Identification of a novel non-AT1, non-AT2 angiotensin binding site in the rat brain
Identification of a novel non-AT1, non-AT2 angiotensin binding site in the rat brain
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DOI：
10.1016/j.brainres.2007.01.051
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发表时间：
2007-04-27
期刊：
影响因子：
2.9
通讯作者：
Speth, Robert C.
中科院分区：
文献类型：
--
作者：
Karamyan, Vardan T.;Speth, Robert C.
在受体结合试验中保护放射性标记的血管紧张素免于代谢的努力可以追溯到30多年前。然而，这仍然是一个问题。本研究主要研究蛋白酶抑制剂对氯汞苯甲酸盐（PCMB）对I-125-Ang Ⅱ与大鼠脑细胞膜结合的影响。将PCMB添加到孵育培养基中揭示了I-125-Ang II在脑膜中的高亲和力结合位点（Kd = 1-4 nM），其结合量大于先前脑Ang II受体研究中所揭示的结合量。对这种结合的进一步表征表明，它对氯沙坦（一种AT（2）受体拮抗剂）和PD 123319（一种AT（2）受体拮抗剂）的抑制不敏感。这种非AT 1、非AT 2结合位点不存在于肝或肾上腺膜中。它被激活的PCMB的浓度范围有限，在0.3-1 mM的最大激活。这种结合位点是同样丰富的大脑皮层（脑区与几个血管紧张素II受体）和下丘脑（脑区与丰富的血管紧张素II受体）。结合位点也存在于小鼠脑中，但不存在于小鼠肝中。该结合位点显示出对Ang I、Ang II和Ang III的高亲和力（Ki类似于40-100 nM），但对较小的血管紧张素片段和其他神经肽的亲和力较低。该结合位点与肝细胞溶质Ang II结合蛋白共有一些特征，后来被鉴定为内肽酶EC 3.4.24.15和/或EC 3.4.24.16。然而，这种非AT 1，非AT 2结合位点的一些独特的特征表明，它可能是一种新的血管紧张素结合物质。(c)2007 Elsevier B. V.保留所有权利。
Efforts to protect radiolabeled angiotensins from metabolism during receptor binding assays date back more than 30 years. However, this continues to be a problem. This study focused on the effects of a protease inhibitor, p-chloromercuribenzoate (PCMB), on the binding of I-125-Ang II to rat brain membranes. Addition of PCMB to the incubation medium revealed a high affinity binding site for I-125-Ang II in brain membranes (K-d = 1-4 nM) with a greater amount of binding than revealed in previous studies of brain Ang II receptors. Further characterization of this binding, revealed it to be insensitive to inhibition by losartan (an AT, receptor antagonist) and PD123319 (an AT(2) receptor antagonist). This non-AT1, non-AT2 binding site was not present in liver or adrenal membranes. It was activated by a limited range of concentrations of PCMB, with maximal activation at 0.3-1 mM. This binding site was equally abundant in cerebral cortex (a brain region with few Ang II receptors) and the hypothalamus (a brain region with abundant Ang II receptors). The binding site was also present in mouse brain, but not mouse liver. The binding site shows high affinity for Ang I, Ang II and Ang III (K-i similar to 40-100 nM), but lesser affinity for smaller angiotensin fragments and other neuropeptides. This binding site shares some characteristics with the liver cytosolic Ang II binding proteins, later identified as endopeptidases EC 3.4.24.15 and/or EC 3.4.24.16. However, some unique characteristics of this non-AT1, non-AT2 binding site suggest that it may be a novel angiotensin binding substance. (c) 2007 Elsevier B.V. All rights reserved.