indicates that continued research into the genetic underpinnings of these glutamate-associated effects will provide important novel molecular targets for treating alcohol abuse and dependence. PMID:26809998

  14. The neurotransmitters serotonin and glutamate accelerate the heart rate of the mosquito Anopheles gambiae.

    PubMed

    Hillyer, Julián F; Estévez-Lao, Tania Y; Mirzai, Homa E

    2015-10-01

    Serotonin and glutamate are neurotransmitters that in insects are involved in diverse physiological processes. Both serotonin and glutamate have been shown to modulate the physiology of the dorsal vessel of some insects, yet until the present study, their activity in mosquitoes remained unknown. To test whether serotonin or glutamate regulate dorsal vessel physiology in the African malaria mosquito, Anopheles gambiae, live mosquitoes were restrained, and a video of the contracting heart (the abdominal portion of the dorsal vessel) was acquired. These adult female mosquitoes were then injected with various amounts of serotonin, glutamate, or a control vehicle solution, and additional videos were acquired at 2 and 10 min post-treatment. Comparison of the videos taken before and after treatment revealed that serotonin accelerates the frequency of heart contractions, with the cardioacceleration being significantly more pronounced when the wave-like contractions of cardiac muscle propagate in the anterograde direction (toward the head). Comparison of the videos taken before and after treatment with glutamate revealed that this molecule is also cardioacceleratory. However, unlike serotonin, the activity of glutamate does not depend on whether the contractions propagate in the anterograde or the retrograde (toward the posterior of the abdomen) directions. Serotonin or glutamate induces a minor change or no change in the percentage of contractions and the percentage of the time that the heart contracts in the anterograde or the retrograde directions. In summary, this study shows that the neurotransmitters serotonin and glutamate increase the heart contraction rate of mosquitoes. PMID:26099947

  15. Characterization of the venom from the spider, Araneus gemma: search for a glutamate antagonist

    SciTech Connect

    Early, S.L.

    1985-01-01

    Venom from three spiders, Argiope aurantia, Neoscona arabesca, and Araneus gemma have been shown to inhibit the binding of L-(/sup 3/H)glutamate to both GBP and synaptic membranes. The venom from Araneus gemma was shown to be the most potent of the three venoms in inhibiting the binding of L-(/sup 3/H)glutamate to GBP. Therefore, Araneus gemma venom was selected for further characterization. Venom from Araneus gemma appeared to contain two factors which inhibit the binding of L-(/sup 3/H)glutamate to GBP and at least one factor that inhibits L-glutamate-stimulated /sup 35/SCN flux. Factor I is thought to be L-glutamic acid, based on: (1) its similar mobility to glutamic acid in thin-layer chromatography and amino acid analysis, (2) the presence of fingerprint molecular ion peaks for glutamate in the mass spectrum for the methanol:water (17:1) extract and for the fraction from the HPLC-purification of the crude venom, and (3) its L-glutamate-like interaction with the sodium-dependent uptake system. Factor II appears to be a polypeptide, possibly 21 amino acids in length, and does not appear to contain any free amino groups or tryptophan. While the venom does not appear to contain any indoleamines, three catecholamines (epinephrine, epinine, dopamine) and one catecholamine metabolite (DOPAC) were detected.

  16. The neurotransmitters serotonin and glutamate accelerate the heart rate of the mosquito Anopheles gambiae.

    PubMed

    Hillyer, Julián F; Estévez-Lao, Tania Y; Mirzai, Homa E

    2015-10-01

    Serotonin and glutamate are neurotransmitters that in insects are involved in diverse physiological processes. Both serotonin and glutamate have been shown to modulate the physiology of the dorsal vessel of some insects, yet until the present study, their activity in mosquitoes remained unknown. To test whether serotonin or glutamate regulate dorsal vessel physiology in the African malaria mosquito, Anopheles gambiae, live mosquitoes were restrained, and a video of the contracting heart (the abdominal portion of the dorsal vessel) was acquired. These adult female mosquitoes were then injected with various amounts of serotonin, glutamate, or a control vehicle solution, and additional videos were acquired at 2 and 10 min post-treatment. Comparison of the videos taken before and after treatment revealed that serotonin accelerates the frequency of heart contractions, with the cardioacceleration being significantly more pronounced when the wave-like contractions of cardiac muscle propagate in the anterograde direction (toward the head). Comparison of the videos taken before and after treatment with glutamate revealed that this molecule is also cardioacceleratory. However, unlike serotonin, the activity of glutamate does not depend on whether the contractions propagate in the anterograde or the retrograde (toward the posterior of the abdomen) directions. Serotonin or glutamate induces a minor change or no change in the percentage of contractions and the percentage of the time that the heart contracts in the anterograde or the retrograde directions. In summary, this study shows that the neurotransmitters serotonin and glutamate increase the heart contraction rate of mosquitoes.

  17. Glutamate and post-traumatic stress disorder: toward a psychobiology of dissociation.

    PubMed

    Chambers, R A; Bremner, J D; Moghaddam, B; Southwick, S M; Charney, D S; Krystal, J H

    1999-10-01

    Dissociative cognitive and perceptual alterations commonly occur at the time of traumatization and as an enduring feature of post-traumatic stress disorder (PTSD). After stress exposure, dissociative symptoms are a predictor of the development of PTSD. Recent preclinical data suggest that stress stimulates the cortico-limbic release of glutamate. The glutamate that is released during stress in animal models influences behavior, induces a variety of changes in neural plasticity that may have long-lasting effects on brain function and behavior, and contributes to neural toxicity. Antagonist of the N-methyl-D-aspartate (NMDA) subtype of glutamate receptor also stimulates transient cortico-limbic glutamate release in animals. Further, some of the effects of NMDA antagonists in animals are blocked by drugs that attenuate glutamate release. Clinical studies suggest that NMDA antagonists may transiently stimulate glutamate release and produce symptoms resembling dissociative states in humans. A recent study suggests that a drug that reduces glutamate release also attenuates the perceptual effects of the NMDA antagonist, ketamine, in humans. Because of the possible contributions of hyperglutamatergic states to the acute and long-lasting consequences of traumatic stress exposure, the therapeutic and neuroprotective potential of drugs that attenuate glutamate release should be explored in traumatized individuals with dissociative symptoms.

  18. 40 CFR 180.1187 - L-glutamic acid; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 40 Protection of Environment 23 2010-07-01 2010-07-01 false L-glutamic acid; exemption from the requirement of a tolerance. 180.1187 Section 180.1187 Protection of Environment ENVIRONMENTAL PROTECTION... Exemptions From Tolerances § 180.1187 L-glutamic acid; exemption from the requirement of a tolerance....

  19. Supplementing monosodium glutamate to partial enteral nutrition slows gastric emptying in preterm pigs

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Emerging evidence suggests that free glutamate may play a functional role in modulating gastroduodenal motor function. We hypothesized that supplementing monosodium glutamate (MSG) to partial enteral nutrition stimulates gastric emptying in preterm pigs. Ten-day-old preterm, parenterally fed pigs re...

  20. 21 CFR 573.500 - Condensed, extracted glutamic acid fermentation product.

    Code of Federal Regulations, 2011 CFR

    2011-04-01

    ... HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.500 Condensed, extracted glutamic acid... 21 Food and Drugs 6 2011-04-01 2011-04-01 false Condensed, extracted glutamic acid...

  1. 21 CFR 573.500 - Condensed, extracted glutamic acid fermentation product.

    Code of Federal Regulations, 2014 CFR

    2014-04-01

    ... HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.500 Condensed, extracted glutamic acid... 21 Food and Drugs 6 2014-04-01 2014-04-01 false Condensed, extracted glutamic acid...

  2. 21 CFR 573.500 - Condensed, extracted glutamic acid fermentation product.

    Code of Federal Regulations, 2010 CFR

    2010-04-01

    ... HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.500 Condensed, extracted glutamic acid... 21 Food and Drugs 6 2010-04-01 2010-04-01 false Condensed, extracted glutamic acid...

  3. 21 CFR 573.500 - Condensed, extracted glutamic acid fermentation product.

    Code of Federal Regulations, 2012 CFR

    2012-04-01

    ... HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.500 Condensed, extracted glutamic acid... 21 Food and Drugs 6 2012-04-01 2012-04-01 false Condensed, extracted glutamic acid...

  4. 21 CFR 573.500 - Condensed, extracted glutamic acid fermentation product.

    Code of Federal Regulations, 2013 CFR

    2013-04-01

    ... HUMAN SERVICES (CONTINUED) ANIMAL DRUGS, FEEDS, AND RELATED PRODUCTS FOOD ADDITIVES PERMITTED IN FEED AND DRINKING WATER OF ANIMALS Food Additive Listing § 573.500 Condensed, extracted glutamic acid... 21 Food and Drugs 6 2013-04-01 2013-04-01 false Condensed, extracted glutamic acid...

  5. 78 FR 57881 - Monosodium Glutamate from China and Indonesia; Institution of Antidumping and Countervailing Duty...

    Federal Register 2010, 2011, 2012, 2013, 2014

    2013-09-20

    ... took effect on November 7, 2011. See 76 FR 61937 (Oct. 6, 2011) and the newly revised Commission's... COMMISSION Monosodium Glutamate from China and Indonesia; Institution of Antidumping and Countervailing Duty... Indonesia of monosodium glutamate, provided for in subheading 2922.42.10 of the Harmonized Tariff...

  6. Levetiracetam inhibits oligomeric Aβ-induced glutamate release from human astrocytes.

    PubMed

    Sanz-Blasco, Sara; Piña-Crespo, Juan C; Zhang, Xiaofei; McKercher, Scott R; Lipton, Stuart A

    2016-06-15

    A recently identified mechanism for oligomeric Aβ-induced glutamate release from astrocytes involves intracellular Ca elevation, potentially by Ca-dependent vesicular release. Evidence suggests that levetiracetam (LEV; Keppra), an antiepileptic drug, can improve cognitive performance in both humans with mild cognitive impairment and animal models of Alzheimer disease. Because LEV acts by modulating neurotransmitter release from neurons by interaction with synaptic vesicles, we tested the effect of LEV on Aβ-induced astrocytic release of glutamate. We used a fluorescence resonance energy transfer-based glutamate sensor (termed SuperGluSnFR), whose structure is based on the ligand-binding site of glutamate receptors, to monitor glutamate release from primary cultures of human astrocytes exposed to oligomeric amyloid-β peptide 1-42 (Aβ42). We found that LEV (10 µM) inhibited oligomeric Aβ-induced astrocytic glutamate release. In addition, we show that this Aβ-induced glutamate release from astrocytes is sensitive to tetanus neurotoxin, an inhibitor of the vesicle release machinery. Taken together, our evidence suggests that LEV inhibits Aβ-induced vesicular glutamate release from astrocytes and thus may underlie, at least in part, the ability of LEV to reduce hyperexcitability in Alzheimer disease. PMID:27183239

  7. Detection and transfer of the glutamate decarboxylase gene in Streptococcus thermophilus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    GABA (gamma-aminobutyric acid) is generated from glutamate by the action of glutamic acid decarboxylase (GAD) and characterized by hypotensive, diuretic and tranquilizing effects in humans and animals. The production of GABA by lactic acid starter bacteria would enhance the functionality of fermen...

  8. EVALUATING THE NMDA-GLUTAMATE RECEPTOR AS A SITE OF ACTION FOR TOLUENE, IN VIVO

    EPA Science Inventory

    In vitro, toluene disrupts the function of NMDA-glutamate receptors, indicating that effects on NMDA receptor function may contribute to toluene neurotoxicity. NMDA-glutamate receptors are widely present in the visual system and contribute to pattern-elicited visual evoked potent...

  9. Vector-mediated chromosomal integration of the glutamate decarboxylase gene in streptococcus thermophilus

    Technology Transfer Automated Retrieval System (TEKTRAN)

    The integrative vector pINTRS was used to transfer glutamate decarboxylase (GAD) activity to Streptococcus thermophilus ST128, thus allowing for the production of '-aminobutyric acid (GABA). In pINTRS, the gene encoding glutamate decarboxylase, gadB, was flanked by DNA fragments homologous to a S. ...

  10. Molecular analysis of the glutamate decarboxylase locus in Streptococcus thermophilus ST110

    Technology Transfer Automated Retrieval System (TEKTRAN)

    GABA ('-aminobutyric acid) is generated from glutamate by the action of glutamic acid decarboxylase (GAD) and characterized by hypotensive, diuretic and tranquilizing effects in humans and animals. The production of GABA by lactic acid starter bacteria would enhance the functionality of fermented da...

  11. Adenosine A2A receptors modulate glutamate uptake in cultured astrocytes and gliosomes.

    PubMed

    Matos, Marco; Augusto, Elisabete; Santos-Rodrigues, Alexandre Dos; Schwarzschild, Michael A; Chen, Jiang-Fan; Cunha, Rodrigo A; Agostinho, Paula

    2012-05-01

    Glutamate is the primary excitatory neurotransmitter in the central nervous system, where its toxic build-up leads to synaptic dysfunction and excitotoxic cell death that underlies many neurodegenerative diseases. Therefore, efforts have been made to understand the regulation of glutamate transporters, which are responsible for the clearance of extracellular glutamate. We now report that adenosine A(2A) receptors (A(2A) R) control the uptake of D-aspartate in primary cultured astrocytes as well as in an ex vivo preparation enriched in glial plasmalemmal vesicles (gliosomes) from adult rats, whereas A(1) R and A(3) R were devoid of effects. Thus, the acute exposure to the A(2A) R agonist, CGS 21680, inhibited glutamate uptake, an effect prevented by the A(2A) R antagonist, SCH 58261, and abbrogated in cultured astrocytes from A(2A) R knockout mice. Furthermore, the prolonged activation of A(2A) R lead to a cAMP/protein kinase A-dependent reduction of GLT-I and GLAST mRNA and protein levels, which leads to a sustained decrease of glutamate uptake. This dual mechanism of inhibition of glutamate transporters by astrocytic A(2A) R provides a novel candidate mechanism to understand the ability of A(2) (A) R to control synaptic plasticity and neurodegeneration, two conditions tightly associated with the control of extracellular glutamate levels by glutamate transporters.

  12. Ethanol-Associated Changes in Glutamate Reward Neurocircuitry: A Minireview of Clinical and Preclinical Genetic Findings.

    PubMed

    Bell, Richard L; Hauser, Sheketha R; McClintick, Jeanette; Rahman, Shafiqur; Edenberg, Howard J; Szumlinski, Karen K; McBride, William J

    2016-01-01

    Herein, we have reviewed the role of glutamate, the major excitatory neurotransmitter in the brain, in a number of neurochemical, -physiological, and -behavioral processes mediating the development of alcohol dependence. The findings discussed include results from both preclinical as well as neuroimaging and postmortem clinical studies. Expression levels for a number of glutamate-associated genes and/or proteins are modulated by alcohol abuse and dependence. These changes in expression include metabotropic receptors and ionotropic receptor subunits as well as different glutamate transporters. Moreover, these changes in gene expression parallel the pharmacologic manipulation of these same receptors and transporters. Some of these gene expression changes may have predated alcohol abuse and dependence because a number of glutamate-associated polymorphisms are related to a genetic predisposition to develop alcohol dependence. Other glutamate-associated polymorphisms are linked to age at the onset of alcohol-dependence and initial level of response/sensitivity to alcohol. Finally, findings of innate and/or ethanol-induced glutamate-associated gene expression differences/changes observed in a genetic animal model of alcoholism, the P rat, are summarized. Overall, the existing literature indicates that changes in glutamate receptors, transporters, enzymes, and scaffolding proteins are crucial for the development of alcohol dependence and there is a substantial genetic component to these effects. This indicates that continued research into the genetic underpinnings of these glutamate-associated effects will provide important novel molecular targets for treating alcohol abuse and dependence.

  13. 40 CFR 180.1187 - L-glutamic acid; exemption from the requirement of a tolerance.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 40 Protection of Environment 24 2011-07-01 2011-07-01 false L-glutamic acid; exemption from the requirement of a tolerance. 180.1187 Section 180.1187 Protection of Environment ENVIRONMENTAL PROTECTION... Exemptions From Tolerances § 180.1187 L-glutamic acid; exemption from the requirement of a tolerance....

  14. Biochemical and immunological changes on oral glutamate feeding in male albino rats

    NASA Astrophysics Data System (ADS)

    Kumar, D.; Bansal, Anju; Thomas, Pauline; Sairam, M.; Sharma, S. K.; Mongia, S. S.; Singh, R.; Selvamurthy, W.

    High altitude stress leads to lipid peroxidation and free radical formation which results in cell membrane damage in organs and tissues, and associated mountain diseases. This paper discusses the changes in biochemical parameters and antibody response on feeding glutamate to male albino Sprague Dawley rats under hypoxic stress. Exposure of rats to simulated hypoxia at 7576 m, for 6 h daily for 5 consecutive days, in an animal decompression chamber at 32+/-2° C resulted in an increase in plasma malondialdehyde level with a concomitant decrease in blood glutathione (reduced) level. Supplementation of glutamate orally at an optimal dose (27 mg/kg body weight) in male albino rats under hypoxia enhanced glutathione level and decreased malondialdehyde concentration significantly. Glutamate feeding improved total plasma protein and glucose levels under hypoxia. The activities of serum glutamate oxaloacetate transaminase (SGOT) and serum glutamate pyruvate transaminase (SGPT) and the urea level remained elevated on glutamate supplementation under hypoxia. Glutamate supplementation increased the humoral response against sheep red blood cells (antibody titre). These results indicate a possible utility of glutamate in the amelioration of hypoxia-induced oxidative stress.

  15. Neuroprotection by glutamate oxaloacetate transaminase in ischemic stroke: an experimental study.

    PubMed

    Campos, Francisco; Sobrino, Tomás; Ramos-Cabrer, Pedro; Argibay, Bárbara; Agulla, Jesús; Pérez-Mato, María; Rodríguez-González, Raquel; Brea, David; Castillo, José

    2011-06-01

    As ischemic stroke is associated with an excessive release of glutamate into the neuronal extracellular space, a decrease in blood glutamate levels could provide a mechanism to remove it from the brain tissue, by increasing the brain-blood gradient. In this regard, the ability of glutamate oxaloacetate transaminase (GOT) to metabolize glutamate in blood could represent a potential neuroprotective tool for ischemic stroke. This study aimed to determine the neuroprotective effects of GOT in an animal model of cerebral ischemia by means of a middle cerebral arterial occlusion (MCAO) following the Stroke Therapy Academic Industry Roundtable (STAIR) group guidelines. In this animal model, oxaloacetate-mediated GOT activation inhibited the increase of blood and cerebral glutamate after MCAO. This effect is reflected in a reduction of infarct size, smaller edema volume, and lower sensorimotor deficits with respect to controls. Magnetic resonance spectroscopy confirmed that the increase of glutamate levels in the brain parenchyma after MCAO is inhibited after oxaloacetate-mediated GOT activation. These findings show the capacity of the GOT to remove glutamate from the brain by means of blood glutamate degradation, and suggest the applicability of this enzyme as an efficient and novel neuroprotective tool against ischemic stroke.

  16. Conditional deletion of the glutamate transporter GLT-1 reveals that astrocytic GLT-1 protects against fatal epilepsy while neuronal GLT-1 contributes significantly to glutamate uptake into synaptosomes.

    PubMed

    Petr, Geraldine T; Sun, Yan; Frederick, Natalie M; Zhou, Yun; Dhamne, Sameer C; Hameed, Mustafa Q; Miranda, Clive; Bedoya, Edward A; Fischer, Kathryn D; Armsen, Wencke; Wang, Jianlin; Danbolt, Niels C; Rotenberg, Alexander; Aoki, Chiye J; Rosenberg, Paul A

    2015-04-01

    GLT-1 (EAAT2; slc1a2) is the major glutamate transporter in the brain, and is predominantly expressed in astrocytes, but at lower levels also in excitatory terminals. We generated a conditional GLT-1 knock-out mouse to uncover cell-type-specific functional roles of GLT-1. Inactivation of the GLT-1 gene was achieved in either neurons or astrocytes by expression of synapsin-Cre or inducible human GFAP-CreERT2. Elimination of GLT-1 from astrocytes resulted in loss of ∼80% of GLT-1 protein and of glutamate uptake activity that could be solubilized and reconstituted in liposomes. This loss was accompanied by excess mortality, lower body weight, and seizures suggesting that astrocytic GLT-1 is of major importance. However, there was only a small (15%) reduction that did not reach significance of glutamate uptake into crude forebrain synaptosomes. In contrast, when GLT-1 was deleted in neurons, both the GLT-1 protein and glutamate uptake activity that could be solubilized and reconstituted in liposomes were virtually unaffected. These mice showed normal survival, weight gain, and no seizures. However, the synaptosomal glutamate uptake capacity (Vmax) was reduced significantly (40%). In conclusion, astrocytic GLT-1 performs critical functions required for normal weight gain, resistance to epilepsy, and survival. However, the contribution of astrocytic GLT-1 to glutamate uptake into synaptosomes is less than expected, and the contribution of neuronal GLT-1 to synaptosomal glutamate uptake is greater than expected based on their relative protein expression. These results have important implications for the interpretation of the many previous studies assessing glutamate uptake capacity by measuring synaptosomal uptake. PMID:25834045

  17. Conditional Deletion of the Glutamate Transporter GLT-1 Reveals That Astrocytic GLT-1 Protects against Fatal Epilepsy While Neuronal GLT-1 Contributes Significantly to Glutamate Uptake into Synaptosomes

    PubMed Central

    Petr, Geraldine T.; Sun, Yan; Frederick, Natalie M.; Zhou, Yun; Dhamne, Sameer C.; Hameed, Mustafa Q.; Miranda, Clive; Bedoya, Edward A.; Fischer, Kathryn D.; Armsen, Wencke; Wang, Jianlin; Danbolt, Niels C.; Rotenberg, Alexander; Aoki, Chiye J.

    2015-01-01

    GLT-1 (EAAT2; slc1a2) is the major glutamate transporter in the brain, and is predominantly expressed in astrocytes, but at lower levels also in excitatory terminals. We generated a conditional GLT-1 knock-out mouse to uncover cell-type-specific functional roles of GLT-1. Inactivation of the GLT-1 gene was achieved in either neurons or astrocytes by expression of synapsin-Cre or inducible human GFAP-CreERT2. Elimination of GLT-1 from astrocytes resulted in loss of ∼80% of GLT-1 protein and of glutamate uptake activity that could be solubilized and reconstituted in liposomes. This loss was accompanied by excess mortality, lower body weight, and seizures suggesting that astrocytic GLT-1 is of major importance. However, there was only a small (15%) reduction that did not reach significance of glutamate uptake into crude forebrain synaptosomes. In contrast, when GLT-1 was deleted in neurons, both the GLT-1 protein and glutamate uptake activity that could be solubilized and reconstituted in liposomes were virtually unaffected. These mice showed normal survival, weight gain, and no seizures. However, the synaptosomal glutamate uptake capacity (Vmax) was reduced significantly (40%). In conclusion, astrocytic GLT-1 performs critical functions required for normal weight gain, resistance to epilepsy, and survival. However, the contribution of astrocytic GLT-1 to glutamate uptake into synaptosomes is less than expected, and the contribution of neuronal GLT-1 to synaptosomal glutamate uptake is greater than expected based on their relative protein expression. These results have important implications for the interpretation of the many previous studies assessing glutamate uptake capacity by measuring synaptosomal uptake. PMID:25834045

  18. Effects of Amoxicillin and Augmentin on Cystine-Glutamate Exchanger and Glutamate Transporter 1 Isoforms as well as Ethanol Intake in Alcohol-Preferring Rats

    PubMed Central

    Hakami, Alqassem Y.; Hammad, Alaa M.; Sari, Youssef

    2016-01-01

    Alcohol dependence is associated with alteration of glutamate transport and glutamate neurotransmission. Glutamate transporter 1 (GLT-1) is a major transporter that regulates the majority of extracellular glutamate concentration, which is also regulated by cystine-glutamate exchanger (xCT). Importantly, we recently reported that amoxicillin and Augmentin (amoxicillin/clavulanate) upreglulated GLT-1 expression in nucleus accumbens (NAc) and prefrontal cortex (PFC) as well as reduced ethanol consumption in male P rats. In this study, we examined the effects of amoxicillin and Augmentin on GLT-1 isoforms (GLT-1a and GLT-1b), xCT, and glutamate/aspartate transporter (GLAST) expression in NAc and PFC as well as ethanol intake in male P rats. We found that both compounds significantly reduced ethanol intake, and increased GLT-1a, GLT-1b, and xCT expression in NAc. However, only Augmentin increased GLT-1a, GLT-1b, and xCT expression in PFC. There were no effects of these compounds on GLAST expression in NAc and PFC. These findings demonstrated that Augmentin and amoxicillin have the potential to upregulate GLT-1 isoforms and xCT expression, and consequently attenuate ethanol dependence. PMID:27199635

  19. Arctigenin protects cultured cortical neurons from glutamate-induced neurodegeneration by binding to kainate receptor.

    PubMed

    Jang, Young P; Kim, So R; Choi, Young H; Kim, Jinwoong; Kim, Sang G; Markelonis, George J; Oh, Tae H; Kim, Young C

    2002-04-15

    We previously reported that arctigenin, a lignan isolated from the bark of Torreya nucifera, showed significant neuroprotective activity against glutamate-induced toxicity in primary cultured rat cortical cells. In this study, the mode of action of arctigenin was investigated using primary cultures of rat cortical cells as an in vitro system. Arctigenin significantly attenuated glutamate-induced neurotoxicity when added prior to or after an excitotoxic glutamate challenge. The lignan protected cultured neuronal cells more selectively from neurotoxicity induced by kainic acid than by N-methyl-D-aspartate. The binding of [(3)H]-kainate to its receptors was significantly inhibited by arctigenin in a competitive manner. Furthermore, arctigenin directly scavenged free radicals generated by excess glutamate and successfully reduced the level of cellular peroxide in cultured neurons. These results suggest that arctigenin exerted significant neuroprotective effects on glutamate-injured primary cultures of rat cortical cells by directly binding to kainic acid receptors and partly scavenging of free radicals.

  20. Production of mesaconate in Escherichia coli by engineered glutamate mutase pathway.

    PubMed

    Wang, Jingyu; Zhang, Kechun

    2015-07-01

    Mesaconate is an intermediate in the glutamate degradation pathway of microorganisms such as Clostridium tetanomorphum. However, metabolic engineering to produce mesaconate has not been reported previously. In this work, two enzymes involved in mesaconate production, glutamate mutase and 3-methylaspartate ammonia lyase from C. tetanomorphum, were recombinantly expressed in Escherichia coli. To improve mesaconate production, reactivatase of glutamate mutase was discovered and adenosylcobalamin availability was increased. In addition, glutamate mutase was engineered to improve the in vivo activity. These efforts led to efficient mesaconate production at a titer of 7.81 g/L in shake flask with glutamate feeding. Then a full biosynthetic pathway was constructed to produce mesaconate at a titer of 6.96 g/L directly from glucose. In summary, we have engineered an efficient system in E. coli for the biosynthesis of mesaconate.

  1. An essential role of acetylcholine-glutamate synergy at habenular synapses in nicotine dependence.

    PubMed

    Frahm, Silke; Antolin-Fontes, Beatriz; Görlich, Andreas; Zander, Johannes-Friedrich; Ahnert-Hilger, Gudrun; Ibañez-Tallon, Ines

    2015-12-01

    A great deal of interest has been focused recently on the habenula and its critical role in aversion, negative-reward and drug dependence. Using a conditional mouse model of the ACh-synthesizing enzyme choline acetyltransferase (Chat), we report that local elimination of acetylcholine (ACh) in medial habenula (MHb) neurons alters glutamate corelease and presynaptic facilitation. Electron microscopy and immuno-isolation analyses revealed colocalization of ACh and glutamate vesicular transporters in synaptic vesicles (SVs) in the central IPN. Glutamate reuptake in SVs prepared from the IPN was increased by ACh, indicating vesicular synergy. Mice lacking CHAT in habenular neurons were insensitive to nicotine-conditioned reward and withdrawal. These data demonstrate that ACh controls the quantal size and release frequency of glutamate at habenular synapses, and suggest that the synergistic functions of ACh and glutamate may be generally important for modulation of cholinergic circuit function and behavior.

  2. Glutamate transport and xanthan gum production in the plant pathogen Xanthomonas axonopodis pv. citri.

    PubMed

    Rojas, Robert; Nishidomi, Sabrina; Nepomuceno, Roberto; Oshiro, Elisa; de Cassia Café Ferreira, Rita

    2013-11-01

    L-glutamate plays a central role in nitrogen metabolism in all living organisms. In the genus Xanthomonas, the nitrogen nutrition is an important factor involved in the xanthan gum production, an important exopolysaccharide with various industrial and biotechnological applications. In this report, we demonstrate that the use of L-glutamate by the phytopathogen Xanthomonas axonopodis pv. citri as a nitrogen source in defined medium significantly increases the production of xanthan gum. This increase is dependent on the L-glutamate concentration. In addition, we have also characterized a glutamate transport system that is dependent on a proton gradient and on ATP and is modulated by amino acids that are structurally related to glutamate. This is the first biochemical characterization of an energy substrate transport system observed in a bacterial phytopathogen with a broad economic and industrial impact due to xanthan gum production. PMID:23719672

  3. Glutamate system, amyloid ß peptides and tau protein: functional interrelationships and relevance to Alzheimer disease pathology.

    PubMed

    Revett, Timothy J; Baker, Glen B; Jhamandas, Jack; Kar, Satyabrata

    2013-01-01

    Alzheimer disease is the most prevalent form of dementia globally and is characterized premortem by a gradual memory loss and deterioration of higher cognitive functions and postmortem by neuritic plaques containing amyloid ß peptide and neurofibrillary tangles containing phospho-tau protein. Glutamate is the most abundant neurotransmitter in the brain and is essential to memory formation through processes such as long-term potentiation and so might be pivotal to Alzheimer disease progression. This review discusses how the glutamatergic system is impaired in Alzheimer disease and how interactions of amyloid ß and glutamate influence synaptic function, tau phosphorylation and neurodegeneration. Interestingly, glutamate not only influences amyloid ß production, but also amyloid ß can alter the levels of glutamate at the synapse, indicating that small changes in the concentrations of both molecules could influence Alzheimer disease progression. Finally, we describe how the glutamate receptor antagonist, memantine, has been used in the treatment of individuals with Alzheimer disease and discuss its effectiveness.

  4. On the Role of Glutamate in Presynaptic Development: Possible Contributions of Presynaptic NMDA Receptors

    PubMed Central

    Fedder, Karlie N.; Sabo, Shasta L.

    2015-01-01

    Proper formation and maturation of synapses during development is a crucial step in building the functional neural circuits that underlie perception and behavior. It is well established that experience modifies circuit development. Therefore, understanding how synapse formation is controlled by synaptic activity is a key question in neuroscience. In this review, we focus on the regulation of excitatory presynaptic terminal development by glutamate, the predominant excitatory neurotransmitter in the brain. We discuss the evidence that NMDA receptor activation mediates these effects of glutamate and present the hypothesis that local activation of presynaptic NMDA receptors (preNMDARs) contributes to glutamate-dependent control of presynaptic development. Abnormal glutamate signaling and aberrant synapse development are both thought to contribute to the pathogenesis of a variety of neurodevelopmental disorders, including autism spectrum disorders, intellectual disability, epilepsy, anxiety, depression, and schizophrenia. Therefore, understanding how glutamate signaling and synapse development are linked is important for understanding the etiology of these diseases. PMID:26694480

  5. Selective recognition of Glutamate based on fluorescence enhancement of graphene quantum dot

    NASA Astrophysics Data System (ADS)

    Hosseini, Morteza; Khabbaz, Hossein; Dezfoli, Amin Shiralizadeh; Ganjali, Mohammad Reza; Dadmehr, Mehdi

    2015-02-01

    Graphene quantum dots (GQDs) have successfully been utilized as an efficient nano-sized fluorescence chemosensor to detect selectively Glutamate (Glu) in Tris-HCl buffer solution (pH = 9). The fluorescence emission spectrum of graphene quantum dots was at about 430 nm. The study showed that fluorescence intensity of the quantum dot gradually enhanced with increase in concentration of Glutamate and any change in fluorescence intensity was directly proportional to the concentration of Glutamate. Under optimum conditions, the linear range for the detection of Glutamate was 1.6 × 10-7 M to 1.0 × 10-5 M with a detection limit of 5.2 × 10-8 M. The sensor showed high selectivity toward Glutamate in comparison with other amino acids.

  6. Sonic hedgehog is a regulator of extracellular glutamate levels and epilepsy.

    PubMed

    Feng, Shengjie; Ma, Shaorong; Jia, Caixia; Su, Yujuan; Yang, Shenglian; Zhou, Kechun; Liu, Yani; Cheng, Ju; Lu, Dunguo; Fan, Liu; Wang, Yizheng

    2016-05-01

    Sonic hedgehog (Shh), both as a mitogen and as a morphogen, plays an important role in cell proliferation and differentiation during early development. Here, we show that Shh inhibits glutamate transporter activities in neurons, rapidly enhances extracellular glutamate levels, and affects the development of epilepsy. Shh is quickly released in response to epileptic, but not physiological, stimuli. Inhibition of neuronal glutamate transporters by Shh depends on heterotrimeric G protein subunit Gαi and enhances extracellular glutamate levels. Inhibiting Shh signaling greatly reduces epileptiform activities in both cell cultures and hippocampal slices. Moreover, pharmacological or genetic inhibition of Shh signaling markedly suppresses epileptic phenotypes in kindling or pilocarpine models. Our results suggest that Shh contributes to the development of epilepsy and suppression of its signaling prevents the development of the disease. Thus, Shh can act as a modulator of neuronal activity, rapidly regulating glutamate levels and promoting epilepsy. PMID:27113760

  7. Effect of aspartame and sucrose loading in glutamate-susceptible subjects.

    PubMed

    Stegink, L D; Filer, L J; Baker, G L

    1981-09-01

    It has been postulated that individuals reporting an idiosyncratic symptom response after glutamate ingestion might also experience such symptoms after aspartame ingestion. Such sensitive subjects might have been missed in earlier studies of aspartame. In the present study, six subjects reporting various symptoms after glutamate ingestion, but not after placebo, were administered aspartame (34 mg/kg body weight) or sucrose (1 g/kg body weight) dissolved in orange juice in a randomized, cross-over, double-blind study. No subject reported symptoms typical of a glutamate response after either sucrose or aspartame loading. One subject reported slight nausea approximately 1.5 h after aspartame ingestion, but indicated that the symptoms were not those of a glutamate response. Plasma phenylalanine and aspartate levels were similar to those noted in normal subjects administered identical doses of aspartame. The data indicate no effect of aspartame loading in glutamate-susceptible subjects.

  8. An amperometric biosensor for L-glutamate determination prepared from L-glutamate oxidase immobilized in polypyrrole-polyvinylsulphonate film.

    PubMed

    Şimşek, Şule; Aynacı, Elif; Arslan, Fatma

    2016-01-01

    In this paper, a novel amperometric L-glutamate (Glu) biosensor with immobilization of L-glutamate oxidase (L-GlOx) on polypyrrole-polyvinylsulphonate (PPy-PVS) film has been successfully developed. L-GlOx enzyme was immobilized on PPy-PVS film by cross-linking with glutaraldehyde (GA) and bovine serum albumin (BSA). Determination of Glu was carried out by oxidation of enzymatically produced H2O2 at 0.3 V versus Ag/AgCl. The optimum pH and temperature parameters were found to be 9.0 and 55 °C, respectively. There were three linear parts in the regions between 1.0 × 10(-9) and 1.0 × 10(-8) M (R(2) = 0.847), 5.0 × 10(-8) and 5.0 × 10(-7) M (R(2) = 0.997), 5.0 × 10(-7) and 5.0 × 10(-5) M (R(2) = 0.994). Storage stability, operation stability of the enzyme electrode were also studied.

  9. Pharmacological inhibitions of glutamate transporters EAAT1 and EAAT2 compromise glutamate transport in photoreceptor to ON- bipolar cell synapses

    PubMed Central

    Tse, Dennis Y.; Chung, Inyoung; Wu, Samuel M.

    2015-01-01

    To maintain reliable signal transmission across a synapse, free synaptic neurotransmitters must be removed from the cleft in a timely manner. In the first visual synapse, this critical task is mainly undertaken by glutamate transporters (EAATs). Here we study the differential roles of the EAAT1, EAAT2 and EAAT5 subtypes in glutamate (GLU) uptake at the photoreceptor-to-depolarizing bipolar cell synapse in intact dark-adapted retina. Various doses of EAAT blockers and/or GLU were injected into the eye before the electroretinogram (ERG) was measured. Their effectiveness and potency in inhibiting the ERG b-wave were studied to determine their relative contributions to the GLU clearing activity at the synapse. The results showed that EAAT1 and EAAT2 plays different roles. Selectively blocking glial EAAT1 alone using UCPH101 inhibited the b-wave 2–24 hours following injection, suggesting a dominating role of EAAT1 in the overall GLU clearing capacity in the synaptic cleft. Selectively blocking EAAT2 on photoreceptor terminals had no significant effect on the b-wave, but increased the potency of exogenous GLU in inhibiting the b-wave. These suggest that EAAT2 play a secondary yet significant role in the GLU reuptake activity at the rod and the cone output synapses. Additionally, we have verified our electrophysiological findings with double-label immunohistochemistry, and extend the literature on the spatial distribution of EAAT2 splice variants in the mouse retina. PMID:25152321

  10. Conceptual convergence: increased inflammation is associated with increased basal ganglia glutamate in patients with major depression.

    PubMed

    Haroon, E; Fleischer, C C; Felger, J C; Chen, X; Woolwine, B J; Patel, T; Hu, X P; Miller, A H

    2016-10-01

    Inflammation and altered glutamate metabolism are two pathways implicated in the pathophysiology of depression. Interestingly, these pathways may be linked given that administration of inflammatory cytokines such as interferon-α to otherwise non-depressed controls increased glutamate in the basal ganglia and dorsal anterior cingulate cortex (dACC) as measured by magnetic resonance spectroscopy (MRS). Whether increased inflammation is associated with increased glutamate among patients with major depression is unknown. Accordingly, we conducted a cross-sectional study of 50 medication-free, depressed outpatients using single-voxel MRS, to measure absolute glutamate concentrations in basal ganglia and dACC. Multivoxel chemical shift imaging (CSI) was used to explore creatine-normalized measures of other metabolites in basal ganglia. Plasma and cerebrospinal fluid (CSF) inflammatory markers were assessed along with anhedonia and psychomotor speed. Increased log plasma C-reactive protein (CRP) was significantly associated with increased log left basal ganglia glutamate controlling for age, sex, race, body mass index, smoking status and depression severity. In turn, log left basal ganglia glutamate was associated with anhedonia and psychomotor slowing measured by the finger-tapping test, simple reaction time task and the Digit Symbol Substitution Task. Plasma CRP was not associated with dACC glutamate. Plasma and CSF CRP were also associated with CSI measures of basal ganglia glutamate and the glial marker myoinositol. These data indicate that increased inflammation in major depression may lead to increased glutamate in the basal ganglia in association with glial dysfunction and suggest that therapeutic strategies targeting glutamate may be preferentially effective in depressed patients with increased inflammation as measured by CRP. PMID:26754953

  11. Conceptual convergence: increased inflammation is associated with increased basal ganglia glutamate in patients with major depression

    PubMed Central

    Haroon, E; Fleischer, C C; Felger, J C; Chen, X; Woolwine, B J; Patel, T; Hu, X P; Miller, A H

    2016-01-01

    Inflammation and altered glutamate metabolism are two pathways implicated in the pathophysiology of depression. Interestingly, these pathways may be linked given that administration of inflammatory cytokines such as interferon-α to otherwise non-depressed controls increased glutamate in the basal ganglia and dorsal anterior cingulate cortex (dACC) as measured by magnetic resonance spectroscopy (MRS). Whether increased inflammation is associated with increased glutamate among patients with major depression is unknown. Accordingly, we conducted a cross-sectional study of 50 medication-free, depressed outpatients using single-voxel MRS, to measure absolute glutamate concentrations in basal ganglia and dACC. Multivoxel chemical shift imaging (CSI) was used to explore creatine-normalized measures of other metabolites in basal ganglia. Plasma and cerebrospinal fluid (CSF) inflammatory markers were assessed along with anhedonia and psychomotor speed. Increased log plasma C-reactive protein (CRP) was significantly associated with increased log left basal ganglia glutamate controlling for age, sex, race, body mass index, smoking status and depression severity. In turn, log left basal ganglia glutamate was associated with anhedonia and psychomotor slowing measured by the finger-tapping test, simple reaction time task and the Digit Symbol Substitution Task. Plasma CRP was not associated with dACC glutamate. Plasma and CSF CRP were also associated with CSI measures of basal ganglia glutamate and the glial marker myoinositol. These data indicate that increased inflammation in major depression may lead to increased glutamate in the basal ganglia in association with glial dysfunction and suggest that therapeutic strategies targeting glutamate may be preferentially effective in depressed patients with increased inflammation as measured by CRP. PMID:26754953

  12. Molecular Determinants of Substrate Specificity in Sodium-coupled Glutamate Transporters.

    PubMed

    Silverstein, Nechama; Ewers, David; Forrest, Lucy R; Fahlke, Christoph; Kanner, Baruch I

    2015-11-27

    Crystal structures of the archaeal homologue GltPh have provided important insights into the molecular mechanism of transport of the excitatory neurotransmitter glutamate. Whereas mammalian glutamate transporters can translocate both glutamate and aspartate, GltPh is only one capable of aspartate transport. Most of the amino acid residues that surround the aspartate substrate in the binding pocket of GltPh are highly conserved. However, in the brain transporters, Thr-352 and Met-362 of the reentrant hairpin loop 2 are replaced by the smaller Ala and Thr, respectively. Therefore, we have studied the effects of T352A and M362T on binding and transport of aspartate and glutamate by GltPh. Substrate-dependent intrinsic fluorescence changes were monitored in transporter constructs containing the L130W mutation. GltPh-L130W/T352A exhibited an ~15-fold higher apparent affinity for l-glutamate than the wild type transporter, and the M362T mutation resulted in an increased affinity of ~40-fold. An even larger increase of the apparent affinity for l-glutamate, around 130-fold higher than that of wild type, was observed with the T352A/M362T double mutant. Radioactive uptake experiments show that GltPh-T352A not only transports aspartate but also l-glutamate. Remarkably, GltPh-M362T exhibited l-aspartate but not l-glutamate transport. The double mutant retained the ability to transport l-glutamate, but its kinetic parameters were very similar to those of GltPh-T352A alone. The differential impact of mutation on binding and transport of glutamate suggests that hairpin loop 2 not only plays a role in the selection of the substrate but also in its translocation.

  13. Post-synaptic calcium influx at the giant synapse of the squid during activation by glutamate.

    PubMed Central

    Eusebi, F; Miledi, R; Parker, I; Stinnakre, J

    1985-01-01

    Changes in free calcium were monitored in the post-synaptic axon of the giant synapse of the squid, using the calcium indicators aequorin and Arsenazo III. The peak size of the calcium-dependent optical signals recorded from aequorin and Arsenazo III both showed a linear relation with the amount of calcium injected ionophoretically into the axon, but the Arsenazo signal had a slower time course than the aequorin. Ionophoretic application of glutamate to the post-synaptic axon depolarized the axon and caused a rise in intracellular free calcium. Aequorin signals were detected in natural sea water, and their size increased when the calcium concentration in the sea water was raised. Arsenazo signals could be detected only in high-calcium (55 mM) sea water. Intracellular calcium signals were detected also during bath application of several glutamate analogues, including kainate, ibotenate, and aspartate. The peak amplitude of the intracellular calcium signal, monitored with both indicators, increased with increasing ionophoretic glutamate dose, and varied linearly with the integral of the glutamate-induced membrane depolarization. No calcium signals were detected when depolarizations, similar to those produced by glutamate, were induced by current injection in the absence of glutamate. We conclude that glutamate increases the calcium permeability of the post-synaptic membrane, independently of the glutamate-induced depolarization. The glutamate-induced depolarization and the rise in intracellular free calcium increased roughly linearly as the membrane potential was made more negative. Extrapolation of these data indicated that the glutamate depolarization would reduce to zero at about -30 mV, while the calcium signals would be suppressed at about +50 mV. PMID:2869144

  14. Altered acetylation and succinylation profiles in Corynebacterium glutamicum in response to conditions inducing glutamate overproduction.

    PubMed

    Mizuno, Yuta; Nagano-Shoji, Megumi; Kubo, Shosei; Kawamura, Yumi; Yoshida, Ayako; Kawasaki, Hisashi; Nishiyama, Makoto; Yoshida, Minoru; Kosono, Saori

    2016-02-01

    The bacterium Corynebacterium glutamicum is utilized during industrial fermentation to produce amino acids such as L-glutamate. During L-glutamate fermentation, C. glutamicum changes the flux of central carbon metabolism to favor L-glutamate production, but the molecular mechanisms that explain these flux changes remain largely unknown. Here, we found that the profiles of two major lysine acyl modifications were significantly altered upon glutamate overproduction in C. glutamicum; acetylation decreased, whereas succinylation increased. A label-free semi-quantitative proteomic analysis identified 604 acetylated proteins with 1328 unique acetylation sites and 288 succinylated proteins with 651 unique succinylation sites. Acetylation and succinylation targeted enzymes in central carbon metabolic pathways that are directly related to glutamate production, including the 2-oxoglutarate dehydrogenase complex (ODHC), a key enzyme regulating glutamate overproduction. Structural mapping revealed that several critical lysine residues in the ODHC components were susceptible to acetylation and succinylation. Furthermore, induction of glutamate production was associated with changes in the extent of acetylation and succinylation of lysine, suggesting that these modifications may affect the activity of enzymes involved in glutamate production. Deletion of phosphotransacetylase decreased the extent of protein acetylation in nonproducing condition, suggesting that acetyl phosphate-dependent acetylation is active in C. glutamicum. However, no effect was observed on the profiles of acetylation and succinylation in glutamate-producing condition upon disruption of acetyl phosphate metabolism or deacetylase homologs. It was considered likely that the reduced acetylation in glutamate-producing condition may reflect metabolic states where the flux through acid-producing pathways is very low, and substrates for acetylation do not accumulate in the cell. Succinylation would occur more

  15. New 4-Functionalized Glutamate Analogues Are Selective Agonists at Metabotropic Glutamate Receptor Subtype 2 or Selective Agonists at Metabotropic Glutamate Receptor Group III.

    PubMed

    Huynh, Tri H V; Erichsen, Mette N; Tora, Amélie S; Goudet, Cyril; Sagot, Emmanuelle; Assaf, Zeinab; Thomsen, Christian; Brodbeck, Robb; Stensbøl, Tine B; Bjørn-Yoshimoto, Walden E; Nielsen, Birgitte; Pin, Jean-Philippe; Gefflaut, Thierry; Bunch, Lennart

    2016-02-11

    The metabotropic glutamate (Glu) receptors (mGluRs) play key roles in modulating excitatory neurotransmission in the brain. In all, eight subtypes have been identified and divided into three groups, group I (mGlu1,5), group II (mGlu2,3), and group III (mGlu4,6-8). In this article, we present a L-2,4-syn-substituted Glu analogue, 1d, which displays selective agonist activity at mGlu2 over the remaining mGluR subtypes. A modeling study and redesign of the core scaffold led to the stereoselective synthesis of four new conformationally restricted Glu analogues, 2a-d. Most interestingly, 2a retained a selective agonist activity profile at mGlu2 (EC50 in the micromolar range), whereas 2c/2d were both selective agonists at group III, subtypes mGlu4,6,8. In general, 2d was 20-fold more potent than 2c and potently activated mGlu4,6,8 in the low-mid nanomolar range.

  16. Structure and function of glutamate receptor ion channels.

    PubMed

    Mayer, Mark L; Armstrong, Neali

    2004-01-01

    A vast number of proteins are involved in synaptic function. Many have been cloned and their functional role defined with varying degrees of success, but their number and complexity currently defy any molecular understanding of the physiology of synapses. A beacon of success in this medieval era of synaptic biology is an emerging understanding of the mechanisms underlying the activity of the neurotransmitter receptors for glutamate. Largely as a result of structural studies performed in the past three years we now have a mechanistic explanation for the activation of channel gating by agonists and partial agonists; the process of desensitization, and its block by allosteric modulators, is also mostly explained; and the basis of receptor subtype selectivity is emerging with clarity as more and more structures are solved. In the space of months we have gone from cartoons of postulated mechanisms to hard fact. It is anticipated that this level of understanding will emerge for other synaptic proteins in the coming decade.

  17. Molecular Insights into Metabotropic Glutamate Receptor Allosteric Modulation

    PubMed Central

    Gregory, Karen J.

    2015-01-01

    The metabotropic glutamate (mGlu) receptors are a group of eight family C G protein–coupled receptors that are expressed throughout the central nervous system (CNS) and periphery. Within the CNS the different subtypes are found in neurons, both pre- and/or postsynaptically, where they mediate modulatory roles and in glial cells. The mGlu receptor family provides attractive targets for numerous psychiatric and neurologic disorders, with the majority of discovery programs focused on targeting allosteric sites, with allosteric ligands now available for all mGlu receptor subtypes. However, the development of allosteric ligands remains challenging. Biased modulation, probe dependence, and molecular switches all contribute to the complex molecular pharmacology exhibited by mGlu receptor allosteric ligands. In recent years we have made significant progress in our understanding of this molecular complexity coupled with an increased understanding of the structural basis of mGlu allosteric modulation. PMID:25808929

  18. Loss of Astrocytic Glutamate Transporters in Wernicke Encephalopathy

    PubMed Central

    Hazell, Alan S.; Sheedy, Donna; Oanea, Raluca; Aghourian, Meghmik; Sun, Simon; Jung, Jee Yong; Wang, Dongmei; Wang, Chunlei

    2012-01-01

    Wernicke encephalopathy (WE), a neurological disorder caused by thiamine deficiency (TD), is characterized by structural damage in brain regions that include the thalamus and cerebral cortex. The basis for these lesions is unclear, but may involve a disturbance of glutamatergic neurotransmission. We have therefore investigated levels of the astrocytic glutamate transporters EAAT1 and EAAT2 in order to evaluate their role in the pathophysiology of this disorder. Histological assessment of the frontal cortex revealed a significant loss of neurons in neuropathologically confirmed cases of WE compared with age-matched controls, concomitant with decreases in α-internexin and synaptophysin protein content of 67 and 52% by immunoblotting. EAAT2 levels were diminished by 71% in WE, with levels of EAAT1 also reduced by 62%. Loss of both transporter sites was confirmed by immunohistochemical methods. Development of TD in rats caused a profound loss of EAAT1 and EAAT2 in the thalamus accompanied by decreases in other astrocyte-specific proteins. Treatment of TD rats with N-acetylcysteine prevented the downregulation of EAAT2 in the medial thalamus, and ameliorated the loss of several other astrocyte proteins, concomitant with increased neuronal survival. Our results suggest that (1) loss of EAAT1 and EAAT2 glutamate transporters is associated with structural damage to the frontal cortex in patients with WE, (2) oxidative stress plays an important role in this process, and (3) TD has a profound effect on the functional integrity of astrocytes. Based on these findings, we recommend that early treatment using a combination of thiamine AND antioxidant approaches should be an important consideration in cases of WE. PMID:19565658

  19. 13N as a tracer for studying glutamate metabolism

    PubMed Central

    Cooper, Arthur J. L.

    2010-01-01

    This mini-review summarizes studies my associates and I carried out that are relevant to the topic of the present volume [i.e. glutamate dehydrogenase (GDH)] using radioactive 13N (t½ 9.96 min) as a biological tracer. These studies revealed the previously unrecognized rapidity with which nitrogen is exchanged among certain metabolites in vivo. For example, our work demonstrated that a) the t½ for conversion of portal vein ammonia to urea in the rat liver is ~10–11 sec, despite the need for five enzyme-catalyzed steps and two mitochondrial transport steps, b) the residence time for ammonia in the blood of anesthetized rats is ≤7–8 sec, c) the t½ for incorporation of blood-borne ammonia into glutamine in the normal rat brain is <3 sec, and d) equilibration between glutamate and aspartate nitrogen in rat liver is extremely rapid (seconds), a reflection of the fact that the components of the hepatic aspartate aminotransferase reaction are in thermodynamic equilibrium. Our work emphasizes the importance of the GDH reaction in rat liver as a conduit for dissimilating or assimilating ammonia as needed. In contrast, our work shows that the GDH reaction in rat brain appears to operate mostly in the direction of ammonia production (dissimilation). The importance of the GDH reaction as an endogenous source of ammonia in the brain and the relation of GDH to the brain glutamine cycle is discussed. Finally, our work integrates with the increasing use of positron emission tomography (PET) and nuclear magnetic resonance (NMR) to study brain ammonia uptake and brain glutamine, respectively, in normal individuals and in patients with liver disease or other diseases associated with hyperammonemia. PMID:21108979

  20. Glutamate dehydrogenases: the why and how of coenzyme specificity.

    PubMed

    Engel, Paul C

    2014-01-01

    NAD(+) and NADP(+), chemically similar and with almost identical standard oxidation-reduction potentials, nevertheless have distinct roles, NAD(+) serving catabolism and ATP generation whereas NADPH is the biosynthetic reductant. Separating these roles requires strict specificity for one or the other coenzyme for most dehydrogenases. In many organisms this holds also for glutamate dehydrogenases (GDH), NAD(+)-dependent for glutamate oxidation, NADP(+)-dependent for fixing ammonia. In higher animals, however, GDH has dual specificity. It has been suggested that GDH in mitochondria reacts only with NADP(H), the NAD(+) reaction being an in vitro artefact. However, contrary evidence suggests mitochondrial GDH not only reacts with NAD(+) but maintains equilibrium using the same pool as accessed by β-hydroxybutyrate dehydrogenase. Another complication is the presence of an energy-linked dehydrogenase driving NADP(+) reduction by NADH, maintaining the coenzyme pools at different oxidation-reduction potentials. Its coexistence with GDH makes possible a futile cycle, control of which is not yet properly explained. Structural studies show NAD(+)-dependent, NADP(+)-dependent and dual-specificity GDHs are closely related and a few site-directed mutations can reverse specificity. Specificity for NAD(+) or for NADP(+) has probably emerged repeatedly during evolution, using different structural solutions on different occasions. In various GDHs the P7 position in the coenzyme-binding domain plays a key role. However, whereas in other dehydrogenases an acidic P7 residue usually hydrogen bonds to the 2'- and 3'-hydroxyls, dictating NAD(+) specificity, among GDHs, depending on detailed conformation of surrounding residues, an acidic P7 may permit binding of NAD(+) only, NADP(+) only, or in higher animals both.

  1. Ionotropic GABA and Glutamate Receptor Mutations and Human Neurologic Diseases.

    PubMed

    Yuan, Hongjie; Low, Chian-Ming; Moody, Olivia A; Jenkins, Andrew; Traynelis, Stephen F

    2015-07-01

    The advent of whole exome/genome sequencing and the technology-driven reduction in the cost of next-generation sequencing as well as the introduction of diagnostic-targeted sequencing chips have resulted in an unprecedented volume of data directly linking patient genomic variability to disorders of the brain. This information has the potential to transform our understanding of neurologic disorders by improving diagnoses, illuminating the molecular heterogeneity underlying diseases, and identifying new targets for therapeutic treatment. There is a strong history of mutations in GABA receptor genes being involved in neurologic diseases, particularly the epilepsies. In addition, a substantial number of variants and mutations have been found in GABA receptor genes in patients with autism, schizophrenia, and addiction, suggesting potential links between the GABA receptors and these conditions. A new and unexpected outcome from sequencing efforts has been the surprising number of mutations found in glutamate receptor subunits, with the GRIN2A gene encoding the GluN2A N-methyl-d-aspartate receptor subunit being most often affected. These mutations are associated with multiple neurologic conditions, for which seizure disorders comprise the largest group. The GluN2A subunit appears to be a locus for epilepsy, which holds important therapeutic implications. Virtually all α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor mutations, most of which occur within GRIA3, are from patients with intellectual disabilities, suggesting a link to this condition. Similarly, the most common phenotype for kainate receptor variants is intellectual disability. Herein, we summarize the current understanding of disease-associated mutations in ionotropic GABA and glutamate receptor families, and discuss implications regarding the identification of human mutations and treatment of neurologic diseases. PMID:25904555

  2. Ionotropic GABA and Glutamate Receptor Mutations and Human Neurologic Diseases.

    PubMed

    Yuan, Hongjie; Low, Chian-Ming; Moody, Olivia A; Jenkins, Andrew; Traynelis, Stephen F

    2015-07-01

    The advent of whole exome/genome sequencing and the technology-driven reduction in the cost of next-generation sequencing as well as the introduction of diagnostic-targeted sequencing chips have resulted in an unprecedented volume of data directly linking patient genomic variability to disorders of the brain. This information has the potential to transform our understanding of neurologic disorders by improving diagnoses, illuminating the molecular heterogeneity underlying diseases, and identifying new targets for therapeutic treatment. There is a strong history of mutations in GABA receptor genes being involved in neurologic diseases, particularly the epilepsies. In addition, a substantial number of variants and mutations have been found in GABA receptor genes in patients with autism, schizophrenia, and addiction, suggesting potential links between the GABA receptors and these conditions. A new and unexpected outcome from sequencing efforts has been the surprising number of mutations found in glutamate receptor subunits, with the GRIN2A gene encoding the GluN2A N-methyl-d-aspartate receptor subunit being most often affected. These mutations are associated with multiple neurologic conditions, for which seizure disorders comprise the largest group. The GluN2A subunit appears to be a locus for epilepsy, which holds important therapeutic implications. Virtually all α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor mutations, most of which occur within GRIA3, are from patients with intellectual disabilities, suggesting a link to this condition. Similarly, the most common phenotype for kainate receptor variants is intellectual disability. Herein, we summarize the current understanding of disease-associated mutations in ionotropic GABA and glutamate receptor families, and discuss implications regarding the identification of human mutations and treatment of neurologic diseases.

  3. Modification of Glutamate Receptor Channels: Molecular Mechanisms and Functional Consequences

    NASA Astrophysics Data System (ADS)

    Hatt, Hanns

    Of the many possible mechanisms for modulating the efficiency of ion channels, the phosphorylation of receptor channel proteins may be the primary one. Changes in the set of molecular subunits of which the channels are composed are also important, especially for long-term regulation. In the central nervous system synaptic plasticity may be altered by modulating the ligand-activated neuronal ion channels involved in synaptic transmission; among them are channels gated directly by glutamate, the regulation of which we are only beginning to understand. This paper focuses on modulation of these channels [α-amino-3-hydroxy-5-methyl-4-isoxazoleprionic acid (AMPA), kainate, and N-methyl-d-aspartate (NMDA) types] by phosphorylation and changes in subunit composition. AMPA- and kainate-activated receptors are modulated by adenosine 3, 5-monophosphate (cAMP) dependent protein kinase A (PKA) coupled via D1 dopamine receptors. An increase in the intracellular concentration of cAMP and protein kinase A potentiates kainate-activated currents in α-motoneurons of the spinal cord by increasing the affinity of the ligand (glutamate) for the phosphorylated receptor protein (GluR6 and 7). The rapid desensitization of AMPA-evoked currents normally observed in horizontal cells of the retina is completely blocked by increasing the intracellular concentration of cAMP. The effects of changes in subunit composition were examined in rat hippocampal neurons. The subunit composition of the NMDA receptor determines the kinetic properties of synaptic currents and can be regulated by the type of innervating neuron. Similar changes also occur during development. An important determinant here is the activity of the system. Dynamic regulation of excitatory receptors by both mechanisms may well be associated with some forms of learning and memory in the mammalian brain.

  4. Ionotropic GABA and Glutamate Receptor Mutations and Human Neurologic Diseases

    PubMed Central

    Yuan, Hongjie; Low, Chian-Ming; Moody, Olivia A.; Jenkins, Andrew

    2015-01-01

    The advent of whole exome/genome sequencing and the technology-driven reduction in the cost of next-generation sequencing as well as the introduction of diagnostic-targeted sequencing chips have resulted in an unprecedented volume of data directly linking patient genomic variability to disorders of the brain. This information has the potential to transform our understanding of neurologic disorders by improving diagnoses, illuminating the molecular heterogeneity underlying diseases, and identifying new targets for therapeutic treatment. There is a strong history of mutations in GABA receptor genes being involved in neurologic diseases, particularly the epilepsies. In addition, a substantial number of variants and mutations have been found in GABA receptor genes in patients with autism, schizophrenia, and addiction, suggesting potential links between the GABA receptors and these conditions. A new and unexpected outcome from sequencing efforts has been the surprising number of mutations found in glutamate receptor subunits, with the GRIN2A gene encoding the GluN2A N-methyl-d-aspartate receptor subunit being most often affected. These mutations are associated with multiple neurologic conditions, for which seizure disorders comprise the largest group. The GluN2A subunit appears to be a locus for epilepsy, which holds important therapeutic implications. Virtually all α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptor mutations, most of which occur within GRIA3, are from patients with intellectual disabilities, suggesting a link to this condition. Similarly, the most common phenotype for kainate receptor variants is intellectual disability. Herein, we summarize the current understanding of disease-associated mutations in ionotropic GABA and glutamate receptor families, and discuss implications regarding the identification of human mutations and treatment of neurologic diseases. PMID:25904555

  5. Connexin-Mediated Cardiac Impulse Propagation: Connexin 30.2 Slows Atrioventricular Conduction in Mouse Heart

    PubMed Central

    Kreuzberg, Maria M.; Willecke, Klaus; Bukauskas, Feliksas F.

    2013-01-01

    In mouse heart, four connexins (Cxs), Cx30.2, Cx40, Cx43, and Cx45, form gap junction (GJ) channels for electric and metabolic cell-to-cell signaling. Extent and pattern of Cx isoform expression together with cytoarchitecture and excitability of cells determine the velocity of excitation spread in different regions of the heart. In the SA node, cell– cell coupling is mediated by Cx30.2 and Cx45, which form lowconductance (approximately 9 and 32 pS, respectively) GJ channels. In contrast, the working cardiomyocytes of atria and ventricles express mainly Cx40 and Cx43, which form GJ channels of high conductance (approximately 180 and 115 pS, respectively) that facilitate the fast conduction necessary for efficient mechanical contraction. In the AV node, cell–cell coupling is mediated by abundantly expressed Cx30.2 and Cx45 and Cx40, which is expressed to a lesser extent. Cx30.2 and Cx45 may determine higher intercellular resistance and slower conduction in the SA- and AV-nodal regions than in the ventricular conduction system or the atrial and ventricular working myocardium. Cx30.2 and its putative human ortholog, Cx31.9, under physiologic conditions form unapposed hemichannels in nonjunctional plasma membrane; these hemichannels have a conductance of approximately 20 pS and are permeable to cationic dyes up to approximately 400 Da in molecular mass. Genetic ablation of Cxs confirmed that Cx40 and Cx43 are important in determining the high conduction velocities in atria and ventricles, whereas the deletion of the Cx30.2 complementary DNA led to accelerated conduction in the AV node and reduced the Wenckebach period. We suggest that these effects are caused by (1) a dominant-negative effect of Cx30.2 on junctional conductance via formation of low-conductance homotypic and heterotypic GJ channels, and (2) open Cx30.2 hemichannels in non-junctional membranes, which shorten the space constant and depolarize the excitable membrane. PMID:17055382

  6. Maternal inflammation leads to impaired glutamate homeostasis and up-regulation of glutamate carboxypeptidase II in activated microglia in the fetal/newborn rabbit brain.

    PubMed

    Zhang, Zhi; Bassam, Bassam; Thomas, Ajit G; Williams, Monica; Liu, Jinhuan; Nance, Elizabeth; Rojas, Camilo; Slusher, Barbara S; Kannan, Sujatha

    2016-10-01

    Astrocyte dysfunction and excessive activation of glutamatergic systems have been implicated in a number of neurologic disorders, including periventricular leukomalacia (PVL) and cerebral palsy (CP). However, the role of chorioamnionitis on glutamate homeostasis in the fetal and neonatal brains is not clearly understood. We have previously shown that intrauterine endotoxin administration results in intense microglial 'activation' and increased pro-inflammatory cytokines in the periventricular region (PVR) of the neonatal rabbit brain. In this study, we assessed the effect of maternal inflammation on key components of the glutamate pathway and its relationship to astrocyte and microglial activation in the fetal and neonatal New Zealand white rabbit brain. We found that intrauterine endotoxin exposure at gestational day 28 (G28) induced acute and prolonged glutamate elevation in the PVR of fetal (G29, 1day post-injury) and postnatal day 1 (PND1, 3days post-injury) brains along with prominent morphological changes in the astrocytes (soma hypertrophy and retracted processes) in the white matter tracts. There was a significant increase in glutaminase and N-Methyl-d-Aspartate receptor (NMDAR) NR2 subunit expression along with decreased glial L-glutamate transporter 1 (GLT-1) in the PVR at G29, that would promote acute dysregulation of glutamate homeostasis. This was accompanied with significantly decreased TGF-β1 at PND1 in CP kits indicating ongoing neuroinflammation. We also show for the first time that glutamate carboxypeptidase II (GCPII) was significantly increased in the activated microglia at the periventricular white matter area in both G29 and PND1 CP kits. This was confirmed by in vitro studies demonstrating that LPS activated primary microglia markedly upregulate GCPII enzymatic activity. These results suggest that maternal intrauterine endotoxin exposure results in early onset and long-lasting dysregulation of glutamate homeostasis, which may be mediated by

  7. Engineering of recombinant Escherichia coli cells co-expressing poly-γ-glutamic acid (γ-PGA) synthetase and glutamate racemase for differential yielding of γ-PGA.

    PubMed

    Cao, Mingfeng; Geng, Weitao; Zhang, Wei; Sun, Jibin; Wang, Shufang; Feng, Jun; Zheng, Ping; Jiang, Anna; Song, Cunjiang

    2013-11-01

    Poly-γ-glutamic acid (γ-PGA) is a promising environmental-friendly material with outstanding water solubility, biocompatibility and degradability. However, it is tough to determine the relationship between functional synthetic enzyme and the strains' yield or substrate dependency. We cloned γ-PGA synthetase genes pgsBCA and glutamate racemase gene racE from both L-glutamate-dependent γ-PGA-producing Bacillus licheniformis NK-03 and L-glutamate-independent B. amyloliquefaciens LL3 strains. The deduced RacE and PgsA from the two strains shared the identity of 84.5% and 78.53%, while PgsB and PgsC possessed greater similarity with 93.13% and 93.96%. The induced co-expression of pgsBCA and racE showed that the engineered Escherichia coli strains had the capacity of synthesizing γ-PGA, and LL3 derived PgsBCA had higher catalytic activity and enhanced productivity than NK-03 in Luria-Bertani medium containing glucose or L-glutamate. However, the differential effect was weakened when providing sufficient immediateness L-glutamate substrate, that is, the supply of substrate could be served as the ascendance upon γ-PGA production. Furthermore, RacE integration could enhance γ-PGA yield through improving the preferred d-glutamate content. This is the first report about co-expression of pgsBCA and racE from the two Bacillus strains, which will be of great value for the determination of the biosynthetic mechanism of γ-PGA.

  8. Real-time imaging of glutamate clearance reveals normal striatal uptake in Huntington disease mouse models

    PubMed Central

    Parsons, Matthew P.; Vanni, Matthieu P.; Woodard, Cameron L.; Kang, Rujun; Murphy, Timothy H.; Raymond, Lynn A.

    2016-01-01

    It has become well accepted that Huntington disease (HD) is associated with impaired glutamate uptake, resulting in a prolonged time-course of extracellular glutamate that contributes to excitotoxicity. However, the data supporting this view come largely from work in synaptosomes, which may overrepresent nerve-terminal uptake over astrocytic uptake. Here, we quantify real-time glutamate dynamics in HD mouse models by high-speed imaging of an intensity-based glutamate-sensing fluorescent reporter (iGluSnFR) and electrophysiological recordings of synaptically activated transporter currents in astrocytes. These techniques reveal a disconnect between the results obtained in synaptosomes and those in situ. Exogenous glutamate uptake is impaired in synaptosomes, whereas real-time measures of glutamate clearance in the HD striatum are normal or even accelerated, particularly in the aggressive R6/2 model. Our results highlight the importance of quantifying glutamate dynamics under endogenous release conditions, and suggest that the widely cited uptake impairment in HD does not contribute to pathogenesis. PMID:27052848

  9. Glutamate-dependent transcriptional regulation of GLAST/EAAT1: a role for YY1.

    PubMed

    Rosas, Sandra; Vargas, Miguel A; López-Bayghen, Esther; Ortega, Arturo

    2007-05-01

    Glutamate is the major excitatory transmitter in the vertebrate brain and its extracellular levels are tightly regulated to prevent excitotoxic effects. The Na(+)-dependent glutamate/aspartate transporter GLAST/EAAT1 is regulated in the short and in the long term by glutamate. A receptors-independent change in its membrane translocation rate, accounts for an acute modulation in GLAST/EAAT1 transport. In contrast, activation of the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate subtype of glutamate receptors represses the transcription of the chick glast gene. A glutamate responsive element has been mapped to the promoter region of this gene containing a bonafide binding site for the transcription factor Ying-Yang 1. Using cultured chick cerebellar Bergmann glia cells, glutamate elicited a time and dose-dependent increase in Ying-Yang 1 DNA binding consistent with the negative response generated in a reporter gene construct controlled for Ying-Yang 1. Over-expression of this transcription factor leads to a substantial reduction in GLAST/EAAT1 transporter uptake and an important decrease in mRNA levels, all associated with the transcriptional repression of the chick glast promoter activity. These results provide evidence for an involvement of Ying-Yang 1 in the transcriptional response to glutamate in glial cells and favor the notion of a relevant role of this factor in GLAST/EAAT1 transcriptional control.

  10. Glutamate-induced octamer DNA binding and transcriptional control in cultured radial glia cells.

    PubMed

    López-Bayghen, Esther; Cruz-Solís, Irma; Corona, Matilde; López-Colomé, Ana María; Ortega, Arturo

    2006-08-01

    Glutamate, the main excitatory neurotransmitter in the vertebrate brain, is critically involved in gene expression regulation in neurons and in glia cells. Neuron-glia interactions provide the framework for synaptic plasticity. Retinal and cerebellar radial glia cells surround glutamatergic excitatory synapses and sense synaptic activity through glutamate receptors expressed in their membranes. Several glutamate-dependent membrane to nuclei signaling cascades have been described in these cells. Octamer DNA binding factors, namely Oct-1 and Oct-2 recognize similar DNA sequences on regulatory regions, but their final transcriptional effect depends on several factors. By these means, different responses can be achieved in different cell types. Here, we describe a comparison between the glutamate-induced DNA binding of octamer factors and their functional activities in two important types of radial glia, retinal Müller and cerebellar Bergmann glial cells. While Oct-1 is expressed in both cell types and in both glutamate treatments results in an increase in Oct-1 DNA binding, this complex is capable of transactivating a reporter gene only in Müller glia cells. In contrast, Oct-2 expression is restricted to Bergmann glia cells in which glutamate treatment results in an augmentation of Oct-2 DNA binding complexes and the repression of kainate binding protein gene transcription. Our present findings demonstrate a differential role for Oct-1 and Oct-2 transcription factors in glial glutamate signaling, and further strengthen the notion of an important role for glial cells in glutamatergic transactions in the central nervous system.

  11. Glutamate-dependent transcriptional regulation in bergmann glia cells: involvement of p38 MAP kinase.

    PubMed

    Zepeda, Rossana C; Barrera, Iliana; Castelán, Francisco; Soto-Cid, Abraham; Hernández-Kelly, Luisa C; López-Bayghen, Esther; Ortega, Arturo

    2008-07-01

    Glutamate (Glu) is the major excitatory neurotransmitter in the Central Nervous System (CNS). Ionotropic and metabotropic glutamate receptors (GluRs) are present in neurons and glial cells and are involved in gene expression regulation. Mitogen-activated proteins kinases (MAPK) are critical for all the membrane to nuclei signaling pathways described so far. In cerebellar Bergmann glial cells, glutamate-dependent transcriptional regulation is partially dependent on p42/44 MAPK activity. Another member of this kinase family, p38 MAPK is activated by non-mitogenic stimuli through its Thr180/Tyr182 phosphorylation and phosphorylates cytoplasmic and nuclear protein targets involved in translational and transcriptional events. Taking into consideration that the role of p38MAPK in glial cells is not well understood, we demonstrate here that glutamate increases p38 MAPK phosphorylation in a time and dose dependent manner in cultured chick cerebellar Bergmann glial cells (BGC). Moreover, p38 MAPK is involved in the glutamate-induced transcriptional activation in these cells. Ionotropic as well as metabotropic glutamate receptors participate in p38 MAPK activation. The present findings demonstrate the involvement of p38 MAPK in glutamate-dependent gene expression regulation in glial cells.

  12. Phenotypic Characterization of Mice Heterozygous for a Null Mutation of Glutamate Carboxypeptidase II

    PubMed Central

    Han, Liqun; Picker, Jonathan D.; Schaevitz, Laura R.; Tsai, Guochuan; Feng, Jiamin; Jiang, Zhichun; Chu, Hillary C.; Basu, Alo C.; Berger-Sweeney, Joanne; Coyle, Joseph T.

    2009-01-01

    Glutamate is the major excitatory neurotransmitter in the mammalian central nervous system. Disturbed glutamate signaling resulting in hypofunction of NMDA receptors has been implicated in the pathophysiology of schizophrenia. Glutamate Carboxypeptidase II (GCP II) hydrolyzes N-acetyl-alpha L-aspartyl-L-glutamate (NAAG) into glutamate and N-acetyl-aspartate (NAA). NAAG is a neuropeptide that is an NMDA receptor antagonist as well as an agonist for the metabotropic glutamate receptor-3 (mGluR3), which inhibits glutamate release. The aggregate effect of NAAG is thus to attenuate NMDA receptor activation. To manipulate the expression of GCP II, loxP sites were inserted flanking exon 1 and 2, which were excised by crossing with a Cre-expressing mouse. The mice heterozygous for this deletion showed a 50% reduction in the expression level of protein and functional activity of GCP II in brain samples. Heterozygous mutant crosses did not yield any homozygous null animals at birth or as embryos (N >200 live births and fetuses). These data are consistent with the previous report that GCP II homozygous mutant mice generated by removing exons 9 and 10 of GCP II gene were embryonically lethal and confirm our hypothesis that GCP II plays an essential role early in embryonic development. Heterozygous mice, however, developed normally to adulthood and exhibited increased locomotor activity, reduced social interaction, and a subtle cognitive deficit in working memory. PMID:19347959

  13. The Excitatory Neurotransmitter Glutamate Stimulates DNA Repair to Increase Neuronal Resiliency

    PubMed Central

    Yang, Jenq-Lin; Sykora, Peter; Wilson, David M.; Mattson, Mark P.; Bohr, Vilhelm A.

    2012-01-01

    Glutamate is the most abundant excitatory neurotransmitter in the vertebrate central nerve system and plays an important role in synaptic plasticity required for learning and memory. Activation of glutamate ionotropic receptors promptly triggers membrane depolarization and Ca2+ influx, resulting in the activation of several different protein kinases and transcription factors. For example, glutamate-mediated Ca2+ influx activates Ca2+/calmodulin-dependent kinase, protein kinase C, and mitogen activated protein kinases resulting in activation of transcription factors such as cyclic AMP response element binding protein (CREB). Abnormally prolonged exposure to glutamate causes neuronal injury, and such “excitotoxicity” has been implicated in many acute and chronic diseases including ischemic stroke, hypoglycemia, epilepsy, amyotrophic lateral sclerosis, Alzheimer’s, Huntington’s and Parkinson’s diseases. Interestingly, although glutamate-induced Ca2+ influx can cause DNA damage by a mitochondrial reactive oxygen species-mediated mechanism, the Ca2+ simultaneously activates CREB, resulting in up-regulation of the DNA repair and redox protein apurinic/apyrimidinic endonuclease 1. Here, we review connections between physiological or aberrant glutamate receptor activation, Ca2+-mediated signaling, oxidative DNA damage and repair efficiency, and neuronal vulnerability. We conclude that glutamate signaling involves an adaptive cellular stress response pathway that enhances DNA repair capability, thereby protecting neurons against injury and disease. PMID:21729715

  14. Tianeptine modulates amygdalar glutamate neurochemistry and synaptic proteins in rats subjected to repeated stress.

    PubMed

    Piroli, Gerardo G; Reznikov, Leah R; Grillo, Claudia A; Hagar, Janel M; Fadel, Jim R; Reagan, Lawrence P

    2013-03-01

    Stress is a common environmental factor associated with depressive illness and the amygdala is thought to be integral for this association. For example, repeated stress impairs amygdalar neuroplasticity in rodents and these defects parallel amygdalar deficits in depressive illness patients. Because the excitatory neurotransmitter glutamate is important in neuroplasticity, we hypothesized that alterations in amygdalar glutamatergic systems may serve as key players in depressive illness. Moreover, restoration of amygdalar glutamatergic systems may serve as important therapeutic targets in the successful management of multiple stress-related mood disorders. To address these hypotheses, we measured glutamate efflux in the basolateral and central amygdalar complexes via in vivo microdialysis, as well as the expression of synaptic proteins that regulate vesicular glutamate packaging and release, in rats subjected to repeated stress and treated daily with saline or the antidepressant tianeptine. Glutamate efflux was significantly reduced in the central amygdalar complex of animals subjected to repeated stress. In addition, repeated stress nearly eliminated amygdalar vGLUT2 expression, thereby proving a potential mechanism through which repeated stress impairs amygdalar glutamate neurochemistry. These stress-induced changes in glutamate efflux and vGLUT2 expression were inhibited by daily tianeptine administration. Moreover, tianeptine administration increased the vesicular localization of SNAP-25, which could account for the ability of tianeptine to modify glutamatergic tone in non-stressed control rats. Collectively, these results demonstrate that repeated stress differentially affects amygdalar glutamate systems and further supports our previous studies indicating that tianeptine's antidepressant efficacy may involve targeting amygdalar glutatamatergic systems.

  15. Determination of glutamate uptake by high performance liquid chromatography (HPLC) in preparations of retinal tissue.

    PubMed

    Moraes, Edinaldo Rogério da Silva; Grisolia, Alan Barroso Araújo; Oliveira, Karen Renata Matos; Picanço-Diniz, Domingos Luiz Wanderley; Crespo-López, Maria Elena; Maximino, Caio; Batista, Evander de Jesus Oliveira; Herculano, Anderson Manoel

    2012-10-15

    The present study describes a simple and efficient method utilizing high performance liquid chromatography (HPLC) coupled to fluorescence detection for the determination of kinetic parameters of glutamate uptake in nervous tissue. Retinal tissue obtained from 7-day-old chicks was incubated with known concentrations of glutamate (50-2000 μM) for 10 min, and the levels of the o-phtaldehyde (OPA)-derivatized neurotransmitter in the incubation medium were measured. By assessing the difference between initial and final concentrations of glutamate in the medium, a saturable uptake mechanism was characterized (K(m)=8.2 and V(max)=9.8 nmol/mg protein/min). This measure was largely sodium- and temperature-dependent, strongly supporting that the mechanism for concentration decrements is indeed uptake by high-affinity transporters. Added to this, our results also demonstrated that zinc chloride (an inhibitor of glutamate/aspartate transporters) evoked a concentration-dependent decrease in glutamate uptake, demonstrating the specificity of our methodology. Overall, the present work characterizes an alternative methodology to evaluate glutamate uptake in nervous tissue using HPLC. This approach could be an important tool for studies associated to the characterization of minute alterations in glutamate transport related with central nervous system injury.

  16. Glutamate Transporters and the Excitotoxic Path to Motor Neuron Degeneration in Amyotrophic Lateral Sclerosis

    PubMed Central

    Foran, Emily

    2009-01-01

    Abstract Responsible for the majority of excitatory activity in the central nervous system (CNS), glutamate interacts with a range of specific receptor and transporter systems to establish a functional synapse. Excessive stimulation of glutamate receptors causes excitotoxicity, a phenomenon implicated in both acute and chronic neurodegenerative diseases [e.g., ischemia, Huntington's disease, and amyotrophic lateral sclerosis (ALS)]. In physiology, excitotoxicity is prevented by rapid binding and clearance of synaptic released glutamate by high-affinity, Na+-dependent glutamate transporters and amplified by defects to the glutamate transporter and receptor systems. ALS pathogenetic mechanisms are not completely understood and characterized, but excitotoxicity has been regarded as one firm mechanism implicated in the disease because of data obtained from ALS patients and animal and cellular models as well as inferred by the documented efficacy of riluzole, a generic antiglutamatergic drug, has in patients. In this article, we critically review the several lines of evidence supporting a role for glutamate-mediated excitotoxicity in the death of motor neurons occurring in ALS, putting a particular emphasis on the impairment of the glutamate-transport system. Antioxid. Redox Signal. 11, 1587–1602. PMID:19413484

  17. Flux of Nitrogen-13 from L-(N-13)Glutamate in isolated myocardium

    SciTech Connect

    Keen, R.E.; Barrio, J.R.; Krivokapich, J.; Phelps, M.E.

    1985-05-01

    Specific activity of nitrogen-13 containing metabolites in tissue and effluent was determined following an intra-arterial bolus of non-carrier added L-(N-13)glutamate (N-13 GLU) given to isolated rabbit septa under different metabolic states which include pyruvate (2 mM), transaminase inhibition (aminooxy-acetate, AOA, 2 mM), or pyruvate with AOA superimposed on the insulin and glucose perfused septa. Six minutes after the N-13 GLU bolus administration relative tissue specific activities of glutamine, alanine, aspartate, and glutamate were approximately 3:38:52:100, respectively, in the control and pyruvate perfused septa. The lower alanine specific activity when compared with control tissue indicated that alanine output was from a pool separate from GPT alanine pools. Higher glutamate specific activity suggested that its output is from a pool(s) different than the larger intra-cellular glutamate pool(s). All interventions with AOA blocked N-13 flux through transminases altering tissue and effluent relative specific activities with increase in % N-13 and specific activities for glutamine, glutamate, ammonia, and protein concomittant with disappearance of labeled aspartate and alanine. These results indicate that N-13 distribution in myocardium after N-13 GLU administration is mainly controlled by glutamate interaction with reversible transaminases. The differences in reactant (N-13 GLU) and product specific activities are a consequence of channeling between different cytosolic and mitochondrial glutamate microcompartments.

  18. Excitatory amino acid glutamate: role in peripheral nociceptive transduction and inflammation in experimental and clinical osteoarthritis.

    PubMed

    Wen, Z-H; Chang, Y-C; Jean, Y-H

    2015-11-01

    Although a large proportion of patients with osteoarthritis (OA) show inflammation in their affected joints, the pathological role of inflammation in the development and progression of OA has yet to be clarified. Glutamate is considered an excitatory amino acid (EAA) neurotransmitter in the mammalian central nervous system (CNS). There are cellular membrane glutamate receptors and transporters for signal input modulation and termination as well as vesicular glutamate transporters (VGLUTs) for signal output through exocytotic release. Glutamate been shown to mediate intercellular communications in bone cells in a manner similar to synaptic transmission within the CNS. Glutamate-mediated events may also contribute to the pathogenesis and ongoing processes of peripheral nociceptive transduction and inflammation of experimental arthritis models as well as human arthritic conditions. This review will discuss the differential roles of glutamate signaling and blockade in peripheral neuronal and non-neuronal joint tissues, including bone remodeling systems and their potentials to impact OA-related inflammation and progression. This will serve to identify several potential targets to direct novel therapies for OA. Future studies will further elucidate the role of glutamate in the development and progression of OA, as well as its association with the clinical features of the disease.

  19. Real-time imaging of glutamate clearance reveals normal striatal uptake in Huntington disease mouse models.

    PubMed

    Parsons, Matthew P; Vanni, Matthieu P; Woodard, Cameron L; Kang, Rujun; Murphy, Timothy H; Raymond, Lynn A

    2016-01-01

    It has become well accepted that Huntington disease (HD) is associated with impaired glutamate uptake, resulting in a prolonged time-course of extracellular glutamate that contributes to excitotoxicity. However, the data supporting this view come largely from work in synaptosomes, which may overrepresent nerve-terminal uptake over astrocytic uptake. Here, we quantify real-time glutamate dynamics in HD mouse models by high-speed imaging of an intensity-based glutamate-sensing fluorescent reporter (iGluSnFR) and electrophysiological recordings of synaptically activated transporter currents in astrocytes. These techniques reveal a disconnect between the results obtained in synaptosomes and those in situ. Exogenous glutamate uptake is impaired in synaptosomes, whereas real-time measures of glutamate clearance in the HD striatum are normal or even accelerated, particularly in the aggressive R6/2 model. Our results highlight the importance of quantifying glutamate dynamics under endogenous release conditions, and suggest that the widely cited uptake impairment in HD does not contribute to pathogenesis. PMID:27052848

  20. A role for preoptic glutamate in the regulation of male reproductive behavior.

    PubMed

    Dominguez, Juan M

    2009-02-01

    Although much progress has been made toward understanding the role of the medial preoptic area (MPOA) in the regulation of male reproductive behaviors, the precise mechanisms responsible for its activation during mating are largely unclear. Several studies implicate glutamate in this response. However, not until recently was there direct evidence supporting this hypothesis. Results obtained using in vivo microdialysis showed that levels of glutamate increased in the MPOA during mating, particularly with ejaculation. Levels then decreased rapidly following ejaculation, during a period of sexual quiescence. The magnitude of this decrease correlated with time spent in quiescence. Additionally, central administration of glutamate uptake inhibitors increased levels of glutamate and facilitated behavior. Glutamate activation of N-methyl-D-aspartate (NMDA) receptors in the MPOA is at least partly responsible for behavioral effects evoked by increase glutamate. This is evidenced by histological analysis of the MPOA, which shows that nearly all cells containing mating-induced Fos also contained NMDA receptors. Mating also increased phosphorylation of NMDA receptors, indicating receptor activation. Finally, bilateral microinjections of NMDA receptor antagonists inhibited copulation. This neurochemical, anatomical, and behavioral evidence points to a key role of preoptic glutamate in the regulation of sexual behavior in males. The implications of these findings are discussed.

  1. Diffuse Brain Injury Elevates Tonic Glutamate Levels and Potassium-Evoked Glutamate Release in Discrete Brain Regions at Two Days Post-Injury: An Enzyme-Based Microelectrode Array Study

    PubMed Central

    Hinzman, Jason M.; Currier Thomas, Theresa; Burmeister, Jason J.; Quintero, Jorge E.; Huettl, Peter; Pomerleau, Francois; Gerhardt, Greg A.

    2010-01-01

    Abstract Traumatic brain injury (TBI) survivors often suffer from a wide range of post-traumatic deficits, including impairments in behavioral, cognitive, and motor function. Regulation of glutamate signaling is vital for proper neuronal excitation in the central nervous system. Without proper regulation, increases in extracellular glutamate can contribute to the pathophysiology and neurological dysfunction seen in TBI. In the present studies, enzyme-based microelectrode arrays (MEAs) that selectively measure extracellular glutamate at 2 Hz enabled the examination of tonic glutamate levels and potassium chloride (KCl)-evoked glutamate release in the prefrontal cortex, dentate gyrus, and striatum of adult male rats 2 days after mild or moderate midline fluid percussion brain injury. Moderate brain injury significantly increased tonic extracellular glutamate levels by 256% in the dentate gyrus and 178% in the dorsal striatum. In the dorsal striatum, mild brain injury significantly increased tonic glutamate levels by 200%. Tonic glutamate levels were significantly correlated with injury severity in the dentate gyrus and striatum. The amplitudes of KCl-evoked glutamate release were increased significantly only in the striatum after moderate injury, with a 249% increase seen in the dorsal striatum. Thus, with the MEAs, we measured discrete regional changes in both tonic and KCl-evoked glutamate signaling, which were dependent on injury severity. Future studies may reveal the specific mechanisms responsible for glutamate dysregulation in the post-traumatic period, and may provide novel therapeutic means to improve outcomes after TBI. PMID:20233041

  2. Opposing roles for caspase and calpain death proteases in L-glutamate-induced oxidative neurotoxicity

    SciTech Connect

    Elphick, Lucy M.; Hawat, Mohammad; Toms, Nick J.; Meinander, Annika; Mikhailov, Andrey; Eriksson, John E.; Kass, George E.N.

    2008-10-15

    Oxidative glutamate toxicity in HT22 murine hippocampal cells is a model for neuronal death by oxidative stress. We have investigated the role of proteases in HT22 cell oxidative glutamate toxicity. L-glutamate-induced toxicity was characterized by cell and nuclear shrinkage and chromatin condensation, yet occurred in the absence of either DNA fragmentation or mitochondrial cytochrome c release. Pretreatment with the selective caspase inhibitors either benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (pan-caspase), N-acetyl-Leu-Glu-His-Asp-aldehyde (caspase 9) or N-acetyl-Ile-Glu-Thr-Asp-aldehyde (caspase 8), significantly increased L-glutamate-induced cell death with a corresponding increase in observed nuclear shrinkage and chromatin condensation. This enhancement of glutamate toxicity correlated with an increase in L-glutamate-dependent production of reactive oxygen species (ROS) as a result of caspase inhibition. Pretreating the cells with N-acetyl-L-cysteine prevented ROS production, cell shrinkage and cell death from L-glutamate as well as that associated with the presence of the pan-caspase inhibitor. In contrast, the caspase-3/-7 inhibitor N-acetyl-Asp-Glu-Val-Asp aldehyde was without significant effect. However, pretreating the cells with the calpain inhibitor N-acetyl-Leu-Leu-Nle-CHO, but not the cathepsin B inhibitor CA-074, prevented cell death. The cytotoxic role of calpains was confirmed further by: 1) cytotoxic dependency on intracellular Ca{sup 2+} increase, 2) increased cleavage of the calpain substrate Suc-Leu-Leu-Val-Tyr-AMC and 3) immunoblot detection of the calpain-selective 145 kDa {alpha}-fodrin cleavage fragment. We conclude that oxidative L-glutamate toxicity in HT22 cells is mediated via calpain activation, whereas inhibition of caspases-8 and -9 may exacerbate L-glutamate-induced oxidative neuronal damage through increased oxidative stress.

  3. Novel metabotropic glutamate receptor negatively coupled to adenylyl cyclase in cultured rat cerebellar astrocytes.

    PubMed

    Kanumilli, Srinivasan; Toms, Nick J; Roberts, Peter J

    2004-04-01

    Several excitatory amino acid ligands were found potently to inhibit forskolin-stimulated cAMP accumulation in rat cultured cerebellar astrocytes: L-cysteine sulfinic acid (L-CSA) = L-aspartate > L-glutamate >/= the glutamate uptake inhibitor, L-PDC. This property did not reflect activation of conventional glutamate receptors, since the selective ionotropic glutamate receptor agonists NMDA, AMPA, and kainate, as well as several mGlu receptor agonists [(1S,3R)-ACPD, (S)-DHPG, DCG-IV, L-AP4, L-quisqualate, and L-CCG-I], were without activity. In addition, the mGlu receptor antagonists, L-AP3, (S)-4CPG, Eglu, LY341495, (RS)-CPPG, and (S)-MCPG failed to reverse 30 microM glutamate-mediated inhibitory responses. L-PDC-mediated inhibition was abolished by the addition of the enzyme glutamate-pyruvate transaminase. This finding suggests that the effect of L-PDC is indirect and that it is mediated through endogenously released L-glutamate. Interestingly, L-glutamate-mediated inhibitory responses were resistant to pertussis toxin, suggesting that G(i)/G(o) type G proteins were not involved. However, inhibition of protein kinase C (PKC, either via the selective PKC inhibitor GF109203X or chronic PMA treatment) augmented glutamate-mediated inhibitory responses. Although mGlu3 receptors (which are negatively coupled to adenylyl cyclase) are expressed in astrocyte populations, in our study Western blot analysis indicated that this receptor type was not expressed in cerebellar astrocytes. We therefore suggest that cerebellar astrocytes express a novel mGlu receptor, which is negatively coupled to adenylyl cyclase, and possesses an atypical pharmacological profile. PMID:14999808

  4. Gq-DREADD Selectively Initiates Glial Glutamate Release and Inhibits Cue-induced Cocaine Seeking

    PubMed Central

    Scofield Michael, D.; Boger Heather, A.; Smith Rachel, J.; Li, Hao; Haydon Philip, G.; Kalivas Peter, W.

    2015-01-01

    Background Glial cells of the central nervous system directly influence neuronal activity by releasing neuroactive small molecules, including glutamate. Long-lasting cocaine-induced reductions in extracellular glutamate in the nucleus accumbens core (NAcore) affect synaptic plasticity responsible for relapse vulnerability. Methods We transduced NAcore astrocytes with an AAV viral vector expressing hM3Dq (Gq) DREADD under control of the glial fibrillary acidic protein (GFAP) promoter in 62 male Sprague Dawley rats, 4 dnSNARE mice and 4 wild type littermates. Using glutamate biosensors we measured NAcore glutamate levels following intracranial or systemic administration of clozapine-N-oxide (CNO), and tested the ability of systemic CNO to inhibit reinstated cocaine or sucrose seeking following self-administration (SA) and extinction training. Results Administration of CNO in GFAP-Gq-DREADD transfected animals increased NAcore extracellular glutamate levels in vivo. The glial origin of released glutamate was validated by an absence of CNO-mediated release in mice expressing a dominant-negative SNARE variant in glia. Also, CNO-mediated release was relatively insensitive to N-type calcium channel blockade. Systemic administration of CNO inhibited cue-induced reinstatement of cocaine seeking in rats extinguished from cocaine, but not sucrose SA. The capacity to inhibit reinstated cocaine-seeking was prevented by systemic administration of the group II metabotropic glutamate receptor (mGluR2/3) antagonist LY341495. Conclusions DREADD-mediated glutamate gliotransmission inhibited cue-induced reinstatement of cocaine seeking by stimulating release-regulating mGluR2/3 autoreceptors to inhibit cue-induced synaptic glutamate spillover. PMID:25861696

  5. Effects of Cymbopogon citratus and Ferula assa-foetida extracts on glutamate-induced neurotoxicity.

    PubMed

    Tayeboon, Ghazaleh S; Tavakoli, Fatemeh; Hassani, Shokoufeh; Khanavi, Mahnaz; Sabzevari, Omid; Ostad, S Nasser

    2013-10-01

    Many of CNS diseases can lead to a great quantity of release of glutamate and the extreme glutamate induces neuronal cell damage and death. Here, we wanted to investigate the effects of Cymbopogon citratus essential oil and Ferula assa-foetida extracts treatment on glutamate-induced cell damage in a primary culture of rat cerebellar granule neurons. Cerebellums were collected from 7-d rat brains and cerebellar granule neurons were obtained after 8-d culture. CGN cells were treated with C. citratus essential oil and F. assa-foetida extracts at concentration of 100 μg/ml before, after, and during exposure to 30 μM glutamate. The cellular viability was evaluated by 3-(4, 5-dimethytthiazol-2-yl)-2, 5-diphenyltetrazoliumbromide (MTT) staining. The flow cytometry assay was used to examine cell cycle and apoptosis. MTT assay showed a glutamate-induced reduction in cellular viability while treatment with C. citratus essential oil and F. assa-foetida extracts before, during, and after exposure to glutamate was increased. Flow cytometric analysis indicated that F. assa-foetida extracts treatment significantly (p < 0.001) attenuated glutamate-induced apoptotic/necrotic cell death and the necrotic rate was decreased by C. citratus essential oil treatment compared to glutamate group, significantly (p < 0.001). The results show that C. citratus essential oil and F. assa-foetida extracts display neuroprotective effects in glutamate-induced neurotoxicity. These extracts exert antiapoptotic activity in cerebellar granule neurons due to cell cycle arrest in G0G1 phase, which explain the beneficial effects of C. citratus essential oil and F. assa-foetida extracts as therapies for neurologic disorders.

  6. Structural and biochemical characterization of the folyl-poly-γ-l-glutamate hydrolyzing activity of human glutamate carboxypeptidase II.

    PubMed

    Navrátil, Michal; Ptáček, Jakub; Šácha, Pavel; Starková, Jana; Lubkowski, Jacek; Bařinka, Cyril; Konvalinka, Jan

    2014-07-01

    In addition to its well-characterized role in the central nervous system, human glutamate carboxypeptidase II (GCPII; Uniprot ID Q04609) acts as a folate hydrolase in the small intestine, participating in the absorption of dietary polyglutamylated folates (folyl-n-γ-l-glutamic acid), which are the provitamin form of folic acid (also known as vitamin B9 ). Despite the role of GCPII as a folate hydrolase, nothing is known about the processing of polyglutamylated folates by GCPII at the structural or enzymological level. Moreover, many epidemiologic studies on the relationship of the naturally occurring His475Tyr polymorphism to folic acid status suggest that this polymorphism may be associated with several pathologies linked to impaired folate metabolism. In the present study, we report: (a) a series X-ray structures of complexes between a catalytically inactive GCPII mutant (Glu424Ala) and a panel of naturally occurring polyglutamylated folates; (b) the X-ray structure of the His475Tyr variant at a resolution of 1.83 Å; (c) the study of the recently identified arene-binding site of GCPII through mutagenesis (Arg463Leu, Arg511Leu and Trp541Ala), inhibitor binding and enzyme kinetics with polyglutamylated folates as substrates; and (d) a comparison of the thermal stabilities and folate-hydrolyzing activities of GCPII wild-type and His475Tyr variants. As a result, the crystallographic data reveal considerable details about the binding mode of polyglutamylated folates to GCPII, especially the engagement of the arene binding site in recognizing the folic acid moiety. Additionally, the combined structural and kinetic data suggest that GCPII wild-type and His475Tyr variant are functionally identical. PMID:24863754

  7. Glutamate oxidase biosensor based on mixed ceria and titania nanoparticles for the detection of glutamate in hypoxic environments

    PubMed Central

    Ganesana, Mallikarjunarao; Leiter, J.C.; Andreescu, Silvana

    2013-01-01

    We report on the design and development of a glutamate oxidase (GmOx) microelectrode for measuring L-glutamic acid (GluA) in oxygen-depleted conditions, which is based on the oxygen storage and release capacity of cerium oxides. To fabricate the biosensor, a nanocomposite of oxygen-rich ceria and titania nanoparticles dispersed within a semi-permeable chitosan membrane was co-immobilized with the enzyme GmOx on the surface of a Pt microelectrode. The oxygen delivery capacity of the ceria nanoparticles embedded in a biocompatible chitosan matrix facilitated enzyme stabilization and operation in oxygen free conditions. GluA was measured by amperometry at a working potential of 0.6 V vs Ag/AgCl. Detection limits of 0.594 μM and 0.493 μM and a sensitivity of 793 pA/μM (RSD 3.49%, n=5) and 395 pA/μM (RSD 2.48%, n=5) were recorded in oxygenated and deoxygenated conditions, with response times of 2s and 5s, respectively. The biosensor had good operational stability and selectivity against common interfering substances. Operation of the biosensor was tested in cerebrospinal fluid. Preliminary in vivo recording in Sprague-Dawley rats to monitor GluA in the cortex during cerebral ischemia and reperfusion demonstrate a potential application of the biosensor in hypoxic conditions. This method provides a solution to ensure functionality of oxidoreductase enzymes in oxygen-free environments. PMID:24090755

  8. Glutamate oxidase biosensor based on mixed ceria and titania nanoparticles for the detection of glutamate in hypoxic environments.

    PubMed

    Özel, Rıfat Emrah; Ispas, Cristina; Ganesana, Mallikarjunarao; Leiter, J C; Andreescu, Silvana

    2014-02-15

    We report on the design and development of a glutamate oxidase (GmOx) microelectrode for measuring l-glutamic acid (GluA) in oxygen-depleted conditions, which is based on the oxygen storage and release capacity of cerium oxides. To fabricate the biosensor, a nanocomposite of oxygen-rich ceria and titania nanoparticles dispersed within a semi-permeable chitosan membrane was co-immobilized with the enzyme GmOx on the surface of a Pt microelectrode. The oxygen delivery capacity of the ceria nanoparticles embedded in a biocompatible chitosan matrix facilitated enzyme stabilization and operation in oxygen free conditions. GluA was measured by amperometry at a working potential of 0.6 V vs Ag/AgCl. Detection limits of 0.594 µM and 0.493 µM and a sensitivity of 793 pA/µM (RSD 3.49%, n=5) and 395 pA/µM (RSD 2.48%, n=5) were recorded in oxygenated and deoxygenated conditions, with response times of 2s and 5s, respectively. The biosensor had good operational stability and selectivity against common interfering substances. Operation of the biosensor was tested in cerebrospinal fluid. Preliminary in vivo recording in Sprague-Dawley rats to monitor GluA in the cortex during cerebral ischemia and reperfusion demonstrate a potential application of the biosensor in hypoxic conditions. This method provides a solution to ensure functionality of oxidoreductase enzymes in oxygen-free environments. PMID:24090755

  9. LY354740 is a potent and highly selective group II metabotropic glutamate receptor agonist in cells expressing human glutamate receptors.

    PubMed

    Schoepp, D D; Johnson, B G; Wright, R A; Salhoff, C R; Mayne, N G; Wu, S; Cockerman, S L; Burnett, J P; Belegaje, R; Bleakman, D; Monn, J A

    1997-01-01

    The novel compound LY354740 is a conformationally constrained analog of glutamate, which was designed for interaction at metabotropic glutamate (mGlu) receptors. In this paper the selectivity of LY354740 for recombinant human mGlu receptor subtypes expressed in non-neuronal (RGT) cells is described. At human mGlu2 receptors, LY354740 produced > 90% suppression of forskolin-stimulated cAMP formation with an EC50 of 5.1 +/- 0.3 nM. LY354740 was six-fold less potent in activating human mGlu3 receptors (EC50 = 24.3 +/- 0.5 nM). LY354740 inhibition of forskolin-stimulated cAMP formation in human mGlu2 receptor-expressing cells was blocked by competitive mGlu receptor antagonists, including (+)-alpha-methyl-4-carboxyphenylglycine (MCPG) and LY307452 ((2S,4S)-2-amino-4-(4,4-diphenylbut-1-yl)-pentane-1,5-dioic acid). LY354740 had no agonist or antagonist activities at cells expressing human mGlu4 or mGlu7 (group III mGlu receptors) (EC50 > 100,000 nM). When tested at group I phosphoinositide-coupled human mGlu receptors (mGlu1a and mGlu5a), LY354740 did not activate or inhibit mGlu receptor agonist-evoked phosphoinositide hydrolysis at up to 100,000 nM. Electrophysiological experiments also demonstrated that LY354740 also had no appreciable activity in cells expressing human recombinant AMPA (GluR4) and kainate (GluR6) receptors. Thus, LY354740 is a highly potent, efficacious and selective group II (mGlu2/3) receptor agonist, useful to explore the functions of these receptors in situ. PMID:9144636

  10. Identification of essential residues involved in the glutamate binding pocket of the group II metabotropic glutamate receptor.

    PubMed

    Malherbe, P; Knoflach, F; Broger, C; Ohresser, S; Kratzeisen, C; Adam, G; Stadler, H; Kemp, J A; Mutel, V

    2001-11-01

    Metabotropic glutamate (mGlu) receptors are a family of G-protein-coupled receptors that play central roles as modulators of both glutamatergic and other major neurotransmitter systems in CNS. Using molecular modeling, site-directed mutagenesis, [(3)H]LY354740 binding, [(35)S]GTPgammaS binding, and activation of GIRK current, we have been able to identify residues crucial for the binding of LY354740 and glutamate to rat mGlu2 receptors. Several of the crucial residues located in the binding site (Arg-57, Tyr-144, Tyr-216, Asp-295) have not been identified previously. We propose that the gamma-carboxyl group of LY354740 forms H-bonds to Arg-57, whereas the alpha-carboxyl group forms an H-bond with the hydroxyl group of Ser-145. The alpha-amino group of LY354740 forms H-bonds to Asp-295 and to the side-chain hydroxyl group of Thr-168. In addition, Tyr-144 may establish a hydrophobic (C-H/pi)-interaction with the bicyclo-hexane ring of LY354740. Furthermore, the mutation of residues Ser-148 and Arg-183, which are too remote for a direct interaction, affected the ligand affinity dramatically. These results suggest that Ser-148 and Arg-183 may be important for the 3D structure and/or are involved in closure of the domain. Finally, Asp-146, which is also remote from the binding site, was shown to be involved in the differential binding affinity of [(3)H]LY354740 for mGlu2 versus mGlu3 receptors. All the mGlu receptors except mGlu2 are activated by Ca(2+) and have serine instead of aspartic acid at this position, which suggests a critical role of this aspartic acid residue in the binding properties of this unique receptor. PMID:11641422

  11. Connexin subtype expression during oral carcinogenesis: A pilot study in patients with oral squamous cell carcinoma

    PubMed Central

    BROCKMEYER, PHILLIPP; HEMMERLEIN, BERNHARD; JUNG, KLAUS; FIALKA, FLORIAN; BRODMANN, TOBIAS; GRUBER, RUDOLF MATTHIAS; SCHLIEPHAKE, HENNING; KRAMER, FRANZ-JOSEF

    2016-01-01

    Gap junctional intercellular communication (GJIC) and connexin (Cx) expression were reported in association with carcinogenesis in various types of tumours. In an earlier histomorphometric study, the protein levels of Cx subtypes 26, 43 and 45 were differentially expressed in oral squamous cell carcinoma (OSCC), corresponding lymph node metastases and dysplasia-free oral mucosa. Moreover, membrane Cx43 acted as an independent prognostic marker in OSCC tissues. This study aimed to confirm the expression of described Cx subtypes at the mRNA level. Hence, a reverse transcription quantitative polymerase chain reaction (RT-qPCR) analysis of Cx26, Cx43 and Cx45 gene expressions was performed in paired carcinoma and mucosa samples of 15 OSCC patients. Additionally, we assessed the interaction between Cx subtype expression and clinicopathological routine parameters. The RT-qPCR analysis revealed that Cx26 was downregulated in OSCC (P=0.01), while Cx43 was marginally upregulated in cancer tissue (P=0.04). Cx45 was significantly overexpressed in OSCC tissue compared with the intraoral mucosa controls (P<0.01), and remained unchanged at different tumour stages. No significant interactions between differential Cx subtype expression and clinicopathological routine parameters were observed. In conclusion, Cx regulation at the transcriptional level appears to be an early event during the initiation and development of OSCC, and is maintained during further progression. However, the mRNA-protein correlation is variable. This may be indicative of post-transcriptional, translational and degradation regulations being associated with the determination of Cx protein concentration during oral carcinogenesis. PMID:26893879

  12. The effect of chronic ethanol on glutamate binding in human and rat brain

    SciTech Connect

    Cummins, J.T.; Sack, M.; von Hungen, K. Univ. of California School of Medicine, Los Angeles )

    1990-01-01

    Quantitative autoradiographic techniques demonstrate that chronic alcohol administration causes a decrease in ({sup 3}H)-glutamate binding to hippocampal N-methyl-D-aspartate (NMDA) receptors. A 14% decrease in ({sup 3}H)-glutamate binding in the hippocampal CA{sub 1} region is seen both in the rat after five days of ethanol administration and in postmortem hippocampal tissues from alcoholics. In the rat, 24 hr ethanol withdrawal values are intermediate between control and alcohol binding levels. There was no significant effect of ethanol on ({sup 3}H)-glutamate binding in the cortex or caudate.

  13. Search for soliton modes in helical poly-γ-benzyl-l-glutamate

    NASA Astrophysics Data System (ADS)

    Renthal, Robert; Taboada, J.

    1989-07-01

    Solid α-helical poly(γ-benzyl-L-glutamate) was examined at low temperature for evidence of the unusual temperature-dependent vibrational mode found by Careri and co-workers in solid acetanilide and attributed to a soliton wave trapped in protein-like hydrogen bonds. We have confirmed the anomaly in acetanilide, however, a similar temperature-dependent mode was not observed in poly(γ-benzyl-L-glutamate). These results indicate that anharmonic amide modes may only be present in certain α-helical structures. Two new low frequency modes (180 and 90 cm -1) are observed for poly(γ-benzyl-L-glutamate).

  14. Avian Imc-tectal projection is mediated by acetylcholine and glutamate.

    PubMed

    Wang, S R; Wu, G Y; Felix, D

    1995-03-27

    In the bird, biochemical and histochemical data suggest that the neurotransmitter between nucleus isthmi pars magnocellularis (Imc) and tectum is either acetylcholine or glutamate. There are, however, discrepancies regarding the functional role of acetylcholine. In the present study we investigated the action of acetylcholine and glutamate and their specific antagonists on excitatory isthmo-tectal synaptic transmission using electrophysiological and microiontophoretic techniques. The results show two different population of cells: (1) excitatory cholinergic input, blocked by atropine sulphate but not by glutamate antagonist; (2) excitatory glutamatergic input of NMDA or non-NMDA receptor type, which is blocked or reduced by CPP or CNQX but not by atropine sulphate.

  15. High protein diet induces pericentral glutamate dehydrogenase and ornithine aminotransferase to provide sufficient glutamate for pericentral detoxification of ammonia in rat liver lobules.

    PubMed

    Boon, L; Geerts, W J; Jonker, A; Lamers, W H; Van Noorden, C J

    1999-06-01

    The liver plays a central role in nitrogen metabolism. Nitrogen enters the liver as free ammonia and as amino acids of which glutamine and alanine are the most important precursors. Detoxification of ammonia to urea involves deamination and transamination. By applying quantitative in situ hybridization, we found that mRNA levels of the enzymes involved are mainly expressed in periportal zones of liver lobules. Free ammonia, that is not converted periportally, is efficiently detoxified in the small rim of hepatocytes around the central veins by glutamine synthetase preventing it from entering the systemic circulation. Detoxification of ammonia by glutamine synthetase may be limited due to a shortage of glutamate when the nitrogen load is high. Adaptations in metabolism that prevent release of toxic ammonia from the liver were studied in rats that were fed diets with different amounts of protein, thereby varying the nitrogen load of the liver. We observed that mRNA levels of periportal deaminating and transaminating enzymes increased with the protein content in the diet. Similarly, mRNA levels of pericentral glutamate dehydrogenase and ornithine aminotransferase, the main producers of glutamate in this zone, and pericentral glutamine synthetase all increased with increasing protein levels in the diet. On the basis of these changes in mRNA levels, we conclude that: (a) glutamate is produced pericentrally in sufficient amounts to allow ammonia detoxification by glutamine synthetase and (b) in addition to the catalytic role of ornithine in the periportally localized ornithine cycle, pericentral ornithine degradation provides glutamate for ammonia detoxification.

  16. Distribution of Vesicular Glutamate Transporter 2 and Ionotropic Glutamate Receptors in the Auditory Ganglion and Cochlear Nuclei of Pigeons (Columba livia).

    PubMed

    Karim, M R; Atoji, Y

    2016-02-01

    Glutamate is a principal excitatory neurotransmitter in the auditory system. Our previous studies revealed localization of glutamate receptor mRNAs in the pigeon cochlear nuclei, suggesting the existence of glutamatergic input from the auditory nerve to the brainstem. This study demonstrated localization of mRNAs for vesicular glutamate transporter 2 (vGluT2) and ionotropic glutamate receptors (AMPA, kainate and NMDA) in the auditory ganglion (AG) and cochlear nuclei (magnocellular, angular and laminar nuclei). VGluT2 mRNA was intensely expressed in AG and intensely or moderately in the cochlear nuclei. The AG and cochlear nuclei showed intense-to-moderate mRNA signals for GluA2, GluA3, GluA4, GluK4 and GluN1. These results suggest that the pigeon AG neurons receives glutamatergic input from hair cells and in turn projects to the magnocellular and angular nuclei. Glutamate may play a pivotal role in the excitatory synapse transmission in the peripheral auditory pathway of birds.

  17. Prolonged time course of glutamate-operated single channel currents in neuromuscular preparations of small crayfish and a membrane current triggered by glutamate channel gating.

    PubMed

    Finger, W; Martin, C; Pareto, A

    1988-08-31

    Single channel currents activated by glutamate were recorded by means of the patch-clamp technique in the abdominal superficial extensor muscle and the claw opener muscle of small (1-3 months old) and large (greater than 16 months old) crayfish. It was found that in small crayfish the time course of glutamate-operated single channel currents was prolonged by a factor of about 4 in these two preparations. In the abdominal superficial extensor muscle, single channel currents activated by 5 mmol/l glutamate had a mean burst length of tau = 2-3 ms in large crayfish and a mean burst length of tau = 8-9 ms in small crayfish. In the claw opener, for large crayfish tau congruent to 0.5 ms and for small crayfish tau = 1.5-2.5 ms resulted (500 mumol/l glutamate). Moreover, single channel currents with long time courses often slowly increased their amplitudes during the open time of the channel and current amplitudes did not decline completely to the baseline after channel closing. In addition, single channel currents with relatively constant amplitude were often followed by a small increasing and decreasing membrane current. The latter results suggest that glutamate channel gating might trigger a membrane current.

  18. Effects of glutamate decarboxylase and gamma-aminobutyric acid (GABA) transporter on the bioconversion of GABA in engineered Escherichia coli.

    PubMed

    Le Vo, Tam Dinh; Kim, Tae Wan; Hong, Soon Ho

    2012-05-01

    Gamma-aminobutyric acid (GABA) is a non-essential amino acid and a precursor of pyrrolidone, a monomer of nylon 4. GABA can be biosynthesized through the decarboxylation of L: -glutamate by glutamate decarboxylase. In this study, the effects of glutamate decarboxylase (gadA, gadB), glutamate/GABA antiporter (gadC) and GABA aminotransferase (gabT) on GABA production were investigated in Escherichia coli. Glutamate decarboxylase was overexpressed alone or with the glutamate/GABA antiporter to enhance GABA synthesis. GABA aminotransferase, which redirects GABA into the TCA cycle, was knock-out mutated. When gadB and gadC were co-overexpressed in the gabT mutant strain, a final GABA concentration of 5.46 g/l was obtained from 10 g/l of monosodium glutamate (MSG), which corresponded to a GABA yield of 89.5%.

  19. Changes in NAD(P)H fluorescence and membrane current produced by glutamate uptake into salamander Müller cells.

    PubMed Central

    Barbour, B; Magnus, C; Szatkowski, M; Gray, P T; Attwell, D

    1993-01-01

    1. Glutamate uptake into isolated, whole-cell patch-clamped glial cells was studied by monitoring the increase of cell fluorescence generated as glutamate and NAD(P) were converted into alpha-ketoglutarate and NAD(P)H by glutamate dehydrogenase. The current generated by the glutamate uptake carrier was recorded simultaneously. 2. L-Glutamate evoked an increase of cell fluorescence and an inward uptake current. L- and D-aspartate generated an uptake current but no fluorescence response, consistent with the amino acid specificity of glutamate dehydrogenase. 3. In the absence of external sodium the glutamate-evoked fluorescence response and uptake current were abolished, showing that there is no sodium-independent glutamate uptake across the cell membrane. 4. Varying the glutamate concentration altered both the fluorescence response and the uptake current. The fluorescence response saturated at a lower glutamate concentration than the uptake current, and depended in a Michaelis-Menten fashion on the uptake current. 5. The fluorescence response and the uptake current were reduced by membrane depolarization, and also by removal of intracellular potassium. 6. The dependence of the fluorescence response on uptake current when membrane potential was altered or intracellular potassium was removed was the same as that seen when the external glutamate concentration was altered. 7. These fluorescence studies show that glutamate uptake is inhibited by depolarization and by removal of intracellular potassium, consistent with the conclusion of earlier work in which uptake was monitored solely as a membrane current. The data are consistent with high-affinity electrogenic sodium- and potassium-dependent glutamate uptake with fixed stoichiometry being the only significant influx route for glutamate. Other possible interpretations of the data are also discussed. PMID:8105078

  20. Hydrochloric acid alters the effect of L-glutamic acid on cell viability in human neuroblastoma cell cultures.

    PubMed

    Croce, Nicoletta; Bernardini, Sergio; Di Cecca, Stefano; Caltagirone, Carlo; Angelucci, Francesco

    2013-07-15

    l-Glutamic acid (l-glutamate) is used to induce excitotoxicity and test neuroprotective compounds in cell cultures. However, because l-glutamate powder is nearly insoluble in water, many manufacturers recommend reconstituting l-glutamate in hydrochloric acid (HCl) prior to successive dilutions. Nevertheless, HCl, even at low concentrations, may alter the pH of the cell culture medium and interfere with cell activity. Thus, the aim of this study was to evaluate whether the reconstitution of l-glutamate powder in HCl alters its capacity to induce neurotoxicity in different human neuroblastoma cell lines. SH-SY5Y, IMR-32 and SK-N-BE(2) cells were exposed to various concentrations of l-glutamate, which was either reconstituted in HCl (1M) or post re-equilibrated to the pH of the culture medium (7.5). After 24 and 48h of incubation, changes in the cell viability of treated versus untreated cells were evaluated. The effect of an identical amount of HCl present in the l-glutamate dilutions on neuroblastoma cell survival was also investigated. Our data showed that the neurotoxicity of glutamate reconstituted in HCl was comparable to that of HCl alone. Moreover, the pH variations induced by glutamate or HCl in the culture medium were similar. When the pH of the glutamate stock solution was re-equilibrated, l-glutamate induced variation in cell viability to a lower extent and after a longer incubation time. This study demonstrated that HCl used to reconstitute l-glutamate powder might alter the effect of glutamate itself in neuroblastoma cell cultures. Thus, this information might be useful to scientists who use l-glutamate to induce excitotoxicity or to test neuroprotective agents.

  1. A kinetic study of the oxidative deamination of L-glutamate by Peptostreptococcus asaccharolyticus glutamate dehydrogenase using a variety of coenzymes.

    PubMed

    Hornby, D P; Engel, P C

    1984-09-17

    The NAD+-specific glutamate dehydrogenase from Peptostreptococcus asaccharolyticus follows Michaelis-Menten kinetics in contrast to the enzyme from several other sources, and thus gives linear double-reciprocal plots of initial-rate data. The initial-rate parameters have been determined for this bacterial dehyrogenase in the direction of oxidative deamination. The use of alternative coenzymes leads to some conclusions about the order of substrate addition. An investigation of the pH dependence of this reaction reveals that the binding of oxidised coenzyme is independent of pH over the range 6-9. The kinetic data are consistent with an ordered addition of coenzyme prior to glutamate, the reverse of the mechanism derived with ox glutamate dehydrogenase in the presence of ADP.

  2. Efficient production of gamma-aminobutyric acid using Escherichia coli by co-localization of glutamate synthase, glutamate decarboxylase, and GABA transporter.

    PubMed

    Dung Pham, Van; Somasundaram, Sivachandiran; Lee, Seung Hwan; Park, Si Jae; Hong, Soon Ho

    2016-01-01

    Gamma-aminobutyric acid (GABA) is an important bio-product, which is used in pharmaceutical formulations, nutritional supplements, and biopolymer monomer. The traditional GABA process involves the decarboxylation of glutamate. However, the direct production of GABA from glucose is a more efficient process. To construct the recombinant strains of Escherichia coli, a novel synthetic scaffold was introduced. By carrying out the co-localization of glutamate synthase, glutamate decarboxylase, and GABA transporter, we redirected the TCA cycle flux to GABA pathway. The genetically engineered E. coli strain produced 1.08 g/L of GABA from 10 g/L of initial glucose. Thus, with the introduction of a synthetic scaffold, we increased GABA production by 2.2-fold. The final GABA concentration was increased by 21.8% by inactivating competing pathways.

  3. VGLUTs and Glutamate Synthesis—Focus on DRG Neurons and Pain

    PubMed Central

    Malet, Mariana; Brumovsky, Pablo R.

    2015-01-01

    The amino acid glutamate is the principal excitatory transmitter in the nervous system, including in sensory neurons that convey pain sensation from the periphery to the brain. It is now well established that a family of membrane proteins, termed vesicular glutamate transporters (VGLUTs), serve a critical function in these neurons: they incorporate glutamate into synaptic vesicles. VGLUTs have a central role both under normal neurotransmission and pathological conditions, such as neuropathic or inflammatory pain. In the present short review, we will address VGLUTs in the context of primary afferent neurons. We will focus on the role of VGLUTs in pain triggered by noxious stimuli, peripheral nerve injury, and tissue inflammation, as mostly explored in transgenic mice. The possible interplay between glutamate biosynthesis and VGLUT-dependent packaging in synaptic vesicles, and its potential impact in various pain states will be presented. PMID:26633536

  4. Adsorption dynamics of L-glutamic acid copolymers at a heptane/water interface.

    PubMed

    Beverung, C J; Radke, C J; Blanch, H W

    1998-02-16

    Random copolymers of glutamic acid (glu-ala, glu-leu, glu-phe, glu-tyr) were employed to investigate the relationship between side chain structure and peptide charge on adsorption behavior at an oil/water boundary. Adsorption of a series of glutamate copolymers at a heptane/water interface was examined by the dynamic pendant-drop method to determine interfacial tension. Incorporation of leucine or phenylalanine into a glutamate copolymer results in greater tension reduction than incorporation of alanine or tyrosine. These effects are amplified at pH values near the isoelectric point of glutamate, where macroscopic adsorbed films of glu-leu and glu-phe exhibit gel-like properties in response to interfacial area compression. Differences in interfacial tension behavior of glu-tyr and glu-phe indicate the importance of the tyrosine p-hydroxyl group on adsorption and aggregation at the oil/water interface. PMID:9540205

  5. Localization of glutamate receptors at a complex synapse. The mammalian photoreceptor synapse.

    PubMed

    Brandstätter, J H; Hack, I

    2001-01-01

    A key feature of signal processing in the mammalian retina is parallel processing, where the segregation of visual information, e.g., brightness, darkness, and color, starts at the first synapse in the retina, the photoreceptor synapse. These various aspects are transmitted in parallel from the input neurons of the retina, the photoreceptor cells, through the interconnecting bipolar cells, to the output neurons, the ganglion cells. The photoreceptors and bipolar cells release a single excitatory neurotransmitter, glutamate, at their synapses. This parsimony is contrasted by the expression of a plethora of glutamate receptors, receptor subunits, and isoforms. The detailed knowledge of the synaptic distribution of glutamate receptors thus is of major importance in understanding the mechanisms of retinal signal processing. This review intends to highlight recent studies on the distribution of glutamate receptors at the photoreceptor synapses of the mammalian retina.

  6. Chronic treatment with anti-bipolar drugs suppresses glutamate release from astroglial cultures.

    PubMed

    Liu, Zhuo; Song, Dan; Yan, Enzhi; Verkhratsky, Alexei; Peng, Liang

    2015-05-01

    Astroglial cells are fundamental elements of most neurological diseases, including bipolar disorders in which astrocytes show morphological and functional deficiency. Here we report the suppression of astroglial glutamate release by chronic treatment with three anti-bipolar drugs, lithium salt (Li(+)), carbamazepine (CBZ) and valproic acid (VPA). Release of glutamate was triggered by transient exposure of astrocytes to ATP (which activated purinoceptors) and 45 mM K(+) (which depolarised cell membrane to ~-30 mV). In both types of stimulation glutamate release was regulated by Ca(2+) entry through plasmalemmal channels and by Ca(2+) release from the endoplasmic reticulum (ER) intracellular stores. Exposure of astroglial cultures to Li(+), CBZ and VPA for 2 weeks led to a significant (more than 2 times) inhibition of glutamate release, which may alleviate the hyperactivity of the glutamatergic transmission in the brain of patients with bipolar disorders and thus contribute the underlying mechanism of drug action. PMID:25676933

  7. Chronic exercise dampens hippocampal glutamate overflow induced by kainic acid in rats.

    PubMed

    Holmes, Philip V; Reiss, Jenny I; Murray, Patrick S; Dishman, Rod K; Spradley, Jessica M

    2015-05-01

    Our laboratory has previously reported that chronic, voluntary exercise diminishes seizure-related behaviors induced by convulsant doses of kainic acid. The present experiments tested the hypothesis that exercise exerts this protective effect through a mechanism involving suppression of glutamate release in the hippocampal formation. Following three weeks of voluntary wheel running or sedentary conditions, rats were injected with 10 mg/kg of kainic acid, and hippocampal glutamate was measured in real time using a telemetric, in vivo voltammetry system. A separate experiment measured electroencephalographic (EEG) activity following kainic acid treatment. Results of the voltammetry experiment revealed that the rise in hippocampal glutamate induced by kainic acid is attenuated in exercising rats compared to sedentary controls, indicating that the exercise-induced protection against seizures involves regulation of hippocampal glutamate release. The findings reveal the potential benefit of regular exercise in the treatment and prevention of seizure disorders and suggest a possible neurobiological mechanism underlying this effect. PMID:25668513

  8. Photoreceptor Ablation Initiates the Immediate Loss of Glutamate Receptors in Postsynaptic Bipolar Cells in Retina

    PubMed Central

    2015-01-01

    Structural changes underlying neurodegenerative diseases include dismantling of synapses, degradation of circuitry, and even massive rewiring. Our limited understanding of synapse dismantling stems from the inability to control the timing and extent of cell death. In this study, selective ablation of cone photoreceptors in live mouse retina and tracking of postsynaptic partners at the cone-to-ON cone bipolar cell synapse reveals that early reaction to cone loss involves rapid and local changes in postsynaptic glutamate receptor distribution. Glutamate receptors disappear with a time constant of 2 h. Furthermore, binding of glutamate receptors by agonists and antagonists is insufficient to rescue glutamate receptor loss, suggesting that receptor allocation depends on the physical presence of cones. These findings demonstrate that the initial step in synapse disassembly involves postsynaptic receptor loss rather than dendritic retraction, providing insight into the early stages of neurodegenerative disease. PMID:25673837

  9. Reconsolidation of Reminder-Induced Amnesia: Role of NMDA and AMPA Glutamate Receptors.

    PubMed

    Nikitin, V P; Kozyrev, S A; Solntseva, S V

    2015-11-01

    We studied the role of glutamate receptors and reminder in the mechanisms of amnesia maintenance caused by disruption of conditioned food aversion reconsolidation with an antagonist of NMDA glutamate receptor in snails. At the early stage of amnesia (day 3 after induction), injection or NMDA of AMPA glutamate receptor antagonists prior to reminder (presentation of the conditioned food stimulus) led to memory recovery. Reminder alone or injection of antagonists without reminder or after reminder was ineffective. At the late stage of amnesia (day 10), antagonists/reminder had no effect on amnesia maintenance. It was hypothesized that reminder at the early stage of amnesia led to reactivation and reconsolidation of the molecular processes of amnesia including activation NMDA and AMPA glutamate receptors. Injection of antagonists of these receptors prior to reminder led to disruption of reactivation/reconsolidation of amnesia and recovery of the conditioned food aversion memory.

  10. A selective review of glutamate pharmacological therapy in obsessive–compulsive and related disorders

    PubMed Central

    Grados, Marco A; Atkins, Elizabeth B; Kovacikova, Gabriela I; McVicar, Erin

    2015-01-01

    Glutamate, an excitatory central nervous system neurotransmitter, is emerging as a potential alternative pharmacological treatment when compared to gamma-aminobutyric acid (GABA)-, dopamine-, and serotonin-modulating treatments for neuropsychiatric conditions. The pathophysiology, animal models, and clinical trials of glutamate modulation are explored in disorders with underlying inhibitory deficits (cognitive, motor, behavioral) including obsessive–compulsive disorder, attention deficit hyperactivity disorder, Tourette syndrome, trichotillomania, excoriation disorder, and nail biting. Obsessive–compulsive disorder, attention deficit hyperactivity disorder, and grooming disorders (trichotillomania and excoriation disorder) have emerging positive data, although only scarce controlled trials are available. The evidence is less supportive for the use of glutamate modulators in Tourette syndrome. Glutamate-modulating agents show promise in the treatment of disorders of inhibition. PMID:25995654

  11. Importance of Glutamate Dehydrogenase (GDH) in Clostridium difficile Colonization In Vivo

    PubMed Central

    Girinathan, Brintha Parasumanna; Braun, Sterling; Sirigireddy, Apoorva Reddy; Lopez, Jose Espinola; Govind, Revathi

    2016-01-01

    Clostridium difficile is the principal cause of antibiotic-associated diarrhea. Major metabolic requirements for colonization and expansion of C. difficile after microbiota disturbance have not been fully determined. In this study, we show that glutamate utilization is important for C. difficile to establish itself in the animal gut. When the gluD gene, which codes for glutamate dehydrogenase (GDH), was disrupted, the mutant C. difficile was unable to colonize and cause disease in a hamster model. Further, from the complementation experiment it appears that extracellular GDH may be playing a role in promoting C. difficile colonization and disease progression. Quantification of free amino acids in the hamster gut during C. difficile infection showed that glutamate is among preferred amino acids utilized by C. difficile during its expansion. This study provides evidence of the importance of glutamate metabolism for C. difficile pathogenesis. PMID:27467167

  12. Reduction of glutamate content in rat superior colliculus after retino-tectal denervation.

    PubMed

    Sakurai, T; Miyamoto, T; Okada, Y

    1990-02-16

    The effect of afferent lesions on glutamate content was measured in the lamina of the superior colliculus (SC) in the rat. The analysis was performed 12 days after unilateral enucleation (left eye), or ablation of visual cortex (right), or both enucleation and ablation. The glutamate contained in the superficial grey layer (SGL) and deep layer was measured in the sectioned freeze-dried sample using an enzymatic cycling method of NAD-NADH. The upper layer of SGL contralateral to enucleation exhibited a significant reduction (23%) in glutamate content. Combining enucleation and ablation further decreased (35%) glutamate content. Additionally, the synaptic potential evoked in the SGL of SC slices after stimulation of optic layer was blocked by the application of kynurenic acid (3 mM) or DNQX (30 microM). These results indicate that the retino-tectal pathway in the rat can be glutamatergic in nature.

  13. [Imbalance of system of glutamin - glutamic acid in the placenta and amniotic fluid at placental insufficiency].

    PubMed

    Pogorelova, T N; Gunko, V O; Linde, V A

    2014-01-01

    Metabolism of glutamine and glutamic acid has been investigated in the placenta and amniotic fluid under conditions of placental insufficiency. The development of placental insufficiency is characterized by the increased content of glutamic acid and a decrease of glutamine in both placenta and amniotic fluid. These changes changes were accompanied by changes in the activity of enzymes involved in the metabolism of these amino acids. There was a decrease in glutamate dehydrogenase activity and an increase in glutaminase activity with the simultaneous decrease of glutamine synthetase activity. The compensatory decrease in the activity of glutamine keto acid aminotransferase did not prevent a decrease in the glutamine level. The impairments in the system glutamic acid-glutamine were more pronounced during the development of premature labor.

  14. VGLUTs and Glutamate Synthesis-Focus on DRG Neurons and Pain.

    PubMed

    Malet, Mariana; Brumovsky, Pablo R

    2015-12-02

    The amino acid glutamate is the principal excitatory transmitter in the nervous system, including in sensory neurons that convey pain sensation from the periphery to the brain. It is now well established that a family of membrane proteins, termed vesicular glutamate transporters (VGLUTs), serve a critical function in these neurons: they incorporate glutamate into synaptic vesicles. VGLUTs have a central role both under normal neurotransmission and pathological conditions, such as neuropathic or inflammatory pain. In the present short review, we will address VGLUTs in the context of primary afferent neurons. We will focus on the role of VGLUTs in pain triggered by noxious stimuli, peripheral nerve injury, and tissue inflammation, as mostly explored in transgenic mice. The possible interplay between glutamate biosynthesis and VGLUT-dependent packaging in synaptic vesicles, and its potential impact in various pain states will be presented.

  15. Tight linkage of genes that encode the two glutamate synthase subunits of Escherichia coli K-12.

    PubMed Central

    Lozoya, E; Sanchez-Pescador, R; Covarrubias, A; Vichido, I; Bolivar, F

    1980-01-01

    A hybrid deoxyribonucleic acid molecule, plasmid pRSP20, which was isolated from the Clarke and Carbon Escherichia coli gene bank, was shown to complement the gltB31 mutation, which affects the synthesis of glutamate synthase in E. coli strain PA340. We present evidence which demonstrates that plasmid pRSP20 carries an 8-megadalton E. coli chromosomal fragment, including the genes encoding the two unequal glutamate synthase subunits. Polypeptides with molecular weights of about 135,000 and 53,000, which comigrated with purified E. coli glutamate synthase subunit polypeptides and immunoprecipitated with antibodies to E. coli glutamate synthase, were synthesized by minicells carrying the pRSP20 plasmid. Images PMID:6107287

  16. Astrocytes and glutamate homoeostasis in Alzheimer's disease: a decrease in glutamine synthetase, but not in glutamate transporter-1, in the prefrontal cortex.

    PubMed

    Kulijewicz-Nawrot, Magdalena; Syková, Eva; Chvátal, Alexander; Verkhratsky, Alexei; Rodríguez, José J

    2013-10-07

    Astrocytes control tissue equilibrium and hence define the homoeostasis and function of the CNS (central nervous system). Being principal homoeostatic cells, astroglia are fundamental for various forms of neuropathology, including AD (Alzheimer's disease). AD is a progressive neurodegenerative disorder characterized by the loss of cognitive functions due to specific lesions in mnesic-associated regions, including the mPFC (medial prefrontal cortex). Here, we analyzed the expression of GS (glutamine synthetase) and GLT-1 (glutamate transporter-1) in astrocytes in the mPFC during the progression of AD in a triple-transgenic mouse model (3xTg-AD). GS is an astrocyte-specific enzyme, responsible for the intracellular conversion of glutamate into glutamine, whereas the removal of glutamate from the extracellular space is accomplished mainly by astroglia-specific GLT-1. We found a significant decrease in the numerical density (Nv, cells/mm3) of GS-positive astrocytes from early to middle ages (1-9 months; at the age of 1 month by 17%, 6 months by 27% and 9 months by 27% when compared with control animals) in parallel with a reduced expression of GS (determined by Western blots), which started at the age of 6 months and was sustained up to 12 months of age. We did not, however, find any changes in the expression of GLT-1, which implies an intact glutamate uptake mechanism. Our results indicate that the decrease in GS expression may underlie a gradual decline in the vital astrocyte-dependent glutamate-glutamine conversion pathway, which in turn may compromise glutamate homoeostasis, leading towards failures in synaptic connectivity with deficient cognition and memory.

  17. The actions of L-glutamate at the postsynaptic membrane of the squid giant synapse.

    PubMed

    Adams, D J; Gillespie, J I

    1988-11-01

    The actions of L-glutamate on the postsynaptic membrane of the squid giant synapse were investigated using two methods of application: ionophoresis and bath perfusion. Bath perfusion of 10 mmoll-1 sodium glutamate did not produce an appreciable depolarization of the postsynaptic membrane but reversibly blocked the neurally evoked postsynaptic potential (PSP). The postsynaptic membrane depolarized when L-glutamate was applied ionophoretically. The sensitivity to glutamate application was not uniform, but sharply localized to sites which may correspond to synaptic contacts made by branching colaterals from the postsynaptic axon. The relationship between membrane potential and amplitude of the glutamate-activated postsynaptic potential (PSP) examined under current-clamp conditions was linear over the voltage range studied (-110 to -60 mV) with an extrapolated reversal potential of -36 mV. The amplitude of the glutamate-activated PSP was reduced either by replacing Na+ in the external solution with Tris+ (Na+-free) or by raising the extracellular K+ concentration to 20 mmoll-1 and was abolished by removing both Na+ and Ca2+ from the bath solution. The PSP amplitude was insensitive to changes in the extracellular Mg2+ concentration. The extrapolated reversal potential of the glutamate PSP was shifted to more positive potentials in both Na+-free and raised-K+ bathing solutions and was unchanged by anion substitution. The depolarization induced by L-glutamate increased with increasing ionophoretic current and reached a maximum with large pulses. Double logarithmic plots of the coulomb dose-response relationship gave a limiting slope in the range 1.7-2.2, suggesting that two glutamate molecules are required for receptor activation. The time course of desensitization of the glutamate response was studied using a double-pulse method. The initial decrease in the ratio, PSP2/PSP1, is followed by a slower time-dependent recovery of the postsynaptic response with a time constant

  18. Electrochemical synthesis of adiponitrile from the renewable raw material glutamic acid.

    PubMed

    Dai, Jian-Jun; Huang, Yao-Bing; Fang, Chi; Guo, Qing-Xiang; Fu, Yao

    2012-04-01

    Current affairs: Adiponitrile, used to produce nylon 6.6, is prepared from the renewable compound glutamic acid by an electrochemical route, involving electro-oxidative decarboxylation and Kolbe coupling reactions. The new route is an example of the use of glutamic acid as a versatile substrate in the transformation of biomass into chemicals. Also, it highlights the use of electrochemical methods in biomass conversion.

  19. Decreased glial and synaptic glutamate uptake in the striatum of HIV-1 gp120 transgenic mice.

    PubMed

    Melendez, Roberto I; Roman, Cristina; Capo-Velez, Coral M; Lasalde-Dominicci, Jose A

    2016-06-01

    The mechanisms leading to the neurocognitive deficits in humans with immunodeficiency virus type 1 (HIV-1) are not well resolved. A number of cell culture models have demonstrated that the HIV-envelope glycoprotein 120 (gp120) decreases the reuptake of glutamate, which is necessary for learning, memory, and synaptic plasticity. However, the impact of brain HIV-1 gp120 on glutamate uptake systems in vivo remains unknown. Notably, alterations in brain glutamate uptake systems are implicated in a number of neurodegenerative and neurocognitive disorders. We characterized the kinetic properties of system XAG (sodium-dependent) and systems xc- (sodium-independent) [3H]-L-glutamate uptake in the striatum and hippocampus of HIV-1 gp120 transgenic mice, an established model of HIV neuropathology. We determined the kinetic constant Vmax (maximal velocity) and Km (affinity) of both systems XAG and xc- using subcellular preparations derived from neurons and glial cells. We show significant (30-35 %) reductions in the Vmax of systems XAG and xc- in both neuronal and glial preparations derived from the striatum, but not from the hippocampus of gp120 mice relative to wild-type (WT) controls. Moreover, immunoblot analysis showed that the protein expression of glutamate transporter subtype-1 (GLT-1), the predominant brain glutamate transporter, was significantly reduced in the striatum but not in the hippocampus of gp120 mice. These extensive and region-specific deficits of glutamate uptake likely contribute to the development and/or severity of HIV-associated neurocognitive disorders. Understanding the role of striatal glutamate uptake systems in HIV-1 gp120 may advance the development of new therapeutic strategies to prevent neuronal damage and improve cognitive function in HIV patients. PMID:26567011

  20. Glutamate regulates eEF1A phosphorylation and ribosomal transit time in Bergmann glial cells.

    PubMed

    Barrera, Iliana; Flores-Méndez, Marco; Hernández-Kelly, Luisa C; Cid, Luis; Huerta, Miriam; Zinker, Samuel; López-Bayghen, Esther; Aguilera, José; Ortega, Arturo

    2010-12-01

    Glutamate, the major excitatory transmitter in the vertebrate brain, is involved in neuronal development and synaptic plasticity. Glutamatergic stimulation leads to differential gene expression patterns in neuronal and glial cells. A glutamate-dependent transcriptional control has been established for several genes. However, much less is known about the molecular events that modify the translational machinery upon exposure to this neurotransmitter. In a glial model of cerebellar cultured Bergmann cells, glutamate induces a biphasic effect on [(35)S]-methionine incorporation into proteins that suggests that the elongation phase of protein biosynthesis is the target for regulation. Indeed, after a 15 min exposure to glutamate a transient increase in elongation factor 2 phosphorylation has been reported, an effect mediated through the activation of the elongation factor 2 kinase. In this contribution, we sought to characterize the phosphorylation status of the eukaryotic elongation factor 1A (eEF1A) and the ribosomal transit time under glutamate exposure. A dose-dependent increase in eEF1A phosphorylation was found after a 60 min glutamate treatment; this phenomenon is Ca(2+)/CaM dependent, blocked with Src and phosphatidyl-inositol 3-kinase inhibitors and with rapamicyn. Concomitantly, the ribosomal transit time was increased with a 15 min glutamate exposure. After 60 more minutes, the average time used by the ribosomes to complete a polypeptide chain had almost returned to its initial level. These results strongly suggest that glutamate exerts an exquisite time-dependent translational control in glial cells, a process that might be critical for glia-neuron interactions.

  1. Mammalian folylpoly-. gamma. -glutamate synthetase. 2. Substrate specificity and kinetic properties

    SciTech Connect

    Cichowicz, D.J.; Shane, B.

    1987-01-27

    The specificity of hog liver folylpolyglutamate synthetase for folate substrates and for nucleotide and L-(/sup 14/C)glutamate substrates and analogues has been investigated. The kinetic mechanism, determined by using aminopterin as the folate substrate, is ordered Ter-Ter with MgATP binding first, folate second, and glutamate last. This mechanism precludes the sequential addition of glutamate moieties to enzyme-bound folate. Folate, dihydrofolate, and tetrahydrofolate possess the optimal configurations for catalysis while 5- and 10-position substitutions of the folate molecule impair catalysis. k/sub cat/ values decrease with increasing glutamate chain length, and the rate of decrease varies depending on the state of reduction and substitution of the folate molecule. Folate binding, as assessed by on rates, is slow. Dihydrofolate exhibits the fastest rate, and the rates are slightly reduced for tetrahydrofolate and 10-formyltetrahydrofolate and greatly reduced for 5-methyltetrahydrofolate and folic acid. Tetrahydrofolate polyglutamates are the only long glutamate chain length folates with detectable substrate activity. The specificity of the L-glutamate binding site is very narrow. L-Homocysteate and 4-threo-fluoroglutamate are alternate substrates and act as chain termination inhibitors in that their addition to the folate molecule prevents or severely retards the further addition of glutamate moieties. The K/sub m/ for glutamate is dependent on the folate substrate used. MgATP is the preferred nucleotide substrate, and ..beta..,..gamma..-methylene-ATP, ..beta..,..gamma..-imido-ATP, adenosine 5'-O-(3-thiotriphosphate), P/sup 1/,P/sup 5/-di(adenosine-5') pentaphosphate, and free ATP/sup 4 -/ are potent inhibitors of the reaction.

  2. Gabapentin increases extracellular glutamatergic level in the locus coeruleus via astroglial glutamate transporter-dependent mechanisms.

    PubMed

    Suto, Takashi; Severino, Amie L; Eisenach, James C; Hayashida, Ken-ichiro

    2014-06-01

    Gabapentin has shown to be effective in animals and humans with acute postoperative and chronic pain. Yet the mechanisms by which gabapentin reduces pain have not been fully addressed. The current study performed in vivo microdialysis in the locus coeruleus (LC) in normal and spinal nerve ligated (SNL) rats to examine the effect of gabapentin on extracellular glutamate concentration and its mechanisms of action with focus on presynaptic GABA-B receptors, astroglial glutamate transporter-1 (GLT-1), and interactions with α2δ subunits of voltage-gated Ca(2+) channels and endogenous noradrenaline. Basal extracellular concentration and tissue content of glutamate in the LC were greater in SNL rats than normal ones. Intravenously administered and LC-perfused gabapentin increased extracellular glutamate concentration in the LC. The net amount of glutamate increased by gabapentin is larger in SNL rats compared with normal ones, although the percentage increases from the baseline did not differ. The gabapentin-related α2δ ligand pregabalin increased extracellular glutamate concentration in the LC, whereas another α2δ ligand, 3-exo-aminobicyclo [2.2.1] heptane-2-exo-carboxylic acid (ABHCA), did not. Selective blockade by the dihydrokainic acid or knock-down of GLT-1 by the small interfering RNA abolished the gabapentin-induced glutamate increase in the LC, whereas blockade of GABA-B receptors by the CGP-35348 and depletion of noradrenalin by the dopamine-β-hydroxylase antibody conjugated to saporin did not. These results suggest that gabapentin induces glutamate release from astrocytes in the LC via GLT-1-dependent mechanisms to stimulate descending inhibition. The present study also demonstrates that this target of gabapentin in astrocytes does not require interaction with α2δ subunits in neurons.

  3. Electrochemical synthesis of adiponitrile from the renewable raw material glutamic acid.

    PubMed

    Dai, Jian-Jun; Huang, Yao-Bing; Fang, Chi; Guo, Qing-Xiang; Fu, Yao

    2012-04-01

    Current affairs: Adiponitrile, used to produce nylon 6.6, is prepared from the renewable compound glutamic acid by an electrochemical route, involving electro-oxidative decarboxylation and Kolbe coupling reactions. The new route is an example of the use of glutamic acid as a versatile substrate in the transformation of biomass into chemicals. Also, it highlights the use of electrochemical methods in biomass conversion. PMID:22441826

  4. Effects of phosphoenolpyruvate carboxylase desensitization on glutamic acid production in Corynebacterium glutamicum ATCC 13032.

    PubMed

    Wada, Masaru; Sawada, Kazunori; Ogura, Kotaro; Shimono, Yuta; Hagiwara, Takuya; Sugimoto, Masakazu; Onuki, Akiko; Yokota, Atsushi

    2016-02-01

    Phosphoenolpyruvate carboxylase (PEPC) in Corynebacterium glutamicum ATCC13032, a glutamic-acid producing actinobacterium, is subject to feedback inhibition by metabolic intermediates such as aspartic acid and 2-oxoglutaric acid, which implies the importance of PEPC in replenishing oxaloacetic acid into the TCA cycle. Here, we investigated the effects of feedback-insensitive PEPC on glutamic acid production. A single amino-acid substitution in PEPC, D299N, was found to relieve the feedback control by aspartic acid, but not by 2-oxoglutaric acid. A simple mutant, strain R1, having the D299N substitution in PEPC was constructed from ATCC 13032 using the double-crossover chromosome replacement technique. Strain R1 produced glutamic acid at a concentration of 31.0 g/L from 100 g/L glucose in a jar fermentor culture under biotin-limited conditions, which was significantly higher than that of the parent, 26.0 g/L (1.19-fold), indicative of the positive effect of desensitized PEPC on glutamic acid production. Another mutant, strain DR1, having both desensitized PEPC and PYK-gene deleted mutations, was constructed in a similar manner using strain D1 with a PYK-gene deleted mutation as the parent. This mutation had been shown to enhance glutamic acid production in our previous study. Although marginal, strain D1 produced higher glutamic acid, 28.8 g/L, than ATCC13032 (1.11-fold). In contrast, glutamic acid production by strain DR-1 was elevated up to 36.9 g/L, which was 1.42-fold higher than ATCC13032 and significantly higher than the other three strains. The results showed a synergistic effect of these two mutations on glutamic acid production in C. glutamicum. PMID:26168906

  5. Glutamate-glutamine cycle and exchange in the placenta-fetus unit during late pregnancy.

    PubMed

    Wu, Xin; Xie, Chunyan; Zhang, Yuzhe; Fan, Zhiyong; Yin, Yulong; Blachier, Francois

    2015-01-01

    The present review focuses on the physiological functions of glutamate-glutamine exchange involving placental amino acid transport and umbilical amino acid uptake in mammals (particularly in sows), with special emphasis on the associated regulating mechanisms. Glutamate plus glutamine are among the most abundant and the most utilized amino acids in fetus during late gestation. During pregnancy, amino acids, notably as precursors of macromolecules including proteins and nucleotides are involved in fetal development and growth. Amino acid concentrations in fetus are generally higher than in the mother. Among amino acids, the transport and metabolism of glutamate and glutamine during fetal development exhibit characteristics that clearly emphasize the importance of the interaction between the placenta and the fetal liver. Glutamate is quite remarkable among amino acids, which originate from the placenta, and is cleared from fetal plasma. In addition, the flux of glutamate through the placenta from the fetal plasma is highly correlated with the umbilical glutamate delivery rate. Glutamine plays a central role in fetal carbon and nitrogen metabolism and exhibits one of the highest fetal/maternal plasma ratio among all amino acids in human and other mammals. Glutamate is taken up by placenta from the fetal circulation and then converted to glutamine before being released back into the fetal circulation. Works are required on the glutamate-glutamine metabolism during late pregnancy in physiological and pathophysiological situations since such works may help to improve fetal growth and development both in humans and other mammals. Indeed, glutamine supplementation appears to ameliorate fetal growth retardation in sows and reduces preweaning mortality of piglets.

  6. Glutamic acid decarboxylase isoform distribution in transgenic mouse septum: an anti-GFP immunofluorescence study.

    PubMed

    Verimli, Ural; Sehirli, Umit S

    2016-09-01

    The septum is a basal forebrain region located between the lateral ventricles in rodents. It consists of lateral and medial divisions. Medial septal projections regulate hippocampal theta rhythm whereas lateral septal projections are involved in processes such as affective functions, memory formation, and behavioral responses. Gamma-aminobutyric acidergic neurons of the septal region possess the 65 and 67 isoforms of the enzyme glutamic acid decarboxylase. Although data on the glutamic acid decarboxylase isoform distribution in the septal region generally appears to indicate glutamic acid decarboxylase 67 dominance, different studies have given inconsistent results in this regard. The aim of this study was therefore to obtain information on the distributions of both of these glutamic acid decarboxylase isoforms in the septal region in transgenic mice. Two animal groups of glutamic acid decarboxylase-green fluorescent protein knock-in transgenic mice were utilized in the experiment. Brain sections from the region were taken for anti-green fluorescent protein immunohistochemistry in order to obtain estimated quantitative data on the number of gamma-aminobutyri