Simple blood-feeding method for live imaging of gut tube remodeling in regenerating planarians

Live cell imaging is a powerful technique to study cellular dynamics in vivo during animal development and regeneration. However, few live imaging methods have been reported for studying planarian regeneration. Here, we developed a simple method for steady visualization of gut tube remodeling during regeneration of a living freshwater planarian, Dugesia japonica. When planarians were fed blood several times, gut branches were well-visualized in living intact animals under normal bright-field illumination. Interestingly, tail fragments derived from these colored planarians enabled successive observation of the processes of the formation of a single anterior gut branch in the prepharyngeal region from the preexisting two posterior gut branches in the same living animals during head regeneration. Furthermore, we combined this method and RNA interference (RNAi) and thereby showed that a D. japonica raf-related gene (DjrafA) and mek-related gene (DjmekA) we identified both play a major role in the activation of extracellular signal-regulated kinase (ERK) signaling during planarian regeneration, as indicated by their RNAi-induced defects on gut tube remodeling in a time-saving initial screening using blood-feeding without immunohistochemical detection of the gut. Thus, this blood-feeding method is useful for live imaging of gut tube remodeling, and provides an advance for the field of regeneration study in planarians.