Cellular and Molecular Mechanisms of Forebrain Axon Pathfinding Defects in FASD

项目摘要 尽管开展了提高公众认识的运动，但胎儿酒精谱系障碍（FASD）仍然普遍存在， 孕妇或育龄妇女饮酒。很好地理解， 和运动信息处理在FASD中被扭曲，但酒精对丘脑的影响， 调节这些功能，尚未得到广泛研究。产前酒精暴露（1）破坏轴突 丘脑和皮质之间的连接，需要适当地接收和响应 环境中的感觉线索，（2）损害腹侧端脑（vTel），一个关键的中间目标 这些连接的正确发展，（3）改变指导线索和信号通路， 是细胞和轴突迁移的基础。我们假设适度的酒精暴露会破坏Slit/Robo 通路，一个家庭的指导线索和受体，可以有直接和间接的影响前脑 轴突形成在这个建议中，我们将研究神经细胞介导的丘脑皮层轴突（TCA）的指导 使用FASD小鼠模型进入vTel和vTel内。我们的实验目的是（1）确定TCA的性质 和走廊细胞的引导错误，（2）建立Slit和Robo作为FASD的分子靶点 体内神经病理学为了分析TCA引导错误，我们将使用染料示踪和免疫染色， 沿着沿着想象这些轴突。为了评估走廊单元，我们将计算并确定 通过免疫染色观察这些细胞的分布。分析酒精引起的Slit和Robo扰动 为了在体内表达，我们将对每种同种型进行蛋白质印迹和RT-PCR。我们还将在体外使用 外植体共培养以测试Slit和Robo功能。在FASD的大脑中，我们预计TCA会不适当地投射 到中间和最终目标以及变形的走廊。我们还预测，Slit/Robo表达 功能将被抑制。我们的研究结果将揭示适度饮酒对指导的影响 神经系统正常布线所需的机制，这是目前研究不足的领域。这 这项工作还将提高对可能参与的细胞和分子机制的理解， 在人类FASD中观察到的感觉运动处理缺陷。拟议的实验将在很大程度上进行 由本科生指导，为他们提供成功所需的指导和专业知识 在科学事业和研究生工作中。

PROJECT SUMMARY Despite public awareness campaigns, Fetal Alcohol Spectrum Disorders (FASD) remain prevalent due to alcohol consumption by women who are pregnant or of child-bearing age. It is well appreciated that sensory and motor information processing is distorted in FASD, but alcohol effects on the thalamus, which likely mediates these functions, have not been widely investigated. Prenatal alcohol exposure (1) disrupts axonal connections between the thalamus and cortex which are required to appropriately receive and respond to sensory cues in the environment, (2) damages the ventral telencephalon (vTel), a critical intermediate target for proper development of these connections, and (3) alters guidance cues and signaling pathways that underlie cell and axon migration. We hypothesize that moderate alcohol exposure disrupts the Slit/Robo pathway, a family of guidance cues and receptors that can have both direct and indirect effects on forebrain axon formation. In this proposal we will investigate corridor-cell mediated thalamocortical axon (TCA) guidance into and within the vTel using a mouse model of FASD. Our experiments aim to (1) define the nature of TCA and corridor cell guidance errors in FASD and (2) establish Slit and Robo as molecular targets of FASD neuropathology in vivo. To analyze TCA guidance errors, we will use dye tracing and immunostaining to visualize these axons along their trajectory. To evaluate corridor cells, we will count and determine the distribution of these cells via immunostaining. To analyze alcohol-induced perturbations of Slit and Robo expression in vivo, we will perform western blotting and RT-PCR for each isoform. We will also use in vitro explant co-cultures to test Slit and Robo function. In the FASD brain, we expect TCAs to project inappropriately to intermediate and final targets and the corridor to be malformed. We also predict that Slit/Robo expression and function will be suppressed. Our findings will reveal the impact of moderate alcohol exposure on guidance mechanisms that are required for proper nervous system wiring, an area that is currently understudied. This work will also enhance understanding of cellular and molecular mechanisms that are likely to be involved in sensorimotor processing defects observed in human FASD. The proposed experiments will largely be carried out by undergraduates who will be mentored to provide them the guidance and expertise needed for success in science careers and graduate work.