SUMMARY We developed a technology (Capturing Activated Neural Ensembles, or CANE) to label, manipulate, and trans-synaptically trace neural circuits that are transiently activated in behavioral contexts with high efficiency and temporal precision. CANE consists of a knock-in mouse and engineered viruses designed to specifically infect activated neurons. Using CANE, we selectively labeled neurons that were activated by either fearful or aggressive social encounters in a hypothalamic subnucleus previously known as a locus for aggression, and discovered that social fear and aggression neurons are intermixed but largely distinct. Optogenetic stimulation of CANE-captured social fear neurons (SFNs) is sufficient to evoke fear-like behaviors in normal social contexts, whereas silencing SFNs resulted in reduced social avoidance. CANE-based mapping of axonal projections and presynaptic inputs to SFNs further revealed a highly distributed and recurrent neural network. CANE is a broadly applicable technology for dissecting causality and connectivity of spatially intermingled but functionally distinct ensembles.

Humans often rank craniofacial pain as more severe than body pain. Evidence suggests that a stimulus of the same intensity induces stronger pain in the face than the body. However, the underlying neural circuitry for the differential processing of facial versus bodily pain remains unknown. Interestingly, the lateral parabrachial nucleus (PBL), a critical node in the affective pain circuit, is activated more strongly by noxious stimulation of the face than the hindpaw. Using a novel activity-dependent technology called CANE developed in our lab, we identified and selectively labeled noxious stimuli-activated PBL neurons, and performed comprehensive anatomical input-output mapping. Surprisingly, a hitherto uncharacterized monosynaptic connection between cranial sensory neurons and the PBL-nociceptive neurons was uncovered. Optogenetic activation of this monosynaptic craniofacial-to-PBL projection induced robust escape/avoidance behaviors and stress calls, whereas optogenetic silencing specifically reduced facial nociception. The monosynaptic circuit revealed here provides a neural substrate for heightened craniofacial affective pain.

Postnatal/adult SVZ neurogenesis is believed to be primarily controlled by neural stem cell (NSC)-intrinsic mechanisms, interacting with extracellular/niche-driven cues. Although behavioral paradigms and disease states have suggested possibilities for higher-level inputs, it is currently unknown if neural activity patterns from discrete circuits can directly regulate SVZ neurogenesis. We have identified a previously undescribed population of ChAT+ neurons residing within the rodent SVZ neurogenic niche. These neurons showed morphological and functional differences from neighboring striatal counterparts, and released acetylcholine locally in activity-dependent fashion. Optogenetic inhibition and stimulation of subependymal ChAT+ neurons in vivo showed that they are necessary and sufficient to control neurogenic proliferation. Furthermore, whole-cell recordings and biochemical experiments revealed direct SVZ NSC responses to local acetylcholine release, synergizing with FGF receptor activation to increase neuroblast production. These results uncovered an unknown gateway connecting SVZ neurogenesis to neuronal activity-dependent control, and possibilities for modulating neuroregenerative capacities in health and disease.