Hervieu G, Emson P C
Laboratory of Cognitive and Molecular Neuroscience, Department of Neurobiology, Babraham Institute, Cambridge, UK.
Brain Res Mol Brain Res. 1999 Aug 25;71(2):290-303. doi: 10.1016/s0169-328x(99)00201-6.
Somatostatin actions are mediated through G-protein coupled receptors named sst(1) to sst(5). We used an affinity-purified polyclonal antibody AS-69, directed against a specific N-terminal peptide sequence of sst(3) to determine the immunohistochemical distribution of the sst(3) receptor in the rat and human brain. The specificity of the antibody was shown by Western blotting experiments using an N-terminal sst(3) fusion protein. Enzymatic deglycosylation experiments were combined to blotting experiments on a sst(3)-transfected cell line and rat brain membrane proteins and with immunocytochemistry on the sst(3)-transfected cell line. These studies showed that the antibody detected the deglycosylated sst(3) receptor protein. Immunohistochemical staining showed that sst(3) immunoreactivity recognised by this N-terminal antiserum was widely distributed throughout the brain with cells and processes labelled in the cerebral cortex, regions of the limbic system (including the hippocampal formation, some amygdaloid regions, some basal ganglia nuclei and regions from the nucleus basalis complex), the habenula, the hypothalamus, the thalamus, different mesencephalic structures (substantia nigra, zona incerta, superior colliculus), the reticular formation, the cerebellum. The distribution of immunoreactivity was in good general agreement with that predicted from the localisation of sst(3) mRNA and radio-ligand binding studies; however, due to the preference of AS-69 towards the deglycosylated receptor, it appears that the sst(3) immunoreactivity detected may correspond largely to the deglycosylated receptor. This study on the immunohistochemical distribution of the sst(3) receptor in the brain may provide a better understanding of the central actions of somatotropin release-inhibiting factor (SRIF).
生长抑素的作用是通过名为sst(1)至sst(5)的G蛋白偶联受体介导的。我们使用了一种针对sst(3)特定N端肽序列的亲和纯化多克隆抗体AS-69，来确定sst(3)受体在大鼠和人脑中的免疫组织化学分布。通过使用N端sst(3)融合蛋白的蛋白质印迹实验证明了该抗体的特异性。酶促去糖基化实验与对sst(3)转染细胞系和大鼠脑膜蛋白的印迹实验以及对sst(3)转染细胞系的免疫细胞化学实验相结合。这些研究表明该抗体可检测到去糖基化的sst(3)受体蛋白。免疫组织化学染色显示，这种N端抗血清识别的sst(3)免疫反应性广泛分布于整个大脑，大脑皮层、边缘系统区域（包括海马结构、部分杏仁核区域、部分基底神经节核以及基底核复合体区域）、缰核、下丘脑、丘脑、不同的中脑结构（黑质、未定带、上丘）、网状结构、小脑中的细胞和突起均有标记。免疫反应性的分布与根据sst(3) mRNA定位和放射性配体结合研究预测的结果总体上吻合良好；然而，由于AS-69对去糖基化受体的偏好，似乎检测到的sst(3)免疫反应性可能在很大程度上对应于去糖基化受体。这项关于sst(3)受体在脑中免疫组织化学分布的研究可能有助于更好地理解生长抑素释放抑制因子（SRIF）的中枢作用。
