Understanding the connecting structure of brain network is the basis to reveal the principle of the brain function and elucidate the mechanism of brain diseases. Trans-synaptic tracing with neurotropic viruses has become one of the most effective technologies to dissect the neural circuits. Although the retrograde trans-synaptic tracing for analyzing the input neural networks with recombinant rabies and pseudorabies virus has been broadly applied in neuroscience, viral tools for analyzing the output neural networks are still lacking. The recombinant vesicular stomatitis virus (VSV) has been used for the mapping of synaptic outputs. However, several drawbacks, including high neurotoxicity and rapid lethality in experimental animals, hinder its application in long-term studies of the structure and function of neural networks. To overcome these limitations, we generated a recombinant VSV with replication-related N gene mutation, VSV-NR7A, and examined its cytotoxicity and efficiency of trans-synaptic spreading. We found that by comparison with the wild-type tracer of VSV, the NR7A mutation endowed the virus lower rate of propagation and cytotoxicity in vitro, as well as significantly reduced neural inflammatory responses in vivo and much longer animal survival when it was injected into the nucleus of the mice brain. Besides, the spreading of the attenuated VSV was delayed when injected into the VTA. Importantly, with the reduced toxicity and extended animal survival, the number of brain regions that was trans-synaptically labeled by the mutant VSV was more than that of the wild-type VSV. These results indicated that the VSV-NR7A, could be a promising anterograde tracer that enables researchers to explore more downstream connections of a given brain region, and observe the anatomical structure and the function of the downstream circuits over a longer time window. Our work could provide an improved tool for structural and functional studies of neurocircuit.

中文翻译：

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一种突变的水泡性口炎病毒，具有降低的细胞毒性和增强的顺行突触效率。

了解脑网络的连接结构是揭示脑功能原理和阐明脑疾病机理的基础。用嗜神经病毒进行突触追踪已成为剖析神经回路的最有效技术之一。尽管逆向反式突触追踪用于分析带有重组狂犬病和伪狂犬病病毒的输入神经网络已广泛应用于神经科学，但仍缺乏用于分析输出神经网络的病毒工具。重组水泡性口炎病毒（VSV）已用于映射突触输出。但是，一些缺点，包括实验动物的高神经毒性和快速致死性，阻碍了其在神经网络的结构和功能的长期研究中的应用。为了克服这些限制，我们产生了具有复制相关N基因突变的重组VSV，VSV-NR7A，并检查了其细胞毒性和反突触传播的效率。我们发现，与野生型VSV示踪剂相比，NR7A突变使病毒在体外的繁殖和细胞毒性降低，并显着降低了体内的神经炎性反应，并且将其注射到小鼠体内时，动物的存活时间更长。小鼠脑核。此外，衰减的VSV的扩散在注入VTA时被延迟。重要的是，随着毒性降低和动物存活期延长，突变型VSV反式突触标记的脑区数量比野生型VSV多。这些结果表明，VSV-NR7A 可能是一种有前途的顺行示踪剂，使研究人员能够探索给定大脑区域的更多下游连接，并在更长的时间范围内观察下游回路的解剖结构和功能。我们的工作可以为神经回路的结构和功能研究提供改进的工具。