Substance P-induced diacylglycerol formation in rat parotid acinar cells

The mechanisms underlying the ability of substance P, to stimulate the sn-1,2-diacylglycerol (DAG) formation were studied using rat parotid acinar cells. During a 60 s stimulation, 1 microM substance P caused a rapid rise in DAG accumulation at 5 s, whereas a low (0.1 microM) concentration of agonist did not. During long term stimulation for 30 min, DAG accumulation induced by 1 microM substance P reached near maximal levels at 5 min and remained elevated for at least 20 min. In contrast, DAG formation induced by 0.1 microM substance P exhibited a peak at 5 min, gradually declined and returned to near basal levels at 30 min. Furthermore, DAG accumulation in response to substance P at 5 and 20 min increased in a dose-dependent manner. The breakdown of both [32P]phosphatidylinositol 4-monophosphate ([32P]PIP) and [32P]phosphatidylinositol 4,5-bisphosphate ([32P]PIP2) stimulated by 1 microM substance P significantly increased from 5 to 20 min and returned to basal levels by 30 min; however, the breakdown of [32P]PIP2 was greater than that of [32P]PIP. At a low concentration of substance P, [32P]PIP2 breakdown reached maximal levels at 5 min followed by a progressive decrease and returned to basal levels at 30 min, whereas the breakdown of [32P]PIP reached maximal levels at 5 min and returned to near basal levels at 10 min. Both concentrations of substance P caused some [32P]phosphatidylinositol breakdown at 5 min. Changes in [3H]inositol trisphosphate induced by substance P were similar to those in [32P]PIP2. In addition, substance P (1 microM) did not stimulate the release of [3H]choline or [3H]ethanolamine metabolites into the medium. Substance P-induced DAG formation was not inhibited by staurosporine, a protein kinase C inhibitor. These results suggest that DAG formation caused by substance P is closely associated with the hydrolysis of phosphatidylinositides but not that of phosphatidylcholine or phosphatidylethanolamine, and is not regulated by protein kinase C-dependent mechanism(s).