Abstract
Neurotropic viral transsynaptic tracing is an increasingly powerful technique for dissecting the structure and function of neural circuits. Herpes simplex virus type 1 strain H129 has been widely used as an anterograde tracer. However, HSV tracers still have several shortcomings, including high toxicity, low sensitivity and non-specific retrograde labeling. Here, we aimed to construct high-brightness HSV anterograde tracers by increasing the expression of exogenous genes carried by H129 viruses. Using a Trojan horse-like strategy, a HSV/AAV (adeno-associated virus) chimaera termed H8 was generated to enhance the expression of a fluorescent marker. In vitro and in vivo assays showed that the exogenous gene was efficiently replicated and amplified by the synergism of the HSV vector and introduced AAV replication system. H8 reporting fluorescence was brighter than that of currently available H129 tracers, and H8 could be used for fast and effective anterograde tracing without additional immunostaining. These results indicated that foreign gene expression in HSV tracers could be enhanced by integrating HSV with AAV replication system. This approach may be useful as a general enhanced expression strategy for HSV-based tracing tools or gene delivery vectors.
Highlights
Transneuronal tracing of neural circuits using neurotropic viral tools is increasingly being recognized as a powerful approach for defining the synaptic organization of neural networks
Construction and characterization of the H8 chimeric virus in vitro The H8 chimeric virus was generated by the introduction of the associated virus (AAV) replication system into the HSV1 strain H129 genome to enhance the expression of an exogenous gene of interest (GOI), which, in this case, was egfp
Western blotting experiments showed that the H8 virus effectively expressed the AAV Rep protein, while no Rep expression was detected in H1- or wild type H129-infected cells (Fig. 1c)
Summary
Transneuronal tracing of neural circuits using neurotropic viral tools is increasingly being recognized as a powerful approach for defining the synaptic organization of neural networks. Neurotropic viruses have the innate ability to invade neurons and produce infectious progeny that spread along synaptically connected neuronal networks [1,2,3]. Mapping neural circuits requires both anterograde and retrograde tracers [4]. Rabies virus and pseudorabies virus (PRV) strains have been (2020) 13:5 tracer with increased brightness would be more convenient for anterograde transsynaptic tracing. Decreasing the sampling time could increase the likelihood that labeled brain regions are directly related to the injection site. Enhancing the expression of exogenous genes is one of the core objectives for improving HSV1 H129-based viral tracers
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