Two-photon manipulation of neuronal activity and behavior in Hydra vulgaris

The introduction of calcium imaging has rendered cnidarians, such as Hydra vulgaris, valuable model organisms for investigating neuronal activity and behavior. Here, we present a comprehensive protocol to image and manipulate neuronal activity and behavior of Hydra. We describe steps for wide-field imaging and two-photon simulation and ablation of neurons. We then detail imaging behavior and post-ablation analysis. We address challenges that may arise during the preparation and execution of the experiments.