Connective tissue cells of liver parenchyma (perisinusoidal myofibroblasts) can be induced to express the lipocyte (Ito cell) phenotype. We have studied phospholipid synthesis and phosphate incorporation during this in vitro conversion, induced by insulin and/or indomethacin, in the established murine cell line GRX. Phospholipid synthesis, measured by [14C]acetate incorporation, was increased after a full induction of the lipocyte phenotype. The 32Pi incorporation into phospholipids was increased from the beginning of induction. Phosphatidic acid and phosphatidylinositol synthesis were increased early in the induction, whilst the increase of major constitutive phospholipids was significant only after the full lipocyte phenotype induction. The presence of unsaturated fatty acids in phospholipids was increased in lipocytes. Linoleic acid was present only in diacylglycerols and in phosphatidylinositol. Since we have shown previously that linoleic acid was not present in triacylglycerols, this result indicates the importance of future studies on activation of phosphatidylinositol cycles in induction of lipocyte phenotype in liver connective tissue cells.