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Dynamics of membrane channels as revealed by freeze-fracture replica labeling

Research Project

Project/Area Number 09670008
Research Category

Grant-in-Aid for Scientific Research (C)

Allocation TypeSingle-year Grants
Section一般
Research Field General anatomy (including Histology/Embryology)
Research InstitutionKYOTO UNIVERSITY

Principal Investigator

FUJIMOTO Kazushi  Kyoto univ., Dept.of Anatomy, Lecturer, 医学研究科, 講師 (50159125)

Project Period (FY) 1997 – 1998
Project Status Completed (Fiscal Year 1998)
Budget Amount *help
¥3,400,000 (Direct Cost: ¥3,400,000)
Fiscal Year 1998: ¥1,500,000 (Direct Cost: ¥1,500,000)
Fiscal Year 1997: ¥1,900,000 (Direct Cost: ¥1,900,000)
Keywordschannels / cell membranes / electron microscopy / freeze-fracture / immuno-labeling / astrocytes / 免疫電顕 / アクアポリン / 生体膜 / 凍結割断レプリカ / チャネル / レセプター
Research Abstract

Glutamate is known to cause swelling of astrocytes both in situ and in vitro. Although several different types of ionotropic and metabotropic glutamate receptors are expressed on astrocytes, their functional role in the glutamate-induced cell swelling is poorly understood. Using freeze-fracture replica immunoelectron microscopy, we have found a distinctive co-localization of metabotropic glutamate receptor 3 (mGluR3) and water channel aquaporn 4 (AQP4) on a specialized membrane domain, called orthogonal arrays of particles (OAPs), of astrocyte membranes in situ. To examine whether mGluR3 and AQP4 assemble to form OAPs, their cDNAs were introduced into Chinese hamster ovary cells. Replica immunoelectron microscopy revealed that both molecules were highly concentrated in OAPs-like structure. Agonists of mGluR3, glutamate and DCG-IV, disrupted co-localization of mGluR3 and AQP4, and decreased the density of OAPs-like structure in these cells. Similar effects of the agonists were also observed in astrocytes in situ. Functional analysis using the transfected cells showed that the agonists increased water permeability across cell membranes leading to cell swelling, probably in a cAMP-dependent manner. These findings suggest that mGluR3 and AQP4 assemble to form OAPs, and changes in the structural and functional states are governed by the activity of mGluR3. Therefore, we propose a pathway, mGluR3-AQP4 coupling, of osmotic cell volume regulation, of astrocytes.

Report

(3 results)
  • 1998 Annual Research Report   Final Research Report Summary
  • 1997 Annual Research Report
  • Research Products

    (35 results)

All Other

All Publications (35 results)

  • [Publications] Fujimoto, K.et al.: "A simple and reliable quick-freezing/freeze-fracturing procedure." Histochem. Cell Biol.,. 107. 81-84 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Fujimoto, K.: "SDS-digested freeze-fracture replica labeling electron microscopy to study the two-dimensional distribution of integral-membrane protein and phospholipids in biomembranes : practical procedure, interpretation and application." Histochem. Cell Biol.,. 107. 87-96 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Fujimoto, K.: "mmunoelectron microscopy of protein kinase C in resting and phagocytosing macrophages." Acta histochem. cytochem.30. 105-111 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Fujimoto, K.: "Dynamics of connexins, E-cadherin and a-catenin on cell membranes during gap junction formation." J.Cell Sci.110. 311-322 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] 藤本 和: "SDS処理凍結割断レプリカ標識法-補講- : 急速凍結/凍結割断法、ならびに膜蛋白質のトポロジー解析と膜リン脂質の検出について" 電子顕微鏡. 32. 48-51 (1997)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Fujimoto. K.: "Immuno-labeling of freeze-fracture replica." Micron. in press. (1999)

    • Description
      「研究成果報告書概要(和文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Fujimoto, K.et al.: "A simple and reliable quick-freezing/freeze-fracturing procedure." Histochem.Cell Biol.107. 81-84 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Fujimoto.K.: "SDS-digested freeze-fracture replica labeling electron microscopy to study the two-dimensional distribution of integral membrane proteins and phospholipids in biomembranes : practical procedure, interpretation and application." Histochem.Cell Biol.107. 87-96 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Fujimoto, K.et al.: "Dynamics of connexins, E-cadherin and a-catenin on cell membranes during gap junction formation." J.Cell Sci.110. 311-322 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Fujimoto, T.and Fujimoto, K.: "Metal sandwich method to quick-freeze monolayer cultured cells for freeze-fracture." J.Histochem.Cytochem.34. 595-598 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Fujimoto, K.et al.: "Immunoelectron microscopy of protein kinase C in resting and phagocytosing macrophages." Acta histochem.cytochem.30. 105-111 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Hirase, T.et al.: "Occludin as a possible determinant of tight junction permeability in endothelial cells." J.Cell Sci.110. 1603-1613 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Saitou, M.et al.: "Mammalian occludin in epithelial cells : its expression and subcellular distribution." Eur.J.Cell Biol.73. 222-231 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Moller, J.V.et al.: "Probing of the membrane topology of sarco-plasmic reticulum Ca2+-ATPase with sequence specific antibodies : Evidence for plasticity of the C-terminal domain." J.Biol.Chem.272. 29015-29032 (1997)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Saitou, M.et al.: "Occludin-deficient embryonic stem cells can differentiate into polarized epithelial cells bearing tight junctions." J.Cell Biol.141. 397-408 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Moroi, S.et al.: "Occludin is concentrated at tight junctions of mouse/rat but not human/guinea pig Sertoli cells in testes." Am.J.Physiol.274. C1708-17 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Morita, K.et al.: "Subcellular distribution of tight junction-associated proteins (Occludin, ZO-1 and ZO-2) in rodent skin." J.Invest.Dermatol.110. 862-866 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Furuse, M.et al.: "Claudin-1 and -2 : novel integral membrane proteins localizing at tight junctions with no sequence similarity to occludin." J.Cell Biol.141. 1539-1550 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Watabe-Uchida, M.et al.: "a-catenin-vinculin interaction functions to organize the apical junctional complex in epithelial cells." J.Cell Biol.142. 847-857 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Furuse, M.et al.: "A single gene product, claudin-1 or-2, reconstitutes tight junction strands and recruits occludin in fibroblasts." J.Cell Biol.143. 391-401 (1998)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Morita, K.et al.: "Claudin multigene family encoding four-trans-membrane domain protein components of tight junction strands." Proc.Natl.Acad.Sci.USA,96. 511-516 (1999)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Fujimoto. K.: "Immuno-labeling of freeze-fracture replica." Micron. (in press). (1999)

    • Description
      「研究成果報告書概要(欧文)」より
    • Related Report
      1998 Final Research Report Summary
  • [Publications] Fujimoto,K.et al.: "A simple and reliable quick-freezing/freezu-fracturing procedure." Histochem.Cell Biol.107. 81-84 (1997)

    • Related Report
      1998 Annual Research Report
  • [Publications] Fujimoto,K.: "SDS-digested freeze-fracture replica labeling electron microscopy to study the two-dimensional distribution of integral membrane proteins and phospholipids in biomembranes:practical procedure,interpretation and application." Histochem.Cell Biol.107. 87-96 (1997)

    • Related Report
      1998 Annual Research Report
  • [Publications] Fujimoto,K.: "Immunoelectron microscopy of protein kinase C in resting and phagocytosing macrophages." Acta histochem.cytochem.30. 105-111 (1997)

    • Related Report
      1998 Annual Research Report
  • [Publications] Fujimoto,K.: "Dynamics of connexins,E-cadherin and a-catenin on cell membranes during gap junction formation." J.Cell Sci.110. 311-322 (1997)

    • Related Report
      1998 Annual Research Report
  • [Publications] 藤本 和: "SDS処理凍結割断レプリカ標識法-補講-:急速凍結/凍結割断法、ならびに膜蛋白質のトポロジー解析と膜・リン脂質の検出について" 電子顕微鏡. 32. 48-51 (1997)

    • Related Report
      1998 Annual Research Report
  • [Publications] Fujimoto,K.: "Immuno-labeling of freeze-fracture replica." Micron. (in press). (1999)

    • Related Report
      1998 Annual Research Report
  • [Publications] Fujimoto.K.et al.: "A simple and reliable quick-freezingl freeza-fracturing procedure." Histochem.Cell Biol.107・1. 81-84 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Fujimoto,K.et al.: "Dynamics of connexins,E-cadherin and α-catenin on cell membranes during gap junction formation." J.Cell.Sci.110・3. 311-322 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Fujimoto,K.: "SDS-digested freeza-Fracture replica labeling electron micro scopy to study the two-dimensional distribution of integral membrame protetus and Phospholipid." Histochem,Cell Biol.107・2. 87-96 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Hirase,T.et al,: "Occludin as a posstble,. determinart of tight Jumcton permeability in andothelral cells." J.Cell Sci.110・17. 1603-1613 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Fujimoto,K.et al.: "Immunoelectron microscopy of protein kinase C in resting and phagocytosing marophages." Acta histochem.cytochem.30・1. 105-111 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] Muller,J.V.et al.: "Probing of the membrane topdogy of sarcoplasmic reticulum Ca-ATPase with sequence specific antibodies." J.Biol.Chem.272・9. 29015-29032 (1997)

    • Related Report
      1997 Annual Research Report
  • [Publications] 藤本 和: "組織細胞化学1997" 日本組織細胞化学会, 11 (1997)

    • Related Report
      1997 Annual Research Report

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Published: 1997-04-01   Modified: 2016-04-21  

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