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| DC 欄位 | 值 | 語言 |
|---|---|---|
| dc.contributor.advisor | 陳示國(Shih-Kuo Chen) | |
| dc.contributor.author | Yu-Yau Shan | en |
| dc.contributor.author | 單禹堯 | zh_TW |
| dc.date.accessioned | 2023-03-20T00:06:24Z | - |
| dc.date.copyright | 2022-08-31 | |
| dc.date.issued | 2022 | |
| dc.date.submitted | 2022-08-08 | |
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| dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/86611 | - |
| dc.description.abstract | 地球上的生物依靠體內的晝夜節律以應對二十四小時的日夜週期變化，而在哺乳類動物中，下視丘的視交叉上核(Suprachiasmatic nucleus, SCN)作為中樞節律器調控眾多生理功能。現代社會中人們頻繁地到世界各地旅遊並跨越不同時區，因此常受時差(jet lag)影響產生睡眠障礙。過去對於時差的研究使用活體外腦片紀錄的方式，發現視交叉上核中的神經細胞的時鐘基因在經歷時差後變得較不同步，並需要幾天的時間逐漸重新適應到新的時區。然而究竟在活體內視交叉上核內的神經活性如何被時差影響仍然未知。藉由在梯度折射率透鏡(gradient index lens, GRIN Lens)、鈣離子影像技術及雙光子顯微鏡，我們得以在清醒小鼠內紀錄視交叉上核內的神經活性。我們發現視交叉上核內神經的鈣離子活動相關性會迅速被外界光線提升，且不會在經歷時差後有所降低；另一方面視交叉上核神經的基礎鈣離子濃度原本具有日夜週期的變化，但卻會在經歷時差後被擾亂，且部分神經會重新適應到新時區。而無法重新適應的神經們彼此之間又具有較強的鈣離子活動相關性，因此他們可能是屬於同一未知功能的神經迴路。綜上所述我們的結果顯示鈣離子活動和基礎鈣離子濃度應該是被兩種不同機制調控，其中後者可能是由生理時鐘基因調控，並負責同步行為與外界光週期。 | zh_TW |
| dc.description.abstract | Living creatures on Earth rely on their intrinsic circadian rhythms to anticipate the external day and night cycle. The suprachiasmatic nucleus (SCN) in the mammalian hypothalamus serves as the central pacemaker to coordinate individual periphery oscillators and govern various physiological functions critical to health. When rapidly crossing several time zones by flight, people often experience jet lag and suffer unsettled sleep-wake cycles. A previous study using in vitro single-cell imaging has revealed that the phases of PER2, a core clock gene, expressions in the SCN neurons become less synchronized after jet lag and require a couple of days to gradually be re-entrained to the new light-dark cycle. However, further in vivo evidence about how jet lag affects SCN neuronal activities is still deficient. In this study, by a combination of gradient index (GRIN) lenses, Ca2+ imaging, and two-photon microscopy, we were able to record single-cell SCN neuronal activities in head-fixed awake mice. We found that the synchrony of transient Ca2+ activities of SCN neurons was acutely enhanced by external light and not altered after jet lag exposure; on the other hand, basal Ca2+ levels of most neurons originally exhibiting daily fluctuations higher in the daytime were disturbed by jet lag, and some neurons achieved re-entrainment of their basal Ca2+ fluctuations to the new light-dark cycle. Moreover, those SCN neurons incapable of re-entrainment exhibited stronger correlations of transient Ca2+ activities to each other, implying they might belong to the same circuit with unknown functions instead of photoentrainment. Together, our findings demonstrate that transient Ca2+ activities and basal Ca2+ levels could be controlled by different mechanisms, with the former acutely responding to light input and the latter probably mediated by clock genes and accounting for photoentrainment of behavior rhythms. | en |
| dc.description.provenance | Made available in DSpace on 2023-03-20T00:06:24Z (GMT). No. of bitstreams: 1 U0001-0208202217072500.pdf: 5173743 bytes, checksum: 9ec177eba958b76f56d11f47cb9c5ccf (MD5) Previous issue date: 2022 | en |
| dc.description.tableofcontents | 論文口試委員審定書 i 誌 謝 ii 摘 要 iv Abstract v Contents vii Chapter I Introduction 1 1.1 Circadian rhythms 1 1.1.1 Molecular clock 2 1.2 Suprachiasmatic nucleus: central pacemaker of mammals 4 1.2.1 Structure and subpopulations 4 1.2.2 Rhythmicity of the SCN 5 1.2.3 Synchrony of the SCN 7 1.2.4 Retinal input and photoentrainment 8 1.2.5 Output 9 1.3 Jet lag 10 1.3.1 Circadian disruption in jet lag 11 1.4 In vivo Ca2+ imaging 11 1.4.1 GCaMP 11 1.4.2 Two-photon microscopy 12 1.4.3 Gradient index (GRIN) lenses 13 Statement of Purpose 15 Chapter II Materials and Methods 17 2.1 Animals 17 2.2 Genotyping 17 2.2.1 DNA extraction 17 2.2.2 Polymerase chain reaction (PCR) 18 2.3 Stereotaxic surgery 18 2.4 Experimental design 20 2.5 Two-photon microscopic image acquisition 21 2.6 Image analysis and statistics 21 2.6.1 Motion correction and ROI definition 22 2.6.2 Production of fluorescence intensity traces, detrend and ΔF/F0 definition 22 2.6.3 Correlation analysis 23 2.6.4 K-means clustering analysis on basal Ca2+ fluctuations 24 Chapter III Results 25 3.1 SCN neuronal activities were successfully visualized by in vivo Ca2+ imaging through GRIN lenses with two-photon microscopy 25 3.2 The correlations among transient Ca2+ activities of SCN neurons showed variation relative higher during the day and reducing right after nightfall but were not affected by jet lag 26 3.3 Basal Ca2+ levels of SCN neurons exhibited fluctuations which were altered after jet lag with some neurons successfully re-entrained to the new LD schedule 27 3.4 The neuron group failing to accomplish re-entrainment displayed significantly stronger intra-cluster correlations than inter-cluster correlations 29 3.5 The SCN neurons within the same groups seemed to be spatially close to each other 30 Chapter IV Discussion 32 4.1 The transient Ca2+ activities and basal Ca2+ fluctuations in SCN neurons could be modulated by distinct mechanisms 32 4.2 The SCN neurons achieved re-entrainment of their basal Ca2+ rhythms might drive the photoentrainment of behaviors 33 4.3 The SCN neurons with basal Ca2+ fluctuations unable to be re-entrained might belong to a dissociable circuit with distinct functions instead of photoentrainment 35 Significance of the work 37 References 38 Figures 52 Figure 1. Experimental design. 52 Figure 2. Schematic of GRIN lens implantation and AAV injection stereotaxic surgery. 53 Figure 3. Average images though GRIN lenses in VIPcre/+; and AVPcre/+; tdTomato/+ mice. 54 Figure 4. Illustrations for data analysis. 55 Figure 5. Population correlation coefficients among SCN neuronal transient Ca2+ activities before and after jet lag. 56 Figure 6. Heat map of basal Ca2+ levels on Baseline. 57 Figure 7. Heat maps of basal Ca2+ levels on Baseline2 and Day1-4. 58 Figure 8. Heap maps of correlations between Baseline2 and Day1-4. 59 Figure 9. K-means clustering heat maps of basal Ca2+ levels on Baseline2 and Day1-4. 60 Figure 10. Correlation coefficients among transient Ca2+ activities of SCN neurons within C1 to C5 before and after jet lag. 61 Figure 11. Averages of daytime or nighttime correlation coefficients among transient Ca2+ activities of SCN neurons within C1 to C5 before and after jet lag. 62 Figure 12. Correlation coefficients among transient Ca2+ activities of SCN neurons within same group and between another group before and after jet lag. 63 Figure 13. Intra- or inter-group correlation coefficients of transient Ca2+ activities during daytime and nighttime. 65 Figure 14. Spatial distributions of different clusters and groups. 66 Figure 15. VIP and AVP neurons distributions in different clusters and groups. 67 Figure 16. Graphic summary. 68 Tables 69 Table 1. List of primers for genotyping. 69 Appendix 70 Poster for 2020 Taiwan Society for Neuroscience (TSFN) 70 Poster for 2022 Poster Contest, Department of Life Science, National Taiwan University 71 Programing design for detrend, ΔF/F0 and PCCs calculation 72 Programing design for K-means clustering analysis 75 | |
| dc.language.iso | en | |
| dc.title | 藉由內視鏡在活體內觀察經歷時差時視交叉上核之神經活性 | zh_TW |
| dc.title | Visualizing the suprachiasmatic nucleus in vivo during jet lag by intracranial endoscope implantation | en |
| dc.type | Thesis | |
| dc.date.schoolyear | 110-2 | |
| dc.description.degree | 碩士 | |
| dc.contributor.author-orcid | 0000-0003-4157-9250 | |
| dc.contributor.oralexamcommittee | 吳順吉(Shun-Chi Wu),吳玉威(Yu-Wei Wu),黃雯華(Wendy Hwang-Verslues) | |
| dc.subject.keyword | 晝夜節律,時差,視交叉上核,活體鈣離子影像,梯度折射率透鏡,雙光子顯微鏡, | zh_TW |
| dc.subject.keyword | circadian rhythms,jet lag,suprachiasmatic nucleus,in vivo Ca2+ imaging,GRIN lenses,two-photon microscopy, | en |
| dc.relation.page | 79 | |
| dc.identifier.doi | 10.6342/NTU202201980 | |
| dc.rights.note | 同意授權(全球公開) | |
| dc.date.accepted | 2022-08-08 | |
| dc.contributor.author-college | 生命科學院 | zh_TW |
| dc.contributor.author-dept | 生命科學系 | zh_TW |
| dc.date.embargo-lift | 2027-08-26 | - |
| 顯示於系所單位： | 生命科學系 | |
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