Coexistence of phosphatidylcholine-specific phospholipase C and phospholipase D activities in rat cerebral cortex synaptosomes

Abstract

DAG derived from phosphatidylcholine (PtdCho) acts as a lipid second messenger. It can be generated by the activation of phospholipase D (PLD) and the phosphatidic acid phosphohydrolase type 2 (PAP2) pathway or by a PtdCho‐specific phospholipase C (PtdCho‐PLC). Our purpose was to study PtdCho‐PLC activity in rat cerebral cortex synaptosomes (CC Syn). DAG production was highly stimulated by detergents such as Triton X‐100 and sodium deoxycholate. Ethanol and tricyclodecan‐9‐yl‐xanthate potassium salt decreased DAG generation by 42 and 61%, respectively, at 20 min of incubation. These data demonstrate that both the PLD/PAP2 pathway and PtdCho‐PLC contribute to DAG generation in CC Syn. PtdCho‐PLC activity remained located mainly in the synaptosomal plasma membrane fraction. Kinetic studies showed Km and Vmax values of 350 μM and 3.7 nmol DAG × (mg protein × h)−1, respectively. Western blot analysis with anti‐PtdCho‐PLC antibody showed a band of 66 KDa in CC Syn. Our results indicate the presence of a novel DAG‐generating pathway in CC Syn in addition to the known PLD/PAP2 pathway.