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A study on the developmental plasticity of neurons using isolated spinal cord and cultures visual cortex.

Research Project

Project/Area Number 03044055
Research Category

Grant-in-Aid for international Scientific Research

Allocation TypeSingle-year Grants
SectionJoint Research
Research InstitutionTokyo Medical and Dental University

Principal Investigator

OTSUKA Masanori  Tokyo Medical and Dental University Faculty of Medicine, Professor, 医学部, 教授 (60013801)

Co-Investigator(Kenkyū-buntansha) BAUGHMAN Rob  ハーバード医科大学, 医学部, 副教授
SUZUKI Hidenori  Tokyo Medical and Dental University Faculty of Medicine, Research Associate, 医学部, 助手 (30221328)
MURAKOSHI Takayuki  Tokyo Medical and Dental University Faculty of Medicine, Research Associate, 医学部, 助手 (60190906)
YOSHIOKA Koichi  Tokyo Medical and Dental University Faculty of Medicine, Assistant Professor, 医学部, 助教授 (00143579)
YANAGISAWA Mitsuhiko  Tokyo Medical and Dental University Faculty of Medicine, Lecturer, 医学部, 講師 (90159252)
BAUGHMAN robert W.  Harvard Medical School Faculty of Medicine, Associate Professor
ROBERT Baugh  ハーバード医科大学, 医学部, 副教授
斎藤 公司 (斉藤 公司)  東京医科歯科大学, 医学部, 助教授 (20002082)
Project Period (FY) 1991 – 1993
Project Status Completed (Fiscal Year 1993)
Budget Amount *help
¥6,000,000 (Direct Cost: ¥6,000,000)
Fiscal Year 1993: ¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1992: ¥2,000,000 (Direct Cost: ¥2,000,000)
Fiscal Year 1991: ¥2,000,000 (Direct Cost: ¥2,000,000)
KeywordsNeural Plasticity / Glutamate Receptor / Acetylcholine Receptor / Tissue Culture / Substance P / Cerebral Cortex / Spinal Cord / アセチルコリン受容体 / c-fos / substance P / phorbol ester / cーfos
Research Abstract

The present research project aimed at studying the neural plasticity by means of examining the interaction between neural activities and synapse formation during development with chronically maintained central nervous system(CNS) preparations. In particular, a special emphasis was made upon investigating the modulatory effects of neurotransmitters on the synaptic plasticity. For this purpose, the following 4 in vitro CNS preparations were employed to perform electrophysiological and cell biological/biochemical analyses : 1) an isolated spinal cord preparation from newborn rats ; 2) a slice (400 mum thick) preparation from rat cerebral cortex ; 3) a dissociated neuronal cell culture, in which neurons were enzymatically isolated from the cerebral cortex, spinal cord, and spinal dorsal root ganglia of newborn rats, and maintained for 3 to 5 weeks ; 4) an organotypic culture of thin slices (100- 200 mum thick) maintained on a membrane for several days. We have been using the semi-acute pre … More parations of 1) and 2) for many years. The culture systems of 3) and 4), suitable for more chronic experiments, have been established by Dr Robert Baughman, a collaboration partner in the present project.
First, the effects of acetylcholine which is considered to promote the plasticity in developing CNS neurons were studied. A whole-cell patch-clamp recording from single neurons in the dissociated cell culture, an extracellular recording from spinal motoneurons of the isolated spinal cord preparation, and an intracellular recording from acute slice preparations all showed that acetylcholine caused a direct excitatory action on the cortical pyramidal neurons and the spinal motoneurons via activation of the muscarinic m_3 type receptor. Furthermore muscarinic agonist suppressed excitatory and inhibitory postsynaptic potentials (EPSPs & IPSPs) in the cortex, and also in the spinal cord suppressed EPSPs by a presynaptic mechanism. Next, the action of the metabotropic glutamate receptor was examined in the cortex, where glutamate is a major excitatory transmitter. Trans ACPD, an agonist for this receptor subtype, caused a membrane depolarization at the concentration of 10-100 muM.
The induction of gene expression is expected to occur during the chronic synaptic changes in response to external stimuli in order to initiate morphological and metabolic changes of the neurons. Expression of c-fos, a member of the immediate early genes, is reported in the spinal neurons after peripheral nociception, and after seizure activity in the cortical and hippocampal neurons. We examined weather this gene expression is induced by activation of the transmitter receptors using dissociated and slice culture preparations from the cortex, hippocampus, and the spinal cord. The neurons were stimulated by 1 hour application of the following agents to the medium, 1) high K+(50 mM), 2) substance P(10 muM), 3) phorbol ester (phorbol 2,3 di-acetate, 1 muM). When examined by immunocytochemistry using anti-c-fos protein, phorbol ester induced a significant increase in the c-fos expression in the dissociated cortical neurons while other stimuli did not. Substance P induced the c-fos expression in the dissociated spinal neurons. In the organotypic slice preparations, phorbol ester again triggered the c-fos expression in the cortex, hippocampus and the spinal cord neurons, however, substance P did not induce significant expression in all the neurons. Considering that substance P is one of the transmitters of the primary afferent neurons which conduct pain sensation to the spinal cord, the positive result in the dissociated spinal neurons may suggest that the present experimental paradigm is suitable for studying participation of external stimuli in the plastic changes in the CNS. Less

Report

(3 results)
  • 1993 Final Research Report Summary
  • 1992 Annual Research Report
  • 1991 Annual Research Report
  • Research Products

    (26 results)

All Other

All Publications (26 results)

  • [Publications] Otsuka M.and Yoshioka K.: "Neurotransmitter functions of mammalian tachykinins." Physiol.Rev.73. 229-308 (1993)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Kurihara T.et al.: "Muscarinic excitatory and inhibitorymechanisms involved in afferent fibre-evoked depolarization of motoneurones in the neonatal rat spinal cord." Br.J.Pharmacol.110. 61-70 (1993)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Suzuki H.et al.: "Pharmacological characteristics of tachykinin receptors mediating acetylcholine release from neonatal rat spinal cord." Eur.J.Pharmacol.241. 105-110 (1993)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Murakoshi T.et al.: "Electrophysiological identification of horizontal synaptic connections in rat visual cortex in vitro." Neurosci.Lett.163. 211-214 (1993)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Yomono H.S.et al.: "Sertonergic fibers induce a long-lasting inhibition of monosynaptic reflex in the neonatal rat spinal cord." Neuroscience. 47. 521-531 (1992)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Sakuma M.et al.: "Substance P-evoked release of GABA from isolated spinal cord of the newborn rat." Neuroscience. 45. 323-330 (1991)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Suzuki H.and Otsuka M.: "Methods in neurotransmitter and neuropeptide research." Elsevier, 443-455 (1993)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Otsuka M.et al.: "The Tachykinin Receptors." Humana Press, in press (1994)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Otsuka M. and Yoshioka K.: "Neurotransmitter functions of mammalian tachykinins." Physiol.Rev.73. 229-308 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Kurihara T. et al.: "Muscarinic excitatory and inhibitory mechanisms involved in afferent fibre-evoked depolarization of motoneurones in the neonatal rat spinal cord." Br.J.Pharmacol.110. 61-70 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Suzuki H. et al.: "Pharmacological characteristics of tachykinin receptors mediating acetylcholine release from neonatal rat spinal cord." Eur.J.Pharmacol.241. 105-110 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Murakoshi T. et al.: "Electrophysiological identification of horizontal synaptic connections in rat visual cortex in vitro." Neurosci.Lett.163. 211-214 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Yomono H.S. et al.: "Sertonergic fibers induce a long-lasting inhibition of monosynaptic reflex in the neonatal rat spinal cord." Neuroscience. 47. 521-531 (1992)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Sakuma M. et al.: "Substance P-evoked release of GABA from isolated spinal cord of the newborn rat." Neuroscience. 45. 323-330 (1991)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Suzuki H. and Otsuka M.: Elsevier. Methods in neurotransmitter and neuropeptide research., 443-455 (1993)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] Otsuka M. et al.: Humana Press(in press). The Tachykinin Receptors., (1994)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1993 Final Research Report Summary
  • [Publications] ISHIZUKI M.and YANAGISAWA M.: "Antinociceptive effects of tizanidine,diazepam and eperisone in isolated spinal cord-tail preparations of newborn rat." Pain. 48. 101-106 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] SAITO K,et al.: "Sensitivity of cultured and skinned chich myotube to calcium,strontium,and barium ions examined by recording isometric contractions." J.Cell.Physiol.150. 45-51 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] YANAGISAWA M.et al.: "The isolated spinal cord-skin preparation of the newborn rat and effects of some algogenic and analgesic substances." Eur.J.Pharmacol.220. 111-117 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] YANAGISAWA M.et al.: "Enzymatic inactivation of tachykinin neurotransmitters in the isolated spinal cord of the newborn rat." Neurosci.Res.15. 289-292 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] YOMONO H.S.et al.: "Serotonergic fibers induce a long-lasting inhibition of monosynaptic reflex in the neonatal rat spinal cord." Neuroscience. 47. 521-531 (1992)

    • Related Report
      1992 Annual Research Report
  • [Publications] YANAGISAWA M.et al.: "Inhibitory effects of human,porcine and rat galanin examined in the isolated spinal cord of the newborn rat." Biomed.Res.

    • Related Report
      1992 Annual Research Report
  • [Publications] Otsuka M.et al.: "Physiological characterization of tachykinkin receptor subtypes in the spinal cord of the newborn ret. In Tachykinin Receptors" Humana Press, (1993)

    • Related Report
      1992 Annual Research Report
  • [Publications] Kobayashi,N.: "Substance P-evoked release of acetylcholine from isolated spinal cord of the newborn rat." Neuroscience. 45. 331-337 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] Sakuma,M.: "Substance P-evoked release of GABA from isolated spinal cord of the newborn rat." Neuroscience. 45. 323-330 (1991)

    • Related Report
      1991 Annual Research Report
  • [Publications] Otsuka,M.: "Tachykinin-evoked release of neurotransmitters from isolated spinal cord of the newborn rat." Annals of the New York Academy of Science. 632. 212-219 (1991)

    • Related Report
      1991 Annual Research Report

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Published: 1991-04-01   Modified: 2016-04-21  

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