Application of Recombinant Rabies Virus to Xenopus Tadpole Brain
Application of Recombinant Rabies Virus to Xenopus Tadpole Brain
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DOI：
10.1523/eneuro.0477-20.2021
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发表时间：
2021-07-01
期刊：
影响因子：
3.4
通讯作者：
Cline, Hollis T.
中科院分区：
文献类型：
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作者：
Faulkner, Regina L.;Wall, Nicholas R.;Cline, Hollis T.
非洲爪蟾的实验系统为神经回路的发育和可塑性提供了重要的见解。如果有更多的工具来研究非洲爪蟾的神经回路结构和功能，将会加强对非洲爪蟾神经科学的研究。狂犬病毒是标记和操纵神经回路的有力工具，已广泛用于中尺度连接组学的研究。狂犬病毒能否在爪蟾中转导神经元并表达转基因，目前还没有系统的研究。糖蛋白缺失狂犬病毒在轴突末端转导神经元并逆行标记其细胞体。我们发现，当直接注射到爪蟾蝌蚪脑组织时，糖蛋白缺失的狂犬病毒感染局部和投射神经元。带EnvA的糖蛋白缺失狂犬病假分型限制了外源性表达EnvA受体（TVA）的细胞感染。EnvA假型病毒通过启动子驱动的TVA表达特异性感染蝌蚪神经元，证明了其标记目标神经元群体的效用。神经细胞类型是由包括解剖位置、遗传标记的表达、轴突投射位置、形态和生理特性在内的特征组合来定义的。我们发现，驱动一个半球的TVA表达并将EnvA假型病毒注射到对侧半球，可以逆行标记由细胞体位置和轴突投射位置定义的神经元。利用这种方法，狂犬病可以用来识别非洲爪蟾大脑中的细胞类型，同时表达转基因，从而能够监测或操纵神经元活动。这使得狂犬病成为研究非洲爪蟾神经回路结构和功能的宝贵工具。
The Xenopus laevis experimental system has provided significant insight into the development and plasticity of neural circuits. Xenopus neuroscience research would be enhanced by additional tools to study neural circuit structure and function. Rabies viruses are powerful tools to label and manipulate neural circuits and have been widely used to study mesoscale connectomics. Whether rabies virus can be used to transduce neurons and express transgenes in Xenopus has not been systematically investigated. Glycoprotein-deleted rabies virus transduces neurons at the axon terminal and retrogradely labels their cell bodies. We show that glyco-protein-deleted rabies virus infects local and projection neurons in the Xenopus tadpole when directly injected into brain tissue. Pseudotyping glycoprotein-deleted rabies with EnvA restricts infection to cells with exogenous expression of the EnvA receptor, TVA. EnvA pseudotyped virus specifically infects tadpole neurons with promoter-driven expression of TVA, demonstrating its utility to label targeted neuronal populations. Neuronal cell types are defined by a combination of features including anatomic location, expression of genetic markers, axon projection sites, morphology, and physiological properties. We show that driving TVA expression in one hemisphere and injecting EnvA pseudotyped virus into the contralateral hemisphere, retrogradely labels neurons defined by cell body location and axon projection site. Using this approach, rabies can be used to identify cell types in Xenopus brain and simultaneously to express transgenes which enable monitoring or manipulation of neuronal activity. This makes rabies a valuable tool to study the structure and function of neural circuits in Xenopus.