Insulin Action on Polyunsaturated Phosphatidic Acid Formation in Rat Brain: An “In Vitro” Model with Synaptic Endings from Cerebral Cortex and Hippocampus

The highly efficient formation of phosphatidic acid from exogenous 1-stearoyl-2-arachidonoyl-sn-glycerol (SAG) in rat brain synaptic nerve endings (synaptosomes) from cerebral cortex and hippocampus is reported. Phosphatidic acid synthesized from SAG or 1,2-dipalmitoylsn-glycerol (DPG) was 17.5 or 2.5 times higher, respectively, than from endogenous synaptosomal diacylglycerides. Insulin increased diacylglycerol kinase (DAGK) action on endogenous substrate in synaptic terminals from hippocampus and cerebral cortex by 199 and 97%, respectively. Insulin preferentially increased SAG phosphorylation from hippocampal membranes. In CC synaptosomes insulin increased phosphatidic acid (PA) synthesis from SAG by 100% with respect to controls. Genistein (a tyrosine kinase inhibitor) inhibited this stimulatory insulin effect. Okadaic acid or cyclosporine, used as Ser/Threo protein phosphatase inhibitors, failed to increase insulin effect on PA formation. GTPcS and particularly NaF were potent stimulators of PA formation from polyunsaturated diacylglycerol but failed to increase this phosphorylation when added after 5 min of insulin exposure. GTPcS and NaF increased phosphatidylinositol 4,5 bisphosphate (PIP2) labeling with respect to controls when SAG was present. On the contrary, they decreased polyphosphoinositide labeling with respect to controls in the presence of DPG. Our results indicate that a DAGK type 3 (DAGKe) which preferentially, but not selectively, utilizes 1-acyl-2-arachidonoyl-sn-glycerol and which could be associated with polyphosphoinositide resynthesis, participates in synaptic insulin signaling. GTPcS and NaF appear to be G protein activators related to insulin and the insulin receptor, both affecting the signaling mechanism that augments phosphatidic acid formation.Fil: Zulian, Sandra Edith. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; ArgentinaFil: Ilincheta, Monica Graciela. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; ArgentinaFil: Giusto, Norma Maria. Consejo Nacional de Investigaciones Científicas y Técnicas. Centro Científico Tecnológico Conicet - Bahía Blanca. Instituto de Investigaciones Bioquímicas de Bahía Blanca. Universidad Nacional del Sur. Instituto de Investigaciones Bioquímicas de Bahía Blanca; Argentin