Bolger G T, Liard F, Garneau M, Jaramillo J
Department of Pharmacology, Bio-Mega Inc., Laval, Que., Canada.
Can J Physiol Pharmacol. 1992 Mar;70(3):377-84. doi: 10.1139/y92-047.
The contractile activity of and binding sites for endothelin-1 (ET-1) were investigated in isolated guinea-pig ileal longitudinal smooth muscle (GPILM). ET-1 produced concentration-dependent contractions of GPILM that either slowly subsided in the continued presence of ET-1 or rapidly subsided following washing of the tissue. The ED50 value for ET-1 contractions was 4.2 +/- 1.3 x 10(-9) M. The removal of extracellular calcium or pretreatment with nifedipine produced a complete inhibition of the contractions to ET-1. The IC50 value of nifedipine for inhibition of ET-1 mediated contractions was 3.0 +/- 0.8 x 10(-8) M. ET-1 produced a marked prolonged homologous desensitization of its contractile response but did not affect the responses mediated by carbachol, histamine, serotonin, substance P, and PLA2. High-affinity binding sites for 125I-labelled ET-1 were identified on microsomal membranes prepared from GPILM with Kd and Bmax values obtained by Scatchard analysis of 3.5 +/- 0.6 x 10(-10) M and 2138 +/- 159 fmol/mg protein, respectively. The binding of 125I-labelled ET-1 to GPILM microsomes was characterized by a rapid association (kob value of 0.077 min-1 at a radioligand concentration of 0.45 nM and an extremely slow dissociation (k1 value of 0.011 min-1; t1/2 value of 793 min). The binding was unaffected by the calcium channel antagonists nifedipine, verapamil, and diltiazem (10(-6) M); the receptor antagonists phenoxybenzamine, atropine, and naloxone (10(-6) M) and propranolol; and the peripheral benzodiazepine receptor antagonists Ro 5-4864 and PK 11195 and psychotomimetic drug phencyclidine (10(-5) M).(ABSTRACT TRUNCATED AT 250 WORDS)
在离体豚鼠回肠纵行平滑肌（GPILM）中研究了内皮素-1（ET-1）的收缩活性和结合位点。ET-1可引起GPILM浓度依赖性收缩，这种收缩在ET-1持续存在时缓慢消退，或在组织冲洗后迅速消退。ET-1收缩的半数有效浓度（ED50）值为4.2±1.3×10⁻⁹M。去除细胞外钙或用硝苯地平预处理可完全抑制对ET-1的收缩。硝苯地平抑制ET-1介导收缩的半数抑制浓度（IC50）值为3.0±0.8×10⁻⁸M。ET-1对其收缩反应产生明显的长时间同源脱敏，但不影响由卡巴胆碱、组胺、5-羟色胺、P物质和磷脂酶A2介导的反应。在从GPILM制备的微粒体膜上鉴定出125I标记的ET-1的高亲和力结合位点，通过Scatchard分析获得的解离常数（Kd）和最大结合容量（Bmax）值分别为3.5±0.6×10⁻¹⁰M和2138±159 fmol/mg蛋白质。125I标记的ET-1与GPILM微粒体的结合特点是快速结合（在放射性配体浓度为0.45 nM时结合速率常数kob值为0.077 min⁻¹）和极缓慢解离（解离速率常数k1值为0.011 min⁻¹；半衰期t1/2值为793 min）。该结合不受钙通道拮抗剂硝苯地平、维拉帕米和地尔硫䓬（10⁻⁶M）、受体拮抗剂酚苄明、阿托品和纳洛酮（10⁻⁶M）以及普萘洛尔、外周苯二氮䓬受体拮抗剂Ro 5-4864和PK 11195以及拟精神病药物苯环己哌啶（10⁻⁵M）的影响。（摘要截短于250字）
