Arachidonic Acid Turnover Research Articles

Calcium dependency of arachidonic acid incorporation into cellular phospholipids of different cell types

Ca 2+-independent phospholipase A 2 (iPLA 2) is involved in the incorporation of arachidonic acid (AA) into resting macrophages by the generation of the lysophospholipid acceptor. The role of iPLA 2 in AA remodeling in different cells was evaluated by studying the Ca 2+ dependency of AA uptake from the medium, the incorporation into cellular phospholipids, and the effect of the iPLA 2 inhibitor bromoenol lactone on these events. Uptake and esterification of AA into phospholipids were not affected by Ca 2+ depletion in human polymorphonuclear neutrophils and rat fibroblasts. The uptake was Ca 2+ independent in chick embryo glial cells, but the incorporation into phospholipids was partially dependent on extracellular Ca 2+. Both events were fully dependent on extra and intracellular Ca 2+ in human platelets. In human polymorphonuclear neutrophils, the kinetics of incorporation in several isospecies of phospholipids was not affected by the absence of Ca 2+ at short times (<30 min). The involvement of iPLA 2 in the incorporation of AA from the medium was confirmed by the selective inhibition of this enzyme with bromoenol lactone, which reduced ≤50% of the incorporation of AA into phospholipids of human neutrophils. These data provide evidence that suggests iPLA 2 plays a major role in regulating AA turnover in different cell types.

Dynamics of docosahexaenoic acid metabolism in the central nervous system: lack of effect of chronic lithium treatment.

Using a method and model developed in our laboratory to quantitatively study brain phospholipid metabolism, in vivo rates of incorporation and turnover of docosahexaenoic acid in brain phospholipids were measured in awake rats. The results suggest that docosahexaenoate incorporation and turnover in brain phospholipids are more rapid than previously assumed and that this rapid turnover dilutes tracer specific activity in brain docoshexaenoyl-CoA pool due to release and recycling of unlabeled fatty acid from phospholipid metabolism. Fractional turnover rates for docosahexaenoate within phosphatidylinositol, choline glycerophospholipids, ethanolamine glycerophospholipids and phosphatidylserine were 17.7, 3.1, 1.2, and 0.2 %.h(-1), respectively. Chronic lithium treatment, at a brain level considered to be therapeutic in humans (0.6 micromol.g(-1)), had no effect on turnover of docosahexaenoic acid in individual brain phospholipids. Consistent with previous studies from our laboratory that chronic lithium decreased the turnover of arachidonic acid within brain phospholipids by up to 80% and attenuated brain phospholipase A2 activity, the lack of effect of lithium on docosahexaenoate recycling and turnover suggests that a target for lithium's action is an arachidonic acid-selective phospholipase A2.

Agonist-stimulated glycerophospholipid acyl turnover in alveolar macrophages

The inflammatory compounds β-glucan, a particulate agonist, and tannin, a soluble agonist, are present in cotton dust and, when inhaled, cause massive arachidonic acid release from alveolar macrophages. Earlier work had shown that these agonists exhibit different effects on arachidonate liberation and release, and that only tannin inhibits the uptake and incorporation of exogenous arachidonic acid, suggesting inhibition of reacylation. Here we have used the time-dependent incorporation of 18O from H 2 18O-containing media into glycerophospholipid acyl groups as an indicator of acyl turnover in resting and agonist-treated rabbit alveolar macrophages. Highest turnover rates were seen in phosphatidylinositol (∼30% per hour) and in choline phospholipids (10–20% per hour). Both β-glucan and tannin stimulated acyl turnover, especially arachidonic acid turnover, in these and other lipid classes by a factor of 2 or more. We conclude that neither agonist promotes arachidonic acid accumulation in and release from alveolar macrophages by inhibiting reacylation.

Detection of acyl-CoA synthetase, acyl-CoA:lysophospholipid acyltransferase and phospholipase A 2 activities in non-pregnant and pregnant guinea-pig uterine tissues

Acyl-CoA synthetase (ACS), acyl-CoA:lysophospholipid acyltransferase (ACLAT) and phospholipase (PL) A 2 activities were detected in guinea-pig endometrium on days 7 and 15 of the cycle, and on days 15, 29 and 36 of pregnancy. Ovariectomy of non-pregnant animals resulted in an increase in the apparent activities of these three enzymes which was reversed by treatment with oestradiol and/or progesterone. ACS, ACLAT and PLA 2 activities were detected in day 15 conceptuses, and in the placenta, sub-placenta, chorion and amnion on days 29 and 36 of pregnancy. Apparent activities of the enzymes were generally higher in the fetal membranes than in the placental tissue. This study has established that the enzymes involved in turnover of arachidonic acid in phospholipids are present in tissues in the non-pregnant and pregnant guinea-pig uterus. The higher apparent activities of enzymes (ACS and ACLAT) involved in arachidonic acid uptake compared to the enzyme (PLA 2) involved in arachidonic acid release is in agreement with there being very low concentrations of free arachidonic acid in tissues.

Tachykinins as enhancers of prostaglandin E2-induced intraocular inflammation

The effects of tachykinins on prostaglandin E 2 (PGE 2 )-induced intraocular inflammation were investigated. PGE 2 (0.01% or 0.1%) instillation induced iridal hyperemia and protein leakage into the aqueous humor in rabbits, but caused minimal miosis. Intravitreally injected substance P (SP) or neurokinin A (NKA), on the other hand, did not induce protein leakage into the aqueous humor in normal rabbits, but they (SP 10 µg/eye or NKA 50 µg/eye) did induce long-lasting miosis. The miotic activity of SP was about fivefold stronger than that of NKA. Intravitreally injected SP (10 µg/eye) but not NKA (50 µg/eye) increased PGE 2 concentration in the aqueous humor in normal rabbits. In addition, SP (10 µg/eye) or NKA (50 µg/eye) markedly enhanced protein leakage into the aqueous humor induced by PGE 2 instillation. Pretreatment with indomethacin partially blocked the enhancing effect of SP on protein leakage, while it did not block that of NKA. These results suggest that SP or NKA may enhance intraocular inflammation in vivo. However, the mechanisms of these effects of SP and NKA may be different. The enhancing effect of SP in eye inflammation may be partially due to an increased turnover of arachidonic acid into PGE 2 caused by activation of the enzyme cyclooxygenase.

Arecoline stimulation of radiolabeled arachidonate incorporation from plasma into brain microvessels of awake rat.

The cholinergic agonist, arecoline, was used to examine the effects of cholinergic stimulation upon incorporation of radiolabeled arachidonic acid from blood into cerebral microvessels of awake rats. Animals received a single i.p. injection of arecoline (1 mg/kg) followed 3 to 5 minutes later by a 5 minute intravenous infusion of [1-14C]arachidonic acid (AA) (170 microCi/kg) via the femoral vein. Timed arterial blood samples were collected over 20 minutes following the start of infusion, after which the animal was killed, and the brain was removed. The incorporation coefficient k* for [1-14C]AA was approximately 2-fold higher in microvessels isolated from arecoline-injected than from sham-injected animals. The data demonstrate in an in vivo paradigm, that activation of cholinergic pathways within the rat CNS stimulates arachidonic acid turnover in cerebral microvessels. This suggests a direct involvement of this fatty acid in second messenger function within microvessel endothelial cells and possibly attached pericytes.

Dynamics of arachidonic acid turnover upon differentiation of human monocytoid cells: D.A. Namgaladze, M.G. Sergeeva, I.A. Yudushkin, M.V. Gonchar, and A.T. Mevkh, A.N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow, Russia

Dynamics of arachidonic acid turnover upon differentiation of human monocytoid cells: D.A. Namgaladze, M.G. Sergeeva, I.A. Yudushkin, M.V. Gonchar, and A.T. Mevkh, A.N. Belozersky Institute of Physico-Chemical Biology, Moscow State University, Moscow, Russia

Identification of two molecular species of rat brain phosphatidylcholine that rapidly incorporate and turn over arachidonic acid in vivo.

In vivo rates of arachidonic acid incorporation and turnover were determined for molecular species of rat brain phosphatidylcholine (PtdCho) and phosphatidylinositol (PtdIns). [3H]Arachidonic acid was infused intravenously in pentobarbital-anesthetized rats at a programmed rate to maintain constant plasma specific activity for 2-10 min. At the end of infusion, animals were killed by microwave irradiation, and brain phospholipids were isolated, converted to diacylglycerobenzoates, and resolved as molecular species by reversed-phase HPLC. Most [3H] arachidonate (> 87%) was incorporated into PtdCho and PtdIns, with arachidonic acid at the sn-2 position and with oleic acid (18:1), palmitic acid (16:0), or stearic acid (18:0) at the sn-1 position. However, 10-15% of labeled brain PtdCho eluted in a small peak containing two molecular species with arachidonic acid at the sn-2 position and palmitoleic acid (16:1) or linoleic acid (18:2) at the sn-1 position. Analysis demonstrated that tracer was present in both the 16:1-20:4 and 18:2-20:4 PtdCho species at specific activities 10-40 times that of the other phospholipids. Based on the measured mass of arachidonate in each phospholipid molecular species, half-lives were calculated for arachidonate of < 10 min in 16:1-20:4 and 18:2-20:4 PtdCho and 1-3 h in 16:0-20:4, 18:1-20:4 PtdCho and PtdIns. The very short half-lives for arachidonate in the 16:1-20:4 and 18:2-20:4 PtdCho molecular species suggest important roles for these molecules in brain phospholipid metabolism and signal transduction.

Effect of 17β-estradiol on chondrocyte membrane fluidity and phospholipid metabolism is membrane-specific, sex-specific, and cell maturation-dependent

In this study we examined the hypothesis that 17β-estradiol exerts both rapid and direct, nongenomic effects on cells in the endochondral pathway. To do this, we used a cell culture model in which chondrocytes at two distinct stages of cell maturation are isolated from the costochondral cartilage of male and female rats, and examined the short-term effect of 17α- and 17β-estradiol on [ 14C]arachidonic acid turnover in the cell layer and phospholipase A 2 specific activity in plasma membranes and extracellular matrix vesicles isolated from similarly prepared cultures. In addition, the effect of 17α- and 17β-estradiol on plasma membrane and matrix vesicle membrane fluidity was assessed. The effect of hormone on arachidonic acid turnover was rapid, time- and concentration-dependent, stereo-specific, and cell maturation-specific. Only resting zone cells from female rats were affected, and only 17β-estradiol elicited a response. Similarly, only female rat resting zone chondrocytes exhibited a change in phospholipase A 2 activity after a 24 h exposure to hormone, causing an increase in enzyme activity in the matrix vesicles, but not plasma membranes. When isolated membranes were incubated directly with hormone, membrane fluidity was decreased in both plasma membranes and matrix vesicles isolated from female rat resting zone chondrocyte cultures. This nongenomic effect was dose-dependent and stereo-specific and differentially expressed in the two membrane fractions with respect to time course and magnitude of response. These results support the hypothesis that 17β-estradiol has a rapid action on chondrocyte membrane lipid metabolism and suggest that specific membrane components, characteristic of a particular sex and state of cell maturation, are involved in the nongenomic effects of this sex hormone on isolated matrix vesicles and plasma membranes.

Arachidonate metabolism in human placenta, fetal membranes, decidua and myometrium: lipoxygenase and cytochrome p450 metabolites as main products in HPLC profiles

Eicosanoids play a key role in pregnancy maintenance and parturition. We investigated the metabolism of arachidonic acid (AA) in short-term tissue cultures of placenta, fetal membranes, decidua and myometrium. Tissues were obtained from caesarean sections before the onset of labour after uncomplicated pregnancies. The released metabolites were analysed by high performance liquid chromatography (HPLC) and specific immunoassays. In radiotracer experiments tissues were labelled with [ 3H]-AA and metabolites released after incubation with calcium ionophore A 23187 were profiled by HPLC. Decidua was more active in metabolizing AA (turnover 34 per cent) than myometrium (28 per cent), placenta (21 per cent) and fetal membranes (17 per cent). Main product in placenta, decidua and myometrium was 12-hydroxyeicosatetraeinoic (12-HETE) (decidua: 19 per cent of released radioactivity, myometrium 14 per cent, placenta 7 per cent). Fetal membranes formed 5-HETE as main product. Another major metabolite in placenta, fetal membranes and decidua was characterized by HPLC as 5(6)-epoxyeicosatrienoic acid. Only myometrium released appreciable amounts of prostaglandins in form of 6-keto-prostaglandin F 1α. In non-radioactive experiments formation of eicosanoids from endogenous AA was investigated by HPLC (fluorescence- and UV-detection) and immunoassays. These experiments confirmed the high production of 12-HETE and the low formation of prostaglandins. Our results suggest that the biological role of AA-metabolites, other than prostaglandins, have as yet been underestimated.

Omega‐3 fatty acids suppress the enhanced production of 5‐Iipoxygenase products from polymorph neutrophil granulocytes in cystic fibrosis

Pulmonary damage in cystic br osis (CF) is associated with chronic inflammation mediated in part by proinflammatory 5-lipoxygenase products (5-LOP, leukotrienes and 5-hydroxyeicosatetraenoic acid) from polymorph neutrophil granulocytes (PMN). The authors studied 5-LOP formation of PMN from CF patients and in vitro effects of added eicosapentaenoic acid (EPA) and fish oil. Circulating PMN were isolated from 10 CF patients without acute infections and 10 control persons of the same age (4-20 years). Total 5-LOP liberation from PMN of CF patients was significantly increased over controls after incubation with the calcium ionophore A23 (1 mumol L-1) without arachidonic acid (AA) (380 +/- 24 vs. 294 +/- 28 pmol mL-1) and with 10 mumol L-1 AA (1303 +/- 104 vs. 1015 +/- 104 pmol mL-1), and there were nonsignificant trends to high values after incubation with 5 mumol L-1 platelet activating factor (PAF, 134% of controls) and 1 mumol L-1 formyl-methionylleucyl-phenylalanine (FMLP, 125%). The addition of 100 micrograms mL-1 fish oil to PMN of CF patients challenged with A23 completely suppressed synthesis of proinflammatory 5-LOP of the 4-series, while inactive 5-LOP metabolites of the 5-series were produced. Added EPA (10 mumol L-1) also suppressed 4-series 5-LOP and significantly reduced leukotriene B4 concentration by 48% from 39.9 +/- 3.2 to 20.6 +/- 11.4 pmol L-1, again with a concomittant increase of inactive 5-series metabolites. The authors conclude that the turnover of endogenous and exogenous AA is enhanced in CF, possibly due to stimulated phospholipase A2 activity. The relatively small effect of the receptor dependent stimuli PAF and FMLP may be caused by a down-regulation of PMN receptors in CF. Supplementation of long-chain omega-3-fatty acids may be beneficial for reducing excessive inflammation in CF patients and should be further evaluated.

Regulation of myo-inositol transport during the growth and differentiation of thyrocytes: a link with thyroid-stimulating hormone-induced phospholipase A2 activity.

The Vmax of myo-inositol transport increased 3-fold during epidermal growth factor (EGF)-induced growth and thyroid-stimulating hormone. (TSH)-induced differentiation in primary cultures of sheep and human thyrocytes. The Km remained unaltered. This up-regulation required the presence of insulin. The TSH-induced rise in myo-inositol transport commenced 8 to 16 h after the initial stimulus and achieved a plateau at 24 h. In human thyrocytes the change in Vmax was accompanied by an increase in the steady-state levels of mRNA for the myo-inositol transporter following treatment with either ligand. Examination of the metabolites of myo-inositol showed few significant changes after treatment of sheep thyrocytes with EGF for 24 h. This is consistent with maintenance of the intracellular concentration of myo-inositol as the cells enlarge in preparation for cell division. In TSH-treated cells, however, up-regulation of myo-inositol transport was linked with increased myo-inositol cycling across the cell membrane, increased phospholipase A2-mediated turnover of phosphatidylinositol and a concomitant increase in arachidonic acid turnover. Increased levels of myo-inositol phosphates were also noted 24 h after TSH treatment. These results indicate the initiation of secondary signalling events many hours after the primary stimulus.

Lipid alterations following impact spinal cord injury in the rat

A computer-controlled impactor was used to produce a severe spinal cord injury in the rat thoracic spinal cord. Cords were rapidly frozen in situ at 5, 15, 30, and 60 min and 6, 12, and 24 h postinjury. Control cords were noninjured cords from animals having undergone a laminectomy and allowed to recover for 90 min postlaminectomy. The cords were assayed for alterations in lipid metabolism. Specifically, there were rapid increases in prostaglandin F2 alpha and thromboxane, with a peak increase in thromboxane levels at 30 min. Prostaglandin F2 alpha levels peaked at 15 min with levels remaining nearly constant for 12 h. There were no detectable changes in phospholipid levels, although diacylglycerol levels and free fatty acid levels were increased. Total free fatty acids were increased at 12 and 24 h postinjury by 2.3- and 3.2-fold over control levels, respectively. Arachidonic acid levels were not significantly elevated at early time points, however, these early time points correspond to elevated eicosanoid synthesis and this may account for the lack of early detectable increases in arachidonic acid. After 6 h postinjury, arachidonic acid levels were 20-fold greater than control levels and remained elevated at 24 h. There were minimal decreases in cholesterol and no decrease in either choline or ethanolamine plasmalogen levels. These results suggest a rapid turnover of arachidonic acid following spinal cord injury with a concomitant increase in vasoconstrictive eicosanoid synthesis. The lack of changes in major membrane constituents suggests the mechanisms may not involve general membrane degradation, but an over-stimulation of phospholipase A2-linked membrane receptors.

Diabetic embryopathy.

Infants of diabetic mothers have three to four times the incidence of congenital malformations than that in the general population. These anomalies include defects of the neural tube, heart, urogenital system, skeleton and alimentary tract, and the caudal regression syndrome. The anomalies are considered to result from the maternal metabolic derangements. The mechanism has been suggested to involve a diminished turnover of phosphoinositide or arachidonic acid, or an excess of free oxygen radicals. As most of the anomalies in infants of diabetic mothers occur in the first few weeks of pregnancy, strict glycemic control beginning before conception appears mandated.

Serotonin, a potent modulator of arachidonic acid turnover, interaction with glutamatergic receptor in brain cortex

Brain cortex synaptoneurosomes actively incorporated [14C]arachidonic acid (AA) into lipids. Serotonin (5-HT), at a concentration range of 10 μM-1 mM, significantly stimulates the incorporation of AA mainly into phosphatidylinositol (PI) of brain cortex synaptoneurosomes. The stimulation rate of AA incorporation by 5-HT was the same in the presence and absence of lysophosphatidylinositol (LPI). However, in the absence of LPI some stimulation of AA uptake was also observed into phosphatidylcholine, phosphatidylethanolamine and phosphatidic acid. Buspirone, an agonist of 5-HT1A receptor, has a similar effect on AA incorporation into membrane lipids as serotonin itself. Moreover, ketanserin, an antagonist of 5-HT2 receptor, also induces activation of AA incorporation into membrane lipids. On the other hand, glutamate, in a concentration dependent manner, significantly inhibits AA uptake into PI and also has some inhibitory action on AA uptake into the other lipids. Serotonin itself and the agonist of 5-HT1A receptor through the activation of AA turnover counteract glutamate-induced inhibition of AA uptake into lipids of brain cortex. Our results indicated that serotonin directly, through the specific receptors, or indirectly, through the interaction with glutamatergic receptors, modulates turnover and the level of arachidonic acid in the brain.

Photodynamic drug action on isolated rat pancreatic acini: Mobilization of arachidonic acid and prostaglandin production

Chloro-aluminium phthalocyanine sulphonate (SALPC) when photon-activated generates singlet oxygen, elicits amylase release and causes plasma membrane permeabilization of pancreatic acinar cells (Matthews and Cui, Biochem Pharmacol 39: 1444–1457, 1990). Amylase release precedes membrane permeabilization suggesting that the initial release of amylase may be due to direct stimulation by singlet oxygen of secretagogue receptors or their coupled guanine nucleotide binding proteins (G-proteins) and effector systems including phospholipase A 2 (PLA 2). The aim of the experiments reported here was to establish the extent to which PLA 2 activation, arachidonic acid mobilization, and prostaglandin production are involved in the photon-induced action of SALPC on dispersed, perifused acini isolated from the rat pancreas. The mobilization of arachidonic acid by a major secretory stimulant of pancreatic exocrine cells, cholecystokinin octapeptide, was also assessed: it produced a time- and concentration-dependent (10 −10–10 −6 M) stimulation of arachidonic acid output from acini prelabelled with [1- 14C]arachidonic acid. In contrast, the kinetics of arachidonic acid mobilization with photon-activated SALPC 1μM, 4500 or 18,400 lux light intensity (λ>570 mm), was biphasic, an intensity-dependent stimulation being preceded by a more immediate initial inhibition of output. Light activation of SALPC and singlet oxygen generation may evoke the stimulatory phase of aarachidonic acid release by an action on G-proteins, or by PLA 2 activated directly, or via calcium influx, because NaF 20 mM, mellitin 2 mg/mL and the calcium ionophore A23187 1 μM caused a 2.9-, 33- and 5-fold increase, respectively, in arachidonic acid output. However, not only was the arachidonate stimulation delayed in response to SALPC but in other experiments designed to gain more insight into the turnover of arachidonic acid and its metabolites, the photodynamic release of amylase preceded maximum prostaglandin E 2 (PGE 2) output and amylase release was completely unaffected when PGE 2 production was blocked by the cyclo-oxygenase inhibitor, indomethacin 10 μM. It is therefore likely that the rapid initial photodynamic release of amylase from pancreatic acini induced by SALPC is mediated by activation of the signal transduction pathway involving the release of intracellulat calcium; arachidonic acid mobilization and prostanoid production may then linked to the longer-term, cytolytic action of SALPC, especially in tumour cells.

Increase of bradykinin-stimulated arachidonic acid release in a ΔF508 cystic fibrosis epithelial cell line

Modification of chloride conductanse by bradykinin in epithelial cells has been attributed to an activation of protein kinase A resulting from adenylcyclase stimulation by arachidonic acid cyclooxygenase products. The results presented here compare tracheal epithelial cell lines from one control and two cystic fibrosis patients which were immortalized by transfection with the SV40 large T oncogene. The three cell lines presented the same arachidonic acid content, turnover and mobilisation under basal conditions. Bradykinin stimulated the release of arachidonic acid and the synthesis of cyclooxygenase derivatives (mainly PGE 2). The cell line from the cystic fibrosis patient bearing a phenylalanine 508 deletion, which is the major form of the disease, showed a higher bradkinin-induced arachidonic acid release than either control cells or cells from a patient presenting a minor form of the disease. This higher sensitivity suggests a dysregulation of phospholipase A 2 stimulation in cystic fibrosis cells and was confirmed on non-immortalized tracheal epithelial cells in primary culture and on skin fibroblasts from patients bearing the same mutation. This defect is associated with a potentiation of cholera toxin pretreatment on cAMP content of ΔF508 cell line. The impaired control of arachidonic acid release cannot be attributed to an increased number of bradykinin binding sites, since this increase was similar in the two cystic fibrosis cell lines.

Arachidonic acid turnover and phospholipase A2 activity in neuronal growth cones.

We analyzed de novo synthesis and local turnover of phospholipids in the growing neuron and the isolated nerve growth cone. The metabolism of phosphatidylinositol (PI) was studied with regard to the incorporation of saturated and unsaturated fatty acids and inositol. A comparison of de novo phospholipid synthesis in the intact neuron (whole brain, cell cultures) versus local turnover in isolated growth cone particles (GCPs) from fetal rat brain revealed different incorporation patterns and, in particular, high arachidonic acid (AA) turnover in PI of GCPs. These observations, together with elevated levels of free AA (2.5% of total AA content) in GCPs, demonstrate the predominance of acylation/deacylation in the sn-2 position of PI. GCP phospholipase A2 (PLA2) activity was demonstrated using [3H]-or [14C]AA-phosphatidylcholine (PC) or -PI as the substrate in vitro and GCPs or a cytosolic GCP extract as the source of enzyme. In contrast to PC, which is hydrolyzed very slowly, PI is a very good GCP PLA2 substrate. PLA2 activity is much higher in GCPs than that of phospholipase C, as demonstrated by the comparison of AA and inositol turnover, by the low levels of 1,2-diacylglycerol generated by GCPs, and by the resistance of AA release to treatment of GCPs with RHC-80267, a specific inhibitor of diacylglycerol lipase. The predominance of PLA2 activity in GCPs raises questions regarding its regulation and the functional roles of PI metabolites, especially lysocompounds, in growth cones.

Changes in Lipid Composition and Arachidonic Acid Turnover during the Life Cycle of the Yeast Dipodascopsis Uninucleata

Ungerminated ascospores of Dipodascopsis uninucleata contained 18 times more lipid (5.5 % dry wt) than germinated cells; the lipid comprised 58 % (w/w) glycolipids, 28 % (w/w) neutral lipids (mainly triacylglycerols)and 14 % (w/w) phospholipids (mainly phosphatidylcholine and phosphatidylethanolamine). During germination the absolute amounts of all three lipid fractions fell sharply but, during the subsequent initiation of hyphal growth,the amount of phospholipids increased. As these hyphae began to differentiate for the sexual stage of the life cycle,the amount of neutral lipid then increased. The fatty acyl groups of the glyco-, neutral and phospholipid fractions throughout the life cycle were mainly palmitate (16:0), oleate (18: 1) and linoleate (18:2). The percentage of 16:O remained constant during the life cycle while the relative amounts of 18:2 plus alpha-linolenate (18:3) in the glyco-,neutral and phospholipid fractions first increased during initiation of growth and then decreased during the onset of differentiation. The opposite trend occurred with 18: 1. When [(3)H]arachidonic acid (ARA) and [1-(14)C]181: were fed separately to D. uninucleata, both were rapidly incorporated into phospholipids. Highest incorporation of ARA was in the growth phase; during the onset and remainder of the differentiation phase, the amount of ARA decreased in this fraction. Incorporation of 18: 1 increased during growth and differentiation, with a significant proportion(49% to 57%) being incorporated into triacylglycerols compared to a much smaller proportion (12% to 17%) of ARA.

Metipranolol-induced changes in membrane phospholipids of isolated mast cells

Metipranolol significantly decreased32P incorporation into membrane phosphatidylcholine but increased the radioactivity in phosphatidylinositol of nonstimulated and 48/80-stimulated rat mast cells. In stimulated cells metipranolol significantly decreased3H-arachidonic acid liberation from total membrane phospholipids as well as thromboxane B2 formation. The effect of metipranolol on stimulated rat mast cells is most probably a result of its direct effect on membrane phospholipid and arachidonic acid turnover.