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完整後設資料紀錄
DC 欄位 | 值 | 語言 |
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dc.contributor.advisor | 黃憲松(Hsien-Sung Huang) | |
dc.contributor.author | De-Fong Huang | en |
dc.contributor.author | 黃得峰 | zh_TW |
dc.date.accessioned | 2021-06-15T11:21:40Z | - |
dc.date.available | 2021-08-26 | |
dc.date.copyright | 2016-08-26 | |
dc.date.issued | 2016 | |
dc.date.submitted | 2016-08-18 | |
dc.identifier.citation | 1 Crick, F. Central dogma of molecular biology. Nature 227, 561-563 (1970).
2 Nilsen, T. W. & Graveley, B. R. Expansion of the eukaryotic proteome by alternative splicing. Nature 463, 457-463, doi:10.1038/nature08909 (2010). 3 Ahmari, S. E. et al. Repeated cortico-striatal stimulation generates persistent OCD-like behavior. Science 340, 1234-1239, doi:10.1126/science.1234733 (2013). 4 Darnell, R. B. RNA protein interaction in neurons. Annu Rev Neurosci 36, 243-270, doi:10.1146/annurev-neuro-062912-114322 (2013). 5 Doxakis, E. RNA binding proteins: a common denominator of neuronal function and dysfunction. Neurosci Bull 30, 610-626, doi:10.1007/s12264-014-1443-7 (2014). 6 Zhou, H., Mangelsdorf, M., Liu, J., Zhu, L. & Wu, J. Y. RNA-binding proteins in neurological diseases. Sci China Life Sci 57, 432-444, doi:10.1007/s11427-014-4647-9 (2014). 7 Zheng, S. & Black, D. L. Alternative pre-mRNA splicing in neurons: growing up and extending its reach. Trends Genet 29, 442-448, doi:10.1016/j.tig.2013.04.003 (2013). 8 Raj, B. & Blencowe, B. J. Alternative Splicing in the Mammalian Nervous System: Recent Insights into Mechanisms and Functional Roles. Neuron 87, 14-27, doi:10.1016/j.neuron.2015.05.004 (2015). 9 Jensen, K. B. et al. Nova-1 regulates neuron-specific alternative splicing and is essential for neuronal viability. Neuron 25, 359-371 (2000). 10 DeBoer, E. M. et al. Prenatal deletion of the RNA-binding protein HuD disrupts postnatal cortical circuit maturation and behavior. J Neurosci 34, 3674-3686, doi:10.1523/JNEUROSCI.3703-13.2014 (2014). 11 Duan, W. et al. Novel Insights into NeuN: from Neuronal Marker to Splicing Regulator. Mol Neurobiol 53, 1637-1647, doi:10.1007/s12035-015-9122-5 (2016). 12 Kim, K. K., Adelstein, R. S. & Kawamoto, S. Identification of neuronal nuclei (NeuN) as Fox-3, a new member of the Fox-1 gene family of splicing factors. J Biol Chem 284, 31052-31061, doi:10.1074/jbc.M109.052969 (2009). 13 Kuroyanagi, H. Fox-1 family of RNA-binding proteins. Cell Mol Life Sci 66, 3895-3907, doi:10.1007/s00018-009-0120-5 (2009). 14 Lovci, M. T. et al. Rbfox proteins regulate alternative mRNA splicing through evolutionarily conserved RNA bridges. Nat Struct Mol Biol 20, 1434-1442, doi:10.1038/nsmb.2699 (2013). 15 Gehman, L. T. et al. The splicing regulator Rbfox1 (A2BP1) controls neuronal excitation in the mammalian brain. Nat Genet 43, 706-711, doi:10.1038/ng.841 (2011). 16 Gehman, L. T. et al. The splicing regulator Rbfox2 is required for both cerebellar development and mature motor function. Genes Dev 26, 445-460, doi:10.1101/gad.182477.111 (2012). 17 Dredge, B. K. & Jensen, K. B. NeuN/Rbfox3 nuclear and cytoplasmic isoforms differentially regulate alternative splicing and nonsense-mediated decay of Rbfox2. PLoS One 6, e21585, doi:10.1371/journal.pone.0021585 (2011). 18 Kim, K. K., Nam, J., Mukouyama, Y. S. & Kawamoto, S. Rbfox3-regulated alternative splicing of Numb promotes neuronal differentiation during development. J Cell Biol 200, 443-458, doi:10.1083/jcb.201206146 (2013). 19 Lal, D. et al. RBFOX1 and RBFOX3 mutations in rolandic epilepsy. PLoS One 8, e73323, doi:10.1371/journal.pone.0073323 (2013). 20 Lucas, C. H., Calvez, M., Babu, R. & Brown, A. Altered subcellular localization of the NeuN/Rbfox3 RNA splicing factor in HIV-associated neurocognitive disorders (HAND). Neurosci Lett 558, 97-102, doi:10.1016/j.neulet.2013.10.037 (2014). 21 Utami, K. H. et al. Detection of chromosomal breakpoints in patients with developmental delay and speech disorders. PLoS One 9, e90852, doi:10.1371/journal.pone.0090852 (2014). 22 Amin, N. et al. Genetic variants in RBFOX3 are associated with sleep latency. Eur J Hum Genet, doi:10.1038/ejhg.2016.31 (2016). 23 Cooper, G. M. et al. A copy number variation morbidity map of developmental delay. Nat Genet 43, 838-846, doi:10.1038/ng.909 (2011). 24 Al Sufiani, F. & Ang, L. C. Neuropathology of temporal lobe epilepsy. Epilepsy Res Treat 2012, 624519, doi:10.1155/2012/624519 (2012). 25 Brenner, R. et al. BK channel beta4 subunit reduces dentate gyrus excitability and protects against temporal lobe seizures. Nat Neurosci 8, 1752-1759, doi:10.1038/nn1573 (2005). 26 Kralic, J. E., Ledergerber, D. A. & Fritschy, J. M. Disruption of the neurogenic potential of the dentate gyrus in a mouse model of temporal lobe epilepsy with focal seizures. Eur J Neurosci 22, 1916-1927, doi:10.1111/j.1460-9568.2005.04386.x (2005). 27 van Groen, T., Miettinen, P. & Kadish, I. The entorhinal cortex of the mouse: organization of the projection to the hippocampal formation. Hippocampus 13, 133-149, doi:10.1002/hipo.10037 (2003). 28 Amaral, D. G., Scharfman, H. E. & Lavenex, P. The dentate gyrus: fundamental neuroanatomical organization (dentate gyrus for dummies). Prog Brain Res 163, 3-22, doi:10.1016/S0079-6123(07)63001-5 (2007). 29 Hsu, D. The dentate gyrus as a filter or gate: a look back and a look ahead. Prog Brain Res 163, 601-613, doi:10.1016/S0079-6123(07)63032-5 (2007). 30 Scharfman, H. E. & Myers, C. E. Hilar mossy cells of the dentate gyrus: a historical perspective. Front Neural Circuits 6, 106, doi:10.3389/fncir.2012.00106 (2012). 31 Hosp, J. A. et al. Morpho-physiological criteria divide dentate gyrus interneurons into classes. Hippocampus 24, 189-203, doi:10.1002/hipo.22214 (2014). 32 Dudek, F. E. & Shao, L. R. Loss of GABAergic Interneurons in Seizure-induced Epileptogenesis. Epilepsy Curr 3, 159-161, doi:10.1046/j.1535-7597.2003.03503.x (2003). 33 Hofmann, G., Balgooyen, L., Mattis, J., Deisseroth, K. & Buckmaster, P. S. Hilar somatostatin interneuron loss reduces dentate gyrus inhibition in a mouse model of temporal lobe epilepsy. Epilepsia 57, 977-983, doi:10.1111/epi.13376 (2016). 34 Wang, H. Y. et al. RBFOX3/NeuN is Required for Hippocampal Circuit Balance and Function. Sci Rep 5, 17383, doi:10.1038/srep17383 (2015). 35 Casanova, E. et al. A CamKIIalpha iCre BAC allows brain-specific gene inactivation. Genesis 31, 37-42 (2001). 36 Harris, J. A. et al. Anatomical characterization of Cre driver mice for neural circuit mapping and manipulation. Front Neural Circuits 8, 76, doi:10.3389/fncir.2014.00076 (2014). 37 Judson, M. C. et al. GABAergic Neuron-Specific Loss of Ube3a Causes Angelman Syndrome-Like EEG Abnormalities and Enhances Seizure Susceptibility. Neuron 90, 56-69, doi:10.1016/j.neuron.2016.02.040 (2016). 38 McHugh, T. J. et al. Dentate gyrus NMDA receptors mediate rapid pattern separation in the hippocampal network. Science 317, 94-99, doi:10.1126/science.1140263 (2007). 39 Coulter, D. A. & Carlson, G. C. Functional regulation of the dentate gyrus by GABA-mediated inhibition. Prog Brain Res 163, 235-243, doi:10.1016/S0079-6123(07)63014-3 (2007). 40 Paz, J. T. & Huguenard, J. R. Microcircuits and their interactions in epilepsy: is the focus out of focus? Nat Neurosci 18, 351-359, doi:10.1038/nn.3950 (2015). 41 Savanthrapadian, S. et al. Synaptic properties of SOM- and CCK-expressing cells in dentate gyrus interneuron networks. J Neurosci 34, 8197-8209, doi:10.1523/JNEUROSCI.5433-13.2014 (2014). 42 Liu, Y. C., Cheng, J. K. & Lien, C. C. Rapid dynamic changes of dendritic inhibition in the dentate gyrus by presynaptic activity patterns. J Neurosci 34, 1344-1357, doi:10.1523/JNEUROSCI.2566-13.2014 (2014). 43 Hsu, T. T., Lee, C. T., Tai, M. H. & Lien, C. C. Differential Recruitment of Dentate Gyrus Interneuron Types by Commissural Versus Perforant Pathways. Cereb Cortex 26, 2715-2727, doi:10.1093/cercor/bhv127 (2016). | |
dc.identifier.uri | http://tdr.lib.ntu.edu.tw/jspui/handle/123456789/49272 | - |
dc.description.abstract | 二十年以來神經元特異性核抗原(Neuronal nuclei, NeuN)被廣泛地使用於神經元的標定。直到2009年,Kim等人確定它是RBFOX3,為具RNA結合能力的FOX蛋白質家族的其中一員。FOX蛋白質家族是一群調控前信使RNA(pre-mRNA)選擇性剪切的蛋白質。近幾年來已經有數個研究指出FOX蛋白質家族的RBFOX1和RBFOX2在腦中所扮演的角色,而過去人類疾病的臨床研究也發現RBFOX3的突變和癲癇、認知障礙、語言及發展遲緩、長睡眠遲滯期和一些自閉症症狀相關。雖然NeuN已經被確認是RBFOX3,它在腦中的生理功能還未能清楚,因此我們使用Rbfox3基因剔除小鼠來研究它在腦中所扮演的角色。我們實驗室過去發現全身性Rbfox3基因剔除小鼠比起野生型小鼠容易引發癲癇症狀,以及有異常的突觸神經傳遞。Rbfox3基因剔除小鼠的海馬迴齒狀回顆粒細胞的微小興奮性和抑制性突觸後電流(mEPSC和mIPSC)表現出頻率的增加,而其電流大小不變。神經突觸傳遞對於神經訊號的傳遞是相當重要的,因此我試著深入研究RBFOX3在神經迴路中所扮演的角色並想找出過去發現Rbfox3基因剔除缺陷的細胞和分子機制。我發現Rbfox3基因剔除小鼠齒狀回顆粒細胞的樹突棘、興奮性神經突觸數量增加,以及樹突長度增加、型態更加複雜。此外,藉由在特定類型神經細胞的Rbfox3基因剔除小鼠,可以讓我們進行Rbfox3基因的遺傳剖析。在抑制性神經細胞特異性地剔除Rbfox3基因的小鼠會有嚴重的自發性癲癇症狀及未成熟時的死亡現象。抑制性神經細胞特異性地Rbfox3基因剔除小鼠的齒狀回顆粒細胞之微小抑制性突觸後電流表現出頻率的增加,而其電流大小不變。興奮性神經細胞和齒狀回顆粒細胞特異性地Rbfox3基因剔除小鼠的齒狀回顆粒細胞之微小抑制性突觸後電流也表現出頻率些微的增加。這些結果說明了RBFOX3在齒狀回迴路的突觸前和突觸後細胞均在正常神經突觸傳遞功能上扮演著一定的角色。另外在生理情況下,抑制性神經細胞特異性地失去Rbfox3基因會造成神經迴路的興奮性過高。這個研究可以幫助我們更加了解RBFOX3在海馬迴神經迴路、Rbfox3基因缺失造成的癲癇症狀之產生所扮演的角色,以及RNA選擇性剪切調控蛋白(如RBFOX3)如何在腦中進行功能。 | zh_TW |
dc.description.abstract | Neuronal nuclei (NeuN), a neuronal nuclear antigen, is widely used to label neurons for over 20 years. In 2009, Kim et al. identified that NeuN corresponds to a protein known as RBFOX3 (RNA binding protein fox-1 homolog 3), one of the FOX family proteins. FOX proteins are a family of regulators that control the pre-mRNA alternative splicing. In recent studies, the roles in the brain of other FOX family members, RBFOX1 and RBFOX2, have been investigated. Previous research also found that RBFOX3 mutations are linked to human epilepsy, cognitive impairment, speech disorder, developmental delay, long sleep latency and some autistic features. Although NeuN has been identified as RBFOX3, it is still unclear what physiological roles it plays in the brain. Therefore, we use Rbfox3 knockout mice model to investigate the roles of RBFOX3 in the brain. The previous findings in our lab indicated that conventional Rbfox3-/- mice had increased seizure susceptibility and abnormal synaptic transmission. The dentate gyrus granule cells (DGGCs) of Rbfox3-/- mice displayed both increased frequency, but not amplitude, of excitatory and inhibitory postsynaptic current (mEPSC and mIPSC). Synaptic transmission is very important to neuronal signal transduction. For this reason, I attempt to further study the roles of RBFOX3 in neuronal circuitry and to find out the cellular and molecular mechanisms underline previous findings. I found that the DGGCs of Rbfox3-/- mice exhibited increased spine density, excitatory synapse number, and dendritic complexity. Furthermore, with the different kinds of cell type-specific conditional Rbfox3 knockout mice, this can allow us to enable genetic dissection of Rbfox3. Specific deletion of Rbfox3 in inhibitory interneurons caused spontaneous seizure and premature death. The DGGCs of Gad2-Cre::Rbfox3loxP/loxP conditional KO mice displayed increased frequency, but normal amplitude, of mIPSC. Specific deletion of RBFOX3 in excitatory neurons and specific deletion of RBFOX3 in the DGGCs also displayed slight increased mIPSC frequency in DGGCs. These results indicated that the RBFOX3 in presynaptic and postsynaptic neurons of the dentate gyrus circuitry play a role in normal function of synaptic transmission. Moreover, in the physiological condition, GABAergic neuron- specific loss of Rbfox3 cause circuitry hyper-excitability. This study can help us more understand the roles of RBFOX3 in hippocampal circuitry, seizure pathogenesis of Rbfox3 defect, as well as how the splicing regulators such as RBFOX3 operate in the brain. | en |
dc.description.provenance | Made available in DSpace on 2021-06-15T11:21:40Z (GMT). No. of bitstreams: 1 ntu-105-R03454012-1.pdf: 5937340 bytes, checksum: dbd1e45aa891c00362320ed070d5820b (MD5) Previous issue date: 2016 | en |
dc.description.tableofcontents | 口 試 委 員 會 審 定 書 i
誌 謝 ii 中 文 摘 要 iii Abstract iv Table of Contents vi List of Figures ix Chapter 1: Introduction 1 1.1 RNA alternative splicing and alternative splicing regulators 1 1.2 Background knowledge of RBFOX3 2 1.3 The hippocampal circuitry and the dentate gyrus granule cell (DGGC) 3 1.4 Previous studies and its limitations 4 1.5 Objectives of this study 4 Chapter 2: Material and Methods 6 2.1 Mice 6 2.2 Golgi-Cox staining 8 2.3 Immunofluorescence staining 9 2.4 Synapse number counting 9 2.5 Fluorescence-activated cell sorting (FACS) 10 2.6 RNA isolation 11 2.7 Quantitative real-time PCR (Q-RT-PCR) 11 2.8 Spontaneous seizure behavior experiment 12 2.9 Hippocampal slice preparation 12 2.10 Electrophysiology 12 2.11 Statistical analysis 15 Chapter 3: Results 16 3.1 Rbfox3 deletion didn’t affect the mRNA expression level of other Rbfox family members and glutamate ionotropic receptor NMDA type subunit 2b (Grin2b) 16 3.2 The dendritic spine density of the DGGCs was increased in Rbfox3 conventional knockout mice 16 3.3 The dendritic length of DGGCs was increased in Rbfox3 conventional knockout mice 17 3.4 Rbfox3 conventional knockout mice had more excitatory synapses in the DGGCs area 17 3.5 Validation of cell type-specific Rbfox3 conditional knockout (Rbfox3 cKO) mice 18 3.6 Low survival rate in Gad2-Cre::Rbfox3loxP/loxP cKO mice 18 3.7 Gad2-Cre::Rbfox3loxP/loxP mice exhibited spontaneous seizure 19 3.8 Miniature excitatory postsynaptic currents of the DGGCs of Rbfox3 cKO mice 19 3.9 Miniature inhibitory postsynaptic currents of the DGGCs of Rbfox3 cKO mice 20 3.10 Intrinsic excitability of the DGGCs of Gad2-Cre::Rbfox3loxP/loxP cKO mice 20 3.11 Intrinsic excitability of the DGGCs of Rbfox3 cKO mice in physiological condition 21 3.12 Evoked postsynaptic potential (ePSP) of the DGGCs of Rbfox3 cKO mice 21 Chapter 4: Discussion 23 Reference 64 | |
dc.language.iso | en | |
dc.title | 探索RBFOX3 (NeuN)在小鼠海馬迴神經迴路中所扮演的角色 | zh_TW |
dc.title | Investigating the Roles of RBFOX3 (NeuN) in the Mouse Hippocampal Circuitry | en |
dc.type | Thesis | |
dc.date.schoolyear | 104-2 | |
dc.description.degree | 碩士 | |
dc.contributor.oralexamcommittee | 李立仁(Li-Jen Lee),郭鐘金(Chung-Chin Kuo),薛一蘋(Yi-Ping Hsueh) | |
dc.subject.keyword | RBFOX3,海馬迴神經迴路,神經突觸傳遞,自發性癲癇,興奮性過高, | zh_TW |
dc.subject.keyword | RBFOX3 (NeuN),Hippocampal circuitry,Synaptic transmission,Spontaneous seizure,Hyperexcitability, | en |
dc.relation.page | 68 | |
dc.identifier.doi | 10.6342/NTU201603354 | |
dc.rights.note | 有償授權 | |
dc.date.accepted | 2016-08-19 | |
dc.contributor.author-college | 醫學院 | zh_TW |
dc.contributor.author-dept | 腦與心智科學研究所 | zh_TW |
顯示於系所單位: | 腦與心智科學研究所 |
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