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Whole-brain serial-section electron microscopy in larval zebrafish

机译：幼虫斑马鱼全脑连续切片电子显微镜

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High-resolution serial-section electron microscopy (ssEM) makes it possible to investigate the dense meshwork of axons, dendrites, and synapses that form neuronal circuits(1). However, the imaging scale required to comprehensively reconstruct these structures is more than ten orders of magnitude smaller than the spatial extents occupied by networks of interconnected neurons(2), some of which span nearly the entire brain. Difficulties in generating and handling data for large volumes at nanoscale resolution have thus restricted vertebrate studies to fragments of circuits. These efforts were recently transformed by advances in computing, sample handling, and imaging techniques(1), but high-resolution examination of entire brains remains a challenge. Here, we present ssEM data for the complete brain of a larval zebrafish (Danio rerio) at 5.5 days post-fertilization. Our approach utilizes multiple rounds of targeted imaging at different scales to reduce acquisition time and data management requirements. The resulting dataset can be analysed to reconstruct neuronal processes, permitting us to survey all myelinated axons (the projectome). These reconstructions enable precise investigations of neuronal morphology, which reveal remarkable bilateral symmetry in myelinated reticulospinal and lateral line afferent axons. We further set the stage for whole-brain structure-function comparisons by co-registering functional reference atlases and in vivo two-photon fluorescence microscopy data from the same specimen. All obtained images and reconstructions are provided as an open-access resource.
机译：高分辨率的串行截面电子显微镜（ssEM）使研究轴突，树突和形成神经元回路的突触的密集网状结构成为可能（1）。但是，全面重建这些结构所需的成像比例要比互连神经元网络所占据的空间范围小十个数量级（2），其中一些神经元几乎覆盖了整个大脑。因此，在纳米级分辨率下产生和处理大量数据的困难将脊椎动物研究限制在电路的片段上。这些努力最近通过计算，样品处理和成像技术的发展而改变（1），但是对整个大脑进行高分辨率检查仍然是一个挑战。在这里，我们介绍了受精后5.5天的幼虫斑马鱼（Danio rerio）完整大脑的ssEM数据。我们的方法利用不同规模的多轮目标成像来减少采集时间和数据管理要求。可以分析所得的数据集以重建神经元过程，从而使我们能够调查所有有髓轴突（轴突）。这些重建使得能够精确研究神经元形态，揭示出髓质网状脊髓和侧线传入轴突中明显的双侧对称性。我们通过共同注册功能参考图谱和来自同一标本的体内双光子荧光显微镜数据，进一步为全脑结构功能比较奠定了基础。所有获得的图像和重建图像均作为开放获取资源提供。

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