Research on mechanisms of development and collapse of the pial and glial limiting membranes, which comprise the cerebrospinal fluid-brain barrier

组成脑脊液-脑屏障的软脑膜和神经胶质限制膜的发育和塌陷机制研究

基本信息

批准号：
16500219
负责人：
KIKKAWA Satoshi
金额：
$ 1.92万
依托单位：
Kobe University
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2004
资助国家：
日本
起止时间：
2004 至 2005
项目状态：
已结题

项目摘要

We anterogradely labeled entorhinodentate axons in wild type and reeler mice at adult and neonatal stages. The results revealed that pioneer axons of the entrhinodentate tract develop similarly and pass through the hippocampal fissure at postnatal day 1(P1) in wild type and reeler mice. In the reeler mutant, however, follower axons can not pass through the fissure and detour around it, whereas those in wild type mice follow their pioneer and continue to pass through the fissure. Immunohistochemical analysis of postnatal development of the hippocampal area using anti-GFAP antibody demonstrated aberrant accumulation of GFAP-positive astroglia along the hippocampal fissure in the reeler but not wild type brain. This accumulation of astroglia seems to prevent the entorhinohippocampal axons from passing through the hippocampal fissure and forces them to detour along the fissure in the reeler mutant.Next, we double-labeled neocortical neurons in the fukutin-deficient chimeric mouse with antero- or retro-grade axonal tracers and anti-laminin antibody to investigate relationship between collapse of the glial limiting membranes due to fukutin-deficiency and malpositioning of neocortical neurons. The results showed that a population of neocortical neurons positioned ectopically in the area just beneath the disrupted limiting membranes which lack laminin immunoreactivity, suggesting that the glial limiting membranes have certain effects on cortical neuronal migration.Thirdly, we cloned and sequenced the complete coding region of zebrafish fukutin. Zebrafish fukutin showed a significant degree of sequence conservation. Fukutin mRNA was detected a wide variety of tissues including the brain and the spinal cord. We also tried knocking-down fukutin protein expression by antisense morpholino oligonucleotide microinjection into fertilized eggs. Further investigation of effects of fukutin knock-down on development of the central nervous system is currently being continued.

我们在成年和新生儿阶段的野生型和卷轴小鼠中顺行标记内鼻齿状轴突。结果显示，野生型和卷轴小鼠的内齿齿道先锋轴突发育相似，并在出生后第 1 天（P1）穿过海马裂。然而，在卷轴突变体中，跟随轴突不能穿过裂隙并绕道绕行，而野生型小鼠中的跟随轴突则跟随它们的先驱并继续穿过裂隙。使用抗 GFAP 抗体对海马区出生后发育进行的免疫组织化学分析表明，在卷取器中，GFAP 阳性星形胶质细胞沿着海马裂隙异常积累，但野生型大脑中没有。这种星形胶质细胞的积累似乎阻止了内鼻海马轴突穿过海马裂隙，并迫使它们沿着 reeler 突变体的裂隙绕行。接下来，我们用前向或后向双标记 fukutin 缺陷嵌合小鼠的新皮质神经元。级轴突示踪剂和抗层粘连蛋白抗体，用于研究由于 fukutin 缺乏而导致的神经胶质限制膜塌陷之间的关系和新皮质神经元的错位。结果表明，在缺乏层粘连蛋白免疫反应性的破坏的限制膜正下方区域有一群异位的新皮质神经元，这表明胶质限制膜对皮质神经元的迁移有一定的影响。第三，我们克隆并测序了完整的编码区斑马鱼福亭。斑马鱼 fukutin 显示出显着程度的序列保守性。 Fukutin mRNA 在多种组织中均检测到，包括大脑和脊髓。我们还尝试通过将反义吗啉寡核苷酸显微注射到受精卵中来敲低 fukutin 蛋白的表达。目前正在继续进一步研究 fukutin 敲除对中枢神经系统发育的影响。