The identity of phosphoinositol isomers accumulating on stimulation of primary cultures of bovine adrenocortical zona fasciculata/reticularis cells with angiotensin II (AII), in the presence of Li+, has been established by chromatographic separation on a MonoQ HR5/5 column. The metabolism of inositol 1,4,5-trisphosphate (Ins(1,4,5)P3) in a broken cell preparation has also been studied in the absence or presence of added ATP. Our results show that Ins(1,4,5)P3 is formed within 5 s of stimulation of whole cells, but is rapidly converted to Ins(1,3,4)P3 through an Ins(1,3,4,5)P4 intermediate. All the phosphoinositol products accumulating on prolonged (15 min) stimulation of whole cells (Ins1P, Ins4P, Ins(1,3)P2, Ins(1,4)P2, Ins(1,3,4)P3, Ins(1,4,5)P3, Ins(1,3,4,5)P4) can be accounted for by the metabolism of Ins(1,4,5)P3 in broken cells, either through direct dephosphorylation in the absence of added ATP (Ins(1,4)P2, Ins4P) or through dephosphorylation of Ins(1,3,4,5)P4 formed in the presence of added ATP (Ins(1,3,4)P3, Ins(1,3)P2 and Ins1P). Our results provide further evidence to suggest that AII stimulates the rapid and sustained breakdown of phosphatidylinositol 4,5-diphosphate (PtdIns(4,5)P2) to form Ins(1,4,5)P3.