Mas-relate G protein-coupled receptor X2 (MRGPRX2) and its mouse ortholog Mrgprb2 are specifically expressed in mast cells and involved in pseudoallergic reactions. We previously reported that extracellular ATP augmented Mrgprb2-mediated mast cell activation via P2X4 receptors. In this study, we investigated the mechanism underlying the synergistic effects of co-stimulation of Mrgprb2 and P2X4 receptor on degranulation in mouse peritoneal mast cells (PMCs). Stimulation of Mrgprb2 with compound 48/80 induced degranulation accompanied by an increase in intracellular Ca²⁺ concentration ([Ca²⁺]i). ATP also induced a rapid increase in [Ca²⁺]i. Co-stimulation with ATP and compound 48/80 promoted the sustained increase in [Ca²⁺]i. This synergistic Ca²⁺ response was absent in PMCs prepared from P2X4 receptor deficient mice. In addition, both increased degranulation and synergistic Ca²⁺ response to co-stimulation with ATP and compound 48/80 were inhibited by the PI3K inhibitors wortmannin and AS605240. These results suggested that the P2X4 receptor signal promotes Mrgprb2-induced degranulation and Ca²⁺ response in a PI3K-dependent mechanism.