Identification and Characterization of Endothelin Receptors in Rat Cerebellum

鼠小腦中內皮素受體之鑑定及其性質之探討

陳惠芳(Hwei-Fang Cheng) ； 葉景睿(Jing-Ruey Yeh) ； 張寬仁(Kwen-Jen Chang)

《藥物食品分析》 1卷2期 (1993/05) Pp. 165-173

摘要

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 K(subscript d) value was 110 pM, and the B(subscript max) value was 4.85pmol/mg of membrane protein. Through experiments of affinity cross-linking of ET receptors with 125 I-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 E1 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.

並列摘要

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 K(subscript d) value was 110 pM, and the B(subscript max) value was 4.85pmol/mg of membrane protein. Through experiments of affinity cross-linking of ET receptors with 125 I-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 E1 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.

並列關鍵字

Endothelin receptor ； Rat cerebellum ； Radioligand binding techniques ； Affinity cross-linking method