Mast cells migrate toward histamine through H₄ receptor (H₄R), which involves the activation of Rac1 and Rac2. Transglutaminase catalyzes the incorporation of histamine into cellular proteins, the process called histaminylation. Although several reports suggest possible roles of histaminlylation in mast cell signaling, there is no direct evidence showing its significance in cellular function. To explore the functional significance of histaminylation, we first investigated the effect of cystamine, a transglutaminase inhibitor, on H₄R-Rac pathway in mast cell. Cystamine attenuated histamine-induced migration and Rac activation, suggesting that H₄R-Rac signaling requires transglutaminase. Organic cation transporter 3 (OCT3) transports histamine into cytoplasm from extracellular space. Given the histaminylation occurs and regulates signaling in mast cells, we next examined if OCT3-mediated transport of histamine plays any role in H₄R signaling. Decynium-22, an inhibitor of OCT3, suppressed histamine-induced migration and Rac activation, suggesting the involvement of OCT3 in H₄R-Rac signaling. Our findings support the idea that transported histamine and transglutaminase activity modulate H₄R-Rac pathway.