Directed stepwise tracing of polysynaptic neuronal circuits with replication-deficient pseudorabies virus

Brain functions are accomplished by polysynaptic circuits formed by neurons wired together through multiple orders of synaptic connections. Polysynaptic connectivity has been difficult to examine due to a lack of methods of continuously tracing the pathways in a controlled manner. Here, we demonstrate directed, stepwise retrograde polysynaptic tracing by inducible reconstitution of replication-deficient trans-neuronal pseudorabies virus (PRV^ΔIE) in the brain. Furthermore, PRV^ΔIE replication can be temporally restricted to minimize its neurotoxicity. With this tool, we delineate a wiring diagram between the hippocampus and striatum—two major brain systems for learning, memory, and navigation—that consists of projections from specific hippocampal domains to specific striatal areas via distinct intermediate brain regions. Therefore, this inducible PRV^ΔIE system provides a tool for dissecting polysynaptic circuits underlying complex brain functions.