Interaction between GPR110 (ADGRF1) and tight junction protein occludin implicated in blood-brain barrier permeability.

Activation of adhesion receptor GPR110 by the endogenous ligand synaptamide promotes neurogenesis, neurite growth, and synaptogenesis in developing brains through cAMP signal transduction. However, interacting partners of GPR110 and their involvement in cellular function remain unclear. Here, we demonstrate using chemical crosslinking, affinity purification, and quantitative mass spectrometry that GPR110 interacts with the tight junction adhesion protein occludin. By removing non-specific partners by comparing the binding proteins of GPR110 WT and an inactive mutant exhibiting impaired surface expression, occludin was distinguished as a true binding partner which was further confirmed by reciprocal co-immunoprecipitation assay. Deletion of GPR110 in mice led to the disruption of blood-brain barrier (BBB) and reduced occludin phosphorylation at Y285 in the brain. The Y285 phosphorylation increased upon the ligand-induced activation of GPR110. These data suggest an important role of GPR110-occludin interaction in BBB function and association of previously unknown GPR110-dependent occludin phosphorylation at Y285 with BBB integrity.

Molecular and Signaling Mechanisms for Docosahexaenoic Acid-Derived Neurodevelopment and Neuroprotection.

The neurodevelopmental and neuroprotective actions of docosahexaenoic acid (DHA) are mediated by mechanisms involving membrane- and metabolite-related signal transduction. A key characteristic in the membrane-mediated action of DHA results from the stimulated synthesis of neuronal phosphatidylserine (PS). The resulting DHA-PS-rich membrane domains facilitate the translocation and activation of kinases such as Raf-1, protein kinase C (PKC), and Akt. The activation of these signaling pathways promotes neuronal development and survival. DHA is also metabolized in neural tissues to bioactive mediators. Neuroprotectin D1, a docosatriene synthesized by the lipoxygenase activity, has an anti-inflammatory property, and elovanoids formed from DHA elongation products exhibit antioxidant effects in the retina. Synaptamide, an endocannabinoid-like lipid mediator synthesized from DHA in the brain, promotes neurogenesis and synaptogenesis and exerts anti-inflammatory effects. It binds to the GAIN domain of the GPR110 (ADGRF1) receptor, triggers the cAMP/protein kinase A (PKA) signaling pathway, and activates the cAMP-response element binding protein (CREB). The DHA status in the brain influences not only the PS-dependent signal transduction but also the metabolite formation and expression of pre- and post-synaptic proteins that are downstream of the CREB and affect neurotransmission. The combined actions of these processes contribute to the neurodevelopmental and neuroprotective effects of DHA.

Biosynthesis of N-Docosahexanoylethanolamine from Unesterified Docosahexaenoic Acid and Docosahexaenoyl-Lysophosphatidylcholine in Neuronal Cells.

We investigated the synthesis of N-docosahexaenoylethanolamine (synaptamide) in neuronal cells from unesterified docosahexaenoic acid (DHA) or DHA-lysophosphatidylcholine (DHA-lysoPC), the two major lipid forms that deliver DHA to the brain, in order to understand the formation of this neurotrophic and neuroprotective metabolite of DHA in the brain. Both substrates were taken up in Neuro2A cells and metabolized to N-docosahexaenoylphosphatidylethanolamine (NDoPE) and synaptamide in a time- and concentration-dependent manner, but unesterified DHA was 1.5 to 2.4 times more effective than DHA-lysoPC at equimolar concentrations. The plasmalogen NDoPE (pNDoPE) amounted more than 80% of NDoPE produced from DHA or DHA-lysoPC, with 16-carbon-pNDoPE being the most abundant species. Inhibition of N-acylphosphatidylethanolamine-phospholipase D (NAPE-PLD) by hexachlorophene or bithionol significantly decreased the synaptamide production, indicating that synaptamide synthesis is mediated at least in part via NDoPE hydrolysis. NDoPE formation occurred much more rapidly than synaptamide production, indicating a precursor-product relationship. Although NDoPE is an intermediate for synaptamide biosynthesis, only about 1% of newly synthesized NDoPE was converted to synaptamide, possibly suggesting additional biological function of NDoPE, particularly for pNDoPE, which is the major form of NDoPE produced.

Emergence of omega-3 fatty acids in biomedical research.

Shortly after the discovery that linoleic acid was an essential fatty acid in 1930, α-linolenic acid also was reported to prevent the fatty acid deficiency syndrome in animals. However, several prominent laboratories could not confirm the findings with α-linolenic acid, and as a result there was a loss of interest in omega-3 fatty acids in lipid research. Even the findings that a prostaglandin can be synthesized from eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) is necessary for optimum retinal function generated only limited interest in omega-3 fatty acids. The breakthrough came in the 1970s when Dyerberg and Bang reported that the low incidence of atherosclerotic coronary disease in Greenland Eskimos was due to the high marine lipid content of their diet. They subsequently found that EPA, which was increased in Eskimo plasma, inhibited platelet aggregation, and they concluded that the low incidence of coronary artery disease was due to the anti-thrombotic effect of EPA. This stimulated widespread interest and research in EPA and DHA, leading to the present view that, like their omega-6 counterparts, omega-3 fatty acids have important physiological functions and are essential fatty acids.

N-Docosahexaenoylethanolamine: A neurotrophic and neuroprotective metabolite of docosahexaenoic acid.

N-Docosahexaenoylethanolamine (synaptamide) is an endocannabinoid-like metabolite endogenously synthesized from docosahexaenoic acid (DHA, 22:6n-3), the major omega-3 polyunsaturated fatty acid present in the brain. Although its biosynthetic mechanism has yet to be established, there is a closely linked relationship between the levels of synaptamide and its precursor DHA in the brain. Synaptamide at nanomolar concentrations promotes neurogenesis, neurite outgrowth and synaptogenesis in developing neurons. Synaptamide also attenuates the lipopolysaccharide-induced neuroinflammatory response and reduces the deleterious effects of ethanol on neurogenic differentiation of neural stem cells (NSCs). These actions are mediated by a specific target receptor of synaptamide GPR110 (ADGRF1), a G-protein coupled receptor that is highly expressed in NSCs and the brain during development. Synaptamide binding to GPR110 induces cAMP production and phosphorylation of protein kinase A (PKA) and the cAMP response element binding protein (CREB). This signaling pathway leads to the expression of neurogenic and synaptogenic genes and suppresses the expression of proinflammatory genes. The GPR110-dependent cellular effects of synaptamide are recapitulated in animal models, suggesting that synaptamide-derived mechanisms may have translational implications. The synaptamide bioactivity transmitted by newly deorphanized GPR110 provides a novel target for neurodevelopmental and neuroprotective control as well as new insight into mechanisms for DHA's beneficial effects on the central nervous system.

Cytochrome P450 epoxygenase pathway of polyunsaturated fatty acid metabolism.

Polyunsaturated fatty acids (PUFA) are oxidized by cytochrome P450 epoxygenases to PUFA epoxides which function as potent lipid mediators. The major metabolic pathways of PUFA epoxides are incorporation into phospholipids and hydrolysis to the corresponding PUFA diols by soluble epoxide hydrolase. Inhibitors of soluble epoxide hydrolase stabilize PUFA epoxides and potentiate their functional effects. The epoxyeicosatrienoic acids (EETs) synthesized from arachidonic acid produce vasodilation, stimulate angiogenesis, have anti-inflammatory actions, and protect the heart against ischemia-reperfusion injury. EETs produce these functional effects by activating receptor-mediated signaling pathways and ion channels. The epoxyeicosatetraenoic acids synthesized from eicosapentaenoic acid and epoxydocosapentaenoic acids synthesized from docosahexaenoic acid are potent inhibitors of cardiac arrhythmias. Epoxydocosapentaenoic acids also inhibit angiogenesis, decrease inflammatory and neuropathic pain, and reduce tumor metastasis. These findings indicate that a number of the beneficial functions of PUFA may be due to their conversion to PUFA epoxides. This article is part of a Special Issue entitled "Oxygenated metabolism of PUFA: analysis and biological relevance".

Discovery of essential fatty acids.

Dietary fat was recognized as a good source of energy and fat-soluble vitamins by the first part of the 20th century, but fatty acids were not considered to be essential nutrients because they could be synthesized from dietary carbohydrate. This well-established view was challenged in 1929 by George and Mildred Burr who reported that dietary fatty acid was required to prevent a deficiency disease that occurred in rats fed a fat-free diet. They concluded that fatty acids were essential nutrients and showed that linoleic acid prevented the disease and is an essential fatty acid. The Burrs surmised that other unsaturated fatty acids were essential and subsequently demonstrated that linolenic acid, the omega-3 fatty acid analog of linoleic acid, is also an essential fatty acid. The discovery of essential fatty acids was a paradigm-changing finding, and it is now considered to be one of the landmark discoveries in lipid research.

Phosphatidylserine in the brain: metabolism and function.

Phosphatidylserine (PS) is the major anionic phospholipid class particularly enriched in the inner leaflet of the plasma membrane in neural tissues. PS is synthesized from phosphatidylcholine or phosphatidylethanolamine by exchanging the base head group with serine, and this reaction is catalyzed by phosphatidylserine synthase 1 and phosphatidylserine synthase 2 located in the endoplasmic reticulum. Activation of Akt, Raf-1 and protein kinase C signaling, which supports neuronal survival and differentiation, requires interaction of these proteins with PS localized in the cytoplasmic leaflet of the plasma membrane. Furthermore, neurotransmitter release by exocytosis and a number of synaptic receptors and proteins are modulated by PS present in the neuronal membranes. Brain is highly enriched with docosahexaenoic acid (DHA), and brain PS has a high DHA content. By promoting PS synthesis, DHA can uniquely expand the PS pool in neuronal membranes and thereby influence PS-dependent signaling and protein function. Ethanol decreases DHA-promoted PS synthesis and accumulation in neurons, which may contribute to the deleterious effects of ethanol intake. Improvement of some memory functions has been observed in cognitively impaired subjects as a result of PS supplementation, but the mechanism is unclear.

Synaptamide, endocannabinoid-like derivative of docosahexaenoic acid with cannabinoid-independent function.

Docosahexaenoylethanolamide, the structural analog of the endogenous cannabinoid receptor ligand anandamide, is synthesized from docosahexaenoic acid (DHA) in the brain. Although docosahexaenoylethanolamide binds weakly to cannabinoid receptors, it stimulates neurite growth, synaptogenesis and glutamatergic synaptic activity in developing hippocampal neurons at concentrations of 10-100 nM. We have previously proposed the term synaptamide for docosahexaenoylethanolamide to emphasize its potent synaptogenic activity and structural similarity to anandamide. Synaptamide is subjected to hydrolysis by fatty acid amide hydrolase, and can be oxygenated to bioactive metabolites. The brain synaptamide content is dependent on the dietary DHA intake, suggesting an endogenous mechanism whereby diets containing adequate amounts of omega-3 fatty acids improve synaptogenesis in addition to well-recognized anti-inflammatory effects.

Peroxisome proliferator-activated receptor γ decouples fatty acid uptake from lipid inhibition of insulin signaling in skeletal muscle.

Peroxisome proliferator-activated receptor γ (PPARγ) is expressed at low levels in skeletal muscle, where it protects against adiposity and insulin resistance via unclear mechanisms. To test the hypothesis that PPARγ directly modulates skeletal muscle metabolism, we created two models that isolate direct PPARγ actions on skeletal myocytes. PPARγ was overexpressed in murine myotubes by adenotransfection and in mouse skeletal muscle by plasmid electroporation. In cultured myotubes, PPARγ action increased fatty acid uptake and incorporation into myocellular lipids, dependent upon a 154 ± 20-fold up-regulation of CD36 expression. PPARγ overexpression more than doubled insulin-stimulated thymoma viral proto-oncogene (AKT) phosphorylation during low lipid availability. Furthermore, in myotubes exposed to palmitate levels that inhibit insulin signaling, PPARγ overexpression increased insulin-stimulated AKT phosphorylation and glycogen synthesis over 3-fold despite simultaneously increasing myocellular palmitate uptake. The insulin signaling enhancement was associated with an increase in activating phosphorylation of phosphoinositide-dependent protein kinase 1 and a normalized expression of palmitate-induced genes that antagonize AKT phosphorylation. In vivo, PPARγ overexpression more than doubled insulin-dependent AKT phosphorylation in lipid-treated mice but did not augment insulin-stimulated glucose uptake. We conclude that direct PPARγ action promotes myocellular storage of energy by increasing fatty acid uptake and esterification while simultaneously enhancing insulin signaling and glycogen formation. However, direct PPARγ action in skeletal muscle is not sufficient to account for the hypoglycemic actions of PPARγ agonists during lipotoxicity.

A synaptogenic amide N-docosahexaenoylethanolamide promotes hippocampal development.

Docosahexaenoic acid (DHA), the n-3 essential fatty acid that is highly enriched in the brain, increases neurite growth and synaptogenesis in cultured mouse fetal hippocampal neurons. These cellular effects may underlie the DHA-induced enhancement of hippocampus-dependent learning and memory functions. We found that N-docsahexaenoylethanolamide (DEA), an ethanolamide derivative of DHA, is a potent mediator for these actions. This is supported by the observation that DHA is converted to DEA by fetal mouse hippocampal neuron cultures and a hippocampal homogenate, and DEA is present endogenously in the mouse hippocampus. Furthermore, DEA stimulates neurite growth and synaptogenesis at substantially lower concentrations than DHA, and it enhances glutamatergic synaptic activities with concomitant increases in synapsin and glutamate receptor subunit expression in the hippocampal neurons. These findings suggest that DEA, an ethanolamide derivative of DHA, is a synaptogenic factor, and therefore we suggest utilizing the term 'synaptamide'. This brief review summarizes the neuronal production and actions of synaptamide and describes other N-docosahexaenoyl amides that are present in the brain.

Neuroticism but not omega-3 fatty acid levels correlate with early responsiveness to escitalopram.

BACKGROUND: Omega-3 fatty acid (O3FA) levels and dimensional personality measures have been associated with major depression and the course of depressive illness. We sought to study the utility of O3FA levels and dimensional personality measures as predictors of early improvement with escitalopram. METHODS: Twenty-four participants were enrolled in an open-label trial of escitalopram 10 mg/d for 4 weeks. Baseline erythrocyte O3 levels and dimensional personality assessments were obtained. RESULTS: Using a conservative, intention-to-treat analysis, baseline neuroticism (r = -0.57; P = .007), as measured by the Revised NEO Personality Inventory but not erythrocyte O3 levels, was correlated with improvements on escitalopram. A facet analysis of the neuroticism domain showed the relationship with antidepressant response to be focused on trait anxiety (r = -0.65; P = .002). CONCLUSIONS: Anxiety may have important prognostic implications on subsequent response to selective serotonin reuptake inhibitors, such as escitalopram.

Characterization of transgenic mice with neuron-specific expression of soluble epoxide hydrolase.

Soluble epoxide hydrolase (sEH) is the major enzyme responsible for the metabolism and inactivation of epoxyeicosatrienoic acids (EETs). EETs are produced by the cytochrome P450 (CYP) epoxygenase pathway of arachidonic acid (AA) metabolism and tend to be anti-hypertensive, anti-inflammatory and protective against ischemic injury. Since the metabolism of EETs by sEH reduces or eliminates their bioactivity, inhibition of sEH has become a therapeutic strategy for hypertension and inflammation. sEH is found in nearly all tissues so the systemic application of inhibitors is likely to affect more than blood pressure and inflammation. In the central nervous system, EETs are thought to play a role in the regulation of local blood flow, protection from ischemic injury, inhibition of inflammation, the release of peptide hormones and modulation of fever. However, little is known about region- and cell-specific expression of sEH in the brain. In the mouse brain, expression of sEH was found widely in cortical and hippocampal astrocytes and also in a few specific neuron types in the cortex, cerebellum, and medulla. To assess the functional significance of neuronal sEH, we generated a transgenic mouse model, which over-expresses sEH specifically in neurons. Transgenic mice showed increased neuron labeling in cortex and hippocampus with little change in labeling of other brain regions. Despite a 3-fold increase in sEH activity in the brain, there was no change in arterial pressure. This data provides new information required for studying the central roles of the cytochrome P450 epoxygenase pathway.

Size-dependent waveguide dispersion in nanowire optical cavities: slowed light and dispersionless guiding.

Fundamental understanding of the size dependence of nanoscale optical confinement in semiconductor nanowire waveguides, as expressed by changes in the dispersion of light, is crucial for the optimal design of nanophotonic devices. Measurements of the dispersion are particularly challenging for nanoscale cavities due to difficulties associated with the in- and out-coupling of light resulting from diffraction effects. We report the strong size dependence of optical dispersion and associated group velocities in subwavelength width ZnSe nanowire waveguide cavities, using a technique based on Fabry-Perot resonator modes as probes over a wide energy range. Furthermore, we observed subwavelength (lambda/9) dispersionless waveguiding and significant slowing of the propagating light by 90% (c/8). These results, in addition to providing insights into nanoscale optical transport, will facilitate the rational design of nanowire photonic devices with tailored dispersion and group velocities.

Arachidonic acid cytochrome P450 epoxygenase pathway.

Cytochrome P450 (CYP) epoxygenases convert arachidonic acid to four epoxyeicosatrienoic acid (EET) regioisomers, 5,6-, 8,9-, 11,12-, and 14,15-EET, that function as autacrine and paracrine mediators. EETs produce vascular relaxation by activating smooth muscle large-conductance Ca2+-activated K+ channels (BKCa). In addition, they have anti-inflammatory effects on blood vessels and in the kidney, promote angiogenesis, and protect ischemic myocardium and brain. CYP epoxygenases also convert eicosapentaenoic acid to vasoactive epoxy-derivatives, and endocannabinoids containing 11,12- and 14,15-EET are formed. Many EET actions appear to be initiated by EET binding to a membrane receptor that activates ion channels and intracellular signal transduction pathways. However, EETs also are taken up by cells, are incorporated into phospholipids, and bind to cytosolic proteins and nuclear receptors, suggesting that some functions may occur through direct interaction of the EET with intracellular effector systems. Soluble epoxide hydrolase (sEH) converts EETs to dihydroxyeicosatrienoic acids (DHETs). Because this attenuates many of the functional effects of EETs, sEH inhibition is being evaluated as a mechanism for increasing and prolonging the beneficial actions of EETs.

Docosahexaenoic acid synthesis from n-3 fatty acid precursors in rat hippocampal neurons.

Docosahexaenoic acid (DHA), the most abundant n-3 polyunsaturated fatty acid in the brain, has important functions in the hippocampus. To better understand essential fatty acid homeostasis in this region of the brain, we investigated the contributions of n-3 fatty acid precursors in supplying hippocampal neurons with DHA. Primary cultures of rat hippocampal neurons incorporated radiolabeled 18-, 20-, 22-, and 24-carbon n-3 fatty acid and converted some of the uptake to DHA, but the amounts produced from either [1-14C]alpha-linolenic or [1-14C]eicosapentaenoic acid were considerably less than the amounts incorporated when the cultures were incubated with [1-14C]22:6n-3. Most of the [1-14C]22:6n-3 uptake was incorporated into phospholipids, primarily ethanolamine phosphoglycerides. Additional studies demonstrated that the neurons converted [1-14C]linoleic acid to arachidonic acid, the main n-6 fatty acid in the brain. These findings differ from previous results indicating that cerebral and cerebellar neurons cannot convert polyunsaturated fatty acid precursors to DHA or arachidonic acid. Fatty acid compositional analysis demonstrated that the hippocampal neurons contained only 1.1-2.5 mol% DHA under the usual low-DHA culture conditions. The relatively low-DHA content suggests that some responses obtained with these cultures may not be representative of neuronal function in the brain.

Brain uptake and utilization of fatty acids, lipids and lipoproteins: application to neurological disorders.

Transport, synthesis, and utilization of brain fatty acids and other lipids have been topics of investigation for more than a century, yet many fundamental aspects are unresolved and, indeed, subject to controversy. Understanding the mechanisms by which lipids cross the blood brain barrier and how they are utilized by neurons and glia is critical to understanding normal brain development and function, for the diagnosis and therapy of human diseases, and for the planning and delivery of optimal human nutrition throughout the world. Two particularly important fatty acids, both of which are abundant in neuronal membranes are: (a) the omega3 polyunsaturated fatty acid docosahexaenoic acid, deficiencies of which can impede brain development and compromise optimal brain function, and (b) the omega6 polyunsaturated fatty acid arachidonic acid, which yields essential, but potentially toxic, metabolic products. There is an exciting emerging evidence that modulating dietary intake of these fatty acids could have a beneficial effect on human neurological health. A workshop was held in October, 2004, in which investigators from diverse disciplines interacted to present new findings and to discuss issues relevant to lipid uptake, utilization, and metabolism in the brain. The objectives of this workshop were: (1) to assess the state-of-the-art of research in brain fatty acid/lipid uptake and utilization; (2) to discuss progress in understanding molecular mechanisms and the treatment of neurological diseases related to lipids and lipoproteins; (3) to identify areas in which current knowledge is insufficient; (4) to provide recommendations for future research; and (5) to stimulate the interest and involvement of additional neuroscientists, particularly young scientists, in these areas. The meeting was divided into four sessions: (1) mechanisms of lipid uptake and transport in the brain, (2) lipoproteins and polyunsaturated fatty acids, (3) eicosanoids in brain function, and (4) fatty acids and lipids in brain disorders. In this article, we will provide an overview of the topics discussed in these sessions.

Action of epoxyeicosatrienoic acids on cellular function.

Epoxyeicosatrienoic acids (EETs), which function primarily as autocrine and paracrine mediators in the cardiovascular and renal systems, are synthesized from arachidonic acid by cytochrome P-450 epoxygenases. They activate smooth muscle large-conductance Ca(2+)-activated K(+) channels, producing hyperpolarization and vasorelaxation. EETs also have anti-inflammatory effects in the vasculature and kidney, stimulate angiogenesis, and have mitogenic effects in the kidney. Many of the functional effects of EETs occur through activation of signal transduction pathways and modulation of gene expression, events probably initiated by binding to a putative cell surface EET receptor. However, EETs are rapidly taken up by cells and are incorporated into and released from phospholipids, suggesting that some functional effects may occur through a direct interaction between the EET and an intracellular effector system. In this regard, EETs and several of their metabolites activate peroxisome proliferator-activated receptor alpha (PPARalpha) and PPARgamma, suggesting that some functional effects may result from PPAR activation. EETs are metabolized primarily by conversion to dihydroxyeicosatrienoic acids (DHETs), a reaction catalyzed by soluble epoxide hydrolase (sEH). Many potentially beneficial actions of EETs are attenuated upon conversion to DHETs, which do not appear to be essential under routine conditions. Therefore, sEH is considered a potential therapeutic target for enhancing the beneficial functions of EETs.