Stromal cell-derived factor-1α (SDF-1α) drives endothelial colony-forming cell (ECFC) homing and incorporation within neovessels, thereby restoring tissue perfusion in ischemic tissues and favoring tumor vascularization and metastasis. SDF-1α stimulates ECFC migration by activating the G-protein-coupled receptor, CXCR4, and then engaging the phosphoinositide 3-kinase (PI3K)/AKT signaling pathway. Sporadic evidence showed that SDF-1α may also act through an increase in intracellular Ca concentration ([Ca]) in bone marrow-derived hematopoietic progenitor cells and fully differentiated endothelial cells. Of note, recent evidence demonstrated that intracellular Ca signals play a key role in controlling the proangiogenic activity of ECFCs. The present investigation was, therefore, undertaken to assess whether and how SDF-1α induces ECFC motility by triggering intracellular Ca signals. We found that SDF-1α caused a dose-dependent increase in [Ca] that was inhibited by ADM3100, a selective CXCR4 antagonist. Pharmacological manipulation revealed that the Ca response to [Ca] was shaped by an initial intracellular Ca release through inositol-1,4,5-trisphosphate receptors (InsPRs), followed by a sustained phase of extracellular Ca entry through store-operated Ca channels. InsP-dependent Ca release and store-operated Ca entry (SOCE) were both necessary for SDF-1α-induced extracellular signal-regulated kinases 1/2 (ERK 1/2) and AKT phosphorylation. Finally, SDF-1α employed intracellular Ca signals, ERK 1/2, and PI3K/AKT to promote ECFC migration in vitro and neovessel formation in vivo. These data, therefore, provide the first evidence that SDF-1α induces ECFC migration through the Ca-dependent activation of the ERK 1/2 and PI3K/AKT pathways.