DEVELOPMENT OF OLFACTORY CONNECTIONS

描述(由申请人提供)：该项目的主要目标是确定嗅觉系统中二尖瓣细胞树突的潜在机制。精确的轴突和树突模式的形成是建立大脑功能神经回路的关键。在嗅觉系统中，嗅觉感觉神经元轴突对其靶点的导向机制最近被研究。然而，对于嗅神经突触后伙伴--二尖瓣细胞树突的分化和靶向的分子机制尚缺乏研究。关于二尖瓣/簇状细胞树突发生的控制，我们提出了以下假设：1)二尖瓣/簇状细胞树突的形态发生需要与嗅神经相互作用。嗅神经对二尖瓣细胞突起延伸的影响将通过体内和体外方法进行测试。将表征二尖瓣细胞树突的发育变化。二尖瓣/簇状细胞树突的定向将使用3D胶原基质进行研究。成熟的嗅轴突在二尖瓣细胞树突发育中的作用将通过遗传条件消融来检验。2)嗅神经提供可扩散的分子线索，促进二尖瓣/簇状神经元的树突伸展和分支。这一假设将通过分离的嗅球培养进行直接检验。使用嗅觉上皮条件培养液检测是否存在促进二尖瓣细胞树突状生长的扩散因子。在分离的嗅球神经元培养中，将检测候选分子、信号素、神经营养素和骨形态发生蛋白的树突生长促进活性。嗅觉上皮细胞的活性将通过凝胶过滤和使用FPLC的离子交换层析来表征。OE衍生活性的分子同一性将通过多肽测序获得。我们研究的长期目标是了解对神经系统发育至关重要的精确的细胞和分子相互作用，并深入了解发育性神经疾病和出生缺陷的原因。

DESCRIPTION (provided by applicant): The main objective of the project is to determine the underlying mechanisms that pattern the mitral cell dendrites in the olfactory system. Formation of precise axonal and dendritic pattern is crucial for establishing functional neural circuitry in the brain. In the olfactory system, the guidance mechanisms of the olfactory sensory neuron axons toward their target have been investigated recently. However, studies are still lacking for the molecular mechanisms of the differentiation and the targeting of the mitral cell dendrites, the postsynaptic partner of the olfactory nerve. We propose to investigate the following hypotheses concerning the control of mitral/tufted cell dendritogenesis: 1) Interaction with olfactory nerve is required for the morphogenesis of mitral/tufted cell dendrites. The influence of the olfactory nerve on mitral cell neurite extension will be tested using in vivo and in vitro approaches. The developmental changes of mitral cell dendrites will be characterized. The orientation of the mitral/tufted cell dendrites will be investigated using 3D collagen matrix. The role of the] mature olfactory axons in mitral cell dendritic development will be examined using genetic conditional ablation. 2) Olfactory nerve provides diffusible molecular cues to promote dendritic extension and branching of the mitral/tufted neurons. This hypothesis will be directly tested using dissociated olfactory bulb culture. The presence of the diffusible factors for promoting mitral cell dendritic growth will be examined using olfactory epithelium conditioned medium. Candidate molecules, Semaphorins, neurotrophins and BMPs will be examined for their dendrite growth promoting activity in the dissociated olfactory bulb neuron cultures. Olfactory epithelium derived activity will be characterized by gel filtration and ion exchange chromatography using FPLC. Molecular identity of the OE derived activity will be obtained by peptide sequencing. The long-term goal of our study is to understand the precise cellular and molecular interactions that are critical for the development of the nervous system and to gain insight into the causes of developmental neurological disorder and birth defects.