Abstract
All optical physiology in vivo provides a conduit for investigating the function of neural circuits in 3-D. Here, we report a new strategy for flexible, axially-decoupled photo-stimulation and two photon readout (ADePT) of neuronal activity. To achieve axially-contained widefield optogenetic patterned stimulation, we couple a digital micro-mirror device illuminated by a solid-state laser with a motorized holographic diffuser. In parallel, we use multiphoton imaging of neural activity across different z-planes. We use ADePT to analyze the excitatory and inhibitory functional connectivity of the mouse early olfactory system. Specifically, we control the activity of individual input glomeruli on the olfactory bulb surface, and map the ensuing responses of output mitral and tufted cell bodies in deeper layers. This approach identifies cohorts of sister mitral and tufted cells, whose firing is driven by the same parent glomerulus, and also reveals their differential inhibition by other glomeruli. In addition, selective optogenetic activation of glomerular GABAergic/dopaminergic (DAT+) interneurons triggers dense, but spatially heterogeneous suppression of mitral and tufted cell baseline activity and odor responses, further demonstrating specificity in the inhibitory olfactory bulb connectivity. In summary, ADePT enables high-throughput functional connectivity mapping in optically accessible brain regions.