TRACR: an anterograde transneuronal tracing system for genetic access across synapses and longitudinal circuit analysis

Abstract

Following neural signals as they converge onto and diverge from individual neurons is central to understanding circuit function and disease-related dysfunction. Existing anterograde transneuronal tracers are limited by cytotoxicity and incomplete genetic control over connected partners. To address these limitations, we adapted synthetic Notch designs to create TRanssynaptic Anterograde Circuit Readout (TRACR). Binding of the engineered ligand-receptor across synapses induces TRE-driven reporter transcription, enabling characterization of postsynaptic neurons. TRACR provides segregated genetic access to pre- and postsynaptic populations, and can be combined with markers, sensors, or effectors to expand circuit analysis. By applying TRACR at multiple synapses in the mouse visual system, we show that TRACR labels postsynaptic partners of sensory neurons, long-range projections and local inhibitory interneurons. TRACR signaling is reversible and fails to activate when synapses are absent or disrupted. Together, TRACR is an accessible, AAV-deliverable transneuronal reporting tool for longitudinal analysis of circuit assembly, degeneration, and repair. HIGHLIGHTSO_LITRACR adapts the synNotch system to signal across synapses for tracing postsynaptic targets. C_LIO_LITRACR identifies local and long-range postsynaptic targets in the mouse visual system. C_LIO_LITRACR activation requires intact synaptic connectivity rather than proximity. C_LIO_LITRACR signals are reversible, diminishing upon synapse loss and activating following assembly. C_LI