Marine invertebrate larvae are known to begin metamorphosis in response to environmentally-derived cues. However, little is known about the relationships between the perception of such cues and internal signalling for metamorphosis. To elucidate the mechanism underlying the initiation of metamorphosis in the ascidian, Ciona intestinalis type A (Ciona robusta), we artificially induced ascidian metamorphosis and investigated Ca²⁺ dynamics from pre- to post-metamorphosis. Ca²⁺ transients were observed and consisted of two temporally distinct phases with different durations before tail regression which is the early event of metamorphosis. In the first phase, Phase I, the Ca²⁺ transient in the papillae (adhesive organ of anterior trunk) was coupled with the Ca²⁺ transient in dorsally localised cells and endoderm cells just after mechanical stimulation. The Ca²⁺ transients in Phase I were also observed when applying only short stimulation. In the second phase, Phase II, the Ca²⁺ transient in papillae was observed again and lasted for~5–11 min just after the Ca²⁺ transient in Phase I continued for a few minutes. The impaired papillae by Foxg-knockdown failed to induce the second Ca²⁺ transient in Phase II and tail regression. In Phase II, a wave-like Ca²⁺ propagation was also observed across the entire epidermis. Our results indicate that the papillae sense a mechanical cue and two-round Ca²⁺ transients in papillae transmits the internal metamorphic signals to different tissues, which subsequently induces tail regression. Our study will help elucidate the internal mechanism of metamorphosis in marine invertebrate larvae in response to environmental cues.