KEIO RESEARCHERS INFORMATION SYSTEM

Career 【 Display / hide 】

Career 【 Display / hide 】

• 2014.04

  -

  2016.03

  Japan Society for the Promotion of Science, 特別研究員

• 2016.04

  -

  2018.03

  The University of Tokyo, Graduate School of Medicine, 特任研究員

• 2018.04

  -

  2021.03

  Japan Society for the Promotion of Science, 特別研究員

• 2021.04

  -

  2021.08

  The University of Tokyo, Graduate School of Medicine, 特任研究員

• 2021.09

  -

  Present

  Keio University, School of Medicine, 助教

Academic Background 【 Display / hide 】

Academic Background 【 Display / hide 】

• 2005.04

  -

  2009.03

  Tokyo University of Science, Faculty of Pharmaceutical Sciences

• 2009.04

  -

  2011.03

  Tokyo University of Science, Graduate School of Pharmaceutical Sciences

• 2011.04

  -

  2015.03

  Keio University, 医学系研究科

Research Areas 【 Display / hide 】

Research Areas 【 Display / hide 】

• Life Science / Neuroscience-general

• Life Science / Pharmacology

Research Keywords 【 Display / hide 】

Research Keywords 【 Display / hide 】

• Multiphoton imaging

Papers 【 Display / hide 】

Papers 【 Display / hide 】

• Layer 5 intratelencephalic neurons in the motor cortex stably encode skilled movement

  Takanori Shinotsuka, Yasuhiro R. Tanaka, Shin-Ichiro Terada, Natsuki Hatano, Masanori Matsuzaki

  The Journal of Neuroscience (Society for Neuroscience)     JN - RM 2023.09

  Research paper (scientific journal), Joint Work, Lead author, Accepted,  ISSN  0270-6474

  　View Summary

  The primary motor cortex (M1) and the dorsal striatum play a critical role in motor learning and the retention of learned behaviors. Motor representations of corticostriatal ensembles emerge during motor learning. In the coordinated reorganization of M1 and the dorsal striatum for motor learning, layer 5a (L5a), which connects M1 to the ipsi- and contra-lateral dorsal striatum, should be a key layer. Although M1 L5a neurons represent movement-related activity in the late stage of learning, it is unclear whether the activity is retained as a memory engram. Here, usingTlx3-Cremale transgenic mice, we conducted two-photon calcium imaging of striatum-projecting L5a intratelencephalic (IT) neurons in forelimb M1 during late sessions of a self-initiated lever-pull task, and in sessions after 6 days of non-training following the late sessions. We found that trained male animals exhibited stable motor performance before and after the non-training days. At the same time, we found that M1 L5a IT neurons strongly represented the well-learned forelimb movement, but not uninstructed orofacial movements. A subset of M1 L5a IT neurons consistently coded the well-learned forelimb movement before and after the non-training days. Inactivation of M1 IT neurons after learning impaired task performance when the lever was made heavier or when the target range of the pull distance was narrowed. These results suggest that a subset of M1 L5a IT neurons continuously represent skilled movement after learning and serve to fine-tune the kinematics of well-learned movement.
  
  Significance Statement
  
  Motor memory persists even when it is not used for a while. Intratelencephalic neurons in layer 5a (L5a IT neurons) of the primary motor cortex (M1) gradually come to represent skilled forelimb movements during motor learning. However, it remains to be determined whether these changes persist over a long period and how these neurons contribute to skilled movements. Here, we show that a subset of M1 L5a IT neurons retain information for skilled forelimb movements even after non-training days. Furthermore, suppressing the activity of these neurons during skilled forelimb movements impaired behavioral stability and adaptability. Our results suggest the importance of M1 L5a IT neurons for tuning skilled forelimb movements over a long period.

  • Access to Document (DOI)

• Stimulated Raman scattering microscopy reveals a unique and steady nature of brain water dynamics.

  Takanori Shinotsuka, Tsuyoshi Miyazawa, Keiko Karasawa, Yasuyuki Ozeki, Masato Yasui, Mutsuo Nuriya

  Cell reports methods 3 ( 7 ) 100519 - 100519 2023.07

  Research paper (scientific journal), Joint Work, Lead author, Accepted

  　View Summary

  The biological activities of substances in the brain are shaped by their spatiotemporal dynamics in brain tissues, all of which are regulated by water dynamics. In contrast to solute dynamics, water dynamics have been poorly characterized, owing to the lack of appropriate analytical tools. To overcome this limitation, we apply stimulated Raman scattering multimodal multiphoton microscopy to live brain tissues. The microscopy system allows for the visualization of deuterated water, fluorescence-labeled solutes, and cellular structures at high spatiotemporal resolution, revealing that water moves faster than fluorescent molecules in brain tissues. Detailed analyses demonstrate that water, unlike solutes, diffuses homogeneously in brain tissues without differences between the intra- and the extracellular routes. Furthermore, we find that the water dynamics are steady during development and ischemia, when diffusions of solutes are severely affected. Thus, our approach reveals routes and uniquely robust properties of water diffusion in brain tissues.

  • Access to Document (DOI)

• Norepinephrine induces rapid and long-lasting phosphorylation and redistribution of connexin 43 in cortical astrocytes

  Mutsuo Nuriya, Ayaka Morita, Takanori Shinotsuka, Tomoko Yamada, Masato Yasui

  BIOCHEMICAL AND BIOPHYSICAL RESEARCH COMMUNICATIONS (ACADEMIC PRESS INC ELSEVIER SCIENCE)  504 ( 4 ) 690 - 697 2018.10

  Research paper (scientific journal), Joint Work, Accepted,  ISSN  0006291X

  　View Summary

  Norepinephrine (NE) modulates brain functions depending on both the internal and external environment. While the neuromodulatory actions of NE have been well characterized, the response and involvement of cortical astrocytes to physiological noradrenergic systems remain largely unknown, especially at the molecular level. In this study, we biochemically characterize the action of NE on astrocytes of the murine neocortex. NE stimulation of acute brain slices rapidly increase phosphorylation of connexin 43 (Cx43) at Serine (Ser) 368, in slices from both juvenile and adolescent animals. The phosphorylation is mediated by the protein kinase C (PKC) pathway under the alpha 1-adrenergic receptor and remains elevated for tens of minutes following brief exposure to NE, well after the intracellular calcium level returns to normal level, suggesting the plastic nature of this phosphorylation event. Importantly, this phosphorylation event persists in the absence of neuronal transmissions, suggesting that the effect of NE on Cx43 phosphorylation is induced directly on astrocytes. Furthermore, these NE-induced phosphorylations are associated with biochemical dissociation of Cx43 from gap-junctional plaques to non-junctional compartments. Finally, we show that pharmacological manipulation of the noradrenergic system using psychoactive drugs modulates phosphorylation of Cx43 in the cerebral cortex in vivo. These data suggest that NE acts directly on astrocytes in parallel with neurons and modulates functionally critical connexin channel proteins in a plastic manner. Thus, plasticity of astrocytes induced by the "gliomodulatory" actions of NE may play important roles in their physiological as well as pharmacological actions in the brain. (C) 2018 Elsevier Inc. All rights reserved.

  • Access to Document (DOI)

• Multimodal two-photon imaging using a second harmonic generation-specific dye

  Mutsuo Nuriya, Shun Fukushima, Atsuya Momotake, Takanori Shinotsuka, Masato Yasui, Tatsuo Arai

  NATURE COMMUNICATIONS (NATURE PUBLISHING GROUP)  7   11557 2016.05

  Research paper (scientific journal), Joint Work, Lead author, Accepted,  ISSN  2041-1723

  　View Summary

  Second harmonic generation (SHG) imaging can be used to visualize unique biological phenomena, but currently available dyes limit its application owing to the strong fluorescent signals that they generate together with SHG. Here we report the first non-fluorescent and membrane potential-sensitive SHG-active organic dye Ap3. Ap3 is photostable and generates SH signals at the plasma membrane with virtually no fluorescent signals, in sharp contrast to the previously used fluorescent dye FM4-64. When tested in neurons, Ap3-SHG shows linear membrane potential sensitivity and fast responses to action potentials, and also shows significantly reduced photodamage compared with FM4-64. The SHG-specific nature of Ap3 allows simultaneous and completely independent imaging of SHG signals and fluorescent signals from various reporter molecules, including markers of cellular organelles and intracellular calcium. Therefore, this SHG-specific dye enables true multimodal two-photon imaging in biological samples.

  • Access to Document (DOI)

• Sustained Down-regulation of beta-Dystroglycan and Associated Dysfunctions of Astrocytic Endfeet in Epileptic Cerebral Cortex

  Asako Gondo, Takanori Shinotsuka, Ayaka Morita, Yoichiro Abe, Masato Yasui, Mutsuo Nuriya

  JOURNAL OF BIOLOGICAL CHEMISTRY (AMER SOC BIOCHEMISTRY MOLECULAR BIOLOGY INC)  289 ( 44 ) 30279 - 30288 2014.10

  Research paper (scientific journal), Joint Work, Lead author, Accepted,  ISSN  0021-9258

  　View Summary

  Epilepsy is characterized by the abnormal activation of neurons in the cerebral cortex, but the molecular and cellular mechanisms contributing to the development of recurrent seizures are largely unknown. Recently, the critical involvement of astrocytes in the pathophysiology of epilepsy has been proposed. However, the nature of plastic modulations of astrocytic proteins in the epileptic cortex remains poorly understood. In this study, we utilized the zero magnesium in vitro model of epilepsy and examined the potential molecular changes of cortical astrocytes, focusing specifically on endfeet, where specialized biochemical compartments exist. We find that the continuous epileptic activation of neurons for 1 h decreases the expression level of beta-dystroglycan (beta DG) in acute cortical brain slices prepared from mice. This change is completely abolished by the pharmacological blockade of NMDA-type glutamate receptors as well as by matrix metalloproteinase inhibitors. Consistent with the highly specialized localization of beta DG at astrocytic endfeet, where it plays a pivotal role in anchoring endfeet-enriched proteins in astrocytes, the down-regulation of beta DG is accompanied by a decrease in the expression of AQP4 but not laminin. Importantly, this down-regulation of beta DG persists for at least 1 h, even after the apparent recovery of neuronal activation. Finally, we show that the down-regulation of beta DG is associated with the dysfunction of the endfeet at the blood-brain interface as a diffusion barrier. These results suggest that the sustained down-regulation of beta DG leads to dysfunctions of astrocytic endfeet in the epileptic cerebral cortex and may contribute to the pathogenesis of epilepsy.

  • Access to Document (DOI)

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Papers, etc., Registered in KOARA 【 Display / hide 】

Papers, etc., Registered in KOARA 【 Display / hide 】

• アストロサイトのギャップ結合ネットワークはin vitroの虚血モデルにおいて細胞傷害を軽減する

  Shinotsuka, Takanori

  (慶應義塾大学大学院医学研究科)  2014

• アストロサイトのギャップ結合ネットワークはin vitroの虚血モデルにおいて細胞傷害を軽減する

  Shinotsuka, Takanori

  (慶應義塾大学大学院医学研究科)  2014

• アストロサイトのギャップ結合ネットワークはin vitroの虚血モデルにおいて細胞傷害を軽減する

  Shinotsuka, Takanori

  (慶應義塾大学大学院医学研究科)  2014

Reviews, Commentaries, etc. 【 Display / hide 】

Reviews, Commentaries, etc. 【 Display / hide 】

• Visualization of water dynamics in brain tissue using multiphoton multimodal imaging

  Shinotsuka Takanori, Miyazawa Tsuyoshi, Karasawa Keiko, Ozeki Yasuyuki, Yasui Masato, Nuriya Mutsuo

  Proceedings for Annual Meeting of The Japanese Pharmacological Society (Japanese Pharmacological Society)  96   2-B-O05-1 2022

  　View Summary

  The flow of cerebrospinal fluid is considered to be a critical factor in the clearance of wastes in the brain. The dynamics of fluid has been investigated by several experimental methods such as fluorescence microscopy and magnetic resonance imaging. However, because probes used for these imagings are much larger compared to water itself, the dynamics of the fluid have been poorly understood. Here, we applied a multimodal multiphoton imaging system to the living brain tissue. Combining stimulated Raman scattering and two-photon fluorescent imaging, the system enables us to visualize spatiotemporal dynamics of deuterated water and fluorescent dyes simultaneously at a cellular level. We demonstrate that deuterated water diffuses faster than fluorescent dyes in the brain tissue. Detailed analysis reveals deuterated water rapidly exchanges inside and outside of cells, whereas fluorescent dyes only diffuse through extracellular spaces. Furthermore, we find that the dynamics of deuterated water is robust to changes under physiological and pathophysiological conditions; there is little change in the spatiotemporal dynamics of deuterated water during development and ischemia whereas fluorescent dyes are severely affected. Thus, our new approach reveals unique properties of the dynamics of the fluid in the living brain tissue.

  • Access to Document (DOI)

• Astrocytic gap junctional networks suppress cellular damages in an in vitro model of ischemia

  Takanori Shinotsuka, Masato Yasui, Mutsuo Nuriya

  JOURNAL OF PHARMACOLOGICAL SCIENCES (JAPANESE PHARMACOLOGICAL SOC)  124   74P - 74P 2014

  ISSN  1347-8613

• てんかんモデル条件下におけるアストロサイトの足突起のバリア機能の変化

  篠塚崇徳, 権藤麻子, 権藤麻子, 森田彩加, 森田彩加, 阿部陽一郎, 安井正人, 塗谷睦生, 塗谷睦生

  日本薬理学会関東部会プログラム・要旨集 131st 2014

• Astrocytic gap junctional networks reduce ischemic damages in vitro

  SHINOTSUKA Takanori, NURIYA Mutsuo, YASUI Masato

  日本神経化学会大会抄録集(Web) 56th 2013

• りん脂質修飾ナノ粒子の制御とそのマウス脳内における特異的分布(Controlling and spacific brain distribution of phospholipids-modified nanoparticles)

  篠塚 崇徳, 杉戸 雄四郎, 横田 理, 鈴木 健一郎, 武田 健

  神経化学 ((一社)日本神経化学会)  49 ( 2-3 ) 602 - 602 2010.08

  ISSN  0037-3796

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Research Projects of Competitive Funds, etc. 【 Display / hide 】

Research Projects of Competitive Funds, etc. 【 Display / hide 】

• 大脳皮質運動野の層構造依存的に符号化された運動記憶情報の長期安定性機構の解明

  2018.04

  -

  2021.03

  日本学術振興会, 科学研究費助成事業 特別研究員奨励費, 特別研究員奨励費, No Setting

  　View Summary

  本研究では学習によって形成される習熟運動が特定の神経細胞の神経活動として長期に安定に符号化されているかを検証してきた。その結果、習熟運動が時間経過に対し安定であること、さらにそのときの大脳皮質一次運動野 5a層の神経活動が安定であることを示唆する結果を得た。今年度はこれまでに得られた知見に関連した新たな知見を得るべく検証を進めた。具体的には習熟運動の学習に関連する分子メカニズムに関する検証を行った。また、習熟運動と大脳皮質一次運動野 5a層の神経活動の関係を細胞選択的な抑制・破壊実験を用いて検証した。
  習熟運動の学習に関連する分子メカニズムに関する検証として記憶・学習に重要な要素の一つであることが示唆されているNMDA受容体に着目した検証を行った。マウスが自発レバー引き運動課題を学習する訓練期間にNMDA受容体特異的な阻害薬を投与した結果、課題の学習が阻害された。このことから、習熟した運動を学習する分子メカニズムの一つとしてNMDA受容体が関与していることが示唆される。
  習熟運動と大脳皮質一次運動野 5a層の神経集団活動の関係性を検証するために細胞選択的な抑制・破壊実験を導入した。大脳皮質5a層特異的にCre遺伝子を発現するマウスの運動野前肢領域にアデノ随伴ウイルスによってCre依存的に特定のタンパク質を発現させることによって、光遺伝学による神経活動抑制またはジフテリアトキシンによる細胞死を誘発させる。これらの細胞選択的な抑制・破壊実験によって、大脳皮質一次運動野5a層の神経活動を特異的に抑制・破壊した場合に習熟運動がどのような影響を受けるかについて検証を進めている。
  今後これらの検証をさらに進め、大脳皮質一次運動野 5a層の神経集団活動と習熟運動の長期安定性についての知見を深めて研究成果を報告するための準備を進める。

• グルタミン酸とノルアドレナリンの協調的作用によるグリア細胞の脳血管制御機構の解明

  2014.04

  -

  2016.03

  日本学術振興会, 科学研究費助成事業 特別研究員奨励費, 特別研究員奨励費, No Setting

  　View Summary

  本研究は神経伝達物質のグルタミン酸とノルアドレナリンが、脳を構成する最も主要なグリア細胞であるアストロサイトの生理学的な応答を介して協調的に働き、脳血流制御を担っているのではないかという仮説を検証することを試みた。
  平成26年度としては、グルタミン酸とノルアドレナリンがアストロサイトの生理学的な変化に対して協調的に働くかどうかについて2光子顕微鏡イメージング法とマウス急性脳スライス法を用いたin vitroの実験系で検討を進めた。その結果、グルタミン酸とノルアドレナリンがアストロサイトの生理学的な変化に対し、アドレナリン受容体のサブタイプ依存的な協調作用を示すことが示唆された。一方、ここ数年で急性脳スライス法による実験は血流がないために脳血管の反応を十分に調べられない可能性が示唆され始めており、in vivoの実験系を導入することが急務であると考えられた。そこで、平成27年度としては実験手法として新たに覚醒動物を用いたin vivoの研究手法を導入することで更なる研究の発展を試みた。in vivo の研究手法を導入するに当たり運動学習中の大脳皮質一次運動野（M1）に着目し、マウス右前肢レバー引き学習試験中のM1での2光子顕微鏡イメージングと薬物の局所投与を組み合わせる実験系の開発に取り組んだ。薬物をM1に限定的に投与するために、M1にガイドカニューラを設置したマイクロポンプによる薬物投与を導入した。この時の脳血管の変化を2光子顕微鏡で観察するための開頭手術を行い、ガラスを設置した。カニューラから蛍光物質のAlexa 594をマイクロポンプによって投与した結果、ポンプから排出された蛍光物質の拡散が観察されたことから、M1に対する限定的な薬物処理を行いつつ脳血管の変化の変化をin vivoでリアルタイムに観察することができる可能性を示した。