一种在体标记成体大鼠室管膜/室下区细胞的方法

目的 研究正常成体大鼠侧脑室注射DiI标记室管膜/室下区(SVZ)细胞的方法.方法 50只雄性SD大鼠随机分为5组,每组10只,均接受2 g/L的 DiI 10 μL右侧脑室注射.采用H33258染色和激光共聚焦显微镜检测DiI注射后6 h、12 h、24 h、36 h和48 h时左侧脑室壁的标记细胞以及标记组织厚度.结论 右侧脑室DiI注射后24 h,DiI标记细胞位于左侧脑室的室管膜层; DiI注射后48 h,DiI标记细胞限定于左侧室的管膜层和室下区.此外,左侧室管膜/中隔室下区(SVZspt)和室管膜/神经节隆起的生后对应物(SVZge)部位荧光标记组织的厚度分别在DiI注射12 h和24 h后维持于稳定水平,而且,在相应各时间点上,室管膜/SVZge部位荧光标记组织的厚度都厚于室管膜/SVZspt部位(P＜0.05).结论 2 g/L 的DiI 10 μL注射于正常大鼠侧脑室后24～48 h,可能仅标记室管膜/室下区细胞.

In vivo Labeling of Cells in the Ependyma/Subventricular Zone of Adult Rats

OBJECTIVE: To identify the labeled cells in the ependyma/subventricular zone (SVZ) of normal adult rats by DiI injected into the lateral ventricle. METHODS: Fifty male Sprague-Dawley rats were divided randomly into five groups (10 per group). All of the rats were injected with 10 microL of 2 g/L fluorescence dye DiI into the right lateral ventricle. The five groups of rats were sacrificed at 6 h, 12 h, 24 h, 36 h or 48 h after injection respectively. Hoechst 33258 staining was used to identify nuclei and laser confocal microscopy was used to detect the DiI-labeled cells and to measure the thickness of the tissue with DiI fluorescence in the wall of the left lateral ventricle. RESULTS: After injection of the DiI into the right lateral ventricle, DiI- Hoechst 33258 double positive cells were found in the ependymal layer of the left lateral ventricular wall at 24 h and in the SVZ at 48 h as well. The thickness of the tissue with DiI fluorescence in the left ependyma/septal subventricular zone (SVZspt) and ependyma/postnatal equivalent of the ganglionic eminences (SVZge) remained unchanged at 12 h and 24 h after DiI injection. The thickness of the tissue with DiI fluorescence in the left ependyma/SVZge was significantly greater than that in the ependyma/SVZspt at all of the time points (P<0.05). CONCLUSION: The ependyma/SVZ cells can be labeled by Dil 24-48 h after injection (10 microL of 2 g/L) into the lateral ventricle.