Regeneration-dependent conditional gene knockdown (Readyknock) in planarian: demonstration of requirement for Djsnap-25 expression in the brain for negative phototactic behavior.

Freshwater planarians have a simple and evolutionarily primitive brain structure. Here, we identified the Djsnap-25 gene encoding a homolog of the evolutionarily conserved synaptic protein SNAP-25 from the planarian Dugesia japonica and assessed its role in brain function. Djsnap-25 was expressed widely in the nervous system. To investigate the specific role of Djsnap-25 in the brain, we developed a unique technique of RNA interference (RNAi), regeneration-dependent conditional gene knockdown (Readyknock), exploiting the high regenerative capacity of planarians, and succeeded in selectively eliminating the DjSNAP-25 activity in the head region while leaving the DjSNAP-25 activity in the trunk region intact. These knockdown animals showed no effect on brain morphology or on undirected movement of the trunk itself. Light-avoidance behavior or negative phototaxis was used to quantitatively analyze brain function in the knockdown animals. The results suggested that the DjSNAP-25 activity within the head region is required for two independent sensory-processing pathways that regulate locomotive activity and directional movement downstream of distinct primary sensory outputs coming from the head margin and the eyes, respectively, during negative phototaxis. Our approach demonstrates that planarians are a powerful model organism to study the molecular basis of the brain as an information-processing center.