Department of Anatomy and Molecular Histology, Interdisciplinary Graduate School of Medicine and Engineering, University of Yamanashi, 1110 Shimokato, Chuo-city, Yamanashi, 409-3898, Japan.
Histochem Cell Biol. 2013 Oct;140(4):477-90. doi: 10.1007/s00418-013-1084-y. Epub 2013 Mar 21.
Angiotensin II (AT) receptors, including AT receptor type 1 (AT1R) and type 2 (AT2R), are expressed in the rodent central nervous system, but their distributions and activation states are still unclear. In this study, we have performed immunohistochemical analyses of AT receptors in mouse cerebellum and adrenal gland using our "in vivo cryotechnique" (IVCT). We used antibodies against amino-terminal domains of AT receptors, which are considered to undergo conformational changes upon the binding of AT. Immunoreactivity of AT1R was detected in mouse cerebellum, and was highest in the outer tissue areas of molecular layers using IVCT. The AT1R immunostaining largely overlapped with glial fibrillary acidic protein (GFAP), a marker of Bergmann glia. Surprisingly, the AT1R immunoreactivity in the cerebellar cortex was remarkably reduced following 5 and 10 min of hypoxia or direct administration of an AT1R antagonist, losartan. By contrast, in the adrenal cortex, such AT1R immunoreactivity detected at the zona glomerulosa did not change even after 15 min of hypoxia. The correlation of localization with GFAP and also hypoxia-induced decrease of its immunoreactivity were similarly observed by immunostaining of AT2R in the cerebellar specimens. These findings demonstrated that IVCT is useful to reveal dynamically changing immunoreactivities usually affected by receptor-ligand binding as well as hypoxia, and also suggested that functional activities of AT receptors are time-dependently modulated under hypoxia in the central nervous system in comparison with the adrenal glands.
血管紧张素 II（AT）受体，包括 AT 受体 1 型（AT1R）和 2 型（AT2R），在啮齿动物中枢神经系统中表达，但它们的分布和激活状态仍不清楚。在这项研究中，我们使用我们的“体内冷冻技术”（IVCT）对小鼠小脑和肾上腺中的 AT 受体进行了免疫组织化学分析。我们使用了针对 AT 受体氨基末端结构域的抗体，这些抗体被认为在与 AT 结合时会发生构象变化。使用 IVCT 在小鼠小脑检测到 AT1R 的免疫反应性，在分子层的外组织区域最高。AT1R 免疫染色与神经胶质纤维酸性蛋白（GFAP），一种 Bergmann 胶质细胞的标志物，高度重叠。令人惊讶的是，小脑皮质中的 AT1R 免疫反应性在缺氧 5 分钟和 10 分钟或直接给予 AT1R 拮抗剂洛沙坦后显著降低。相比之下，在肾上腺皮质中，即使在缺氧 15 分钟后，在肾小球带检测到的 AT1R 免疫反应性也没有改变。在小脑标本中对 AT2R 进行免疫染色也观察到了与 GFAP 的定位相关性以及其免疫反应性在缺氧下的减少。这些发现表明，IVCT 可用于揭示通常受受体-配体结合以及缺氧影响的动态变化的免疫反应性，并且还表明，与肾上腺相比，AT 受体的功能活性在中枢神经系统中受到缺氧的时间依赖性调节。
