Serotonin (5‐HT) stimulates smooth muscle cell growth through 5‐HT receptors and the 5‐HT transporter (5‐HTT), and has been associated with pulmonary hypertension (PH). Platelet‐derived growth factor receptors (PDGFR) have also been associated with PH. We present evidence for the first time that 5‐HT transactivates PDGFRβ through the 5‐HTT in pulmonary artery (PA) SMCs. Inhibition of PDGFR kinase with imatinib or AG1296 blocks 5‐HT‐stimulated PDGFRβ phosphorylation. 5‐HTT inhibitors and the Na⁺/K⁺‐ATPase inhibitor ouabain, but not 5‐HT2 and 5‐HT1B/1D receptor inhibitors, block PDGFRβ activation by 5‐HT. Notably, 5‐HTT binds the PDGFRβ upon 5‐HT stimulation and the 5‐HTT inhibitor fluoxetine blocks both the binding and PDGDRβ activation. Activation of PDGFRβ may occur through oxidation of a catalytic cysteine of tyrosine phosphatase. 5‐HT‐acti‐vated PDGFRβ phosphorylation is blocked by the anti‐oxidant N‐acetyl‐L‐cysteine and the NADPH oxidase inhibitor, DPI. Inhibition of PDGFR kinase with ima‐tinib or AG1296 significantly inhibits SMC proliferation and migration induced by 5‐HT in vitro. Infusion of 5‐HT by miniosmotic pumps enhances PDGFRβ activation in mouse lung in vivo. In summary, these results demonstrate that 5‐HT transactivates PDGFRβ in PASMCs leading to SMC proliferation and migration, and may be an important signaling pathway in the production of PH in vivo..—Liu, Y., Li, M., Warburton, R. R., Hill, N. S., Fanburg, B. L. The 5‐HT transporter transactivates the PDGFβ receptor in pulmonary artery smooth muscle cells. FASEB J. 21, 2725–2734 (2007)