MOLECULAR CLONING OF BLOOD/BRAIN BARRIER NUTRIENT TRANSPORTERS

本项目的长期目标是克隆、测序、表达和定位 营养转运蛋白在血脑和血液中的分布， 脊髓液屏障 大脑的内皮细胞 毛细血管调节营养物质进入大脑的吸收， 脉络丛的上皮细胞清除脑脊液， 许多代谢物和药物。 关于这一点，我们所知甚少。 参与这些重要过程的运输者。 我们会克隆血液- 脑屏障转运蛋白使用两种不同的策略。 一是 以确定已经从其他组织克隆的转运蛋白是否 表达于血脑屏障。 克隆大脑的候选人 毛细血管是肾脏Na+/肌肌醇协同转运蛋白，因为分离 脑微血管运输这种重要的磷脂前体， 脉络丛的候选者是肾Na+/核苷 协同转运蛋白 北方和西方印迹，原位杂交，和 免疫细胞化学技术将被用来绘制分布， 血脑界面的候选克隆人 第二个策略是 利用表达克隆技术分离特定的脑毛细血管 和脉络丛转运蛋白，例如L-型氨基酸和Na+- 依赖脯氨酸转运蛋白。 目标是使用以下工具： 细胞和分子生物学，以获得营养素的详细了解 以及血液和大脑之间的代谢物运输。

The long term goal of this project is to clone, sequence, express and map the distribution of nutrient transporters in the blood-brain and blood- cerebrospinal-fluid barriers. The endothelial cells of the brain capillaries regulate the uptake of nutrients into the brain, and the epithelial cells of the choroid plexus clear the cerebrospinal fluid of many metabolites and drugs. Relatively little is known about the transporters involved in these vital processes. We will clone blood- brain barrier transporters using two different strategies. The first is to determine if the transporters already cloned from other tissues are expressed at the blood-brain-barrier. A candidate clone for brain capillaries is the renal Na+/myo-mositol cotransporter, since isolated brain microvessels transport this important phospholipid precursor, and a candidate for the choroid plexus is the renal Na+/nucleoside cotransporter. Northern and Western blots, in situ hybridization, and immunocytochemical techniques will be used to map the distribution of candidate clones at the blood-brain interface. The second strategy is to use expression cloning techniques to isolate specific brain capillary and choroid plexus transporters, e.g. The L-type amino acid and Na+- dependent proline transporters. The objective is to use the tools of cell and molecular biology to obtain a detailed understanding of nutrient and metabolite transport between the blood and brain.