Hydrolysis of exogenous [3H]phosphatidylcholine by brain membranes and cytosol

Phosphatidylcholine, in addition to the widely studied inositol phospholipids, is cleaved to produce second messengers in neuronal signal transduction processes. Because of the difficulty in labelling and measuring the metabolism of endogenous phosphatidylcholine in brain tissue, we investigated the utility of measuring the hydrolysis of exogenous labelled substrate incubated with rat cerebral cortical cytosol and membrane fractions as has been successful in studies of phosphoinositide hydrolysis. In the cytosol ³H]phosphatidylcholine was hydrolyzed at a linear rate for 60 min of incubation and GTPS stimulated hydrolysis by 63%. The products of phospholipase C and phospholipase D, phosphorylcholine and choline, contributed only 44% of the ³H]phosphatidylcholine hydrolytic products in the cytosol, with phospholipase D activity slightly predominating. GTPS stimulated cytosolic phospholipase C and reduced phospholipase D activity. ³H]Phosphatidylcholine was hydrolyzed much more slowly by membranes than by cytosol. In membranes the production of ³H]phosphorylcholine and ³H]choline were approximately equal, contributing 27% of the total ³H]phosphatidylcholine hydrolysis, and GTPS only caused a slight stimulation of phospholipase C activity. Chronic lithium treatment (4 weeks) appeared to slightly reduce ³H]phosphatidylcholine metabolism in the cytosol and in membranes, but no statistically significant reductions were achieved. Cytosol and membrane fractions from postmortem human brain metabolized ³H]phosphatidylcholine slowly, and GTPS had no effects. In summary, exogenous ³H]phosphatidylcholine was hydrolyzed by brain cytosol and membranes, and this was stimulated by GTPS, but the complex contributions of multiple metabolic pathways complicates the application of this method for studying individual pathways, such as phospholipase D which contributes only a fraction of the total processes hydrolyzing exogenous ³H]phosphatidylcholine.