Neural Circuitry in the Developing Zebrafish Spinal Cord

描述（由申请人提供）：斑马鱼胚胎在其存在的第一个24小时内从单细胞变成具有功能神经系统的生物体。受精后24小时，胚胎具有机械感觉介导的反射运动行为，96小时时，鱼是自由游动的幼虫。这些行为的表现是由于在发育中的鱼的脊髓和后脑中建立了一个简单而有效的神经网络。这个简单的神经系统为生物体成熟过程中的进一步扩张提供了支架。阐明斑马鱼早期神经系统形成的机制可能会对我们理解人类出现了多少神经系统疾病产生巨大影响。胚胎/幼体斑马鱼的特性使其非常适合研究神经网络的形成——它是外部受精的，在发育过程中是透明的，并且很容易用于反义处理和转基因等分子技术。使用染料标记的解剖学研究已经获得了这些早期时间点脊髓神经元形态的信息，表明在24-96 HPF之间，脊髓每半段大约存在11-15个神经元。此外，这些研究还提供了一些关于神经元如何相互连接形成网络的见解。然而，这些神经元是如何相互交流的，我们几乎一无所知。这是本提案的目的，以产生转基因鱼线，将使阐明发育中的脊髓的神经化学解剖。这些鱼不仅可以提供关于各种脊髓和棘上通路连接的信息，而且还可以为利用共表达（例如，与钙指示剂）或电生理方法进行功能研究提供标记物。此外，它们可以提供特定神经元类型的标记物，这些标记物可用于前向遗传筛选，旨在识别这些细胞中表型或连接决策所需的基因

DESCRIPTION (provided by applicant): A zebrafish embryo goes from a single-cell to an organism with a functioning nervous system within its first 24 hours of existence. By 24 hours post-fertilization, the embryo has mechanosensory-mediated reflex motor behaviors and by 96 hr the fish is a free-swimming larva. The manifestation of these behaviors is due to the establishment of a simple but effective neural network in the spinal cord and hindbrain of the developing fish. This simple nervous system provides the scaffolding for further expansion during maturation of the organism. Elucidating the mechanisms underlying early nervous system formation in the zebrafish could have a huge impact on our understanding of how many neurological disorders arise in man. The properties of the embryonic/larval zebrafish has made it uniquely suited to study the formation of a neural network- it's fertilized externally, transparent during its development and lends itself easily to molecular techniques such as anti-sense treatment and transgenesis. Anatomical studies using dye labeling have yielded information regarding the morphologies of spinal neurons during these early time points, suggesting the presence of some 11-15 neurons per hemi-segment of the cord between 24-96 HPF. In addition, these studies have provided some insight into how the neurons connect to one another to form a network. However, almost nothing is known about how these neurons communicate amongst each other. It is the purpose of this proposal to generate transgenic fish lines that will enable the elucidation of the neurochemical anatomy of the developing spinal cord. Not only will these fish provide information as to the connectivity of the various spinal and supraspinal pathways, but they will also provide markers for use in functional studies utilizing either co-expression (e.g., with calcium indicators) or electrophysiological approaches. In addition, they can provide markers for specific neuronal types that can be used in forward genetic screens aimed at identifying genes required for either phenotype or connectivity decisions in these cells