臺灣博碩士論文加值系統

樹突棘(dendritic spines)為樹突分支上的棘狀小突起，是大腦興奮性突觸的主要受體。文獻指出記憶、神經疾病與神經樹突棘的型態變化有關，在神經樹突棘的型態研究中，需要輔助分析的軟體，既方便、快速又能減少人為誤差。本研究藉由Matlab撰寫樹突棘分析軟體，此軟體(NUUspine software)藉由樹突棘頭寬、頸寬、棘長度依形態特徵自動進行型態分類，分析後可輸出數量、密度、面積統計圖，同時利用動物誘導老化模型實驗，探討老化過程中是否可以藉由游泳改善記憶能力及對海馬迴區塊樹突棘型態之影響。將Sprague-Dawley大鼠分為不運動+控制組、運動+控制組、不運動+誘導老化組及運動+誘導老化組，進行每日D-半乳糖誘導老化及游泳訓練，最終利用行為學觀察其行為及記憶能力。大鼠腦切片以Golgi-Cox染色法進行染色，利用自製分析軟體對海馬迴DG區塊顆粒神經細胞和CA3區塊、CA1區塊的錐體神經細胞樹突分支上的樹突棘進行分析。水迷宮結果顯示老化確實會降低空間記憶能力，而持續游泳有改善的效果。分析海馬迴CA1區塊結果，觀察到不運動+誘導老化組樹突棘絲狀偽足型態的數量較不運動+控制組多，但是運動+誘導老化組蘑菇型態的數量較不運動+誘導老化組多，絲狀偽足型態的數量相對較少，而樹突棘的總數在各組中並無差異。綜合以上的分析結果老化過程中持續游泳有效改善空間記憶與樹突棘型態，並證實利用Matlab自製的半自動化神經元樹突棘型態分析軟體與已發表的文獻有相似結果，也適用於發育、疾病等樹突棘相關研究。

The dendritic spines are the primary recipients of excitatory synaptic input in the brain. Previous research shows that memory and diseases are correlated with dendritic spine morphology. Dendritic spine analysis software is usually very expensive. If we can construct the dendritic spine analysis method for free, it will benefit many researchers. Our semi-auto software, NUUspine, is written based on Matlab, which could define the dendritic spine types according to manually measured dendritic spine head width neck length and total length by Reconstruct software. The data output includes dendritic spine type, density, and area in excel profile. We used NUUspine to discuss the influence of aging and swimming exercise on the changes of memory and dendritic spine morphology. Experimental groups are divided into control without exercise, control with exercise, induced aging without exercise, and induced aging with exercise. We used behavioral test to assess rat aging. Golgi-Cox staining was applied to visualize spines. The dendritic spines on the dendritic branches of granule cell in hippocampal DG and pyramidal neurons in hippocampal CA3, CA1 were analyzed by using NUUspine software. The Morris water maze test showed aging decrease spatial memory. Continuous swimming improved spatial memory. The classify dendritic spines in hippocampal CA1 showed that the number of filopodium type in induced aging without exercise group was more than induced aging with exercise group. The number of mushroom type in induced aging with exercise group was more than induced aging without exercise group. But total number of spines showed no different between groups. Above all, exercise can improve spatial memory and cause dendritic spine morphology turn to mature type. Moreover, we adopted data from a published study to verify the accuracy of NUUspine software, the result from our software showed no difference to the published one. We successfully establish the software which can analyze dendritic spines.

摘要 I
Abstract II
致謝 III
目錄 IV
圖目錄 VII
表目錄 VIII
第一章 緒論 1
1.1 大腦與記憶 1
1.2 神經細胞與樹突棘 2
1.3 樹突棘型態 3
1.4 老化與誘導老化動物模型 4
1.5 運動 5
1.6 神經退行性疾病 5
1.7 Matlab與圖片分析軟體 6
1.8 實驗目的 6
第二章 材料與方法 8
2.1 動物飼養方式 8
2.2 誘導老化與游泳運動 8
2.3 動物行為學測試 9
2.3.1 曠野測試 9
2.3.2 Morris水迷宮測試 9
2.3.3 新物體辨識系統 9
2.4 高爾基染色 10
2.4.1 組織收集 10
2.4.2 組織包埋與切片 10
2.4.3 高爾基染色 11
2.5 影像處理(Image J、Photozoom) 11
2.6 Reconstruct分析 11
2.7 Matlab 13
2.7.1 軟體分析撰寫 13
2.7.2 樹突棘型態定義 13
2.7.3 操作步驟 13
2.8 軟體驗證 14
第三章 結果 15
3.1 行為學 15
3.1.1 曠野測試 15
3.1.2 Morris水迷宮 15
3.2.3 新物體辨識系統 15
3.2 Reconstruct分析 16
3.3 Matlab 16
3.3.1 多檔案讀取 16
3.3.2 樹突棘型態分類 16
3.3.3 輸出存檔 17
3.3.4 組間數據相比 17
3.4 軟體分析驗證 18
3.5 測量運動是否改善老化所造成的傷害的動物實驗結果 18
第四章 討論 20
第五章 結論 23
第六章 參考文獻 66
第七章 附錄 75

圖一、動物實驗架構 25
圖二、樹突棘分析整體架構圖 27
圖三、曠野測試滯留時間強弱圖 29
圖四、水迷宮學習曲線 31
圖五、水迷宮滯留時間強弱圖 33
圖六、水迷宮經過平台象限之頻率 35
圖七、新物體辨識率 37
圖八、Reconstruct軟體長度分析結果 39
圖九、樹突棘長度分析預定儲存格式 41
圖十、軟體分析驗證 44
圖十一、Dentate gyrus區域的樹突棘分析結果 50
圖十二、Cornu Ammonis 3區域的樹突棘分析結果 56
圖十三、Cornu Ammonis 1區域的樹突棘分析結果 62

表一、分析參考文獻樹突棘平均數量的計算結果與誤差 63
表二、手動與自動分析樹突棘平均數量的計算結果與誤差 64

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