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Regulation of NMDA-type glutamate receptor by endogenous d -serine
Abstracts SY1-D2-1 Progress of the ␦2 glutamate receptor research from the viewpoint of functional domains Hirokazu Hirai, Takashi Torashima, Akira Iizuka Gunma University Graduate School of Medicine, Japan iGluRs share a common membrane topology that consists of three transmembrane domains (TM1, TM3, TM4), an ion-channel-pore-forming loop, an extracellular Nterminal domain (NTD) of approximately 400 amino acids, S1S2 ligand binding domains, and an intracellular C-terminal domain (CTD). Unlike other domains of iGluRs, the functions of NTDs in iGluRs including the ␦2 glutamate receptor (GluR␦2) were not well characterized. We produced various constructs with or without the NTD of GluR␦2, and roles of the NTD were examined by lentivector-based expression of those constructs in Purkinje cells of GluR␦2-null mice. Our results indicate that expression of constructs that contained the NTD, but lacked the S1S2 domains and channel-pore domain in Purkinje cells of immature GluR␦2-null mice reliably rescued ataxia and the expression of LTD. These results suggest the critical roles of the NTD in the function of GluR␦2. We, then, propose novel NTD-mediated and ion-channel-unrelated functions of GluR␦2 by which GluR␦2 regulates cerebellar functions. doi:10.1016/j.neures.2009.09.1517
SY1-D2-2 Roles of glutamate receptor ␦ subfamily in synapse formation Tomoo Hirano, Tomoaki Kuroyanagi, Marie Yokoyama Kyoto University, Japan Roles of ionotoropic glutamate receptor ␦ subfamily in synapse formation were examined. When non-neuronal HEK cells expressing ␦2 were co-cultured with mouse cerebellar neurons, punctate structures expressing marker proteins for glutamatergic presynaptic terminals of granule neurons were accumulated around them. Furthermore, HEK cells expressing both ␦2 and GluR1, a glutamate receptor subunit forming a functional glutamate-gated ion channel, showed postsynaptic current. Deletion of the extracellular LIVBP domain of ␦2 abolished the induction ability for synapse formation, and the LIVBP domain directly fused to a transmembrane sequence was sufficient to induce presynaptic differentiation. Furthermore, a mutant GluR1 whose LIVBP domain was replaced with the delta; 2 LIVBP domain was sufficient by itself to establish synaptic transmission. Another member of ␦ glutamate receptor family ␦1 also induced the presynaptic differentiation Thus, the ␦ glutamate receptor subunit subfamily can induce the differentiation of glutamatergic presynaptic terminals and contribute to the establishment of synaptic transmission. doi:10.1016/j.neures.2009.09.1518
SY1-D2-3 Regulatory mechanism of multiple signaling via the metabotropic glutamate receptor1␣ Michihiro Tateyama 1,2 , Yoshihiro Kubo 1,2 1 Div Biophysics & Neurobiology, National Institute for Phsyiological Sciences, Okazaki, Japan; 2 SORST, JST, Kawaguchi, Japan
The metabotropic glutamate receptor 1␣ (mGluR1␣) is crucial for some forms of synaptic plasticity. The mGluR1␣ is known to couple with different types of G proteins, Gq, Gs and Gi/o, which confers diversity in the mGluR1␣ singling. The multiple signaling has recently been reported to occur in neuronal and immune cells and is regulated by phosphorylation and truncation of the mGluR1␣. Here we would like to report dimeric configuration of mGluR1␣ also provides variety in the receptor activation mechanism that could alter the coupling profile of mGluR1␣. Glutamate binding to one subunit activated Gq pathway either through the glutamate-bound subunit (cis-activation) or the other subunit (trans-activation), as has been reported. In contrast Gi/o pathway was not activated through the cis- or trans-activation alone. The result suggested that the intersubunit interaction within the mGluR1␣ dimer regulates the multiple signaling. Detail of the regulation depending on the intersubunit interaction will be discussed. doi:10.1016/j.neures.2009.09.1519
SY1-D2-4 Regulation of NMDA-type glutamate receptor by endogenous d-serine Hisashi Mori Department of Molecular Neuroscience, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Japan NMDR-type glutamate receptor (GluR) plays critical roles in neural network formation, synaptic plasticity, higher brain functions and neuropathological disorders. In the mammalian brain, d-serine is detected and acts as a coagonist at the “glycine-site”
S9
of the NMDA-type GluR. Although d-serine can be directly produced from L-serine by serine racemase (SR), the relative contributions SR and d-serine for regulation of NMDA receptor in vivo are not known. We have recently produced the SR knockout (KO) mice, and found about 90% decrease d-serine content and a reduced neurotoxicity induced by NMDA- and Ab1-42 peptide injections into the forebrain. These results suggest that SR is the major enzyme for d-serine production in the brain, d-serine is the predominant endogenous coagonist of the NMDA receptor in the forebrain, and d-serine may be involved in controlling the extent of NMDA receptor-mediated neurotoxic insults. Supported by the grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan (507-16047210, 507-18053008 and 021-20022016). doi:10.1016/j.neures.2009.09.1520
SY1-D3-1 Regeneration of peripheral and central nerves Jianwu Dai Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China Injuries to peripheral nerves result in loss of motor, sensory and autonomic functions. Peripheral nervous system injury can result in functional loss and decreased quality of life. We have developed a targeting repair system by creating specific binding between NGF and collagen avoiding diffusion. In animal models such as the subcutaneous implantation model, ear dermal ischemic ulcer model, and sciatic nerve crush model, the targeting repair system could retain NGF at the nerve injured site for a longer time to promote the regeneration of peripheral nerves.Unlike the peripheral nervous system, the regeneration ability is extremely limited in adult mammalian CNS. We constructed an efficient brain-derived neurotrophic factor (BDNF) delivery system including collagen-binding BDNF and linear ordered collagen scaffolds (LOCS). Using the rat hemisection SCI model, we found that LOCS loaded with CBD-BDNF significantly improved the regeneration of CNS and the SCI recovery evaluated by the Basso, Beattie, and Bresnahan (BBB) scale and immunohistochemical staining with anti-neurofilament antibody. doi:10.1016/j.neures.2009.09.1521
SY1-D3-2 Tumor formation in cell replacement therapy using human ES cells Jun Takahashi, Daisuke Doi, Asuka Morizane Kyoto University, Institute for Frontier Medical Sciences, Japan Considering cell replacement therapy with embryonic stem cells (ESCs) for Parkinson’s disease, formation of a graft cell-derived tumor is a major concern about safety of the therapy. To confirm the safety and effectiveness of the graft, long-term observation after cell transplantation into the animal models, especially primate ones, is needed. In the present study, we induced neural progenitor cells (NPCs) from human ES cell lines (KhES-1, KhES-2) by the modified SDIA (stromal cell-derived inducing activity) method. The NPCs were transplanted into the bilateral striatum of monkey models of Parkinson’s disease. The survival and differentiation of the grafted cells were monitored by MRI and PET scans, and histological studies were performed after the animals were sacrificed in nine months. The graft contaminated with undifferentiated ESCs formed large tumors, while that with no Oct4+ cells did not. Purification of neural cells by FACS is expected to prevent tumor formation in the primate brains. doi:10.1016/j.neures.2009.09.1522
SY1-D3-3 Ex vivo gene therapy for central nervous system disorders Takao Yasuhara, Isao Date Okayama University Graduate School of Medicine, Japan Regenerative medicine for diseases of the central nervous systems (CNS) is developing in accordance with molecular biological advancement. Genetic engineering is one of the essential techniques and gene therapy is a hopeful modality for CNS disorders. Gene therapy is divided into 2 methods, ex vivo and in vivo gene therapy. Ex vivo gene therapy is a combined method of cell transplantation after genetic manipulation of donor cells, while in vivo gene therapy is direct gene transduction into host tissue. In this article, we review ex vivo gene therapies for CNS disorders both in laboratory and clinical settings with our recent data. We demonstrate the therapeutic effects of encapsulated cell transplantation after genetic manipulation and transplantation of genetically engineered neural stem/progenitor cells for cerebral infarct and Parkinson’s disease model of animals. doi:10.1016/j.neures.2009.09.1523
SY1-D2-2 Roles of glutamate receptor ␦ subfamily in synapse formation Tomoo Hirano, Tomoaki Kuroyanagi, Marie Yokoyama Kyoto University, Japan Roles of ionotoropic glutamate receptor ␦ subfamily in synapse formation were examined. When non-neuronal HEK cells expressing ␦2 were co-cultured with mouse cerebellar neurons, punctate structures expressing marker proteins for glutamatergic presynaptic terminals of granule neurons were accumulated around them. Furthermore, HEK cells expressing both ␦2 and GluR1, a glutamate receptor subunit forming a functional glutamate-gated ion channel, showed postsynaptic current. Deletion of the extracellular LIVBP domain of ␦2 abolished the induction ability for synapse formation, and the LIVBP domain directly fused to a transmembrane sequence was sufficient to induce presynaptic differentiation. Furthermore, a mutant GluR1 whose LIVBP domain was replaced with the delta; 2 LIVBP domain was sufficient by itself to establish synaptic transmission. Another member of ␦ glutamate receptor family ␦1 also induced the presynaptic differentiation Thus, the ␦ glutamate receptor subunit subfamily can induce the differentiation of glutamatergic presynaptic terminals and contribute to the establishment of synaptic transmission. doi:10.1016/j.neures.2009.09.1518
SY1-D2-3 Regulatory mechanism of multiple signaling via the metabotropic glutamate receptor1␣ Michihiro Tateyama 1,2 , Yoshihiro Kubo 1,2 1 Div Biophysics & Neurobiology, National Institute for Phsyiological Sciences, Okazaki, Japan; 2 SORST, JST, Kawaguchi, Japan
The metabotropic glutamate receptor 1␣ (mGluR1␣) is crucial for some forms of synaptic plasticity. The mGluR1␣ is known to couple with different types of G proteins, Gq, Gs and Gi/o, which confers diversity in the mGluR1␣ singling. The multiple signaling has recently been reported to occur in neuronal and immune cells and is regulated by phosphorylation and truncation of the mGluR1␣. Here we would like to report dimeric configuration of mGluR1␣ also provides variety in the receptor activation mechanism that could alter the coupling profile of mGluR1␣. Glutamate binding to one subunit activated Gq pathway either through the glutamate-bound subunit (cis-activation) or the other subunit (trans-activation), as has been reported. In contrast Gi/o pathway was not activated through the cis- or trans-activation alone. The result suggested that the intersubunit interaction within the mGluR1␣ dimer regulates the multiple signaling. Detail of the regulation depending on the intersubunit interaction will be discussed. doi:10.1016/j.neures.2009.09.1519
SY1-D2-4 Regulation of NMDA-type glutamate receptor by endogenous d-serine Hisashi Mori Department of Molecular Neuroscience, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, Japan NMDR-type glutamate receptor (GluR) plays critical roles in neural network formation, synaptic plasticity, higher brain functions and neuropathological disorders. In the mammalian brain, d-serine is detected and acts as a coagonist at the “glycine-site”
S9
of the NMDA-type GluR. Although d-serine can be directly produced from L-serine by serine racemase (SR), the relative contributions SR and d-serine for regulation of NMDA receptor in vivo are not known. We have recently produced the SR knockout (KO) mice, and found about 90% decrease d-serine content and a reduced neurotoxicity induced by NMDA- and Ab1-42 peptide injections into the forebrain. These results suggest that SR is the major enzyme for d-serine production in the brain, d-serine is the predominant endogenous coagonist of the NMDA receptor in the forebrain, and d-serine may be involved in controlling the extent of NMDA receptor-mediated neurotoxic insults. Supported by the grants from the Ministry of Education, Culture, Sports, Science and Technology of Japan (507-16047210, 507-18053008 and 021-20022016). doi:10.1016/j.neures.2009.09.1520
SY1-D3-1 Regeneration of peripheral and central nerves Jianwu Dai Institute of Genetics and Developmental Biology, Chinese Academy of Sciences, Beijing, China Injuries to peripheral nerves result in loss of motor, sensory and autonomic functions. Peripheral nervous system injury can result in functional loss and decreased quality of life. We have developed a targeting repair system by creating specific binding between NGF and collagen avoiding diffusion. In animal models such as the subcutaneous implantation model, ear dermal ischemic ulcer model, and sciatic nerve crush model, the targeting repair system could retain NGF at the nerve injured site for a longer time to promote the regeneration of peripheral nerves.Unlike the peripheral nervous system, the regeneration ability is extremely limited in adult mammalian CNS. We constructed an efficient brain-derived neurotrophic factor (BDNF) delivery system including collagen-binding BDNF and linear ordered collagen scaffolds (LOCS). Using the rat hemisection SCI model, we found that LOCS loaded with CBD-BDNF significantly improved the regeneration of CNS and the SCI recovery evaluated by the Basso, Beattie, and Bresnahan (BBB) scale and immunohistochemical staining with anti-neurofilament antibody. doi:10.1016/j.neures.2009.09.1521
SY1-D3-2 Tumor formation in cell replacement therapy using human ES cells Jun Takahashi, Daisuke Doi, Asuka Morizane Kyoto University, Institute for Frontier Medical Sciences, Japan Considering cell replacement therapy with embryonic stem cells (ESCs) for Parkinson’s disease, formation of a graft cell-derived tumor is a major concern about safety of the therapy. To confirm the safety and effectiveness of the graft, long-term observation after cell transplantation into the animal models, especially primate ones, is needed. In the present study, we induced neural progenitor cells (NPCs) from human ES cell lines (KhES-1, KhES-2) by the modified SDIA (stromal cell-derived inducing activity) method. The NPCs were transplanted into the bilateral striatum of monkey models of Parkinson’s disease. The survival and differentiation of the grafted cells were monitored by MRI and PET scans, and histological studies were performed after the animals were sacrificed in nine months. The graft contaminated with undifferentiated ESCs formed large tumors, while that with no Oct4+ cells did not. Purification of neural cells by FACS is expected to prevent tumor formation in the primate brains. doi:10.1016/j.neures.2009.09.1522
SY1-D3-3 Ex vivo gene therapy for central nervous system disorders Takao Yasuhara, Isao Date Okayama University Graduate School of Medicine, Japan Regenerative medicine for diseases of the central nervous systems (CNS) is developing in accordance with molecular biological advancement. Genetic engineering is one of the essential techniques and gene therapy is a hopeful modality for CNS disorders. Gene therapy is divided into 2 methods, ex vivo and in vivo gene therapy. Ex vivo gene therapy is a combined method of cell transplantation after genetic manipulation of donor cells, while in vivo gene therapy is direct gene transduction into host tissue. In this article, we review ex vivo gene therapies for CNS disorders both in laboratory and clinical settings with our recent data. We demonstrate the therapeutic effects of encapsulated cell transplantation after genetic manipulation and transplantation of genetically engineered neural stem/progenitor cells for cerebral infarct and Parkinson’s disease model of animals. doi:10.1016/j.neures.2009.09.1523