De Kloet E R, McEwen B S
Biochim Biophys Acta. 1976 Jan 14;421(1):124-32. doi: 10.1016/0304-4165(76)90176-8.
The binding of [3H]corticosterone and [3H]dexamethasone to soluble macromolecules in cytosol of the hippocampal region of the brain has been studied in adrenalectomized male rats. Unlabeled dexamethasone appears to be a less effective competitor than corticosterone in the binding of [3H]corticosterone, while both unlabeled steroids compete equally well for the binding or [3H]dexamethasone. Further investigation of macromolecular complexes with [3H]dexamethasone and [3H]corticosterone revealed that they differ from each other in their behavior during ammonium sulfate precipitation, BioRad A-5M gel permeation chromatography, DE-52 anion exchange chromatography and DNA-cellulose chromatography. (1) After exposure to a 33% ammonium sulfate solution relatively more [3H]dexamethasone complex than [3H]corticosterone complex is precipitated. (2) Treatment of the cytosol with 0.3 M KCl gives disaggregation of the supramolecular 3H-labeled corticoid complexes which are seen eluting with the void volume during gel permeation chromatography on Biorad A-5M at low ionic strength. In 0.3 M KCl, the [3H]dexamethasone complex has an elution volume somewhat smaller than that of bovine serum albumin, while the [3H]-corticosterone complex in 0.3 M KCl is too unstable to survive chromatography with A-5M. (3) Chromatography on DE-52 resolved the 3H-labeled corticoid complexes into three binding components. The complex with [3H]dexamethasone contains a higher percentage (85%) of a component less firmly attached (i.e. eluted by 0.15 M KCl) to the anion exchange resin than is observed for the complex with [3H]corticosterone (49%). (4) The complexes with 3H-labeled corticoids display an enhanced affinity for calf thymus DNA adsorbed to cellulose following "activation", warming to 25 degrees C for 15 min. Concurrently, a fraction of the [3H]dexamethasone complex becomes able to more firmly attach to the DE-52 anion exchange resin. These results with the binding of the cytosol hormone-receptor complexes to DNA-cellulose do not explain the marked in vivo preference of hippocampus for the cell nuclear uptake of [3H] corticosterone. However, the other differences in the properties of the complexes formed with the two labeled glucocorticoids support our previous inference that there may be more than one population of adrenal steroid "receptors" in brain tissue.
在肾上腺切除的雄性大鼠中，研究了[3H]皮质酮和[3H]地塞米松与大脑海马区细胞质中可溶性大分子的结合情况。在[3H]皮质酮的结合中，未标记的地塞米松似乎比皮质酮作为竞争剂的效果要差，而两种未标记的类固醇在与[3H]地塞米松的结合竞争中表现相当。对与[3H]地塞米松和[3H]皮质酮形成的大分子复合物的进一步研究表明，它们在硫酸铵沉淀、BioRad A - 5M凝胶渗透色谱、DE - 52阴离子交换色谱和DNA - 纤维素色谱过程中的行为存在差异。（1）暴露于33%硫酸铵溶液后，沉淀出的[3H]地塞米松复合物比[3H]皮质酮复合物相对更多。（2）用0.3 M KCl处理细胞质会使超分子3H标记的皮质类固醇复合物解聚，在低离子强度下于Biorad A - 5M上进行凝胶渗透色谱时，这些复合物会以空体积洗脱。在0.3 M KCl中，[3H]地塞米松复合物的洗脱体积略小于牛血清白蛋白，而0.3 M KCl中的[3H]皮质酮复合物过于不稳定，无法在A - 5M柱上完成色谱分析。（3）在DE - 52上进行色谱分析可将3H标记的皮质类固醇复合物分离为三个结合组分。与[3H]地塞米松形成的复合物中，与阴离子交换树脂结合较不牢固（即被0.15 M KCl洗脱）的组分所占百分比更高（85%），而与[3H]皮质酮形成的复合物中该组分的比例为49%。（4）“活化”（在25℃加热15分钟）后，3H标记的皮质类固醇复合物对吸附在纤维素上的小牛胸腺DNA的亲和力增强。同时，一部分[3H]地塞米松复合物变得能够更牢固地附着在DE - 52阴离子交换树脂上。细胞质激素 - 受体复合物与DNA - 纤维素结合的这些结果并不能解释海马体在体内对[3H]皮质酮细胞核摄取的明显偏好。然而，与两种标记糖皮质激素形成的复合物在性质上的其他差异支持了我们之前的推断，即脑组织中可能存在不止一种肾上腺类固醇“受体”群体。
