(7), has been increasingly associated with pulmonary hypertension (2, 4, 9, 30), and it is recognized that hyperplasia of SMCs accompanies the pathology of pulmonary hypertension (5, 10, 22). 5-HT initiates its SMC mitogenic action through both 5-HT receptors and the 5-HT transporter (5-HTT) (7) that are present on the cell membrane in close proximity to membrane structural lipids that include phosphatidylcholine. Through the receptor and transporter ligands, 5-HT produces SMC proliferation via combinatorial actions on the small G protein Rho, p 42/44 mitogen-activated protein kinase (MAPK), phosphatidylinositol 3-kinase (PI3-kinase)/protein kinase B (Akt), and platelet-derived growth factor-␤ receptor signaling pathways (17-20).Lipid-derived second messengers generated by the action of phospholipases on membrane-associated phospholipids play an important role in cellular signal transduction in general (3). Phospholipase D (PLD), which catalyzes the hydrolysis of phosphotidylcholine to phosphatidic acid (PA) and choline, has been recognized to be an intermediate in the actions of a wide variety of agonists on cellular functions, including those that cause proliferation (8,33). Agonists identified to activate PLD include neurotransmitters, growth factors, hormones, calcium ionophores, and reactive oxygen species (6, 24). The cell signaling associated with PLD activation occurs at least in part through actions of PA (14).It previously was reported that 5-HT activates PLD in rat mesangial cells (15) and mesenteric SMCs, which associates the activation of PLD with contraction (11). Plevin and coworkers (27) demonstrated in 1994 that 5-HT activates PLD in cultured sheep pulmonary artery SMCs. However, there is no information currently available about the relationship between 5-HT, its receptors and transporter, and PLD in pulmonary artery SMC signaling and proliferation. In the present study, we show that 5-HT activates PLD of pulmonary artery SMCs via the 5-HT 2A receptor, leading to the generation of PA that promotes SMC proliferation through activations of mammalian target of rapamycin (mTOR), S6K1 and MAPK but not the Rho or PI3-kinase/Akt signaling pathways. We hypothesize that these observations link an important lipid component of the cell membrane, i.e., phosphatidyl choline, with cell signaling occurring via 5-HT ligands in SMCs.