Dopamine stimulation of phosphatidylcholine (lecithin) biosynthesis in rat brain neurons.

Abstract

Rat brain synaptosomes contain enzymes, phosphatidylethanolamine N-methyltransferase(s) (EC 2.1.1.17), that catalyze the methylation of endogenous phosphatidylethanolamine to form its mono-, di-, and trimethyl (i.e., phosphatidylcholine) derivatives. We observe that the activity of these enzymes is [hepatocytes (16)]. This process is rapid and not dependent on cAMP synthesis. Little information has been available on the control of PtdEtnMeTase activity within brain neurons. We report here that catecholamine neurotransmitters, especially dopamine, enhance the activities of phosph6lipid methylating enzymes in rat brain synaptosomes. METHODSPreparation of Subcellular Fractions. Male Sprague-Dawley rats (200-300 g; Charles River Breeding Laboratories) were housed under a 12-hr/12-hr light/dark schedule (lights on, 0900-2100 hr) and had free access to water and food (Charles River chow; 22% protein, 0.018% choline). The animals were decapitated at 1000 hr, and the brains were quickly dissected on ice and homogenized in 10 vol of 0.32 M sucrose in a Potter-Elvehjem homogenizer; the homogenate was then centrifuged at 1,000 x g for 15 min to remove tissue debris and nuclei. The supernatant was layered on top of 4 ml of 1.2 M sucrose and centrifuged at 165,000 X -g for 15 min in a Beckman SW 41 Ti rotor. The resulting pellet contained mitochondria; the material at the interface of the gradient (containing the synaptosomes and myelin) was diluted to obtain approximately 0.32 M sucrose, layered on top of 4 ml of 0.8 M sucrose, and centrifuged at 165,000 X g for 15 min (17). The material at the interface, containing myelin, and the pellet, shown to contain synaptosomes by electron microscopy, were collected. In some cases, the postnuclear supernatant was centrifuged at 20,000 x g for 20 min to obtain the P2 pellet (containing synaptosomes, mitochondria, and myelin). The post-P2 supernatant was centrifuged at 100,000 X g for 60 min to obtain a microsomal pellet.PtdEtnMeTase Assay. PtdEtnMeTase activity was assayed by a modification of the method described previously (4). The standard assay medium was changed to 50 mM Tris HCl, pH 7.5/5 mM MgCl2/0.2 mM EDTA/0. 1 mM ATP and tissue preparation (0.2-0.4 mg of protein) in a final volume of 120 1.l. The effect on phospholipid [3Himethyl incorporation of omitting various components of this medium was examined (see Table 1). Norepinephrine-HCI (Sigma), dopamineHCl (Sigma), haloperidol lactate (5 mg/ml, containing 1.8 mg of methylparaben and 0.2 mg of propylparaben; a gift from McNeil, Fort Washington, PA), were added to some tubes before incubation. Reactions were started by addition of 2.5 jCi of S-adenosyl[methyl-3H]methionine (New England Nuclear; 13-15 Ci/mmol; 1 Ci = 37 GBq) to a final AdoMet concentration of 1.2-1.6 ,uM. Samples were incubated for 30 min at 37TC. Reactions were stopped by addition of 3 ml of chloroform/methanol/HCI, 100:50:1 (vol/vol). Water-soluble radioactive. materials were extracted by washing the incubation mixture with two 2-ml portions of 0....

D4 dopamine receptor-mediated phospholipid methylation and its implications for mental illnesses such as schizophrenia

“…They further showed that neuroleptic drugs such as haloperidol blocked this effect, indicating involvement of a D2-like receptor. Subsequently, Leprohon et al 30 showed that dopamine could stimulate PLM in rat brain-derived synaptosomes and this effect was also blocked by haloperidol. We therefore undertook a more detailed study of dopamine receptor involvement in PLM.…”

Insulin stimulation of phospholipid methylation in isolated rat adipocyte plasma membranes.

“…(27), and synaptosomes (28) that suggest a surface membrane localization. The PtdEtn MeTase activity of adipocyte plasma membranes is higher than that reported for adrenal medulla endoplasmic reticulum (19), reticulocyte ghosts (3), or erythrocyte ghosts (28) and comparable to that reported for synaptosomes (5) and liver endoplasmic reticulum (23). Like the PtdEtn MeTase associated with rat liver endoplasmic reticulum (23) and L-929 cells (13), adipocyte plasma membrane PtdEtn MeTase was found to be indepen- A, dent of exogenous Mg2+.…”

“…After this report, investigators examined the possibility that enzymatic methylation of phospholipids plays a role in the transduction of receptormediated signals through the membranes of a variety of cells (2). In addition to agents such as f-agonists (3), benzodiazepine (4), dopamine (5), concanavalin A (6), and IgE antibody (7), several peptide hormones have been shown to alter phospholipid methylation, including glucagon (8), vasopressin (9), angiotensin (10), bradykinin (11), and nerve growth factor (12). This process is rapid and has been localized to a stimulation of plasma membrane methyltransferase activity in certain systems (13).…”

Effects of hyperbaric oxygen on lipid peroxidation and content of phospholipids in rat brain

“…The 2-fold decrease in the content ofphosphatidylethanolamine observed in our experiments was probably due to inhibition of phosphatidylserine decarboxylation and stimulation of phosphatidylcholine synthesis through methylation of phosphatidylethanolamine. Phosphatidylcholine synthesis during stress is induced by accumulation of dopamine (and total catecholamines) in the left hemisphere stimulating phosphatidylethanolamine methyltransferase and, therefore, phosphatidylcholine production [ 11 ]. Intensification of LPO caused by SOD was probably due to the fact that SOD considerably enhances cerebral blood flow in rats exposed to HBO 2 and increases oxygenation of brain tissues, thus promoting generation of reactive oxygen species [14].…”