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Simultaneous measurement for intracellular temperature and activity of thermosensitive TRP channel

Research Project

Project/Area Number 18K15024
Research Category

Grant-in-Aid for Early-Career Scientists

Allocation TypeMulti-year Fund
Review Section Basic Section 48020:Physiology-related
Research InstitutionCenter for Novel Science Initatives, National Institutes of Natural Sciences

Principal Investigator

Ujisawa Tomoyo  大学共同利用機関法人自然科学研究機構(新分野創成センター、アストロバイオロジーセンター、生命創成探究, 生命創成探究センター, 特別研究員 (10816515)

Project Period (FY) 2018-04-01 – 2021-03-31
Project Status Completed (Fiscal Year 2020)
Budget Amount *help
¥4,160,000 (Direct Cost: ¥3,200,000、Indirect Cost: ¥960,000)
Fiscal Year 2019: ¥1,950,000 (Direct Cost: ¥1,500,000、Indirect Cost: ¥450,000)
Fiscal Year 2018: ¥2,210,000 (Direct Cost: ¥1,700,000、Indirect Cost: ¥510,000)
KeywordsTRP channel / 細胞内温度 / TRPチャネル / パッチクランプ / 細胞内温度計測 / TPRチャネル / 温度プローブ / 膜電流計測
Outline of Final Research Achievements

It is hard to determine the accurate temperature thresholds for activation of thermosensitive TRP channels since people measure the temperature in the bath solution closed to the patch-clamped cell expressing thermosensitive TRP channels. Therefore, we attempted to develop the system for the simultaneous measurement of the intracellular temperature and membrane currents using a fluorescent temperature probe. We introduced the fluorescent temperature probe into the HEK293 cells and analyzed the changes in fluorescent ratio upon local stimulation with infrared laser. Then, we combined the patch-clamp system on the confocal microscope with the infrared laser. We clearly observed rat TRPV1-mediated membrane currents with this system upon laser stimulation. Thus, we could establish the system for the simultaneous measurement of intracellular temperature and thermosensitive TRP channel currents.

Academic Significance and Societal Importance of the Research Achievements

温度感受性TRPチャネルの活性化温度閾値は膜電流とパッチクランプされた細胞近傍の外溶液温度の同時計測から決定されている。しかし、細胞外からの熱刺激による細胞内温度、さらにはTRPチャネルが発現する形質膜付近の温度変化とチャネル応答を同時記録した研究はない。さらには、IRレーザーを用いて局所温度変化を与えた際の分子の活性変化を報告したものはなく、今後cell-attached法による単一チャネル電流解析から、チャネルgatingへの影響も解析可能であり、細胞膜温度変化からチャネル開口に至る過程の解明に寄与できると期待される。

Report

(4 results)
  • 2020 Annual Research Report   Final Research Report ( PDF )
  • 2019 Research-status Report
  • 2018 Research-status Report
  • Research Products

    (2 results)

All 2019 2018

All Presentation (2 results) (of which Int'l Joint Research: 2 results)

  • [Presentation] Simultaneous intracellular temperature imaging during patch-clamp recording of TRPV1 activity2019

    • Author(s)
      Tomoyo Ujisawa, Kunitoshi Uchida, Kohki Okabe, Takeharu Nagai, Makoto Tominaga
    • Organizer
      9th FAOPS congress
    • Related Report
      2018 Research-status Report
    • Int'l Joint Research
  • [Presentation] Simultaneous measurement of thermosensitive TRP channels activity and intracellular temperature2018

    • Author(s)
      Tomoyo Ujisawa, Kunitoshi Uchida, Kohki Okabe, Makoto Tominaga
    • Organizer
      11th FENS Forum of Neuroscience
    • Related Report
      2018 Research-status Report
    • Int'l Joint Research

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Published: 2018-04-23   Modified: 2022-01-27  

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