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研究生:黃知偉
研究生(外文):Chi–Wei Huang
論文名稱:藍綠藻水萃物對耳鳴相關接受器表現之研究
論文名稱(外文):The effects of the Spirulina platensis water extract on the expression of related receptors in tinnitus
指導教授:詹吟菁 博士
指導教授(外文):Yin-Ching Chan, Ph. D.
學位類別:碩士
校院名稱:靜宜大學
系所名稱:食品營養研究所
學門:醫藥衛生學門
學類:營養學類
論文種類:學術論文
論文出版年:2011
畢業學年度:99
語文別:中文
論文頁數:98
中文關鍵詞:藍綠藻耳鳴耳鳴相關接受器主動迴避試驗老化促進小鼠
外文關鍵詞:active avoidance testtinnitus relative receptorsenescence accelerated prone miceSpirulinatinnitussalicylate
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耳鳴是體內產生的聲音,非受外來音源刺激所造成。嚴重的耳鳴會影響情緒、注意力甚至是生活品質,目前為止對耳鳴的治療方法仍有待進一步研究。本實驗目的為探討藍綠藻(Spirulina)水萃物對水楊酸誘導老化促進小鼠(senescence accelerated prone mice;SAMP8)自覺性耳鳴動物模式,耳鳴相關接受器表現之影響。實驗選用三月齡雄性SAMP8小鼠隨機分成控制組、腹腔注射300mg/kg BW水楊酸誘發耳鳴組及管灌1000mg/kg BW藍綠藻後腹腔注射水楊酸誘發耳鳴組。實驗期間記錄體重與進行主動迴避試驗。試驗後犧牲,評估血液生化值並進行腦幹、額葉、海馬迴、顳葉及內耳等部位cannabinoid receptor 1(CB1) 、vanilloid receptor type 1(VR1)、dopamine D1A receptor及γ-amino butyric acid(GABA) Aβ3 receptor mRNA表現量之分析。結果顯示,給予水楊酸與藍綠藻水萃物並不會影響小鼠的肝、腎功能。水楊酸的給予會誘發耳鳴使誤判次數增加,成功迴避反應次數則呈下降的趨勢;藍綠藻水萃物,可改善誤判及成功迴避次數。基因表現結果發現,水楊酸會顯著減少腦幹、額葉、海馬迴、顳葉及內耳的CB1 receptor及GABA Aβ3 receptor mRNA表現量,而藍綠藻水萃物可顯著增加其表現量。另水楊酸會顯著增加腦幹、海馬迴、顳葉及內耳中D1A receptor mRNA表現量,藍綠藻水萃物則可顯著降低其表現;此外,水楊酸也會顯著增加腦幹、額葉、海馬迴及顳葉中VR1 receptor mRNA表現量,藍綠藻水萃物亦能顯著降低其表現。綜合上述,藍綠藻水萃物可能藉由調控耳鳴相關接受器的表現,而改善了老化促進小鼠因水楊酸所誘發的自覺性耳鳴。
Tinnitus is a phantom sensation of sound in the absence of external stimulations. Serious tinnitus may affect emotion, concentration, and even the quality of life. However, the treatments of tinnitus need further study. The purpose of this study was to develop a behavioral model of conscious tinnitus with SAMP8 mice induced by salicylate, and evaluated the effects of Spiulina water extract on relative receptors. Three-month-old SAMP8 mice were randomly divided into normal control, tinnitus with intraperitoneal injection of salicylate (300 mg/kg BW), and Spiulina water extract (1000 mg/kg BW) with salicylate-induced tinnitus groups. Body weight were recorded and active avoidance test were performed during the experimental period. The biochemical values was analyzed after sacrificed, and the cannabinoid receptor 1 (CB1 receptor), vanilloid receptor type 1 (VR1 receptor), dopamine D1A receptor and γ-amino butyric acid (GABA) Aβ3 receptor mRNA expressions of brain stem, frontal lobe, hippocampus, temporal lobe and inner ear were also examined. Our results demonstrated that both salicylate and Spiulina water extract did not affect the liver and kidney functions of SAMP8 mice. Salicylate-induced tinnitus led significant increase in the number of false positive response while the percentage of correct response tented to reduce and Spiulina water extract could improve the trend. Salicylate significantly reduced the cannabinoid receptor 1 (CB1 receptor) and γ-amino butyric acid (GABA) Aβ3 receptor mRNA expression in brain stem, frontal lobe, hippocampus, temporal lobe and inner ear, while Spiulina water extract significantly increased their expression. In addition, salicylate significantly increased the dopamine D1A receptor expression in brain stem, hippocampus, temporal lobe and inner ear, and Spiulina water extract could significantly lessen their expression. Salicylate significantly increased the VR1 receptor expression in brain stem, frontal lobe, hippocampus and temporal lobe, and Spiulina water extract also significantly reduced the expression. In summary, we conclude that the Spiulina water extract could improve the salicylate-induced conscious tinnitus in SAMP8 mice, which might due to regulate the expression of tinnitus related receptor.
目錄 頁次
中文摘要 I
英文摘要 II
目錄 IV
圖目錄 VI
表目錄 VIII
縮寫表 IX
第一章 前言 I
第二章 文獻回顧 2
第一節 老化與疾病及免疫之相關性 2
第二節 耳鳴簡介 3
第三節 水楊酸誘發自覺性耳鳴之動物模式 11
第四節 調控耳鳴之相關因子 20
第五節 藍綠藻 29
第六節 老化促進小鼠之簡介 35
第三章 材料與方法 36
第一節 實驗動物 36
第二節 耳鳴動物模式建立 36
第三節 實驗流程與方法 37
壹 實驗流程 37
貳 實驗方法 39
一 主動迴避試驗 39
二 制約學習訓練(Conditioning) 39
三 行為測試(Testing) 39
参 基因表現測定 41
一 總RNA萃取 41
二 反轉錄酶聚合酶連鎖反應(Reverse-transcriptase polymerase 43
三 PCR產物偵測 46
四 統計分析……………..……………………………………………………………………………47
第四章 結果與討論 48
第一節 體重 48
第二節 器官重量 50
第三節 血液生化值評估 54
第四節 主動迴避試驗 55
第五節 調控耳鳴之相關因子分析 60
第五章 結論 72
第六章 參考文獻 74

圖目錄 頁次

圖一、聽覺路徑 5
圖二、水楊酸結構式 11
圖三、多巴胺受器結構圖 25
圖四、TRPV1結構及活化位置 22
圖五、CB1受體調控GABA產生突觸後抑制性電位(IPSP) 28
圖六、實驗流程圖 38
圖七、主動迴避試驗裝置圖 40
圖八、3月齡雄性SAMP8小鼠餵食不同飼料9天後腦部不同部位重量之比較52
圖九、3月齡雄性SAMP8小鼠餵食不同飼料9天後耳蝸重量之比較 53
圖十、3月齡雄性SAMP8小鼠經餵食不同飼料並注射水楊酸後之誤判次數 58
圖十一、3月齡雄性SAMP8小鼠經餵食不同飼料並注射水楊酸後之成功迴避測試比例 59
圖十二、3月齡雄性SAMP8小鼠餵食不同飼料並注射水楊酸後腦幹、額葉及海馬迴之CB1 receptor mRNA 表現 61
圖十三、3月齡雄性SAMP8小鼠餵食不同飼料並注射水楊酸後額葉及內耳之CB1 receptor mRNA 表現 62
圖十四、3月齡雄性SAMP8小鼠餵食不同飼料並注射水楊酸後額葉及內耳之VR1 receptor mRNA表現 64
圖十五、3月齡雄性SAMP8小鼠餵食不同飼料並注射水楊酸後額葉及內耳之VR1 receptor mRNA表現 65
圖十六、3月齡雄性SAMP8小鼠餵食不同飼料並注射水楊酸後腦幹、額葉及海馬迴之dopamine D1A receptor mRNA表現 67
圖十七、3月齡雄性SAMP8小鼠餵食不同飼料並注射水楊酸後額葉及內耳之dopamine D1A receptor mRNA表現 68
圖十八、3月齡雄性SAMP8小鼠餵食不同飼料並注射水楊酸後腦幹、額葉及海馬迴之GABA Aβ3 receptor mRNA表現 70
圖十九、3月齡雄性SAMP8小鼠餵食不同飼料並注射水楊酸後腦幹、額葉及海馬迴之GABA Aβ3 receptor mRNA表現 71

表目錄 頁次

表一、聚合酶連鎖反應之寡核甘酸引子序列及反應條件 45
表二、3月齡雄性SAMP8小鼠餵食不同飼料9天後體重之變化 49
表三、3月齡雄性SAMP8小鼠餵食不同飼料9天後相對器官重量之比較 51
表四、3月齡雄性SAMP8小鼠餵食不同飼料9天後血液生化值之比較 54
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