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Polyunsaturated fatty acyl (PUFA) chains in both the ω3 and ω6 series are essential for normal animal brain development, and cannot be interconverted to compensate for a dietary deficiency of one or the other. Paradoxically, a dietary ω3-PUFA deficiency leads to the accumulation of docosapentaenoate (DPA, 22:5ω6), an ω6-PUFA chain that is normally scarce in the brain. We applied a high-precision LC/MS method to characterize the distribution of DPA chains across phospholipid headgroup classes, the fatty acyl chains with which they were paired, and the extent to which they were oxidatively damaged in the cortical brain of rats on an ω3-deficient diet. Results indicate that dietary ω3-PUFA deficiency markedly increased the concentrations of phospholipids with DPA chains across all headgroup subclasses, including plasmalogen species. The concentrations of phospholipids containing docosahexaenoate chains (22:6ω3) decreased 20-25%, while the concentrations of phospholipids containing arachidonate chains (20:4ω6) did not change significantly. Although DPA chains are more saturated than DHA chains, a larger fraction of DPA chains were monohydroxylated, particularly among diacyl-phosphatidylethanolamines and plasmalogen phosphatidylethanolamines, suggesting that they were disproportionately subjected to oxidative stress. Differences in the pathological significance of ω3 and ω6 oxidation products suggest that greater oxidative damage among the ω6 PUFAs that increase in response to dietary ω3 deficiency may have pathological significance in Alzheimer's disease.

Phosphoinositides, lipid-signaling molecules, participate in diverse brain processes within a wide metabolic cascade.Gene transcriptional networks coordinately regulate the phosphoinositide cascade during human brain Development and Aging.We used the public BrainCloud database for human dorsolateral prefrontal cortex to examine age-related expression levels of 49 phosphoinositide metabolic genes during Development (0 to 20+ years) and Aging (21+ years).We identified three groups of partially overlapping genes in each of the two intervals, with similar intergroup correlations despite marked phenotypic differences between Aging and Development. In each interval, ITPKB, PLCD1, PIK3R3, ISYNA1, IMPA2, INPPL1, PI4KB, and AKT1 are in Group 1, PIK3CB, PTEN, PIK3CA, and IMPA1 in Group 2, and SACM1L, PI3KR4, INPP5A, SYNJ1, and PLCB1 in Group 3. Ten of the genes change expression nonlinearly during Development, suggesting involvement in rapidly changing neuronal, glial and myelination events. Correlated transcription for some gene pairs likely is facilitated by colocalization on the same chromosome band.Stable coordinated gene transcriptional networks regulate brain phosphoinositide metabolic pathways during human Development and Aging.

Down syndrome is determined by the presence of an extra copy of autosome 21 and is expressed by multiple abnormalities, with mental retardation being the most striking feature. The condition results in altered electrical membrane properties of fetal dorsal root ganglia (DRG) neurons, as in the trisomy 16 fetal mouse, an animal model of the human condition. Cultured trisomic DRG neurons from human and mouse fetuses present faster rates of depolarization and repolarization in the action potential compared to normal controls and a shorter spike duration. Also, trisomy 16 brain and spinal cord tissue exhibit reduced acetylcholine secretion. Therefore, we decided to study Ca2+ currents in cultured DRG neurons from trisomy 16 and age-matched control mice, using the whole-cell patch-clamp technique. Trisomic neurons exhibited a 62% reduction in Ca2+ current amplitude and reduced voltage dependence of current activation at -30 and -20 mV levels. Also, trisomic neurons showed slower activation kinetics for Ca2+ currents, with up to 80% increase in time constant values. Kinetics of the inactivation phase were similar in both conditions. The results indicate that murine trisomy 16 alter Ca2+ currents, which may contribute to impaired cell function, including neurotransmitter release. These abnormalities also may alter neural development.

The polyunsaturated arachidonic and docosahexaenoic acids (AA and DHA) participate in cell membrane synthesis during neurodevelopment, neuroplasticity, and neurotransmission throughout life. Each is metabolized via coupled enzymatic reactions within separate but interacting metabolic cascades.AA and DHA pathway genes are coordinately expressed and underlie cascade interactions during human brain development and aging.The BrainCloud database for human non-pathological prefrontal cortex gene expression was used to quantify postnatal age changes in mRNA expression of 34 genes involved in AA and DHA metabolism.Expression patterns were split into Development (0 to 20 years) and Aging (21 to 78 years) intervals. Expression of genes for cytosolic phospholipases A2 (cPLA2), cyclooxygenases (COX)-1 and -2, and other AA cascade enzymes, correlated closely with age during Development, less so during Aging. Expression of DHA cascade enzymes was less inter-correlated in each period, but often changed in the opposite direction to expression of AA cascade genes. Except for the PLA2G4A (cPLA2 IVA) and PTGS2 (COX-2) genes at 1q25, highly inter-correlated genes were at distant chromosomal loci.Coordinated age-related gene expression during the brain Development and Aging intervals likely underlies coupled changes in enzymes of the AA and DHA cascades and largely occur through distant transcriptional regulation. Healthy brain aging does not show upregulation of PLA2G4 or PTGS2 expression, which was found in Alzheimer's disease.

BACKGROUND:Age changes in expression of inflammatory, synaptic, and neurotrophic genes are not well characterized during human brain development and senescence. Knowing these changes may elucidate structural, metabolic, and functional brain processes over the lifespan, as well vulnerability to neurodevelopmental or neurodegenerative diseases. HYPOTHESIS:Expression levels of inflammatory, synaptic, and neurotrophic genes in the human brain are coordinated over the lifespan and underlie changes in phenotypic networks or cascades. METHODS:We used a large-scale microarray dataset from human prefrontal cortex, BrainCloud, to quantify age changes over the lifespan, divided into Development (0 to 21 years, 87 brains) and Aging (22 to 78 years, 144 brains) intervals, in transcription levels of 39 genes. RESULTS:Gene expression levels followed different trajectories over the lifespan. Many changes were intercorrelated within three similar groups or clusters of genes during both Development and Aging, despite different roles of the gene products in the two intervals. During Development, changes were related to reported neuronal loss, dendritic growth and pruning, and microglial events; TLR4, IL1R1, NFKB1, MOBP, PLA2G4A, and PTGS2 expression increased in the first years of life, while expression of synaptic genes GAP43 and DBN1 decreased, before reaching plateaus. During Aging, expression was upregulated for potentially pro-inflammatory genes such as NFKB1, TRAF6, TLR4, IL1R1, TSPO, and GFAP, but downregulated for neurotrophic and synaptic integrity genes such as BDNF, NGF, PDGFA, SYN, and DBN1. CONCLUSIONS:Coordinated changes in gene transcription cascades underlie changes in synaptic, neurotrophic, and inflammatory phenotypic networks during brain Development and Aging. Early postnatal expression changes relate to neuronal, glial, and myelin growth and synaptic pruning events, while late Aging is associated with pro-inflammatory and synaptic loss changes. Thus, comparable transcriptional regulatory networks that operate throughout the lifespan underlie different phenotypic processes during Aging compared to Development.

Dietary long-chain n-3 polyunsaturated fatty acid (PUFA) supplementation may be beneficial for chronic brain illnesses, but the issue is not agreed on. We examined effects of dietary n-3 PUFA deprivation or supplementation, compared with an n-3 PUFA adequate diet (containing alpha-linolenic acid [18:3 n-3] but not docosahexaenoic acid [DHA, 22:6n-3]), on brain markers of lipid metabolism and excitotoxicity, in rats treated chronically with NMDA or saline.Male rats after weaning were maintained on one of three diets for 15 weeks. After 12 weeks, each diet group was injected i.p. daily with saline (1 ml/kg) or a subconvulsive dose of NMDA (25 mg/kg) for 3 additional weeks. Then, brain fatty acid concentrations and various markers of excitotoxicity and fatty acid metabolism were measured.Compared to the diet-adequate group, brain DHA concentration was reduced, while n-6 docosapentaenoic acid (DPA, 22:5n-6) concentration was increased in the n-3 deficient group; arachidonic acid (AA, 20:4n-6) concentration was unchanged. These concentrations were unaffected by fish oil supplementation. Chronic NMDA increased brain cPLA2 activity in each of the three groups, but n-3 PUFA deprivation or fish oil did not change cPLA2 activity or protein compared with the adequate group. sPLA2 expression was unchanged in the three conditions, whereas iPLA2 expression was reduced by deprivation but not changed by supplementation. BDNF protein was reduced by NMDA in N-3 PUFA deficient rats, but protein levels of IL-1β, NGF, and GFAP did not differ between groups.N-3 PUFA deprivation significantly worsened several pathological NMDA-induced changes produced in diet adequate rats, whereas n-3 PUFA supplementation did not affect NMDA induced changes. Supplementation may not be critical for this measured neuropathology once the diet has an adequate n-3 PUFA content.

Chronic alcohol dependence has been associated with disturbed behavior, cerebral atrophy and a low plasma concentration of docosahexaenoic acid (DHA, 22∶6n-3), particularly if liver disease is present. In animal models, excessive alcohol consumption is reported to reduce brain DHA concentration, suggesting disturbed brain DHA metabolism. We hypothesized that brain DHA metabolism also is abnormal in chronic alcoholics.We compared 15 non-smoking chronic alcoholics, studied within 7 days of their last drink, with 22 non-smoking healthy controls. Using published neuroimaging methods with positron emission tomography (PET), we measured regional coefficients (K*) and rates (J(in)) of DHA incorporation from plasma into the brain of each group using [1-(11)C]DHA, and regional cerebral blood flow (rCBF) using [(15)O]water. Data were partial volume error corrected for brain atrophy. Plasma unesterified DHA concentration also was quantified.Mean K* for DHA was significantly and widely elevated by 10-20%, and rCBF was elevated by 7%-34%, in alcoholics compared with controls. Unesterified plasma DHA did not differ significantly between groups nor did whole brain J(in), the product of K* and unesterified plasma DHA concentration.Significantly higher values of K* for DHA in alcoholics indicate increased brain avidity for DHA, thus a brain DHA metabolic deficit vis-à-vis plasma DHA availability. Higher rCBF in alcoholics suggests increased energy consumption. These changes may reflect a hypermetabolic state related to early alcohol withdrawal, or a general brain metabolic change in chronic alcoholics.

Abstract Background Apolipoprotein E4 (APOE4) is associated with a greater response to neuroinflammation and the risk of developing late-onset Alzheimer’s disease (AD), but the mechanisms for this association are not clear. The activation of calcium-dependent cytosolic phospholipase A2 (cPLA2) is involved in inflammatory signaling and is elevated within the plaques of AD brains. The relation between APOE4 genotype and cPLA2 activity is not known. Methods Mouse primary astrocytes, mouse and human brain samples differing by APOE genotypes were collected for measuring cPLA2 expression, phosphorylation, and activity in relation to measures of inflammation and oxidative stress. Results Greater cPLA2 phosphorylation, cPLA2 activity and leukotriene B4 (LTB4) levels were identified in ApoE4 compared to ApoE3 in primary astrocytes, brains of ApoE-targeted replacement (ApoE-TR) mice, and in human brain homogenates from the inferior frontal cortex of patients with AD carrying APOE3/E4 compared to APOE3/E3. Greater cPLA2 phosphorylation was also observed in human postmortem frontal cortical synaptosomes and primary astrocytes after treatment with recombinant ApoE4 ex vivo. In ApoE4 astrocytes, the greater levels of LTB4, reactive oxygen species (ROS), and inducible nitric oxide synthase (iNOS) were reduced after cPLA2 inhibition. Conclusions Our findings implicate greater activation of cPLA2 signaling system with APOE4, which could represent a potential drug target for mitigating the increased neuroinflammation with APOE4 and AD.

This article has been retracted. Please see the Retraction Notice for more detail: https://doi.org/10.1186/s13024-021-00438-3.

Ca2+-independent phospholipase A2β (iPLA2β) selectively hydrolyzes docosahexaenoic acid (DHA, 22:6n-3) in vitro from phospholipid. Mutations in the PLA2G6 gene encoding this enzyme occur in patients with idiopathic neurodegeneration plus brain iron accumulation and dystonia-parkinsonism without iron accumulation, whereas mice lacking PLA2G6 show neurological dysfunction and neuropathology after 13 months. We hypothesized that brain DHA metabolism and signaling would be reduced in 4-month-old iPLA2β-deficient mice without overt neuropathology. Saline or the cholinergic muscarinic M1,3,5 receptor agonist arecoline (30 mg/kg) was administered to unanesthetized iPLA2β−/−, iPLA2β+/−, and iPLA2β+/+ mice, and [1-14C]DHA was infused intravenously. DHA incorporation coefficients k∗ and rates Jin, representing DHA metabolism, were determined using quantitative autoradiography in 81 brain regions. iPLA2β−/− or iPLA2β+/− compared with iPLA2β+/+ mice showed widespread and significant baseline reductions in k∗ and Jin for DHA. Arecoline increased both parameters in brain regions of iPLA2β+/+ mice but quantitatively less so in iPLA2β−/− and iPLA2β+/− mice. Consistent with iPLA2β’s reported ability to selectively hydrolyze DHA from phospholipid in vitro, iPLA2β deficiency reduces brain DHA metabolism and signaling in vivo at baseline and following M1,3,5 receptor activation. Positron emission tomography might be used to image disturbed brain DHA metabolism in patients with PLA2G6 mutations.

Isotope feeding studies report a wide range of conversion fractions of dietary shorter-chain polyunsaturated fatty acids (PUFAs) to long-chain PUFAs, which limits assessing nutritional requirements and organ effects of arachidonic (AA, 20:4n-6) and docosahexaenoic (DHA, 22:6n-3) acids. In this study, whole-body (largely liver) steady-state conversion coefficients and rates of circulating unesterified linoleic acid (LA, 18:2n-6) to esterified AA and other elongated n-6 PUFAs were quantified directly using operational equations, in unanesthetized adult rats on a high-DHA but AA-free diet, using 2 h of intravenous [U-13C]LA infusion. Unesterified LA was converted to esterified LA in plasma at a greater rate than to esterified γ-linolenic (γ-LNA, 18:3n-6), eicosatrienoic acid (ETA, 20:3n-6), or AA. The steady-state whole-body synthesis-secretion (conversion) coefficient to AA equaled 5.4 × 10−3 min−1, while the conversion rate (coefficient × concentration) equaled 16.1 μmol/day. This rate exceeds the reported brain AA consumption rate by 27-fold. As brain and heart cannot synthesize significant AA from circulating LA, liver synthesis is necessary to maintain their homeostatic AA concentrations in the absence of dietary AA. The heavy-isotope intravenous infusion method could be used to quantify steady-state liver synthesis-secretion of AA from LA under different conditions in rodents and in humans.

In vitro studies show that docosahexaenoic acid (DHA) can be released from membrane phospholipid by Ca2+-independent phospholipase A2 (iPLA2), Ca2+-independent plasmalogen PLA2 or secretory PLA2 (sPLA2), but not by Ca2+-dependent cytosolic PLA2 (cPLA2), which selectively releases arachidonic acid (AA). Since glutamatergic NMDA (N-methyl-D-aspartate) receptor activation allows extracellular Ca2+ into cells, we hypothesized that brain DHA signaling would not be altered in rats given NMDA, to the extent that in vivo signaling was mediated by Ca2+-independent mechanisms. Isotonic saline, a subconvulsive dose of NMDA (25 mg/kg), MK-801, or MK-801 followed by NMDA was administered i.p. to unanesthetized rats. Radiolabeled DHA or AA was infused intravenously and their brain incorporation coefficients k∗, measures of signaling, were imaged with quantitative autoradiography. NMDA or MK-801 compared with saline did not alter k∗ for DHA in any of 81 brain regions examined, whereas NMDA produced widespread and significant increments in k∗ for AA. In conclusion, in vivo brain DHA but not AA signaling via NMDA receptors is independent of extracellular Ca2+ and of cPLA2. DHA signaling may be mediated by iPLA2, plasmalogen PLA2, or other enzymes insensitive to low concentrations of Ca2+. Greater AA than DHA release during glutamate-induced excitotoxicity could cause brain cell damage.

Neuroinflammation, caused by 6 days of intracerebroventricular infusion of a low dose of lipopolysaccharide (LPS; 0.5 ng/h), stimulates brain arachidonic acid (AA) metabolism in rats, but 6 weeks of lithium pretreatment reduces this effect. To further understand this action of lithium, we measured concentrations of eicosanoids and docosanoids generated from AA and docosahexaenoic acid (DHA), respectively, in high-energy microwaved rat brain using LC/MS/MS and two doses of LPS. In rats fed a lithium-free diet, low (0.5 ng/h)- or high (250 ng/h)-dose LPS compared with artificial cerebrospinal fluid increased brain unesterified AA and prostaglandin E2 concentrations and activities of AA-selective Ca2+-dependent cytosolic phospholipase A2 (cPLA2)-IV and Ca2+-dependent secretory sPLA2. LiCl feeding prevented these increments. Lithium had a significant main effect by increasing brain concentrations of lipoxygenase-derived AA metabolites, 5- hydroxyeicosatetraenoic acid (HETE), 5-oxo-eicosatetranoic acid, and 17-hydroxy-DHA by 1.8-, 4.3- and 1.9-fold compared with control diet. Lithium also increased 15-HETE in high-dose LPS-infused rats. Ca2+-independent iPLA2-VI activity and unesterified DHA and docosapentaenoic acid (22:5n-3) concentrations were unaffected by LPS or lithium. This study demonstrates, for the first time, that lithium can increase brain 17-hydroxy-DHA formation, indicating a new and potentially important therapeutic action of lithium.

Bipolar disorder (BD) is a progressive psychiatric disorder characterized by recurrent changes of mood and is associated with cognitive decline. There is evidence of excitotoxicity, neuroinflammation, upregulated arachidonic acid (AA) cascade signaling and brain atrophy in BD patients. These observations suggest that BD pathology may be associated with apoptosis as well as with disturbed synaptic function. To test this hypothesis, we measured mRNA and protein levels of the pro-apoptotic (Bax, BAD, caspase-9 and caspase-3) and anti-apoptotic factors (BDNF and Bcl-2) and of pre- and post-synaptic markers (synaptophysin and drebrin), in postmortem prefrontal cortex (Brodmann area 9) from 10 BD patients and 10 age-matched controls. Consistent with the hypothesis, BD brains showed significant increases in protein and mRNA levels of the pro-apoptotic factors and significant decreases of levels of the anti-apoptotic factors and the synaptic markers, synaptophysin and drebrin. These differences may contribute to brain atrophy and progressive cognitive changes in BD.

Dietary docosahexaenoic acid (DHA; 22:6n-3) and eicosapentaenoic acid (EPA; 20:5n-3) are considered important for maintaining normal heart and brain function, but little EPA is found in brain, and EPA cannot be elongated to DHA in rat heart due to the absence of elongase-2. Ingested EPA may have to be converted in the liver to DHA for it to be fully effective in brain and heart, but the rate of conversion is not agreed on. This rate was determined in male adult rats fed a standard n-3 PUFA, containing diet by infusing unesterified albumin-bound [U-13C]EPA intravenously for 2 h and measuring esterified [13C]labeled PUFAs in arterial plasma lipoproteins, as well as the specific activity of unesterified plasma EPA. Whole-body (presumably hepatic) synthesis secretion rates from circulating unesterified EPA, calculated from peak first derivatives of plasma esterified concentration × volume curves, equaled 2.61 μmol/day for docosapentaenoic acid (22:5n-3) and 5.46 μmol/day for DHA. The DHA synthesis rate was 24-fold greater than the reported brain DHA consumption rate in rats. Thus, dietary EPA could help to maintain brain and heart DHA homeostasis because it is converted at a relatively high rate in the liver to circulating DHA.

Docosahexaenoic acid (DHA; 22:6n-3) is a critical constituent of the brain, but its metabolism has not been measured in the human brain in vivo. In monkeys, using positron emission tomography (PET), we first showed that intravenously injected [1-11C]DHA mostly entered nonbrain organs, with ∼0.5% entering the brain. Then, using PET and intravenous [1-11C]DHA in 14 healthy adult humans, we quantitatively imaged regional rates of incorporation (K*) of DHA. We also imaged regional cerebral blood flow (rCBF) using PET and intravenous [15O]water. Values of K* for DHA were higher in gray than white matter regions and correlated significantly with values of rCBF in 12 of 14 subjects despite evidence that rCBF does not directly influence K*. For the entire human brain, the net DHA incorporation rate Jin, the product of K*, and the unesterified plasma DHA concentration equaled 3.8 ± 1.7 mg/day. This net rate is equivalent to the net rate of DHA consumption by brain and, considering the reported amount of DHA in brain, indicates that the half-life of DHA in the human brain approximates 2.5 years. Thus, PET with [1-11C]DHA can be used to quantify regional and global human brain DHA metabolism in relation to health and disease.

Docosahexaenoic acid (DHA) and eicosapentaenoic acid (EPA), long-chain n-3 PUFAs important for brain and heart function, can be obtained from dietary fish products or by liver synthesis from α-linolenic acid (α-LNA). Their daily human dietary requirements are not clear, and their liver synthesis rates in humans and nonhumans are unknown. We estimated whole-body (presumably liver) synthesis rates in unanesthetized rats by infusing [U-13C]α-LNA intravenously for 2 h and measuring labeled and unlabeled n-3 PUFA in arterial plasma using negative chemical ionization GC-MS. Newly synthesized esterified [13C]DHA, [13C]EPA, and [13C]docosapentaenoic acid (DPA) appeared in arterial plasma after 60 min of infusion, then their concentrations rose in an S-shaped manner. Esterified concentration × plasma volume data were fit with a sigmoidal equation, whose peak first derivatives provided synthesis rates of unlabeled EPA, DPA, and DHA equal to 8.40, 6.27, and 9.84 μmol/day, respectively. The DHA synthesis rate exceeded the published daily rat brain DHA consumption rate by 30-fold, suggesting that liver synthesis from α-LNA could maintain brain DHA homeostasis were DHA absent from the diet. This stable isotope infusion method could be used to quantify whole-body DHA synthesis rates in human subjects.

Inflammatory lipid mediators derived from arachidonic acid (AA) and docosahexaenoic acid (DHA) modify the pathophysiology of brain ischemia. The goal of this work was to investigate the formation of eicosanoids and docosanoids generated from AA and DHA, respectively, during no-flow cerebral ischemia. Rats were subjected to head-focused microwave irradiation 5 min following decapitation (complete ischemia) or prior to decapitation (controls). Brain lipids were extracted and analyzed by reverse-phase liquid chromatography-tandem mass spectrometry. After complete ischemia, brain AA, DHA, and docosapentaenoic acid concentrations increased 18-, 5- and 4-fold compared with controls, respectively. Prostaglandin E2 (PGE2) and PGD2 could not be detected in control microwaved rat brain, suggesting little endogenous PGE2/D2 production in the brain in the absence of experimental manipulation. Concentrations of thromboxane B2, E2/D2-isoprostanes, 5-hydroxyeicosatetraenoic acid (5-HETE), 5-oxo-eicosatetraenoic acid, and 12-HETE were significantly elevated in ischemic brains. In addition, DHA products such as mono-, di- and trihydroxy-DHA were detected in control and ischemic brains. Monohydroxy-DHA, identified as 17-hydroxy-DHA and thought to be the immediate precursor of neuroprotectin D1, was 6.5-fold higher in ischemic than in control brain. The present study demonstrated increased formation of eicosanoids, E2/D2-IsoPs, and docosanoids following cerebral ischemia. A balance of these lipid mediators may mediate immediate events of ischemic injury and recovery.

The extent to which the heart can convert α-linolenic acid (α-LNA, 18:3n-3) to longer chain n-3 PUFAs is not known. Conversion rates can be measured in vivo using radiolabeled α-LNA and a kinetic fatty acid model. [1-14C]α-LNA was infused intravenously for 5 min in unanesthetized rats that had been fed an n-3 PUFA-adequate [4.6% α-LNA, no docosahexaenoic acid (DHA, 22:6n-3)] or n-3 PUFA-deficient diet (0.2% α-LNA, nor DHA) for 15 weeks after weaning. Arterial plasma was sampled, as was the heart after high-energy microwaving. Rates of conversion of α-LNA to longer chain n-3 PUFAs were low, and DHA was not synthesized at all in the heart. Most α-LNA within the heart had been β-oxidized. In deprived compared with adequate rats, DHA concentrations in plasma and heart were both reduced by >90%, whereas heart and plasma levels of docosapentaenoic acid (DPAn-6, 22:5n-6) were elevated. Dietary deprivation did not affect cardiac mRNA levels of elongase-5 or desaturases Δ6 and Δ5, but elongase-2 mRNA could not be detected. In summary, the rat heart does not synthesize DHA from α-LNA, owing to the absence of elongase-2, but must obtain its DHA entirely from plasma. Dietary n-3 PUFA deprivation markedly reduces heart DHA and increases heart DPAn-6, which may make the heart vulnerable to different insults.

Whereas antibipolar drug administration to rats reduces brain arachidonic acid turnover, excessive N-methyl-d-aspartate (NMDA) signaling is thought to contribute to bipolar disorder symptoms and may increase arachidonic acid turnover in rat brain phospholipids. To determine whether chronic NMDA would increase brain arachidonic acid turnover, rats were daily administered NMDA (25 mg/kg, ip) or vehicle for 21 days. In unanesthetized rats, on day 21, [1-14C]arachidonic acid was infused intravenously and arterial blood plasma was sampled until the animal was euthanized at 5 min and its microwaved brain was subjected to chemical and radiotracer analysis. Using equations from our in vivo fatty acid model, we found that compared with controls, chronic NMDA increased the net rate of incorporation of plasma unesterified arachidonic acid into brain phospholipids (25–34%) as well as the turnover of arachidonic acid within brain phospholipids (35–58%). These changes were absent at 3 h after a single NMDA injection. The changes, opposite to those after chronic administration of antimanic drugs to rats, suggest that excessive NMDA signaling via arachidonic acid may be a model of upregulated arachidonic acid turnover in brain phospholipids.

Fifteen weeks of dietary n-3 PUFA deprivation increases coefficients of conversion of circulating α-linolenic acid (α-LNA; 18:3n-3) to docosahexaenoic acid (DHA; 22:6n-3) in rat liver but not brain. To determine whether these increases reflect organ differences in enzymatic activities, we examined brain and liver expression of converting enzymes and of two of their transcription factors, peroxisome proliferator-activated receptor α (PPARα) and sterol-regulatory element binding protein-1 (SREBP-1), in rats fed an n-3 PUFA “adequate” (4.6% α-LNA of total fatty acid, no DHA) or “deficient” (0.2% α-LNA, no DHA) diet for 15 weeks after weaning. In rats fed the deficient compared with the adequate diet, mRNA and activity levels of Δ5 and Δ6 desaturases and elongases 2 and 5 were upregulated in liver but not brain, but liver PPARα and SREBP-1 mRNA levels were unchanged. In rats fed the adequate diet, enzyme activities generally were higher in liver than brain. Thus, differences in conversion enzyme expression explain why the liver has a greater capacity to synthesize DHA from circulating α-LNA than does the brain in animals on an adequate n-3 PUFA diet and why liver synthesis capacity is increased by dietary deprivation. These data suggest that liver n-3 PUFA metabolism determines DHA availability to the brain when DHA is absent from the diet.

Rates of conversion of α-linolenic acid (α-LNA, 18:3n-3) to docosahexaenoic acid (DHA, 22:6n-3) by the mammalian brain and the brain's ability to upregulate these rates during dietary deprivation of n-3 polyunsaturated fatty acids (PUFAs) are unknown. To answer these questions, we measured conversion coefficients and rates in post-weaning rats fed an n-3 PUFA deficient (0.2% α-LNA of total fatty acids, no DHA) or adequate (4.6% α-LNA, no DHA) diet for 15 weeks. Unanesthetized rats in each group were infused intravenously with [1-14C]α-LNA, and their arterial plasma and microwaved brains collected at 5 minutes were analyzed. The deficient compared with adequate diet reduced brain DHA by 37% and increased brain arachidonic (20:4n-6) and docosapentaenoic (22:5n-6) acids. Only 1% of plasma [1-14C]α-LNA entering brain was converted to DHA with the adequate diet, and conversion coefficients of α-LNA to DHA were unchanged by the deficient diet. In summary, the brain's ability to synthesize DHA from α-LNA is very low and is not altered by n-3 PUFA deprivation. Because the liver's reported ability is much higher, and can be upregulated by the deficient diet, DHA converted by the liver from circulating αLNA is the source of the brain's DHA when DHA is not in the diet.

Cyclooxygenase (COX)-2 plays an important role in brain arachidonic acid (20:4n-6) metabolism, and its expression is upregulated in animal models of neuroinflammation and excitotoxicity. Our hypothesis was that brain lipid composition would be altered in COX-2 knockout (COX-2−/−) compared with wild-type (COX-2+/+) mice, reflecting the important role of COX-2 in brain lipid metabolism. Concentrations of different lipids were measured in high-energy microwaved brain from COX-2−/− and COX-2+/+ mice. Compared with the COX-2+/+ mouse brain, the brain of the COX-2−/− mouse had a statistically significant 15% increase in phosphatidylserine (PtdSer) and significant 37, 27, and 32% reductions in triacylglycerol and cholesterol concentrations and in the cholesterol-to-phospholipid ratio, respectively. The normalized concentration of palmitic acid (16:0) was increased in PtdSer, as was the brain concentration of unesterified arachidic acid (20:0). A lifetime absence of COX-2 produces multiple changes in brain lipid composition. These changes may be related to reported changes in fatty acid kinetics and in resistance to neuroinflammation and excitotoxicity in the COX-2−/− mouse.

We quantified incorporation rates of plasma-derived α-linolenic acid (α-LNA, 18:3n-3) into “stable” liver lipids and the conversion rate of α-LNA to docosahexaenoic acid (DHA, 22:6n-3) in male rats fed, after weaning, an n-3 PUFA-adequate diet (4.6% α-LNA, no DHA) or an n-3 PUFA-deficient diet (0.2% α-LNA, no DHA) for 15 weeks. Unanesthetized rats were infused intravenously with [1-14C]α-LNA, and arterial plasma was sampled until the liver was microwaved at 5 min. Unlabeled α-LNA and DHA concentrations in arterial plasma and liver were reduced >90% by deprivation, whereas unlabeled arachidonic acid (20:4n-6) and docosapentaenoic acid (22:5n-6) concentrations were increased. Deprivation did not change α-LNA incorporation coefficients into stable liver lipids but increased synthesis-incorporation coefficients of DHA from α-LNA by 6.6-, 8.4-, and 2.3-fold in triacylglycerol, phospholipid, and cholesteryl ester, repectively. Assuming that synthesized-incorporated DHA even tually would be secreted within lipoproteins, calculated liver DHA secretion rates equaled 2.19 and 0.82 μmol/day in the n-3 PUFA-adequate and -deprived rats, respectively. These rates exceed the published rates of brain DHA consumption by 6- and 10-fold, respectively, and should be sufficient to maintain normal and reduced brain DHA concentrations, respectively, in the two dietary conditions.

We quantified the rates of incorporation of α-linolenic acid (α-LNA; 18:3n-3) into “stable” lipids (triacylglycerol, phospholipid, cholesteryl ester) and the rate of conversion of α-LNA to docosahexaenoic acid (DHA; 22: 6n-3) in the liver of awake male rats on a high-DHA-containing diet after a 5-min intravenous infusion of [1-14C]α-LNA. At 5 min, 72.7% of liver radioactivity (excluding unesterified fatty acid radioactivity) was in stable lipids, with the remainder in the aqueous compartment. Using our measured specific activity of liver α-LNA-CoA, in the form of the dilution coefficient λα-LNA-CoA, we calculated incorporation rates of unesterified α-LNA into liver triacylglycerol,phospholipid, and cholesteryl ester as 2,401, 749, and 9.6 nmol/s/g × 10−4, respectively, corresponding to turnover rates of 3.2, 8.7, and 2.9%/min and half-lives of 8–24 min. A lower limit for the DHA synthesis rate from α-LNA equaled 15.8 nmol/s/g × 10−4 (0.5% of the net in corporation rate). Thus, in rats on a high-DHA-containing diet, rates of β-oxidation and esterification of α-LNA into stable liver lipids are high, whereas its conversion to DHA is comparatively low and insufficient to supply significant DHA to the brain. High incorporation and turnover rates likely reflect a high secretion rate by liver of stable lipids within very low density lipoproteins.

Male rat pups at weaning (21 days of age) were subjected to a diet deficient or adequate in n-3 polyunsaturated fatty acids (n-3 PUFAs) for 15 weeks. Performance on tests of locomotor activity, depression, and aggression was measured in that order during the ensuing 3 weeks, after which brain lipid composition was determined. In the n-3 PUFA-deprived rats, compared with n-3 PUFA-adequate rats, docosahexaenoic acid (22:6n-3) in brain phospholipid was reduced by 36% and docosapentaenoic acid (22:5n-6) was elevated by 90%, whereas brain phospholipid concentrations were unchanged. N-3 PUFA-deprived rats had a significantly increased (P = 0.03) score on the Porsolt forced-swim test for depression, and increased blocking time (P = 0.03) and blocking number (P = 0.04) scores (uncorrected for multiple comparisons) on the isolation-induced resident-intruder test for aggression. Large effect sizes (d > 0.8) were found on the depression score and on the blocking time score of the aggression test. Scores on the open-field test for locomotor activity did not differ significantly between groups, and had only small to medium effect sizes. This single-generational n-3 PUFA-deprived rat model, which demonstrated significant changes in brain lipid composition and in test scores for depression and aggression, may be useful for elucidating the contribution of disturbed brain PUFA metabolism to human depression, aggression, and bipolar disorder.

We examined brain phospholipid metabolism in mice in which the cytosolic phospholipase A2 (cPLA2, Type IV, 85 k11111111111111111135) was knocked out (cPLA2−/− mice). Compared with controls, these mice demonstrated altered brain concentrations of several phospholipids, reduced esterified linoleate, arachidonate, and docosahexaenoate in choline glycerophospholipid, and reduced esterified arachidonate in phosphatidylinositol. Unanesthetized cPLA2−/− mice had reduced rates of incorporation of unlabeled arachidonate from plasma and from the brain arachidonoyl-CoA pool into ethanolamine glycerophospholipid and choline glycerophospholipid, but elevated rates into phosphatidylinositol. These differences corresponded to altered turnover and metabolic loss of esterified brain arachidonate. These results suggests that cPLA2 is necessary to maintain normal brain concentrations of phospholipids and of their esterified polyunsaturated fatty acids.Reduced esterified arachidonate and docosahexaenoate may account for the resistance of the cPLA2−/− mouse to middle cerebral artery occlusion, and should influence membrane fluidity, neuroinflammation, signal transduction, and other brain processes.

The rates of synthesis, turnover, and half-lives were determined for brain microsomal ether phospholipids in the awake adult unanesthetized rat. A multicompartmental kinetic model of phospholipid metabolism, based on known pathways of synthesis, was applied to data generated by a 5 min intravenous infusion of [1,1-3H]hexadecanol. At 2 h post-infusion, 29%, 33%, and 31% of the total labeled brain phospholipid was found in the 1-O-alkyl-2-acyl-sn-glycero-3-phosphate, ethanolamine, and choline ether phospholipid fractions, respectively. Autoradiography and membrane fractionation showed that 3% of the net incorporated radiotracer was in myelin at 2 h, compared to 97% in gray matter microsomal and synaptosomal fractions. Based on evidence that ether phospholipid synthesis occurs in the microsomal membrane fraction, we calculated the synthesis rates of plasmanylcholine, plasmanylethanolamine, plasmenylethanolamine, and plasmenylcholine equal to 1.2, 9.3, 27.6, and 21.5 nmol·g−1·min−1, respectively. Therefore, 8% of the total brain ether phospholipids have half-lives of about 36.5, 26.7, 23.1, and 15.1 min, respectively. Furthermore, we clearly demonstrate that there are at least two pools of ether phospholipids in the adult rat brain. One is the static myelin pool with a slow rate of tracer incorporation and the other is a dynamic pool found in gray matter. The short half-lives of microsomal ether phospholipids and the rapid transfer to synaptosomes are consistent with evidence of the marked involvement of these lipids in brain signal transduction and synaptic function. —Rosenberger, T. A., J. Oki, A. D. Purdon, S. I. Rapoport, and E. J. Murphy. Rapid synthesis and turnover of brain microsomal ether phospholipids in the adult rat. J. Lipid Res. 2002. 43: 59–68.

Polyunsaturated fatty acids (PUFAs) are critical to nervous system function and structure, but their rates of incorporation from plasma into brain have not been evaluated. In the adult rat, calculations based on our model show that at least 3–5% of esterified brain arachidonic acid (AA) and 2–8% of esterified brain docosahexaenoic acid (DHA) are replaced daily by unesterified PUFAs in plasma. These rates, when related to unlabeled brain PUFA composition, give half-lives of 1–2 weeks for plasma-brain exchange of AA and DHA. In the human brain, the arachidonate replacement rate is 0.3% per day.Although unesterified plasma PUFA concentrations are low, their rates of incorporation into brain are sufficient to compensate for metabolic and efflux losses, so that PUFA transport from plasma into brain as a component of a lipoprotein is unnecessary. Dietary supplementation, by altering plasma unesterified PUFA concentrations, can regulate brain PUFA content and may help to treat brain diseases involving PUFA imbalance.