Mechanisms of Network-Scale Neural Regeneration in a New Neuroscience Model - Regenerative abilities are widespread, suggesting deeply conserved roles in biology. However, these abilities vary dramatically across species: understanding conserved and novel mechanisms enabling varying degrees of regeneration has important implications for regenerative medicine. An enduring medical challenge is the ability to regenerate extensively damaged nervous systems. This presents unique problems, bridging fields and levels of analysis, from the molecular mechanisms of neural development to the neural activity dynamics driving behavior. Here, we propose to study fundamental principles linking neurodevelopmental processes to the functional organization of neural circuits to ask how they could enable network-scale regeneration. Our strategy is to use a new animal model, Clytia, that is small, transparent, and capable of regenerating entire genetically ablated neural subnetworks. This affords a novel, high-resolution platform to study regenerative processes at their interface with systems-level organization. In preliminary work, we used whole-animal calcium imaging and single-cell RNA- sequencing (scRNAseq) to identify intermingled subnetworks with distinct functional organizations. We found that inducible, genetic ablation of a particular subnetwork led to loss of a specific behavior. However, this subnetwork and behavior completely regenerated within days. These observations have raised exciting questions: how is network injury sensed and translated into regenerative programs? Where do newborn neurons come from and what rules guide them to their final positions? Are all subnetworks regenerative despite their dramatically different architectures? Do regenerative abilities shared between subnetworks reflect numerous solutions to the problem of network-scale regeneration or common, enabling principles? In Aim 1, we propose to systematically ablate genetically defined neural subnetworks to determine which are capable of regeneration, allowing for a powerful, within-animal comparative approach to understand enabling mechanisms. In Aim 2, we focus on intermingled subnetworks controlling feeding versus swimming, both of which are regenerative. We will determine which stem cells give rise to newborn neurons of each type and test models linking neuronal origins to migration patterns to the ultimate position of newborn neurons in regenerated networks. Lastly, in Aim 3, we will use scRNAseq and genetic manipulations to examine molecular mechanisms by which network damage is sensed and initiates these regenerative programs. Together, we expect this work to provide fundamental insights into rules that connect processes such as injury sensing, migration, and functional integration to systems-level organization to enable regeneration.
