| 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 – 2022-03-31
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| キーワード | hippocampus / memory / immediate early genes / Zif268 / functional organization / sleep / spatial navigation / place cells |
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| 研究実績の概要 |
There were two aims to the proposed studies: the first was to determine whether there is an underlying functional neuronal organization in the hippocampus to encode different types of memories. We tested animals behaviorally in spatial, sequential, and fear memory tasks and determined neuronal activity through the use of the immediate early gene Zif268. We found that indeed the hippocampus was functionally organized in 'cell clusters' consisting of a few closely located active cells in both the CA1 and CA3 fields for each of the memory tasks. Two manuscripts have been recently completed and are in the process of being published.
The second aim was to determine mechanisms underlying the 'cluster-type' organization by optogenetically exciting/inhibiting the main inputs to the hippocampus arising from the entorhinal cortex (EC) and determining effects on the cluster formation. For this, we developed a behavioral task which combines spatial (context) and object components which are known to be processed by the medial and lateral entorhinal cortex, respectively. The medial and lateral EC in turn project to distinct parts of the hippocampus which will guide us to search for topographic changes. Thus far, we have performed the majority of the behavioral tasks successfully . Further, we have started the anatomical analysis of both EC activation/inactivation and hippocampal topographic organization using Zif268 expression. This analysis is very time consuming and will require further effort and time to complete.
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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
Significant progress has been made on virus injections/optogenetic stimulation and behavioral testing/results, none of which is trivial - hitting the right MEC/LEC injection sites, recording cellular activity to determine light stimulation effectiveness, etc. proved to be rather challenging. Effective optogenetic stimulation parameters necessary to affect a sufficient area/cells had to also be determined. These aspects of the study have now been successfully accomplished and results have been obtained.
Progress has also been made, although at a slower pace, for the second part of the study, i.e., to determine functional neuronal organization in the hippocampus following inhibition/activation of the MEC/LEC. We have, thus far, determined that light stimulation produces desired effects by showing a suppression in Zif268 expression at the stimulation site. Further, we have collected a number of brain samples from the behavioral experiments and are in the process of performing the immunohistochemistry/histological analysis to determine effects on neuronal topographic organization - i.e., cell clusters. This analysis is very time consuming and will require further effort/time to complete.
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| 今後の研究の推進方策 |
Completion of the studies will require addition of a few animals to the behavioral study for statistical analysis. A good part of the behavioral analysis has already been completed and has produced expected results. We have also worked out all the details with regards to virus injections, stimulation parameters, etc. Still the anatomical analysis will require considerable effort which will be continued in the upcoming year. This will require processing the brains histologically including sectioning, immunohistochemistry, microscopic image capture and analysis as well as determining topographic organization in different parts of the hippocampus and perhaps EC. Our predicted outcome from the behavioral experiments is that inhibition of MEC/LEC should induce a behavioral dissociation with the former affecting spatial while the latter affecting object memory. If we confirm this, we further predict that the restricted MEC/LEC inhibition should differentially affect cell clustering in the hippocampus. Time permitting, we would like to proceed with a double labeling experiment using CatFISH Zif268 immunohistochemistry to more definitively identify different cell clusters.
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| 次年度使用額が生じた理由 |
The carry over amount will be used for the purchase of animals and IHC/image analysis. Immunohistochemistry supplies, including antibodies, reagents, etc are somewhat expensive. Funds have also been set aside for doing the CatFISH immunohistochemistry which will require additional costs both for supplies as well as travel to Hoygo College of Medicine for doing some of the procedures. We will also need funds to hire an assistant (20hr/week, for 8month). We are also anticipating a number of publications which should be completed during next year's budget period which will require substantial publication costs.
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