| 研究課題/領域番号 |
22KF0211
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| 補助金の研究課題番号 |
22F22753 (2022)
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| 研究種目 |
特別研究員奨励費
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| 配分区分 | 基金 (2023)
補助金 (2022) |
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| 応募区分 | 外国 |
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| 審査区分 |
小区分46010:神経科学一般関連
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| 研究機関 | 京都大学 |
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研究代表者 |
林 康紀 京都大学, 医学研究科, 教授 (90466037)
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| 研究分担者 |
YUEN JESSICA 京都大学, 医学研究科, 外国人特別研究員
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| 研究期間 (年度) |
2023-03-08 – 2025-03-31
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| 研究課題ステータス |
交付 (2023年度)
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| 配分額 *注記 |
2,200千円 (直接経費: 2,200千円)
2024年度: 500千円 (直接経費: 500千円)
2023年度: 1,100千円 (直接経費: 1,100千円)
2022年度: 600千円 (直接経費: 600千円)
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| キーワード | 記憶固定化 |
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| 研究開始時の研究の概要 |
I set specific aims of this research project as follows:
SA1. To develop a new optogenetic tool that allows us to specifically erase long-term potentiation (LTP) with a defined spatiotemporal window.
SA2. To identify the time window of offline-LTP required for long-term consolidation of memory.
SA3. To identify the cortical circuit modification induced by offline-LTP.
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| 研究実績の概要 |
Synaptic localization of endogenous cofilin fused to green fluorescent protein (GFP) was visualized with single synapse resolution in hippocampal CA1 neurons of transgenic mice. The expression of our cofilin-GFP is strictly under the control of Cre recombinase, thus circumventing the problem of protein overexpression via exogenously applied cofilin and allows for direct visualization of cofilin localization in neurons. Structural long-term potentiation (sLTP) was induced by two-photon uncaging of glutamate to confirm endogenous cofilin only accumulates in stimulated spines. The results were in line with the laboratory’s previous publications that showed exogenously administered cofilin accumulates in enlarged dendritic spines following induction of LTP and is involved in memory formation and consolidation in vivo. Next, we sparsely labeled hippocampal CA1 neurons by injection of an adeno-associated virus vector containing iCre to activate expression of endogenous cofilin and mRuby3 as a volume filler. We then utilized the unique properties of our endogenous cofilin to quantify online LTP. Mice were sacrificed following inhibitory avoidance learning for brain sectioning and target hippocampal dendritic spines were analyzed for the ratio of cofilin-GFP to mRuby3 labeling. Results were compared against control mice that did not receive footshocks. After normalizing fluorescence across the groups, we identified spines that have enlarged following learning and thus serve as an indicator of online LTP.
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| 現在までの達成度 (区分) |
現在までの達成度 (区分)
3: やや遅れている
理由
Jessica is taking too long time expand animal colonies, which could have been facilitated by using in vitro fertilization. Also, she does not have sufficient knowledge on recombinant DNA technique.
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| 今後の研究の推進方策 |
Presently, we are attempting to establish a link between online LTP and the accumulation of cofilin in dendritic spines in vivo. The goal is to use cofilin as a synaptic tag to chronically track individual spines that have undergone LTP. We will surgically implant microprisms in the CA1 of our transgenic mice for live imaging of hippocampal apical dendrites. Using the quantification measures obtained in our previous experiments, we can identify individual spines that have undergone LTP and track these spines over months. This method not only allows for chronic imaging of spines prior to and after a learning task at various time points, but also captures the ensuing offline LTP after sleep. The research will provide unprecedented spatiotemporal imaging of both online and offline LTP using endogenous cofilin as a synaptic marker, which has yet to be achieved with existing imaging methods.
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