Trophic Interactions During Visual System Development

描述(申请人提供)：netrin-1是一种分泌分子，在轴突引导过程中既可以作为引诱剂，也可以作为驱避剂。在视觉系统中，netrin被认为与眼睛外的视神经轴突导向大脑中的目标有关。我们的长期目标是阐明控制视网膜和其中心靶点之间适当连接发展的调节机制。在这里，我们建议检验这一假设，即netrin-1与视网膜顶盖发育的后期有关，这些方面显著影响视觉功能。具体地说，我们的目的是证明：1)netrin-1通过特定受体亚型上的独立信号，影响靶区的视网膜轴突和它们在大脑中的突触后神经元；2)正确的视网膜顶盖连接需要netrin和脑源性神经营养因子(BDNF)之间的功能相互作用，BDNF是一种调节突触发生的营养因子。这些假说的基础是：i)netrin在视网膜轴突发现后影响视网膜轴突，ii)在结直肠癌中缺失的两个netrin受体(DCC)和UNC-5在视网膜顶盖发育过程中以互补的方式表达，iii)BDNF和netrin共享共同的信号机制来影响轴突引导。我们将在活着的非洲爪哇蝌蚪身上进行实时成像实验以及在空间和时间上控制netrin信号的操作，以检验netrin在视觉系统发育中的作用。有待检验的具体假设是：1.netrin-1通过激活DCC受体，指示视网膜轴突终止于何处，并塑造视轴突杆结构和突触连接。在视顶盖内对Netrin水平和DCC信号的操纵将被用来分析Netrin在轴突靶向、分支和突触前位置分化中的作用。2.netrin-1通过UNC-5受体信号影响突触后顶盖神经元的形态和突触发育。体内成像实验将被用来确定Netrin是否参与早期树突状细胞的发生，控制先锋树突起的方向，以及Netrin是否调节树突分支和突触后位置分化。3.Netrin和BDNF在功能上相互作用，形成视网膜顶盖突触连接。分子和体内成像研究将检查通过DCC受体的netrin信号是否位于BDNF调节视网膜轴突分支和突触形成的上游。BDNF拯救受DCC信号改变影响的视网膜轴突的能力也将得到测试。总之，这些拟议的研究将揭示Netrin-1是视网膜顶盖电路发育的关键角色，并将加深我们对控制视觉通路发育的机制的理解，而视觉通路在维持正常视觉功能方面至关重要。

DESCRIPTION (provided by applicant): Netrin-1 is a secreted molecule that can function as either an attractant or repellent during axon guidance. In the visual system, netrin has been implicated in the guidance of optic axons out of the eye towards their targets in the brain. Our long-term goal is to elucidate the regulatory mechanisms controlling the development of proper connections between the retina and its central targets. Here, we propose to test the hypothesis that netrin-1 is implicated in late aspects of retinotectal development that significantly impact visual function. Specifically, we aim to demonstrate that; 1) netrin-1, through independent signaling on specific receptor subtypes, influences retinal axons at the target and their postsynaptic neurons in the brain, and 2) proper retinotectal wiring requires functional interactions between netrin and brain-derived neurotrophic factor (BDNF), a trophic factor that modulates synaptogenesis. We base these hypotheses on the observations that; i) netrin influences retinal axons past the period of retinal axon pathfinding, ii) the two netrin receptors deleted-in-colorectal cancer (DCC) and UNC-5 are expressed in complementary patterns during retinotectal development and, iii) BDNF and netrin share common signaling mechanisms to influence axon guidance. Real-time imaging experiments and spatially and temporally controlled manipulations in netrin signaling will be performed in living Xenopus tadpoles to examine netrin's role in visual system development. The specific hypotheses to be tested are: 1. Netrin-1, through DCC receptor activation, instructs retinal axons where to terminate and shapes optic axon arbor structure and synaptic connectivity. Manipulations in netrin levels and DCC signaling within the optic tectum will be used to analyze the role of netrin in axon targeting, branching, and presynaptic site differentiation. 2. Netrin-1, through UNC-5 receptor signaling, influences the morphological and synaptic development of postsynaptic tectal neurons. In vivo imaging experiments will be used to establish whether netrin participates in early dendritogenesis, controlling the orientation of pioneer dendritic processes and whether netrin modulates dendritic branching and postsynaptic site differentiation. 3. Netrin and BDNF interact functionally to shape retinotectal synaptic connectivity. Molecular and in vivo imaging studies will examine if netrin signaling through DCC receptors is upstream of BDNF regulation of retinal axon branching and synaptogenesis. The ability of BDNF to rescue retinal axons affected by altered DCC signaling will also be tested. Collectively, the proposed studies will reveal that netrin-1 is a key player in retinotectal circuit development and will further our understanding of the mechanisms that control the development of visual pathways, that are critically important in the maintenance of normal visual function.