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    Title: 鈣離子在斑馬魚早期肌肉發育過程中所扮演的角色
    Other Titles: Calcium Plays a Role during Zebrafish Myogenesis
    Authors: 陳曜鴻
    Contributors: 淡江大學化學學系
    Date: 2012-08
    Issue Date: 2015-05-05 16:47:47 (UTC+8)
    Abstract: 在成體的肌肉細胞中,神經的活性會調節肌肉的活性。D-serine 是一種神經傳導 物質,會活化NMDA 接受體(NMDAR),過多的D-serine 會被D-型胺基酸氧化酶(DAO) 所分解。DAO, D-serine, NMDAR 三者之間的互動,會調控成體肌肉細胞內鈣離子濃度, 進而調節肌肉的生理功能。然而,胚胎時期肌肉形成過程中,肌肉芽細胞是否也受相同 路徑而調控肌肉細胞分化,目前科學界所知有限。最近的研究發現,NMDA 接受體除 了在中樞神經系統扮演重要的功能之外,也被發現會表現在體外培養的小鼠肌肉芽細胞 中,據推測,應該在胚胎時期肌肉發育過程中,扮演重要的角色。這些研究項目十分繁 重,故擬分三年來完成下列目標。 第一年計畫內容涵蓋:(1) DAO 抗體製作;(2) 抗體免疫螢光染色實驗的建立;(3) 苯甲酸鈉的浸泡與D-serine 的注射;(4) 胚胎發育抑制劑的注射;(5)共軛焦顯微鏡影像 分析;(6)電子顯微鏡成像。 第二年計畫內容涵蓋:(1)突變魚種nic-1 的飼育;(2) 胚胎發育抑制劑的注射;(3) 細胞內鈣離子濃度的即時動態偵測;(4) 胚胎運動能力分析;(5) MK801 與NMDA 的浸 泡實驗。 第三年計畫內容涵蓋:(1) 2-APB, 咖啡因與ryanodine 的浸泡實驗;(2)全胚胎原位 雜交實驗;(3) 抗體免疫螢光染色實驗的建立;(4) 切片技術與組織染色;(5) 細胞骨架 phallodin 螢光染劑染色;(6) 成魚游泳能力分析。
    Nerve activity is known to be an important regulator of muscle phenotype in the adult, but its contribution to muscle development during embryogenesis remains unresolved. We try to use the zebrafish embryo and in vivo imaging approaches to address the role of activity generated signals, D-serine, D-amino acids oxidase (DAO), NMDA-R and intracellular calcium, in vertebrate slow muscle development. Our proposal will be including: The first year: (1) producing the anti-DAO antibody; (2) using antibodies staining to study the endogenous protein levels; (3) transferring sodium benzoate and D-serine to zebrafish embryos; (4) injection of DAO morpholino; (5) confocal images analysis; and (6) electromicroscopy analysis. The second year: (1) breeding of zebrafish nic-1 mutant; (2) injection of NMDA-R morpholino; (3) ratio images analysis of intracellular calcium influx; (4) locomotor activity analysis; and (5) MK801, NMDA immersion experiments. The third year: (1) caffeine, 2-APB, ryanodine immersion experiments; (2) whole mount in situ hybridization experiments; (3) using antibodies staining to study the endogenous protein levels; (4) using cryosection and immunohistochemical staining to further confirm the expression domain of each gene transcript; (5) phallodin staining; and (6) swimming test of adult zebrafish.
    Appears in Collections:[化學學系暨研究所] 研究報告

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