The larval 4D standard brain of Drosophila melanogaster on a single cell level

大脑中的行为输出是如何根据外部感官输入、内部动机状态甚至是通过先前经验获得的知识来组织的？理解这些过程是神经科学领域最重要的问题。几个世纪以来，功能脑科学的研究人员一直在将行为属性映射到大脑的各个区域。然而，从简单的地图到一个被普遍接受的模型的步骤已经被证明是极其困难的。在极度简化的模式生物（如秀丽隐杆线虫或灰蛉）中，只有在有限的水平上才能与人类和更复杂的脊椎动物系统（如老鼠、斑马鱼）进行有意义的比较，这些系统具有数量、伦理和技术上的限制，经典的遗传模式生物果蝇已经获得了重要的中间地位。最近的方法要么试图通过连续透射电子显微镜（TEM）建立幼虫大脑的完整连接组，要么基于不同GAL4驱动线的共聚焦显微镜堆栈，为成年大脑的三维脑图谱工作。然而，这两种方法都达到了生物学极限；虽然TEM方法仅限于单个大脑的解剖水平，但允许基于其高分辨率建立连接组，但3D注册允许将解剖数据与功能研究联系起来。然而，到目前为止，由于其大约20万个神经元的数字复杂性，它被限制在成年果蝇大脑的10%左右。因此，我们提出了一种联合方法，以GAL4、LexA和分裂GAL4系为基础，建立了约10000个幼虫大脑神经元的全面4D重建。该项目将通过在神经科学和信息学领域工作的三个小组的独特专业知识来实现；1) Thum实验室将通过免疫组织化学技术和共聚焦显微镜观察这些神经元。2)梅尔霍夫实验室将根据其解剖信息将这些神经元注册到标准大脑上。3) b<s:1> hler实验室将对处理后的数据进行处理、组织和可视化，并将其放入开放存取数据库中。此外，位于HHMI Janelia农场研究园区（美国）的B. Dickson和J. Truman的实验室将通过提供世界上最大的遗传工具集和幼虫大脑解剖的解剖学原始数据来支持这项工作。总之，我们的方法将在4D神经元水平上重建幼虫的大脑；同时，它还将定义一套新的注释（在开放访问数据库中）和免费访问的神经遗传学工具，以研究整个大脑中单个神经元的特定功能。这将是第一次有可能建立一个共同的资源基础，为欧洲快速发展的研究果蝇幼虫的神经生物学社区提供支持。

How is behavior output organized in a brain based on external sensory inputs, internal motivational states or even knowledge gained through prior experience? Understanding these processes is the most essential issue in the field of neuroscience. For centuries, researchers in the functional brain sciences have been mapping properties of behavior to areas of the brain. Yet, the step from simple maps to a generally accepted model has proven exceedingly difficult. Among the extremely reduced model organisms (e.g. C.elegans or Aplysia) that only on a constricted level allow for meaningful comparisons with humans, and more complex vertebrate systems (e.g. mouse, zebra fish) that have numerical, ethical and technical constraints, the classical genetic model organism Drosophila has acquired an important intermediate position. Recent approaches either try to establish the complete connectome of the larval brain by serial transmission electron microscopy (TEM) or work on a 3D brain atlas for its adult counterpart based on confocal microscopy stacks of different GAL4 driver lines. However, both approaches have reached biological limits; whereas the TEM method is restricted to the anatomical level of a single brain but allows for the establishment of the connectome based on its high resolution, the 3D registration allows linking anatomical data to functional studies. However, so far it is restricted to about 10% of the adult Drosophila brain due to its numerical complexity of about 200.000 neurons.Therefore, we propose a joined approach to establish a comprehensive 4D reconstruction of the about 10.000 neurons of the larval brain based on GAL4, LexA and split-GAL4 lines. The project will be realized by the unique expertise of three groups working in the field of neuroscience and informatics; 1) The Thum lab will visualize these neurons by immunohistochemical techniques and confocal microscopy. 2) The Merhof lab will register these neurons based on its anatomical information onto a standard brain. 3) The Bühler lab will process, organize and visualize the processed data in an open access database. In addition, the labs of B. Dickson and J. Truman, both located at the HHMI Janelia Farm Research Campus (USA) will support the endeavor by offering access to the world-wide largest set of genetic tools and anatomical raw data on larval brain anatomy. In summary, our approach will reconstruct the larval brain in 4D on the neuronal level; at the same time it will also define a new set of annotated (in an open access database) and freely accessible neurogenetic tools to investigate the particular function of single neurons within an entire brain. By that - for the first time - it will be possible to establish a common resource base that offers support for the rapidly growing neurobiology community in Europe working on the Drosophila larva.