pituitrin and phosphatidylinositol-4-phosphate

Studies in various cells have led to the idea that agonist-stimulated diacylglycerol (DAG) generation results from an early, transient phospholipase C (PLC)-catalyzed phosphoinositide breakdown, while a more sustained elevation of DAG originates from phosphatidylcholine (PC). We have examined this issue further, using cultured rat hepatocytes, and report here that various G protein-coupled receptor (GPCR) agonists, including vasopressin (VP), angiotensin II (Ang.II), prostaglandin F2alpha, and norepinephrine (NE), may give rise to a prolonged phosphoinositide hydrolysis. Preincubation of hepatocytes with 1-butanol to prevent conversion of phosphatidic acid (PA) did not affect the agonist-induced DAG accumulation, suggesting that phospholipase D-mediated breakdown of PC was not involved. In contrast, the GPCR agonists induced phosphoinositide turnover, assessed by accumulation of inositol phosphates, that was sustained for up to 18 h, even under conditions where PLC was partially desensitized. Pretreatment of hepatocytes with wortmannin, to inhibit synthesis of phosphatidylinositol 4-phosphate and phosphatidylinositol 4,5-bisphosphate (PIP2), prevented agonist-induced inositol phosphate and DAG accumulation. Upon VP stimulation the level of PIP) declined, but only transiently, while increases in inositol 1,4,5-trisphosphate (InsP3) and DAG mass were sustained, suggesting that efficient resynthesis of PIP2 allowed sustained PLC activity. This was confirmed when cells were pretreated with wortmannin to prevent resynthesis of PIP2. Furthermore, metabolism of InsP3 was rapid, compared to that of DAG, with a more than 20-fold difference in half-life. Thus, rapid metabolism of InsP3 and efficient resynthesis of PIP2 may account for the larger amount of DAG generated and the more sustained time course, compared to InsP3. The results suggest that DAG accumulation that is sustained for many hours in response to VP, Ang.II, NE, and prostaglandin F2alpha in hepatocytes is mainly due to phosphoinositide breakdown.

Topics: 1-Butanol; Androstadienes; Angiotensin II; Animals; Cells, Cultured; Diglycerides; Dinoprost; Enzyme Activation; Hepatocytes; Male; Norepinephrine; Oxytocics; Phosphatidic Acids; Phosphatidylinositol 4,5-Diphosphate; Phosphatidylinositol Phosphates; Phosphatidylinositols; Phospholipase D; Protein Kinase Inhibitors; Rats; Rats, Wistar; Receptors, G-Protein-Coupled; Type C Phospholipases; Vasoconstrictor Agents; Vasopressins; Wortmannin

We have previously shown that vasopressin exerts a marked mitogenic effect on adrenal glomerulosa cells. In the present study, we demonstrate that vasopressin (VP) stimulates the formation of inositol monophosphate (IP), inositol diphosphate (IP2) and inositol triphosphate (IP3) in primary cultures of glomerulosa as well as fasciculata cells 5- to 8-fold over the corresponding basal values. In both cell types, the relative stimulations of IP, IP2, and IP3 formation were similar. Angiotensin II (ATII) also induced glomerulosa cells to produce a dose-dependent (up to 10-fold) increase in IP, IP2, and IP3, but had only a small effect on fasciculata cells. The dose dependencies for ATII-induced IP, IP2, and IP3 formation and aldosterone production were nearly the same. We conclude that VP- and ATII-induced formation of inositol phosphates may represent an early step in the action of these peptides on adrenal cells. However, additional elements must be involved to account for the cell specificity of VP and ATII. In glomerulosa cells, VP stimulates mitotic activity and aldosterone secretion, while ATII is only steroidogenic. On fasciculata cells, VP induces a significant increase in the formation of inositol phosphates in spite of the absence of a known biological function in these cells.

Topics: Adrenal Cortex; Angiotensin II; Animals; Dose-Response Relationship, Drug; Inositol 1,4,5-Trisphosphate; Inositol Phosphates; Male; Membranes; Phosphatidylinositol 4,5-Diphosphate; Phosphatidylinositol Phosphates; Phosphatidylinositols; Rats; Time Factors; Vasopressins