Relationship between neural activity and neuronal cell fate in regeneratingHydrarevealed by cell-type specific imaging

Abstract

AbstractUnderstanding how neural circuits are regenerated following injury is a fundamental question in neuroscience.Hydrais a powerful model for studying this process because it has significant and reproducible regenerative abilities, a simple and transparent body that allows for whole nervous system imaging, and established methods for creating transgenics with cell-type-specific expression. In addition, cnidarians such asHydrasplit from bilaterians (the group that encompasses most model organisms used in neuroscience) over 500 million years ago, so similarities with other models likely indicates deeply conserved biological processes.Hydrais a long-standing regeneration model and is an emerging model for neuroscience; however, relatively little is known regarding the restoration of neural activity and behavior following significant injury. In this study, we ask if regenerating neurons reach a terminal cell fate and then reform functional neural circuits, or if neural circuits regenerate first and then guide the constituent cells toward their terminal fate. To address this question, we developed a dual-expression transgenicHydraline that expresses a cell-type-specific red fluorescent protein (tdTomato) in ec5 peduncle neurons, and a calcium indicator (GCaMP7s) in all neurons. This transgenic line allowed us to monitor neural activity while we simultaneously track the reappearance of terminally differentiated ec5 neurons as determined by the expression of tdTomato. Using SCAPE (Swept Confocally Aligned Planar Excitation) microscopy, we tracked both calcium activity and expression of tdTomato-positive neurons in 3D with single-cell resolution during regeneration ofHydra’saboral end. We observed tdTomato expression in ec5 neurons approximately four hours before the neural activity begins to display synchronized patterns associated with a regenerated neural circuit. These data suggest that regenerating neurons undergo terminal differentiation prior to re-establishing their functional role in the nervous system. The combination of dynamic imaging of neural activity and gene expression during regeneration makeHydraa powerful model system for understanding the key molecular and functional processes involved in neuro-regeneration following injury.