Effects of ethanol on phosphorylation of lipids in rat synaptic plasma membranes

Synaptic plasma membranes (SPM) isolated from rat cerebral cortex contain lipid kinases for conversion of phosphatidylinositol (PI), phosphatidylinositol 4-phosphate (PIP), and diacylglycerol (DG) to PIP, phosphatidylinositol 4,5-bisphosphate (PIP2), and phosphatidic acid (PA), respectively. These anionic phospholipids are important in signal transduction mechanisms and are required for synaptic function. The effect of ethanol and other aliphatic alcohols on phosphorylation of these lipids in SPM has not been established. Incubation of SPM with [gamma-32P]ATP resulted in labeling of PIP, lyso-PIP, PIP2, and PA. Ethanol (50-200 mM) added to the incubation system showed a dose-dependent decrease in labeling of PIP2, but not PIP or PA. To a lesser extent, labeling of PIP2 was also inhibited by 1-propanol, but neither isopropanol nor 1-butanol could alter the PIP2 labeling pattern. Under similar incubation conditions, labeling of PIP and PA in SPM was not altered by ethanol, 1-propanol, iso-propanol, but 1-butanol stimulated PIP labeling with a peak at 25 mM. Addition of exogenous PIP to the incubation mixture led to an increase in labeling of PIP2, suggesting that the endogenous PIP pool in SPM is limiting for the synthesis of PIP2 in SPM. Interestingly, when SPM were incubated with exogenous PIP, addition of ethanol (50-100 mM) to this incubation mixture resulted in an increase in PIP2 labeling. Taken together, these results suggest a specific effect of ethanol on PIP kinase in SPM, and this effect seems to be dependent on the location and/or amount of PIP in the membrane.