The First Comprehensive Neural Connectivity Map of Mouse

描述（由申请人提供）：该申请涉及广泛的挑战领域（15）：转化科学和特定的挑战主题，15- MH-103绘制小鼠模型的神经连通性。大脑的功能是由它的电路决定的，但我们对它的线路结构知之甚少：在研究最多的哺乳动物（老鼠）中，估计只有10-30%的远程电路连接被探测到。目前的挑战主题验证了越来越多的共识，即是时候通过生成模型脊椎动物的全脑连接图来缩小这一差距。在过去的两年中，我们组织了几次涉及神经解剖学社区的会议，以深入了解这样一个项目的技术和科学挑战。基于这一经验，我们已经设计并开始建立和测试一个自动化的实验和计算技术管道来实现这一目标。自动化宽视场载玻片扫描显微镜技术的进步、数据存储成本的降低，以及使用注射经典示踪剂和工程病毒建立的途径追踪方法，使我们的建议成为可能。实验方案可以总结如下。根据经典的神经解剖学和区域基因表达数据，将小鼠大脑划分为约200个区域。在每个区域，我们给一只小鼠注射经典示踪剂，另一只小鼠注射病毒示踪剂。示踪剂从注射部位顺行运输到该区域的投射靶，并逆行运输到该区域投射到注射部位的区域。通过这种方式，单个投影被多次显示出来。为了获得这些信息，我们将对每只老鼠的整个大脑进行切片，并使用自动滑动扫描显微镜对切片进行成像。由此产生的二维切片图像将在软件中组合，为每次注射生成三维重建的大脑图像。最后，来自所有个体注射的3D图像将通过空间注册到艾伦参考图集来组合，最终生成统一的全脑神经连接图。在老鼠身上生成第一个无偏见的全脑连接图将具有广泛的神经科学意义。神经发育、神经网络建模、进化神经解剖学以及联想和综合脑功能的研究将从最终拥有这一具有里程碑意义的参考图中受益匪浅，它将有意义地约束理论，并有助于实验设计和结果解释。通过分析Allen研究所生成的基因表达图谱，结合本项目生成的连通性图谱，可以探索基因表达与连通性之间的关系。本研究中生成的基线神经连接图将作为随后研究突变小鼠系之间电路多态性的基础。客观量化神经精神疾病（如自闭症和精神分裂症）小鼠模型中连通性变化的能力将有助于我们了解其病因和病理生理学。最后，我们强调开源软件开发，成本优化和可重复性将导致其他学术实验室能够实施的负担得起的集成仪器，因此这种方法可以快速应用于各种各样的神经科学问题。

DESCRIPTION (provided by applicant): This application addresses broad Challenge Area (15): Translational Science and specific Challenge Topic, 15- MH-103 Mapping the Neural Connectivity of a Mouse Model. Brain function is dictated by its circuitry, yet we know little about its wiring architecture: in the most-studied mammal (rat), only an estimated 10-30% of the long range circuit connections have been probed. The present Challenge Topic validates the growing consensus that it is time to close this gap by generating brainwide connectivity maps for model vertebrates. Over the last two years, we have organized several meetings involving the neuroanatomy community to gain in-depth understanding of the technical and scientific challenges of such a project. Based on this experience, we have designed and have begun to build and test an automated pipeline of experimental and computational techniques for achieving this goal. Our proposal is enabled by advances in automated wide-field slide scanning microscopy, decreasing data-storage costs, and established tract-tracing methods using injections of classical tracers and engineered viruses. The experimental plan can be summarized as follows. The mouse brain is divided into ~200 regions based on classical neuroanatomical and regional gene-expression data. For each region we inject one mouse with classical tracers and one mouse with viral tracers. From the injection site, the tracers are transported anterogradely to the area's projection targets and retrogradely to areas which project to the injection site. In this way, individual projections are revealed multiple times. In order to acquire this information, we will section the entire brain from each mouse and image the sections using an automated slide-scanning microscope. The resulting 2D slice-images will be combined in software to produce a 3D reconstructed brain image for each injection. Finally the 3D images from all of the individual injections will be combined by spatially registering them to the Allen Reference Atlas, ultimately generating a unified brainwide neural connectivity map. Generating the first unbiased, brainwide connectivity map in the mouse will have broad neuroscientific implications. The study of neural development, neural network modeling, evolutionary neuroanatomy, and associative and integrative brain function will benefit tremendously from finally having this landmark reference map to meaningfully constrain theories and aid in experimental design and interpretation of results. Relationships between gene expression and connectivity can be probed by analyzing the gene-expression maps generated by the Allen Institute in combination with the connectivity maps generated by this project. The baseline neural connectivity map generated in the present study will serve as a foundation for subsequently studying circuit polymorphisms across mutant mouse lines. The ability to objectively quantify alterations in connectivity in mouse models of neuropsychiatric disorders such as autism and schizophrenia will aid our understanding of their etiology and pathophysiology. Finally, our emphasis on open source software development, cost optimization and duplicability will result in an affordable, integrated instrument which other academic laboratories will be able to implement, so that this approach can be rapidly applied to a wide variety of neuroscientific problems. NARRATIVE The study of mouse models of neuropsychiatric disorders provides hope for the development of therapies for these burdensome illnesses, but progress has been slow due to the lack of knowledge about how the mouse brain is wired. This project aims to close this gap by generating the first brain-wide wiring diagram of mouse, automating techniques that are known to work but are labor-intensive. If successful, the project has the potential to fundamentally transform our understanding of the architecture of the normal and disordered brain.