Hydrolysis of Endogenous Diacyglycerol and Monoacylglycerol by Lipases in Rat Brain Microsomes

[¹⁴C] Fatty acids were injected into brains of 18-day-old rats and the labeled microsomes were isolated and treated with phospholipase C. This resulted in a degradation of the membranal phospholipids, thus forming radioactivelylabeled, endogenous diacylglycerol. Hydrolysis of this diacylglycerol by the microsomal lipases was followed by incubating the membranes at pH 7.4 and 4.8. After phospholipase C treatment and further incubation at pH 7.4, part of the endogenous diacylglycerol was degraded by the membranal alkaline diacylglycerol lipase. The monoacylglycerol thus formed was further degraded by the membranal monoglyceride lipase. Pretreatment of the membranes with phospholipase C at pH 7.4, followed by incubation at pH 4.8, resulted in a further decrease in the amount of diacylglycerol accompanied by concommitant increases in monoacylglycerol and free fatty acids. These experiments verify the existence of two diacylglycerol lipases in rat brain microsomes, one with an alkaline and the second with an acidic pH optimum, but of only one monoacylglycerol lipase with an alkaline pH optimum (Cabot, M. C., and Gatt, S. (1976), Biochim. Biophys. Acta 431, 105-115). This system provides a method for the study of membrane-bound lipolytic enzymes acting on endogenous substrate, thereby obviating the use of exogenous dispersions of lipid substrates.