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研究生:鍾桂芳
研究生(外文):Kuei-Fang Chung
論文名稱:鋸緣青蟳(Scyllaserrata)滲透壓調節器官之Na+,K+-ATPase活性與蛋白質表現
論文名稱(外文):Na+, K+-ATPase Activity and Protein Level in the Osmoregulatory Organs of Scylla serrata
指導教授:林惠真林惠真引用關係
指導教授(外文):Hui-Chen Lin
學位類別:碩士
校院名稱:東海大學
系所名稱:生物學系
學門:生命科學學門
學類:生物學類
論文種類:學術論文
論文出版年:2002
畢業學年度:90
語文別:中文
論文頁數:56
中文關鍵詞:鋸緣青蟳滲透壓調節器官Na+K+-ATPase
外文關鍵詞:Na+K+-ATPaseScylla serrata
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鋸緣青蟳(Scylla serrata)為廣鹽性之海生蟹,牠可生存在1- 42 ppt的環境鹽度下。環境中鹽度的變化,會使得個體必須面臨環境中滲透壓劇烈變化並影響其生理功能。螃蟹進行滲透壓調節的幾個主要器官包括:鰓、觸角腺與腸道。在過去與螃蟹滲透壓調節相關的研究多著重於鰓的部位,極少同時針對各滲透壓調節器官作一研究與探討。由於Na+, K+-ATPase為帶動離子流動之原動力,因此可作為代表螃蟹滲透壓調節能力的有效指標。由過去的文獻中得知,在低鹽度處理下,不同種螃蟹的後鰓、觸角腺或腸道分別具有比海水處理高的Na+, K+- ATPase活性,但其共同表現目前則尚未明朗。這可能代表了在不同環境下,各滲透壓調節器官所擔負之滲透壓調節功能在比例上並不一致。而此活性的升高是藉由調整現有的Na+, K+-ATPase或是新合成更多蛋白質造成,目前仍不清楚。本實驗將鋸緣青蟳自25 ppt分別轉移至5、25與45 ppt不同時間後,測量體液滲透壓、Na+, K+- ATPase活性,並以西方墨點法分析Na+, K+-ATPase α-次單元的蛋白質表現量。實驗結果指出,鋸緣青蟳在5 ppt與 25 ppt的海水中,其體液滲透壓高於環境滲透壓,因此應被歸類為hyper-osmoregulator。就器官間的比較上,無論是Na+, K+-ATPase活性或Na+, K+-ATPase α-次單元的蛋白質表現量,後鰓與觸角腺均具有較前鰓與腸道高的情形。而針對不同鹽度處理間的比較,轉移至5 ppt海水14天後的鋸緣青蟳,其後鰓的Na+, K+-ATPase活性顯著高於其他鹽度處理組;而這種鹽度處理間活性差異的情形在其他器官則不顯著。在西方墨點法的結果方面,除了腸道在鹽度轉移後第1天Na+, K+-ATPase α-次單元的蛋白質表現量曾出現些許差異外,不同鹽度處理對各器官上Na+, K+- ATPase α-次單元蛋白質表現量沒有顯著影響。當後鰓上Na+, K+- ATPase活性因鹽度變化而改變時,Na+, K+-ATPase α-次單元蛋白質表現量並未隨之改變。綜合以上結果,鋸緣青蟳在低鹽度環境下,後鰓在滲透壓調節上具有最重要的角色,而其酵素活性的變化可能是透過已經存在的Na+, K+-ATPase進行調節。

Scylla serrata is a euryhaline marine crab, which can tolerate a wide range of salinities between 1 ppt and 42 ppt. And the salinity fluctuation may cause serious changes in hemolymph osmolality and influence the physiological functions. The osmoregulatory organs of crab primarily include gills, antennal glands and guts. Most studies on crustacean osmoregulation had their focus on the gills only, but very rarely were these organs investigated simultaneously. Since Na+, K+-ATPase has been known as the driving force of ions transportations, it can be taken as a valid indicator of osmoregulatory ability. From previous literatures, posterior gills, antennal gland and gut respectively showed a higher Na+, K+-ATPase activity when the crabs were kept in diluted seawater. This implies that the relative importance in osmoregulation among these organs is not identical. And whether this increase resulted from the modulation of pre-existing Na+, K+-ATPase or new protein synthesis is still unclear. In this study, after transferring S. serrata from 25 ppt to 5, 25 and 45 ppt respectively, hemolymph osmolality, Na+, K+-ATPase activity and the level of Na+, K+-ATPase α-subunit in western blotting were examined at different time intervals. The results indicated that S. serrata maintained their hemolymph osmolality significantly higher than their respective environments of 5 and 25 ppt, therefore, it should be categorized as a hyper-osmoregulator. The posterior gill (gill 6) and antennal gland were significantly higher than the anterior gill (gill 2) and gut in both the Na+, K+-ATPase activities and their α-subunit levels. Upon 14 days after transfer, the activity of the posterior gill was significantly higher in 5 ppt than in the other two salinity treatments. Such differences among salinity levels were not observed in other organs. In the results of western blotting, salinity did not cause a significant change in the protein levels of Na+, K+-ATPase α-subunit among these osmoregulatory organs, except the gut in the first day after transferring. The protein level of Na+, K+-ATPase α-subunit did not change significantly in the posterior gill while the enzyme activity fluctuated with environmental salinity. In conclusion, the posterior gills played the most important role in osmoregulation when the crabs were in diluted SW and the activity of the enzyme might be modulated by the pre-existing Na+, K+-ATPase.

中文摘要………………………………………………………… 1
Abstract………………………………………………………… 3
前言……………………………………………………………… 5
一、螃蟹的滲透壓調節………………………………………… 5
二、滲透壓調節器官…………………………………………… 6
三、Na+, K+-ATPase…………………………………………… 9
四、鋸緣青蟳…………………………………………………… 11
五、實驗目的…………………………………………………… 12
材料與方法……………………………………………………… 13
一、實驗動物…………………………………………………… 13
二、方法………………………………………………………… 13
(一)馴養與實驗用水的配置………………………………… 13
(二)馴養及轉移水槽的設置………………………………… 13
(三)實驗設計與步驟………………………………………… 14
(四)實驗方法………………………………………………… 14
(五)統計……………………………………………………… 19
實驗結果………………………………………………………… 20
一、滲透壓調節………………………………………………… 20
二、Na+, K+-ATPase活性測定………………………………… 21
三、Na+, K+-ATPase α-subunit蛋白質表現差異…………… 24
討論……………………………………………………………… 27
參考文獻………………………………………………………… 47
圖表……………………………………………………………… 52

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