| 研究課題/領域番号 |
18K03168
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| 研究機関 | 筑波大学 |
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研究代表者 |
Pavlides C 筑波大学, 人間系, 教授 (50712808)
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| 研究期間 (年度) |
2018-04-01 – 2021-03-31
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| キーワード | Hippocampus / fear memory / spatial memory / immediate early genes / Zif268 / functional organization |
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| 研究実績の概要 |
The proposed studies were aimed at determining neuronal functional organization in the hippocampus for fear memory. Previously, we observed a topographic organization in the hippocampus in animals engaged in both spatial as well as sequential order memory tasks. That is, pyramidal cells in the dorsal hippocampus formed clusters of a few adjacent active cells. This is contrary to the prevalent view of a random, distributed organization. However, the previous studies included only the dorsal hippocampus and using 2-D analysis. For the present studies, the behavioral paradigm was fear conditioning (another hippocampus dependent task), and we performed a 3-D reconstruction of the entire hippocampus. Based on previous reports showing that the proximal and distal parts of the hippocampus may be organized differently both in terms of inputs as well as functionally, we analyzed each subregion separately in terms of topographic organization.
To allow for better 3-D imaging we initially tried to use a tissue clearing method (Cubic) that would allow us to visualize the entire hippocampus without sectioning. In our hands, however, this proved not to be a suitable solution. Granted this method has certain advantages, it produced significant tissue swelling which did not allow for precise reconstruction of the topographic organization. Instead we switched back to sectioning of the brain tissue and fluorescent immunohistochemistry (IHC).
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| 現在までの達成度 (区分) |
現在までの達成度 (区分)
2: おおむね順調に進展している
理由
Thus far, we have completed one set of experiments in animals trained in contextual fear conditioning and have processed the dorsal hippocampus for Zif268 IHC. 3-D analysis revealed that a cluster type organization took place both in the dorsal proximal-distal and longitudinal axis (Arai and Pavlides, 2019a; Arai and Pavlides, 2019b). Using a 3-D reconstruction we also observed clustering in the longitudinal axis of the dorsal hippocampus (Arai and Pavlides, 2018). This study is still preliminary in that more subjects will have to be included. Further, we would like to compare functional organization using contextual (hippocampus dependent) and cued (hippocampus independent) tasks.
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| 今後の研究の推進方策 |
During the second year of funding, we will complete the ongoing experiments to include a larger number of subjects as well as cued conditioned groups. Further, we will image both proximal-distal parts of CA1 and CA3 and dorsal-ventral sections. The main objective of the second year will be to use optogenetic stimulation to activate/suppress entorhinal cortex afferents to hippocampus to determine effects on neuronal functional organization and behaviors. We have started with viral injections to optimize parameters - including volume and brain sites. We have also started with microprobe surgeries to determine effectiveness of optogenetic stimulation parameters. The experiments should be completed during the second year.
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| 次年度使用額が生じた理由 |
Part of the funds were to be used for the purchase of Virus constructs as well as confocal microscopy time, both of which are rather expensive. We are getting to that part of the studies in the second year. Therefore, the funds will be used during the second year of funding. Also, both the immunohistochemistry and optogenetic stimulation will require substantial supplies, especially since the number of animals to be used and brain tissue to be processed will be substantially more than in the previous year.
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