Reul J M, de Kloet E R, van Sluijs F J, Rijnberk A, Rothuizen J
Department of Clinical Sciences of Companion Animals, Faculty of Veterinary Medicine, State University of Utrecht, The Netherlands.
Endocrinology. 1990 Aug;127(2):907-15. doi: 10.1210/endo-127-2-907.
A series of studies was started to gain insight into the functioning of the canine hypothalamo-pituitary-adrenocortical axis during normo- and hypercortisolemic states. In this first study, we have focused on the binding characteristics of the mineralocorticoid receptor (MR) and the glucocorticoid receptor (GR) in the brain and pituitary of the adrenalectomized dog. In hippocampal cytosol at 0 C, corticosterone had the highest association rate, followed by cortisol and aldosterone. Cortisol had the most rapid rate of dissociation from MR at 0 C (t1/2 = 45.5 h), followed by aldosterone (70.4 h) and corticosterone (102 h). The selective glucocorticoid RU 28362 associated rapidly with hippocampal GR, attaining maximum binding within 4 h, and dissociated with a t1/2 of 34.8 h. Saturation binding of [3H]cortisol in adrenalectomized dog hippocampal cytosol produced a curvilinear Scatchard plot. After inclusion of RU 28362, [3H]cortisol bound solely to MR [dissociation constant (Kd) = 0.34 nM, Bmax = 72.8 fmol/mg]. GR capacity was determined with [3H]RU 28362 (Kd = 0.39 nM, Bmax = 120 fmol/mg). Competition binding analyses of various steroids for MR and GR revealed markedly different patterns of steroid binding specificity for these receptors. The rank order for displacement of [3H]aldosterone binding of MR was: corticosterone greater than aldosterone = cortisol greater than dexamethasone greater than ZK 91587 greater than RU 26752 greater than spironolactone much greater than RU 38486, and for displacement of [3H]RU 28362 binding of GR: RU 28362 much greater than corticosterone = cortisol greater than dexamethasone greater than aldosterone greater than ZK 91587 greater than RU 26752 = RU 38486 much greater than spironolactone. MR was located in all brain regions examined, with highest levels in the septo-hippocampal complex, whereas GR was rather evenly distributed. Substantial amounts of MR and GR were present in the anterior part of the pituitary as well as in the neurointermediate lobe. Our findings show that the ligand binding specificity of canine MR and GR is remarkably different from that of rodent MR and GR, but is similar to that of recombinant-derived human receptors. Spironolactone and RU 38486 are selective antagonists for MR and GR, respectively. In contrast to other species, the dog has relatively large quantities of MR widely distributed in the brain and pituitary, which makes this species an interesting animal model to study the role of corticosteroid receptor diversity in control of homeostasis.
为深入了解犬下丘脑 - 垂体 - 肾上腺皮质轴在正常和高皮质醇血症状态下的功能，开展了一系列研究。在第一项研究中，我们聚焦于肾上腺切除犬脑和垂体中盐皮质激素受体（MR）和糖皮质激素受体（GR）的结合特性。在0℃的海马细胞溶质中，皮质酮的结合率最高，其次是皮质醇和醛固酮。在0℃时，皮质醇从MR解离的速度最快（t1/2 = 45.5小时），其次是醛固酮（70.4小时）和皮质酮（102小时）。选择性糖皮质激素RU 28362与海马GR迅速结合，4小时内达到最大结合，并以34.8小时的t1/2解离。[3H]皮质醇在肾上腺切除犬海马细胞溶质中的饱和结合产生了曲线型Scatchard图。加入RU 28362后，[3H]皮质醇仅与MR结合[解离常数（Kd）= 0.34 nM，Bmax = 72.8 fmol/mg]。用[3H]RU 28362测定GR容量（Kd = 0.39 nM，Bmax = 120 fmol/mg）。各种类固醇对MR和GR的竞争结合分析显示，这些受体的类固醇结合特异性模式明显不同。MR上[3H]醛固酮结合的置换顺序为：皮质酮＞醛固酮 = 皮质醇＞地塞米松＞ZK 91587＞RU 26752＞螺内酯＞RU 38486，GR上[3H]RU 28362结合的置换顺序为：RU 28362＞皮质酮 = 皮质醇＞地塞米松＞醛固酮＞ZK 91587＞RU 26752 = RU 38486＞螺内酯。在所检查的所有脑区均发现有MR，在隔 - 海马复合体中水平最高，而GR分布较为均匀。垂体前部以及神经中间叶中存在大量的MR和GR。我们的研究结果表明，犬MR和GR的配体结合特异性与啮齿动物的MR和GR明显不同，但与重组衍生的人受体相似。螺内酯和RU 38486分别是MR和GR的选择性拮抗剂。与其他物种不同，犬在脑和垂体中广泛分布有相对大量的MR，这使得该物种成为研究皮质类固醇受体多样性在体内平衡控制中作用的有趣动物模型。
