MacCumber M W, Snyder S H, Ross C A
Department of Neuroscience, Johns Hopkins University School of Medicine, Baltimore, Maryland 21205.
J Neurosci. 1990 Aug;10(8):2850-60. doi: 10.1523/JNEUROSCI.10-08-02850.1990.
Carboxypeptidase E (CPE), also referred to as enkephalin convertase or carboxypeptidase H (EC 3.4.17.10), is present in neurotransmitter secretory granules and can remove C-terminal basic residues following endopeptidase cleavage during peptide processing. Using in situ hybridization with 35S-labeled oligonucleotide probes, we have mapped the localization of CPE mRNA in the rat brain. Specificity for CPE was confirmed by control experiments, which included production of identical patterns hybridization with 3 different antisense oligonucleotide probes, loss of label with RNase pretreatment of sections or co-incubation with excess unlabeled probe, and lack of labeling with sense orientation probes. In addition, the regional distribution of CPE mRNA by Northern blot analysis corresponded with distribution of labeling by in situ hybridization. The highest levels of CPE mRNA were found to be present in the pyramidal cells of the hippocampus, the pituitary anterior and intermediate lobes, the ependymal cells of the lateral ventricle, the endopiriform nucleus, the basolateral amygdala, the supraoptic nucleus, and the paraventricular nucleus. Intermediate levels were present in the thalamus, medial geniculate nucleus, lateral septal nucleus, piriform and entorhinal cortex, nucleus of the tractus solitarius, cerebellar cortex, pontine nuclei, and inferior olive. The lowest levels were found in the hippocampal granule cell layer, lateral hypothalamus, globus pallidus, and brain stem reticular formation. Ibotenic acid lesions of the hippocampus eliminated the majority of the label, which had been present over pyramidal cells, though labeling was increased over areas of reactive gliosis, suggesting that activated astrocytes can also synthesize CPE mRNA. In general, the localization of CPE mRNA in the rat brain corresponded to the distribution of enkephalin and other peptide neurotransmitter-synthesizing neurons, though CPE mRNA was also present in neurons that do not secrete known peptides and in reactive glia. The widespread yet specific localization of CPE mRNA in the rat brain suggests it may be an excellent marker for peptide synthesizing cells in the CNS.
羧肽酶E（CPE），也被称为脑啡肽转换酶或羧肽酶H（EC 3.4.17.10），存在于神经递质分泌颗粒中，在肽加工过程中，可在内肽酶切割后去除C端碱性残基。我们使用35S标记的寡核苷酸探针进行原位杂交，绘制了大鼠脑中CPE mRNA的定位图。对照实验证实了对CPE的特异性，这些实验包括用3种不同的反义寡核苷酸探针产生相同的杂交模式、切片经核糖核酸酶预处理或与过量未标记探针共孵育后信号消失以及正义方向探针无标记。此外，通过Northern印迹分析的CPE mRNA区域分布与原位杂交的标记分布相对应。发现CPE mRNA水平最高的部位是海马锥体细胞、垂体前叶和中间叶、侧脑室室管膜细胞、内梨状核、基底外侧杏仁核、视上核和室旁核。中等水平存在于丘脑、内侧膝状体核、外侧隔核、梨状和内嗅皮质、孤束核、小脑皮质、脑桥核和下橄榄核。最低水平见于海马颗粒细胞层、外侧下丘脑、苍白球和脑干网状结构。海马的鹅膏蕈氨酸损伤消除了锥体细胞上大部分原本存在的标记，不过在反应性胶质增生区域标记增加，这表明活化的星形胶质细胞也能合成CPE mRNA。总体而言，大鼠脑中CPE mRNA的定位与脑啡肽和其他肽类神经递质合成神经元的分布相对应，尽管CPE mRNA也存在于不分泌已知肽的神经元和反应性胶质细胞中。大鼠脑中CPE mRNA广泛而特异的定位表明它可能是中枢神经系统中肽合成细胞的一个优秀标记物。
