Somatostatin effects on the cyclic AMP system and lipid metabolism in mouse brain

The effect of somatostatin on cyclic AMP-protein kinase system and lipid metabolism was studied in mouse brain. Subcutaneous injection of the peptide decreased the cyclic AMP and cyclic GMP levels (70% and 60% respectively) as well as protein kinase and triglyceride lipase activities (30%). Cyclic AMP binding protein activity was not affected. Experiments carried out with [¹⁴C]acetate as precursor of lipids seem to indicate that somatostatin blocks the fatty acid turnover. On the other hand, the general decrease of³²P incorporation into all phospholipids by somatostatin suggests that the peptide interferes with the precursor uptake into phospholipids. The findings reported here indicate that somatostatin has a role on brain metabolism and further add more data in support for its neuromodulating action.     

Accelerated clearance of plasma triglycerides and free fatty acids in omega3-depleted rats.

The present study aims mainly at exploring the effects of a severe depletion in polyunsaturated long-chain omega3 fatty acids upon the fate of circulating lipids. The plasma concentration and fatty acid pattern of triglycerides, diglycerides, free fatty acids, and phospholipids were measured in omega3-depleted and control rats injected intravenously one hour before sacrifice with either saline, a control medium-chain triglyceride:olive oil emulsion or a medium-chain triglyceride:fish oil emulsion recently found to rapidly increase the phospholipid content of C20:5omega3 and C22:6omega3 in different cell types. The estimated fractional removal rate of the injected triglycerides and the clearance of free fatty acids from circulation were both higher in omega3-depleted rats than in control animals. The injection of the lipid emulsions apparently inhibited intracellular lipolysis, this being least pronounced in omega3-depleted rats. The increased clearance of circulating triglycerides and unesterified fatty acids in omega3-depleted rats may favor the cellular accumulation of lipids. In turn, such an accumulation and the lesser regulatory inhibition of tissular lipolysis may match the increased clearance of circulating unesterified fatty acids and, hence, account for the lack of any significant difference in plasma unesterified fatty acid concentration between these and control animals.     

Activation of protein kinase C by cis- and trans-fatty acids and its potentiation by diacylglycerol

Both cis- and trans-unsaturated but not saturated fatty acids activated protein kinase C purified to apparent homogeneity from rat brain. Fatty-acid-induced enzyme activation was not more than additive with that by phospholipids and was potentiated by diacylglycerol. Recently, we demonstrated that cis- and trans-unsaturated fatty acids induced platelet aggregation and phosphorylation of specific proteins. Both events were potentiated by a cell-permeable diacylglycerol [(1987) Biochem. Biophys. Res. Commun. 149, 762-768]. Thus, trans-unsaturated fatty acids may provide useful experimental tools for the study of protein kinase C activation in vitro and in vivo. Our results suggest that fatty acids and diacylglycerol may synergistically be involved in hormonal stimulation of protein kinase C, as certain hormonal stimuli cause release of diacylglycerol and fatty acids from phospholipids by parallel activation of phospholipases C and A2.     

Inhibition of fatty acid synthesis and stimulation of glycogen breakdown by vasopressin in the perfused mouse liver

1. Vasopressin (anti-diuretic hormone, [8-arginine]vasopressin) inhibited the synthesis de novo of fatty acids (measured with (3)H(2)O and U-(14)C-labelled lactate or U-(14)C-labelled glucose) and stimulated glycogen breakdown in the perfused liver of fed mice. 2. The concentration dependence of these effects (range 200-1000muunits/ml, i.e. 0.5-2.5ng/ml) resembled that for the action on glycogen breakdown which was previously reported for rat liver. 3. The appearance of newly synthesized fatty acids in both phospholipids and triglycerides was inhibited by vasopressin, whereas synthesis of cholesterol was less affected. 4. Inhibition of hepatic lipogenesis by vasopressin is the most potent short-term hormonal action on this process yet reported. Aspects of the effect are discussed, including the lack of a role for cyclic AMP, and a possible link with vasopressin action on glycogen metabolism.     

Dose-dependent effects of linear and cyclic somatostatin on ethanol-induced gastric erosions: the role of mast cells and increased vascular permeability in the rat

Somatostatin prevents hemorrhagic gastric erosions produced by ethanol. In this paper we describe studies with linear (reduced) and cyclic (oxidized) synthetic somatostatin-14 in the rat model of ethanol-induced gastric mucosal injury. The linear form of somatostatin was more potent at concentrations of 10(-9) to 10(-8) mol per rat than the cyclic isomere. However, at a concentration of 10(-7) mol per rat i.p. injection of linear somatostatin significantly (P less than 0.01) enhanced gastric erosions caused by the alcohol. The area of hemorrhagic mucosal lesions correlated significantly (r = -0.846) with mast cell depletion in the gastric mucosa of the animals. Increased vascular permeability and mast cell degranulation were also observed after intradermal injection of linear or cyclic somatostatin. The 'cytoprotective' as well as the aggravating potency of linear somatostatin may be connected to gastric mucosal mast cell activity in the rat.     

Differential changes in the fatty acid composition of the main lipid classes of chick plasma induced by dietary coconut oil

For a better understanding of the hyperlipidemic function of saturated fat, we have studied the comparative effects of diet supplementation with 10 and 20% coconut oil on the main lipid classes of chick plasma. Changes in fatty acid composition of free fatty acid and triglyceride fractions were parallel to that of the experimental diet. Thus, the increase in the percentages of 12:0 and 14:0 acids may contribute to the hypercholesterolemic effects of coconut oil feeding. Plasma phospholipids incorporated low levels of 12:0 and 14:0 acids whereas 18:0, the main saturated fatty acid of this fraction, also increased after coconut oil feeding. The percentage of 20:4 n-6 was higher in plasma phospholipids than in the other fractions and was significantly decreased by our dietary manipulations. Likewise, minor increases were found in the percentages of 12:0 and 14:0 acids in plasma cholesterol esters. However, the percentage of 18:2 acid significantly increased after coconut oil feeding. Our results show a relationship between fatty acid composition of diets and those of plasma free fatty acid and triglyceride fractions, whereas phospholipids and cholesterol esters are less sensitive to dietary changes.     

Effect of "marine" phospholipids omega-3 fatty acids on the composition of fatty acids of rat liver microsomal membrane under oxidative stress

As a result of experiments conducted the marine phospholipids preparation enriched by omega-3 fatty acids was defined to modify fatty acids content due to changes of fatty acids level change in the neutral lipids and phospholipids fractions. As well it was identified, that at the oxidative stress induced by administration of CCl4 the growth of arachidonic and docozahexaenoic acids in the neutral lipids fractions was observed if compare with the norm. At the same time, the presented fatty acids in the phospholipids fractions remained unchanged. At oxidative stress the phospholipids fraction reacts to levels of arachidonic and docozahexaenoic acids just only as a result of administrating phospholipids with omega-3 fatty acids. The most attractive is the change of correlation C20:4/C22:6--increasing at administration of CCl4 and decreasing both at phospolipids and vitamin E injection. Thus, at the oxidative stress the first reacting ones are the fatty acids of neutral lipids microsomal membranes.     

Effects of pneumogastric nerve stimulation and injection of a beta 2 stimulant (terbutaline) on the lipid composition of the alveolar lining fluid and on pulmonary compliance in the rat

The effects of vagal stimulation and of terbutaline injection on lipidic composition of alveolar fluid and pulmonary compliance were studied. Three groups of rats were used: control, after right vagus nerve stimulation, after 0.2 mg terbutaline injection. The lungs of the rats were isolated. We studied pulmonary pressure-volume curves with air and we measured pulmonary compliance. We realised an alveolar lavage to obtain alveolar lipids. We observed: Vagus nerve stimulation and beta 2 agonist significantly increased fatty acids of total lipids respectively by 52.5% and 25.5% and phospholipids, respectively by 43.6% and 25.7%. beta 2 agonist did not change fatty acid composition of total lipids and phospholipids. Right vagus nerve stimulation increased the percentage of palmitic acid in phospholipids and decreased the percentage of saturated fatty acids and of palmitic acid in total lipids. Terbutaline injection induced more significant changes in pressure-volume curves and in pulmonary compliance than right vagus nerve stimulation. Our results suggest that both vagal stimulation and beta 2 agonists increase lipid release in alveolar lining, but only vagal stimulation modifies the composition of these lipids. These modifications could be, at least in part, correlated with the changes observed in the pressure-volume curves.     

Incorporation of exogenous fatty acids into phospholipids by cultured hamster fibroblasts. Effect of SV40 transformation

In situ incorporation of two saturated (palmitic, 16:0; stearic, 18:0) and three unsaturated fatty acids (oleic, 18:1; linoleic, 18:2; arachidonic, 20:4) into the four major phospholipids, sphingomyelin, PC, PI and PE, was followed. Transformed cells incorporated unsaturated fatty acids more rapidly, whereas no significant differences were found concerning saturated fatty acids. In vitro determination of phospholipid acylation showed that incorporation of coenzyme A-activated forms of two saturated fatty acids (16:0 and 18:0) and one unsaturated fatty acid (18:1) into phospholipids was increased in transformed cells. Comparison of results obtained in situ and in vitro strongly suggests that incorporation of fatty acids into phospholipids in cultured cells is not limited by acyltransferase activities.     

Effects of honeybee insulin-immunoreactive peptide on haemolymph lipid and carbohydrate: Interaction of vertebrate somatostatin

The haemolymph lipid and carbohydrate concentrations were determined in emerging worker honeybees following injection of [I] saline, [II] a bee insulin-immunoreactive peptide (BIRP) and [III] BIRP + vertebrate somatostatin. BIRP promptly lowered trehalose, glucose, diacylglycerol and phospholipid concentrations, and increased the levels of fatty acid and triacylglycerol. It also induced a positive correlation between 1,2-diacylglycerol and short chain triacylglycerol, and completely reversed a negative correlation between long chain triacylglycerol and fatty acid, as well as a positive correlation between 1,3-diacylglycerol and steroid, both of the latter correlation appearing following treatment [I]. In all correlations except those between fatty acid, phospholipid and diacylglycerol, injection of vertebrates somatostatin together with BIRP partially neutralizes the specific effects of BIRP injected alone, a result similar to the action of somatostatin in vertebrates. BIRP is an endogenous intestinal insulin-like peptide whose physiological activity may be modulated by somatostatin-like molecules.     

Interaction of propranolol and somatostatin with the octopaminergic response of honeybee lipemia in vivo

The injection of DL-octopamine to emerging adult honey bees (1 nanomol per individual) promptly rises the haemolymph levels of steroids, diacyl and triacylglycerols (over 100% increase occurring after 10 min). Free fatty acids are then increased (by 131%) 45 mn after injections. These responses are completely abolished by 2.5 nanomol propranolol and 0.07 nanomol cyclic somatostatin. Octopamine thus seems to be more strongly involved in the lipemia than in the glycemia regulation, by contrast with noradrenaline which is, apparently, more active on carbohydrates.     

Monolayer and calorimetric studies of phosphatidylcholines containing branched-chain fatty acids and of their interactions with cholesterol and with a bacterial hopanoid in model membranes

Although methyl iso- and anteiso-branched fatty acids occur widely in the membrane lipids of prokaryotic microorganisms, relatively little is known about the physical properties of phospholipids containing these fatty acids. We report here a monolayer and differential scanning calorimetric characterization of several synthetic phosphatidylcholines containing branched-chain fatty acids, and describe the interactions of these phospholipids with cholesterol and with a bacterial hopanoid. We find that monolayers as well as bilayers of methyl isobranched- and especially of methyl anteisobranched-fatty-acid-containing phosphatidylcholines exhibit a reduced solid-to-fluid phase transition temperature in comparison with linear saturated fatty acid-containing phosphatidylcholines of comparable chain length. We also find that the liquid-condensed or gel states of branched-chain fatty acid-containing phosphatidylcholines are partially disordered relative to those of phospholipids containing linear saturated fatty acids, although the presence of a methyl branch has only a small effect on hydrocarbon chain packing in the liquid-expanded or liquid-crystalline states. The presence of cholesterol was found to produce a marked condensation of liquid-expanded films and a small condensation of liquid-condensed films, whether the phosphatidylcholine contained linear or branched-chain fatty acyl constituents. The presence of a bacterial hopanoid produced similar, although slightly smaller, monolayer-condensing effects, indicating that these compounds may perform a cholesterol-like function in bacterial membranes.     

Somatostatin immunoneutralization affects plasma metabolite concentrations in the domestic fowl

Young leghorn cockerels were injected with antiserum to somatostatin (anti-SRIF) and plasma glucose, free fatty acids and alpha-amino nitrogen concentrations determined. Plasma glucose concentrations increased rapidly after anti-SRIF and remained high for up to 2 hr. Two different antisera tested had hyperglycaemic activity. Plasma free fatty acids also increased rapidly after administration of the two different anti-SRIFs, and remained high for about 1 hr. Plasma alpha-amino nitrogen increased during the first 30 min after anti-SRIF, then declined to levels significantly lower than control by 1-2 hr after injection. Anaesthesia reduced plasma concentrations of glucose and alpha-amino nitrogen, and also reduced the changes of these metabolites following anti-SRIF. The results show the importance of endogenous somatostatin in the regulation of plasma metabolite concentrations.     

Properties of the cyclic GMP phosphodiesterase from rat lung. Inhibition by unsaturated fatty acids

Catalytic and regulatory properties of the major form of cyclic GMP phosphodiesterase (3':5'-cyclic-GMP 5'-nucleotidohydrolase, EC 3.1.4.35) from rat lung were studied. The enzyme partially purified by a DEAE-Sepharose chromatography displayed a much higher affinity toward cyclic GMP than toward cyclic AMP, the apparent Km values being 5.7 microM and 482 microM for the guanylic and the adenylic cyclic nucleotide, respectively. In contrast, the V value for cyclic AMP was about 3-times higher than the V value for cyclic GMP. Linear double reciprocal plots of initial velocity were observed with each cyclic nucleotide. From 10(-8) to 3.3 X 10(-6) M, cyclic GMP did not change the hydrolysis of 1 or 10 microM cyclic [3H]AMP, while it became inhibitory at higher concentrations. In contrast with a calmodulin-sensitive phosphodiesterase prepared from rat brain, the lung enzyme was not stimulated by a heat-stable Ca2+-dependent factor from rat lung or by rat brain calmodulin or by lipids including fatty acids and lysophosphatidylcholine. Various unsaturated 18- and 20-carbon fatty acids inhibited at varying degrees the cyclic GMP phosphodiesterase from rat lung. The inhibitory potency increased with the number of double bonds in the hydrocarbon chain. In contrast, the methyl esters of the unsaturated fatty acids and the saturated fatty acids of variable hydrocarbon chain lengths had no appreciable effects. A linear Hill plot of phosphodiesterase inhibition with a slope of unity was obtained with arachidonic acid up to 30 microM, suggesting only one type of inhibitory site. In this range of concentrations the inhibition was entirely reversible. Kinetics analysis demonstrated that up to 30 microM arachidonic acid was a purely competitive inhibitor with an apparent Ki of 20 microM. Over 30 microM, the Hill coefficient increased progressively, indicating the binding to other inhibitory sites, while the reversibility disappeared.     

Fatty acid synthesis in Escherichia coli is indirectly inhibited by phenethyl alcohol.

Experiments were performed to determine how phenethyl alcohol inhibits phospholipid synthesis in E. coli. At a nonbacteriostatic concentration, the drug reduces the rate of de novo fatty acid and phospholipid synthesis by 60 to 70%. The inhibition of fatty acid synthesis was found to be a secondary consequence of the inhibition of phospholipid synthesis. Phenethyl alcohol reduces the rate of incorporation of exogenous fatty acids into the phospholipids of a fatty acid auxotroph by 60%. These results indicate that this drug controls phospholipid synthesis beyond the level of fatty acid synthesis. Phenethyl alcohol inhibits the synthesis of phospholipids containing saturated fatty acids to a greater extent than it does the synthesis of phospholipids containing unsaturated fatty acids. It controls the synthesis of phospholipids containing saturated fatty acids at both the level of fatty acid synthesis and the level of incorporation of the saturated fatty acids into phospholipids. The synthesis of phospholipids containing unsaturated fatty acids is inhibited at the level of incorporation of the fatty acids into phospholipids.     

Changes in fatty acid composition during cell differentiation in the small intestine of suckling piglets.

Alterations of phospholipid fatty acid composition in the renewing intestine were studied in the infant piglet. Newborn piglets were fed from birth to 2 weeks of age a concentrated cow's milk which defined a standard supply of dietary fatty acids. Phospholipids were isolated from the whole mucosa, isolated intestinal cells and purified brush border membranes. Intestinal cells were isolated according to their position along the crypt-villus axis and cell phospholipids were extracted at each step of differentiation. Changes in fatty acid composition of cell phospholipids were related to those of lactase activity in the corresponding cell homogenates. In cell phospholipids, the relative content of linoleic and linoleic acids increased about 2-fold from crypt base to villus tip. Substantial contents of alkenylacyl glycerophospholipids (plasmalogens) were found in crypt cell phospholipids and in purified brush border membrane phosphatidylethanolamine (11 and 14% of alkenyl groups by weight of total fatty acids, respectively). The proportion of alkenylacyl glycerophospholipids decreased as cells ascended the villus column and became more differentiated. The results show that fatty acid compositional changes in differentiating cell phospholipids occurred in the immature intestine (before weaning) and suggest that these alterations might be related to the appearance of specific functions.     

A new approach to the study of phospholipid-protein interactions in biological membranes. Synthesis of fatty acids and phospholipids containing photosensitive groups.

In a general approach to the study, in vivo and in vitro, of the interactions between phospholipids and proteins in biological membranes, a variety of fatty acids containing photosensitive groups in different positions in the alkyl chains has been synthesized. The fatty acids synthesized include: 16-azidopalmitelaidic acid, 12-azidooleic acid, 6-, 9-, 11-, and 12-azidostearic acid, 12-omicron-(ethyl-2-diazomalonyl)stearic and -oleic acids, 12-omicron-(4-azido-2-nitrophenyl)stearic and -oleic acids, and 12-oxo-10-octadecenoic acid. Some of the above synthetic fatty acids were also prepared in the radioactively labeled form. For in vitro studies, many of the above fatty acids were used to acylate the 2 position in the preparation of a number of mixed acylphosphatidylcholines and mixed acylphosphatidylethanolamines. On sonication, the synthetic phospholipids formed sealed vesicles. Intermolecular cross-linking of the fatty acyl chains in phospholipids was demonstrated on photolysis of the vesicles.     

Phospholipids and fatty acids of Neisseria gonorrhoeae.

The phospholipids and fatty acids of two strains of Neisseria gonorrhoeae of different penicillin susceptibilities were examined. The phospholipids, which comprise about 8% of the dry weight of the cells, consisted of phosphatidylethanolamine (70%) and phosphatidylglycerol (20%); small amounts of phosphatidylcholine and traces of cardiolipin were also present. Growing and stationary-phase cells were similar in content and composition of phospholipids except for phosphatidylcholine, which increased two- to fivefold in the stationary-phase cells. The fatty acids of the phospholipids were characterized by two major acids, palmitic and a C16:1, with myristic and a C18:1 acid present in smaller amounts. The fatty acids present in purified phospholipid fractions varied considerably in relative proportions from fraction to fraction. No significant difference in the composition of phospholipids from the two strains was evident. Large amounts of beta-hydroxy lauric acid were detected only after saponification of the organisms. Differences in the lipid composition between the gonococcus and other gram-negative bacteria are discussed.     

Indentification and Localization of the Fatty Acids in Haemophilus parainfluenzae

Haemophilus parainfluenzae was capable of synthesizing 22 fatty acids. These fatty acids were equivalent to 4% of the bacterial dry weight. These fatty acids were localized in the membrane-wall complex, which contained the respiratory pigments, the quinone, and the phospholipids. The fatty acids which could be extracted with organic solvents comprised 86% of the total fatty acids of the cell. These fatty acids were distributed as 98% in the phospholipids and 1.9% in the neutral lipids, of which 0.5% were free fatty acids. Palmitic, palmitoleic, oleic, and vaccenic acids comprised 72% of the total fatty acids and were found almost exclusively in the phospholipids. The phospholipids also contained the cyclopropane fatty acids. The neutral lipids contained significant proportions of the odd-numbered branched and straight-chain fatty acids. The principal free fatty acids were n-dodecanoic and pentadecenoic acids. The nonextractable wall complex contained 14% of the total fatty acids. These wall fatty acids were rendered soluble only after saponification. The wall fraction contained all of the beta-hydroxymyristic acid and most of the myristoleic and pentadecenoic acids. The significance of the distribution of fatty acids between nonesterified, neutral lipid, phospholipid, and nonextractible wall remains to be determined.     

Alteration of fatty acid composition of LM cells by lipid supplementation and temperature.

Alteration of the fatty acid composition of monolayer cultures of LM cells grown in chemically defined medium was achieved by supplementation with fatty acids complexed to bovine serum albumin. Phospholipids containing up to 40% linoleate were found in cells grown in medium containing 20 mu g of linoleate/ml. Incorporation of linoleate into phospholipids reached a plateau after 12-24 hr, and cells remained viable for at least 3-4 days. Although linoleic, linolenic, and arachidonic acids were incorporated into LM cells equally well, only the latter was elongated by these cells under these experimental conditions. Nonadecanoic acid was incorporated to a lesser extent than the polyunsaturated fatty acids. Phosphatidylcholine and phosphatidylethanolamine of LM cells had different fatty acid compositions; phosphatidylethanolamine contained more longer chain and unsaturated fatty acids. Cells were also grown in the absence of choline and presence of choline analogs such as N,N-dimethylethanolamine, N-methylethanolamine, 3-amino-1-propanol, and 1-2-amino-1-butanol. The analog phospholipids in these cells had fatty acid compositions which were intermediate between those of phosphatidylethanolamine and phosphatidylcholine of control cells grown in the presence of choline. Linoleate was found in both phosphatidylcholine and phosphatidylethanolamine of cells supplemented with linoleate. The sphingolipid fraction of these cells, however, did not contain significant amounts of linoleate. When linoleate was present in the phospholipids, compensatory decreases in the oleate and palmitoleate content of phospholipids were observed. Lowering of the growth temperature to 28 degrees produced an increase in unsaturate fatty acid content of the phospholipids. When linoleate was supplied to cells grown at 28 degrees, there was no further increase in the unsaturated fatty acid composition of the phospholipids. Using both fatty acid supplementation and lowered growth temperature, LM cell membranes can be produced which have phospholipids with vastly different fatty acid compositions.