Molecular Mechanisms of Trigeminal Development

三叉神经发育的分子机制

• 批准号: 7470073
• 负责人: MARK F JACQUIN
• 金额: $ 117.19万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2005
• 资助国家: 美国
• 起止时间: 2005-04-01 至 2010-12-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7470073
• 关键词: Molecular; Mechanisms; Trigeminal; Development

Description (provided by applicant): During the current funding cycle, we revealed trophic and tropic actions of neurotrophins on specific classes of developing trigeminal (V) primary afferent neurons, and showed that arborization and elongation effects of neurotrophins are mediated by the Rho family GTPases. Proposed experiments address the differentiation of V ganglion cells and their projections in mice with targeted deletion of specific genes known to be necessary for sensory neuron survival. Mice with deletion of a preapoptotic gene (bax) will be crossed with NGF-, NT3-,TrkA-, or TrkC-knockout mice to reveal axonal differentiation effects of neurotrophins on V ganglion cells, independent of their cell survival effects. To address chemotropic actions of NGF and NTS on embryonic V afferent projections, loss-of-function and gain-of-function studies will be performed. Synaptotrophic actions of NGF and NTS will be analyzed in these various preparations by ultrastructural and immunohistochemical studies. Pilot data prompt the following 3 hypotheses and experiments: 1. In Bax/NGF or Bax/NTS double knockout mice, all classes of primary sensory neurons in the V ganglion will survive, but they display deficits in axon differentiation, therein preventing whisker related brainstem pattern formation. Cell death markers, stereologic estimates of cell numbers (Core A), and cell class-specific markers will be used to assess embryonic V ganglion cell survival. Axonal labeling and quantification of projections will reveal changes in axonal differentiation relative to wildtype controls (Core B). Metabolic staining will reveal effects on CNS pattern formation (Core A). 2. Neurotrophins facilitate synaptogenesis in developing V primary afferents. Wholemount cultures of the V pathway subjected to exogenous neurotrophin augmentation regimens will be used to study development of synaptic protein expression and synaptic profiles in V brainstem nuclei (Core A). Complementary analyses will be performed in vivo in TrkA and TrkC knockout embryos to study synaptic development in the absence of high-affinity receptors for NGF and NTS. These 2 experiments will reveal NGF family actions in V system development. We further hypothesize: 3. The GDNF family ligands (GDNF, neurturin, artemin) contribute to perinatal V ganglion cell survival and axon development. V primary afferent survival, differentiation, phenotypes, morphology and patterning will be assessed in comparable gene targeting and factor augmentation wholemount paradigms as they pertain to the GDNF family ligands and receptors (Core A). Chemotropic effects will also be tested by using isolated V ganglia embedded with a localized source of GDNF family ligand. V axon growth towards or away from this GDNF source will be analyzed (Core B).

描述（由申请人提供）： 在当前的融资周期中，我们揭示了神经营养蛋白对特定类型的三叉神经（V）原发性传入神经元的营养和热带作用，并表明神经营养蛋白的弧形和伸长作用是由Rho家族GTPases介导的。提出的实验解决了V神经节细胞的分化及其在小鼠中的投影，其靶向缺失的特定基因已知是感觉神经元存活所必需的。缺失丙型基因（BAX）的小鼠将与NGF-，NT3-，TRKA-或TRKC-KNOCKOUT小鼠交叉，以揭示神经营养蛋白对V神经节细胞的轴突分化效应，与其细胞存活效应无关。为了解决NGF和NTS对胚胎V传入预测的趋化作用，将进行功能丧失和功能障碍研究。 NGF和NTS的突触营养作用将通过超微结构和免疫组织化学研究在这些制剂中进行分析。 PILOT数据促使以下3个假设和实验：1。在Bax/NGF或Bax/NTS双基因敲除小鼠中，V神经节中的所有主要感觉神经元将存活，但它们在轴突差异中显示出缺陷，其中在其中阻止了晶须相关的脑干模式形成。细胞死亡标记，细胞数（核心A）的立体估计和细胞类特异性标记将用于评估胚胎V神经节细胞的存活。轴突标记和投影的定量将揭示相对于野生型对照（核心B）的轴突分化的变化。代谢染色将揭示对中枢神经系统模式形成的影响（核心A）。 2。神经营养蛋白促进开发V主要传入的突触发生。经受外源性神经营养蛋白增强方案的V途径的全长培养物将用于研究V脑干核中突触蛋白表达和突触特征的开发（核心A）。在没有NGF和NTS的高亲和力受体的情况下，将在TRKA和TRKC敲除胚胎中在体内进行互补分析，以研究突触发育。这两个实验将揭示NGF家庭在V系统开发中的行动。我们进一步假设：3。GDNF家族配体（GDNF，Neurturin，Artemin）有助于围产期V神经节细胞存活和轴突发育。 v主要传入的生存，分化，表型，形态和模式将在与GDNF家族配体和受体（核心A）有关的同类基因靶向和因子增强定位范式中进行评估。趋化效应还将通过使用嵌入局部GDNF家族配体来源的分离的V神经节来测试。 V将分析a轴突在该GDNF源方面或远离该GDNF源的生长（核心B）。