Resources for Studying Neural Circuits with G-Deleted Rabies Virus - PROJECT SUMMARY Monosynaptic input tracing with G-deleted rabies virus is a powerful tool for unraveling neural circuits. By targeting initial viral infection to specific, genetically defined cell types, any transgene of interest can be delivered to neurons that provide direct, monosynaptic input to these cells. This approach for identifying and manipulating neural circuits has had a tremendous impact across all areas of neuroscience. Nevertheless, several factors limit its accessibility and impact the quality of data collected via this experimental platform. First, unlike chemical tracers that can be cheaply produced in large quantities, production of the rabies and AAV helper vectors is expensive and therefore cost is often prohibitive. Second, an extremely wide range of AAV helper viruses must be used to tailor vectors for diverse purposes, brain areas, and cell types. The Salk Institute’s Viral Vector Core (VVC) is one of only two core facilities that produces and distributes AAV helper vectors for rabies tracing, but it can only offer a relatively small number of helper AAVs that are not ideally suited to many applications. Thus, limited access to appropriate AAV helper vectors often impedes progress. Finally, determining which combination of vectors is best suited for a particular experiment can be extremely difficult, and thus reagent selection (and the design of appropriate controls) can be a significant barrier to uptake. Here, U24 funds will be leveraged to remove these barriers to progress via three specific aims. (1) The Salk VVC will ensure that a diverse array of neuroscientists can access and utilize the G-deleted rabies circuit-tracing platform by subsidizing viral vector costs and distributing small aliquots to allow researchers to purchase the exact quantity of each vector they need to perform their analyses. (2) To provide widespread access to cutting-edge neural circuit- tracing technologies suitable for use in a wide range of brain areas, the Salk VVC will expand its catalogue of G- deleted rabies and AAV helper vectors by incorporating new neuroscience tools into the rabies genome as they become available and, more critically, adding an array of helper vectors with different combinations of payloads and a range of AAV serotypes to efficiently transduce a large array of cell types and brain areas. (3) The Salk VVC will build and advertise a user-friendly website to promote best practices when using G-deleted rabies reagents. In addition to providing a comprehensive list of available reagents, the VVC website will include: a) detailed protocols for using and generating each reagent, b) virtual kits describing combinations of reagents most suitable for common use scenarios, c) an information warehouse providing examples of appropriately controlled rabies-based experiments and pitfalls to avoid when using each reagent, and d) mechanisms for researchers to contact the VVC for guidance in experimental design and execution. Further, VVC offerings and its website will be broadly advertised so that a wide diversity of researchers learn about and take advantage of these unique circuit-tracing resources. Together, these complementary approaches will democratize access to monosynaptic rabies tracing and improve the rigor and reproducibility of data collected using these critical tools.
