美国麻省理工学院Ian R. Wickersham团队近期取得重要工作进展，他们研究利用双缺失突变狂犬病病毒对体内突触连接的神经网络进行长期标记和成像。相关研究成果2024年1月11日在线发表于《自然—神经科学》杂志上。

据介绍，基于狂犬病病毒的单突触追踪是一种广泛使用的绘制神经回路的技术，但其细胞毒性使其主要局限于解剖学应用。

研究人员提出了一种第二代系统，使用双缺失突变狂犬病病毒，以最小的毒性标记靶向神经元群体的直接输入。病毒传播需要两种缺失的病毒基因在突触后源细胞中的反式表达。在病毒复制的最初阶段用多四环素抑制这种表达可以降低对突触后细胞的毒性。

体内纵向双光子成像表明，超过90%的突触前细胞和源细胞在整个12周的成像过程中存活下来。感染后5周的离体全细胞记录显示，与第一代系统相比，第二代系统对输入和源细胞的干扰要小得多。最后，视觉皮层神经元标记网络的双光子钙成像显示，10周后，它们的视觉反应特性恢复正常，这是我们随访时间最长的一次。

附：英文原文

Title: Long-term labeling and imaging of synaptically connected neuronal networks in vivo using double-deletion-mutant rabies viruses

Author: Jin, Lei, Sullivan, Heather A., Zhu, Mulangma, Lavin, Thomas K., Matsuyama, Makoto, Fu, Xin, Lea, Nicholas E., Xu, Ran, Hou, YuanYuan, Rutigliani, Luca, Pruner, Maxwell, Babcock, Kelsey R., Ip, Jacque Pak Kan, Hu, Ming, Daigle, Tanya L., Zeng, Hongkui, Sur, Mriganka, Feng, Guoping, Wickersham, Ian R.

Issue&Volume: 2024-01-11

Abstract: Rabies-virus-based monosynaptic tracing is a widely used technique for mapping neural circuitry, but its cytotoxicity has confined it primarily to anatomical applications. Here we present a second-generation system for labeling direct inputs to targeted neuronal populations with minimal toxicity, using double-deletion-mutant rabies viruses. Viral spread requires expression of both deleted viral genes in trans in postsynaptic source cells. Suppressing this expression with doxycycline following an initial period of viral replication reduces toxicity to postsynaptic cells. Longitudinal two-photon imaging in vivo indicated that over 90% of both presynaptic and source cells survived for the full 12-week course of imaging. Ex vivo whole-cell recordings at 5weeks postinfection showed that the second-generation system perturbs input and source cells much less than the first-generation system. Finally, two-photon calcium imaging of labeled networks of visual cortex neurons showed that their visual response properties appeared normal for 10weeks, the longest we followed them.

DOI: 10.1038/s41593-023-01545-8

Source: https://www.nature.com/articles/s41593-023-01545-8