Can mitochondria and synaptosomes of guinea-pig brain synthesize phospholipids?

Abstract

1. The use of ‘marker’ enzymes for investigating the contamination by endoplasmic reticulum of mitochondrial and synaptosomal (nerve-ending) fractions isolated from guinea-pig brain was examined. NADPH–cytochrome c reductase appeared to be satisfactory. With the synaptosomal preparation there was a non-occluded enzymic activity believed to arise from contaminating microsomes and an occluded form released by detergent, which probably was derived from some type of intraterminal smooth endoplasmic reticulum. 2. Isolated brain mitochondria, both intact and osmotically shocked, could not synthesize more labelled phosphatidylcholine from CDP-[Me-14C]choline or phosphoryl[Me-14C]choline than could be accounted for by microsomal contamination. They could synthesize only phosphatidic acid and diphosphatidylglycerol from a [32P]Pi precursor and not nitrogen-containing phosphoglycerides or phosphatidylinositol. 3. The synaptosomal outer membrane and the intraterminal mitochondria could not synthesize phosphatidylcholine from CDP-[Me-14C]choline but the synaptic vesicles and probably the intraterminal ‘endoplasmic reticulum’ appeared to be capable of catalysing the incorporation of label from this substrate into their phospholipids. 4. Microsomal fractions and synaptosomes from guinea-pig brain could incorporate [Me-14C]choline into their phospholipids by a non-energy-requiring exchange process, which was catalysed by Ca2+. Fractionation of the synaptosomes after such an exchange had taken place revealed that the label was predominantly in the intraterminal mitochondria and not associated with membranes containing NADPH–cytochrome c reductase. 5. On the intraperitoneal injection of [32P]Pi into guinea pigs, incorporation of radioactivity into phosphatidylinositol and phosphatidic acid was much faster than into the nitrogen-containing phosphoglycerides. Mitochondria and microsomal fractions showed a roughly equivalent incorporation into individual phospholipids, and that into synaptosomes was appreciably less, whereas the phospholipids of myelin showed little 32P incorporation up to 10h.