The stimulatory effects of angiotensin (Ang) I, Ang II, and Ang III on production of diacylglycerol (DAG), a second messenger, were examined in porcine pulmonary artery endothelial cells. Ang I, Ang II, and Ang III provoked rapid increases in [3H]glycerol labeling of DAG. The stimulatory effect on DAG production was maximal after 1 and 5 min. Pretreatment of cells with angiotensin-converting enzyme activity inhibitors prevented the stimulatory effect of Ang I on DAG production, indicating that Ang II but not Ang I is responsible for increased DAG production. The stimulatory effects of Ang II and Ang III on DAG production were concentration dependent and were maximal at a 10-nM concentration of both Ang II and Ang III. Data from further experiments revealed that the Ang II- and Ang III-elicited formation of DAG is derived from the coordinated hydrolysis of membrane phosphatidylinositol and phosphatidylcholine by phospholipase C- and phospholipase D-catalyzed pathways. The angiotensin analogue [Sar1 Ile8] Ang II, an Ang II receptor antagonist, blocked the hydrolysis of phosphatidylinositol and phosphatidylcholine and thus the increased production of DAG by Ang II and Ang III. These results indicate that Ang II- and Ang III-induced stimulation of DAG production in pulmonary artery endothelial cells involves multiple pathways of phospholipid hydrolysis and is mediated by angiotensin receptors.