Ekstrom R C, Hunzicker-Dunn M
Department of Molecular Biology, Northwestern University Medical School, Chicago, Illinois 60611.
Endocrinology. 1990 Feb;126(2):1191-8. doi: 10.1210/endo-126-2-1191.
The hormone responsiveness of the adenylyl cyclase of pig ovarian follicles or corpora lutea was examined. Adenylyl cyclase activity was assayed in 10,000 x g membrane fractions that had been prepared with or without (control) a urea extraction. In control luteal membranes there was little stimulation (less than 2-fold) or adenylyl cyclase by saturating ovine (o) LH, hCG, or (-)isoproterenol in the absence or presence of 10 microM GTP. However, in urea-treated luteal membranes, a 2- to 3-fold stimulation of adenylyl cyclase was caused by saturating oLH or hCG, and a 4- to 5-fold stimulation by (-)isoproterenol; the marked stimulation by the gonadotropins was only observed if 10 microM GTP was added. In follicular membranes, a 3- to 4-fold stimulation of adenylyl cyclase by gonadotropins was observed regardless of whether GTP was added or the membranes had been urea extracted. Stimulation of adenylyl cyclase by (-)isoproterenol was always less than 2-fold in follicular membranes. The binding affinity for [125I]hCG was similar in control follicular and luteal membranes, but there were approximately 10-fold more [125I]hCG-binding sites in follicular compared with luteal membranes. The binding affinities and number of receptor sites were not significantly changed by urea treatment. The ED50 values for hCG or (-)isoproterenol were the same in follicular and luteal membranes and were uneffected by the addition of 10 microM GTP, but the ED50 for oLH was 3-fold lower in follicular than in luteal membranes. GTP caused a dose-dependent increase in adenylyl cyclase activity in luteal and follicular membranes, and both tissues had the same ED50. A saturating hormone concentration resulted in an approximately 2-fold decrease in the ED50 for GTP. In vitro hCG-induced desensitization of the hCG-responsive adenylyl cyclase was 31% in follicular membranes, but only 11-15% in luteal membranes. Hormone-induced desensitization was not increased in incubations of luteal homogenate or membranes plus cytosol. These results establish the existence of a LH/hCG-sensitive adenylyl cyclase in the pig corpus luteum and indicate that the G-protein and catalytic moieties of the follicular and luteal adenylyl cyclase complex are functionally the same, but some difference exists in the way the LH/hCG-receptor in the two tissues interacts with the G-protein/catalytic complex.
对猪卵巢卵泡或黄体的腺苷酸环化酶的激素反应性进行了检测。在经过或未经过（对照）尿素提取制备的10,000×g膜组分中测定腺苷酸环化酶活性。在对照黄体膜中，无论是在不存在还是存在10μM GTP的情况下，饱和的羊（o）促黄体生成素（LH）、人绒毛膜促性腺激素（hCG）或（-）异丙肾上腺素对腺苷酸环化酶的刺激作用都很小（小于2倍）。然而，在经尿素处理的黄体膜中，饱和的oLH或hCG可引起腺苷酸环化酶2至3倍的刺激，（-）异丙肾上腺素可引起4至5倍的刺激；只有添加10μM GTP时，促性腺激素才会产生明显的刺激作用。在卵泡膜中，无论是否添加GTP或膜是否经过尿素提取，促性腺激素对腺苷酸环化酶的刺激作用均为3至4倍。在卵泡膜中，（-）异丙肾上腺素对腺苷酸环化酶的刺激作用始终小于2倍。对照卵泡膜和黄体膜中对[125I]hCG的结合亲和力相似，但卵泡膜中[125I]hCG结合位点的数量比黄体膜中多约10倍。尿素处理后，结合亲和力和受体位点数量没有明显变化。卵泡膜和黄体膜中hCG或（-）异丙肾上腺素的半数有效剂量（ED50）相同，且不受添加10μM GTP的影响，但卵泡膜中oLH的ED50比黄体膜中低3倍。GTP导致黄体膜和卵泡膜中腺苷酸环化酶活性呈剂量依赖性增加，且两种组织的ED50相同。饱和激素浓度导致GTP的ED50降低约2倍。体外hCG诱导的hCG反应性腺苷酸环化酶脱敏在卵泡膜中为31%，但在黄体膜中仅为11 - 15%。在黄体匀浆或膜加细胞溶质的孵育中，激素诱导的脱敏作用没有增强。这些结果证实了猪黄体中存在LH/hCG敏感的腺苷酸环化酶，并表明卵泡和黄体腺苷酸环化酶复合物的G蛋白和催化部分在功能上是相同的，但两种组织中LH/hCG受体与G蛋白/催化复合物相互作用的方式存在一些差异。
