鼠小腦中內皮素受體之鑑定及其性質之探討   全文下載

Identification and Characterization of Endothelin Receptors in Rat Cerebellum

內皮素（endothelins）不但是種強烈血管收縮劑，也是一種位於哺乳動物腦中的神經胜?。本研究主要是利用放射性配位體結合技術以及親和性交互聯結作用之方法，來作為對鼠小腦中內皮素受體之鑑定及其性質之探討。結果發現碘125所標識之內皮素-1可專一性地與鼠小腦上之受體結合，並且呈現量化之關係。由斯可恰圖（Scatchard plot）之結果看出，只有一種具高親和性之接受體存在於鼠小腦中，其解離常數（dissociation constant; Kd）為110 pM，而最大之結合位置（maximum binding site; Bmax）為每毫克之膜蛋白中便有4.85 pmol之接受體存在。以碘125所標識之內皮素-1 與其受體作親和性交互聯結作用後，經由電泳分析發現有兩種分子量大小之內皮素受體，一為47,000，另一為32,000，較小分子量之受體可能乃是較大分子量之受體經由水解蛋白之酵素作用而來的。另亦發現此接受體在電泳中之移動速率並不受是否有還原劑存在之影響，這表示內皮素受體乃是以單一多胜鏈之形式存在。進一步地利用競爭性結合實驗，發現鼠小腦之內皮素體對於內皮素及sarafotoxin 6b （S6b）之親和力為內皮素-1 = 內皮素2 = 內皮素-3 >S6b。由本研究之結果，可在鼠小腦中鑑定到相當量且高親和力之內皮素受體之存在，這支持了內皮素具有調節腦神經功能之假說，另由其受體專一性之結果得知鼠小腦中之內皮素受體是屬於B亞型之內皮素受體。"

Endothelins (ETs), potent vasoconstrictors, are also neuropeptides in mammalian brain. The objectives of this study were to identify and characterize the receptors for ETs in rat cerebellum, using radioligand binding techniques and affinity cross-linking method. Iodinated ET-1 bound specifically to receptors in rat cerebellar membrane in a dose-dependent manner. Scatchard plot indicated a single class of high-affinity binding sites. Apparent KI value was 110 pM, and the Bmax value was 4.85 pmo1/mg of membrane protein. Through experiments of affinity cross-linking of ET receptors with 125 1-ET-1, two forms of endothelin receptors with molecular masses of 47 KD and 32 KD were identified. Electrophoresis conducted in the presence of reducing reagents did not affect the mobilities of specifically labeled bands, suggesting that endothelin receptors exist as a single polypeptide chain. The order of potency for ETs and sarafotoxin 6b (S6b) in displacing the specifically bound iodinated ET-1 from cerebellar membrane was ET-1 = ET-2 = ET-3 ＞S6b.Result from receptor specificity indicated that endothelin receptors in rat cerebellum belong to endothelin receptor subtype B.