A major neurotransmitter dopamine transmits signals via five different seven transmembrane G protein-coupled receptors termed D1–D5. It is now evident that dopamine is released from leukocytes and acts as autocrine or paracrine immune modulator. However, the role of dopamine for dendritic cells (DCs) and Th differentiation remains unclear. We herein demonstrate that human monocyte-derived dendritic cells (Mo-DCs) stored dopamine in the secretary vesicles. The storage of dopamine in Mo-DCs was enhanced by forskolin and dopamine D2-like receptor antagonists via increasing cyclic adenosine 3′,5′-monophosphate (cAMP) formation. Antigen-specific interaction with naive CD4+ T cells induced releasing dopamine-including vesicles from Mo-DCs. In naive CD4+ T cells, dopamine dose dependently increased cAMP levels via D1-like receptors and shifts T-cell differentiation to Th2, in response to anti-CD3 plus anti-CD28 mAb. Furthermore, we demonstrated that dopamine D2-like receptor antagonists, such as sulpiride and nemonapride, induced a significant DC-mediated Th2 differentiation, using mixed lymphocyte reaction between human Mo-DCs and allogeneic naive CD4+ T cells. When dopamine release from Mo-DCs is inhibited by colchicines (a microtubule depolymerizer), T-cell differentiation shifts toward Th1. These findings identify DCs as a new source of dopamine, which functions as a Th2-polarizing factor in DC-naive T-cell interface.