Laboratory of oxylipins

Department of molecular bases of neurosignalization

Head: Vladimir Bezuglov, D.Sc, professor

oxylipins; endocannabinoids; endovanilloids; biochemistry; lipids; neurochemistry; cell biology; neurodegeneration; cancer; stroke

The research of the Laboratory is aimed at revealing the role of lipids as molecules that transmit information in the body regulation system. One of the directions is the study of signaling pathways that mediate the lipids ability to enhance proliferation or induce apoptosis, protect cells of the nervous system from the effects of damaging factors, induce cell differentiation and morphogenesis. Lipids are important regulators of a wide range of normal and pathological processes in the body, so the search for lipid molecules with more selective action is also at the center of the Laboratory's attention.

The main object of the Laboratory's research is natural oxylipins (prostaglandins, etc.), natural neurolypes (conjugates of fatty acids with biogenic amines, neurotransmitters and amino acids) and their synthetic analogues.

Earlier, the Laboratory staff (Bezuglov et al., 1995) suggested that the acylated derivatives of endogenous neurotransmitters can be a new family of bioregulators combining the properties of neuroactive lipids (anandamide) and classical neurotransmitters (dopamine, serotonin, etc.). Acyl-dopamines and derivatives of fatty acids with various amino acids were then found in the nervous and peripheral tissues of various organisms, including mammals. It turned out that these compounds have a wide range of biological effects that continue to be studied at the Laboratory.

The Laboratory discovered new ways of metabolizing acyl-dopamines and clarified some aspects of their biosynthesis. Also, the team showed that acyl-dopamines  has a neuroprotective effect and it is able to induce apoptosis of the oncotransformed cells of various tissue origin in vitro, which indicates the large pharmacological potential of these substances. Now the researchers are investigating the mechanism of their toxic and differentiating effects on cancer cells, and also seek to increase the effectiveness of existing antitumor drugs.

To fight neurodegenerative diseases, an important socially significant problem, the Laboratory developed a strategy for searching for new neuroprotectors and protecting neurons from irreversible changes under the influence of stress. This approach is based on the use of the natural neuroactive lipids, peptides and other low-molecular bioregulators potential. Researchers managed to show that some modified acyl-dopamine are promising candidates for treatment of stroke consequences. In addition, the Laboratory is developing methods of directional delivery compounds to neurons capable to slowing down and reversing the neurodegenerative process.

Researchers of the Laboratory actively use modern methods of cell and molecular biology, fine organic synthesis, analytical chemistry and biochemistry.

The Laboratory cooperates with the Institute departments, as well as the Institute of Molecular Genetics of the Russian Academy of Sciences (RAS), the Mendeleev Russian Chemistry and Technology University, the Zakusov Institute of Pharmacology RAS, the Institute of Physiologically Active Substances RAS, the Koltsov Institute of Developmental Biology RAS, the Institute of Bioorganic Chemistry of the National Academy of Sciences of Belarus, the Central University of Rajasthan (India), the Max Mousseron Institute of Biomolecules (France) etc.

The Laboratory was founded in 1990 on the basis of the Group of Prostaglandins and Leukotrienes.

  • Neuroactive lipids and their role in normal and pathological processes.
  • Mechanisms of lipid signals perception and intracellular transmission.
  • The development of novel molecular instruments based of bioactive lipids (including fluorescent, biotynilated and isotope-labeled compounds).
  • The development of new types of drugs, based on the conception of Information multifunctional compound molecules.
  • The characterization of biosynthetic and catabolic pathways of N-acyl dopamines in rat liver, brain and spinal cord, as well as in freshwater hydra homogenate. The alternate pathways of N-acyl dopamines (sulfation, glucuronidation and catechol group oxidation) were identified for the first time.
  • The identification of N-docosahexaenoyl dopamine in Hydra magnipapillata and H. Attenuate. The morphogenetic effect of acyl-dopamines and their ability to regulate regeneration processes in hydra were shown.
  • It was shown for the first time that neurolypes from the group of N-acyl dopamine interfere in the death of neurons, save their functional properties in models of neurodegenerative processes, reduce the volume of neural tissue damage in focal ischemia, and increase survival in acute hypoxia and contribute to the preservation of cognitive functions in the posthypoxic period in experimental animals.
  • The mechanism of signal transmission during the induction of apoptosis by N-acyl dopamine is established, the main proteins participating in this process are identified, including the cytoplasmic receptor, intracellular regulatory proteins, kinases and transcription factors.
  • The ability of acyl-dopamine to induce the differentiation of oncotransformed cells was detected, new cell lines with increased resistance to acyl dopamine were obtained.
  • A test system was developed to assess the hydrolytic stability of peptides and other compounds when administered orally on the basis of the rat gastrointestinal tract fragments.
  • A new class of hybrid molecules peptolipins was synthesized, combining fragments of bioactive lipids, peptides and biogenic amines, which possess neuroprotective, vasoactive and anti-inflammatory effects.
  • The development of a novel conception for pharmacological drug action prolongation using non-covalent complexes of protein with natural polysialic acid (size <=100 nm). A technology for the manufacturing of such nanocomplexes with genetical engineered interferons α2b and β1b, insulin and human G-CSF was developed, which is quite simple and productive. The experiments with cell cultures and animals confirmed the prolonged activity of the proteins inside nanocomplexes.
Vladimir Bezuglov, D.Sc, professordepart.
Natal'ja Greckaja, Ph.D.s. r.
Mihail Bobrov, Ph.D.s. r. f.+7(495)3306592
Mikhail Akimov, Ph.D.s. r.
Elena Fomina-Ageeva, Ph.D.r. f.+7(495)330-65-92
Galina Zinchenkoj. r.
Alina LavrovaPhD
Alina AshbaPhD

Selected publications

  1. Gholami S., Bordbar A.K., Akvan N., Parastar H., Fani N., Gretskaya N.M., Bezuglov V.V., Haertlé T. (2015). Binding assessment of two arachidonic-based synthetic derivatives of adrenalin with β-lactoglobulin: Molecular modeling and chemometrics approach. Biophys. Chem. 207, 97–106 [+]

    A computational approach to predict the main binding modes of two adrenalin derivatives, arachidonoyl adrenalin (AA-AD) and arachidonoyl noradrenalin (AA-NOR) with the β-lactoglubuline (BLG) as a nano-milk protein carrier is presented and assessed by comparison to the UV-Vis absorption spectroscopic data using chemometric analysis. Analysis of the spectral data matrices by using the multivariate curve resolution-alternating least squares (MCR-ALS) algorithm led to the pure concentration calculation and spectral profiles resolution of the chemical constituents and the apparent equilibrium constants computation. The negative values of entropy and enthalpy changes for both compound indicated the essential role of hydrogen bonding and van der Waals interactions as main driving forces in stabilizing protein-ligand complex. Computational studies predicted that both derivatives are situated in the calyx pose and remained in that pose during the whole time of simulation with no any significant protein structural changes which pointed that the BLG could be considered as a suitable carrier for these catecholamine compounds.

  2. Akimov M.G., Gretskaya N.M., Zinchenko G.N., Bezuglov V.V. (2015). Cytotoxicity of Endogenous Lipids N-acyl Dopamines and their Possible Metabolic Derivatives for Human Cancer Cell Lines of Different Histological Origin. Anticancer Res. 35 (5), 2657–61 [+]

    Dopamine amides of long chain fatty acids are a family of endogenous mammalian lipids with an unknown function; they are anti-proliferative for the C6 glioblastoma cell line. To assess their possible anti-cancer activity we evaluated their cytotoxicity for a set of cancer cell lines.

  3. Nikishin D.A., Milosević I., Gojković M., Rakić L., Bezuglov V.V., Shmukler Y.B. (2015). Expression and functional activity of neurotransmitter system components in sea urchins' early development. Zygote , 1–13 [+]

    Reverse-transcription polymerase chain reaction (RT-PCR) investigation of the expression of the components supposedly taking part in serotonin regulation of the early development of Paracentrotus lividus has shown the presence of transcripts of five receptors, one of which has conservative amino acid residues characteristic of monoaminergic receptors. At the early stages of embryogenesis the expressions of serotonin transporter (SERT) and noradrenaline transporter (NET) were also recognized. The activities of the enzymes of serotonin synthesis and serotonin transporter were shown using immunohistochemistry and incubation with para-chlorophenylalanine (PСРА) and 5-hydroxytryptophan (HTP). Pharmacological experiments have shown a preferential cytostatic activity of ligands characterized as mammalian 5-hydroxytryptamine (5-HT)1-antagonists. On the basis of the sum of the data from molecular biology and embryo physiological experiments, it is suggested that metabotropic serotonin receptors and membrane transporters take part in the regulatory processes of early sea urchin embryogenesis.

  4. TaheriKafrani A., Choiset Y., Faizullin D.A., Zuev Y.F., Bezuglov V.V., Chobert J.M., Bordbar A.K., Haertlé T. (2011). Interactions of β-lactoglobulin with serotonin and arachidonyl serotonin. Biopolymers 95 (12), 871–80 [+]

    β-Lactoglobulin (β-LG) is a lipocalin, which is the major whey protein of cow's milk and the milk of other mammals. However, it is absent from human milk. The biological function of β-LG is not clear, but its potential role in carrying fatty acids through the digestive tract has been suggested. β-LG has been found in complexes with lipids such as butyric and oleic acids and has a high affinity for a wide variety of compounds. Serotonin (5-hydroxytryptamine, 5-HT), an important compound found in animals and plants, has various functions, including the regulation of mood, appetite, sleep, muscle contraction, and some cognitive functions such as memory and learning. In this study, the interaction of serotonin and one of its derivatives, arachidonyl serotonin (AA-5HT), with β-LG was investigated using circular dichroism (CD) and fluorescence intensity measurements. These two ligands interact with β-LG forming equimolar complexes. The binding constant for the serotonin/β-LG interaction is between 10⁵ and 10⁶ M(-1) , whereas for the AA-5HT/β-LG complex it is between 10⁴ and 10⁵ M(-1) as determined by measurements of either protein or ligand fluorescence. The observed binding affinities were higher in hydroethanolic media (25% EtOH). The interactions between serotonin/β-LG and AA-5HT/β-LG may compete with self-association (micellization) of both the ligand and the protein. According to far- and near-UV CD results, these ligands have no apparent influence on β-LG secondary structure, however they partially destabilize its tertiary structure. Their binding by β-LG may be one of the peripheral mechanisms of the regulation of the content of serotonin and its derivatives in the bowel of milk-fed animals.

  5. Timofeev V.I., ChuprovNetochin R.N., Samigina V.R., Bezuglov V.V., Miroshnikov K.A., Kuranova I.P. (2010). X-ray investigation of gene-engineered human insulin crystallized from a solution containing polysialic acid. Acta Crystallogr. Sect. F Struct. Biol. Cryst. Com 66 (Pt 3), 259–63 [+]

    Attempts to crystallize the noncovalent complex of recombinant human insulin with polysialic acid were carried out under normal and microgravity conditions. Both crystal types belonged to the same space group, I2(1)3, with unit-cell parameters a = b = c = 77.365 A, alpha = beta = gamma = 90.00 degrees. The reported space group and unit-cell parameters are almost identical to those of cubic insulin reported in the PDB. The results of X-ray studies confirmed that the crystals obtained were cubic insulin crystals and that they contained no polysialic acid or its fragments. Electron-density maps were calculated using X-ray diffraction sets from earth-grown and microgravity-grown crystals and the three-dimensional structure of the insulin molecule was determined and refined. The conformation and secondary-structural elements of the insulin molecule in different crystal forms were compared.

  6. Buznikov G.A., Nikitina L.A., Bezuglov V.V., Francisco M.E., Boysen G., ObispoPeak I.N., Peterson R.E., Weiss E.R., Schuel H., Temple B.R., Morrow A.L., Lauder J.M. (2010). A putative 'pre-nervous' endocannabinoid system in early echinoderm development. Dev. Neurosci. 32 (1), 1–18 [+]

    Embryos and larvae of sea urchins (Lytechinus variegatus, Strongylocentrotus droebachiensis, Strongylocentrotus purpuratus, Dendraster excentricus), and starfish (Pisaster ochraceus) were investigated for the presence of a functional endocannabinoid system. Anandamide (arachidonoyl ethanolamide, AEA), was measured in early L. variegatus embryos by liquid chromatography/mass spectrometry. AEA showed a strong developmental dynamic, increasing more than 5-fold between the 8-16 cell and mid-blastula 2 stage. 'Perturb-and-rescue' experiments in different sea urchin species and starfish showed that AEA blocked transition of embryos from the blastula to the gastrula stage, but had no effect on cleavage divisions, even at high doses. The non-selective cannabinoid receptor agonist, CP55940, had similar effects, but unlike AEA, also blocked cleavage divisions. CB1 antagonists, AEA transport inhibitors, and the cation channel transient membrane potential receptor V1 (TrpV1) agonist, arachidonoyl vanillic acid (arvanil), as well as arachidonoyl serotonin and dopamine (AA-5-HT, AA-DA) acted as rescue substances, partially or totally preventing abnormal embryonic phenotypes elicited by AEA or CP55940. Radioligand binding of [(3)H]CP55940 to membrane preparations from embryos/larvae failed to show significant binding, consistent with the lack of CB receptor orthologs in the sea urchin genome. However, when binding was conducted on whole cell lysates, a small amount of [(3)H]CP55940 binding was observed at the pluteus stage that was displaced by the CB2 antagonist, SR144528. Since AEA is known to bind with high affinity to TrpV1 and to certain G-protein-coupled receptors (GPCRs), the ability of arvanil, AA-5-HT and AA-DA to rescue embryos from AEA teratogenesis suggests that in sea urchins AEA and other endocannabinoids may utilize both Trp and GPCR orthologs. This possibility was explored using bioinformatic and phylogenetic tools to identify candidate orthologs in the S. purpuratus sea urchin genome. Candidate TrpA1 and TrpV1 orthologs were identified. The TrpA1 ortholog fell within a monophyletic clade, including both vertebrate and invertebrate orthologs, whereas the TrpV1 orthologs fell within two distinct TrpV-like invertebrate clades. One of the sea urchin TrpV orthologs was more closely related to the vertebrate epithelial calcium channels (TrpV5-6 family) than to the vertebrate TrpV1-4 family, as determined using profile-hidden Markov model (HMM) searches. Candidate dopamine and adrenergic GPCR orthologs were identified in the sea urchin genome, but no cannabinoid GPCRs were found, consistent with earlier studies. Candidate dopamine D(1), D(2) or alpha(1)-adrenergic receptor orthologs were identified as potential progenitors to the vertebrate cannabinoid receptors using HMM searches, depending on whether the multiple sequence alignment of CB receptor sequences consisted only of urochordate and cephalochordate sequences or also included vertebrate sequences.

  7. Akimov M.G., Nazimov I.V., Gretskaya N.M., Zinchenko G.N., Bezuglov V.V. (2009). Sulfation of N-acyl dopamines in rat tissues. Biochemistry Mosc. 74 (6), 681–5 [+]

    Sulfation of N-acyl dopamines has been shown for the first time in cytosolic fractions of rat liver and nervous system. Sulfation of dopamine amides of docosahexaenoic and oleic acids occurred in all tissues studied, N-arachidonoyl dopamine was sulfated in the liver and spinal cord, and N-stearoyl dopamine was sulfated only in the liver. Depending on the substrate and tissue, the sulfation activity varied from 0.5 to 3.5 nmol/min per mg total protein. Kinetic parameters of N-docosahexaenoyl dopamine sulfation in the brain were determined. The findings characterize the sulfation system as the most productive metabolic pathway of N-acyl dopamines, but the role of this system in the body is unclear because of high K(m) value.

  8. Akimov M.G., Nazimov I.V., Gretskaia N.M., Deigin V.I., Bezuglov V.V. (2009). [The study of peptide stability during hydrolysis by rat gastroenteric tract fragments]. Bioorg. Khim. 36 (6), 753–9 [+]

    The hydrolytic stability of therapeutic peptides such as dalargin, stemokin and some others, including cyclic tripeptides modified by ibuprofen and aspirin, was studied. Two experimental systems were used, one containing purified enzymes pepsin, trypsin and chymotrypsin and other based on fragments of rat stomach and ileum. It was found that linear peptides without D-aminoacids are hydrolyzed by fragments of stomach and ileum but resistant to hydrolysis with purified enzymes. The peptides with D-aminoacids and cyclic peptides are stable in all experimental conditions used, however, peptides modified with aspirin lost acetyl moiety of aspirin residue in acidic medium, the process is accelerated in presence of pepsin.

  9. Ostroumova T.V., Markova L.N., Akimov M.G., Gretskaia N.M., Bezuglov V.V. (2009). [Docosahexaenoyl dopamine in freshwater hydra: effects on regeneration and metabolic changes]. Ontogenez 41 (3), 199–203 [+]

    The effects of docosahexaenoyl dopamine and docosahexaenoic acid on the regeneration of hydra gastric and basal fragments are studied. Docosahexaenoyl dopamine induced morphogenetic abnormalities such as single ectopic tentacles in the gastric region and projections in the gastric and basal regions. Docosahexaenoic acid had no effect on the morphogenesis except for a mild slowing of the regeneration rate. Since no hydrolysis of docosahexaenoyl dopamine was detected in hydra extract, it was assumed that the morphogenetic effect could be associated with the dopamine component of this complex.

  10. Bezuglov V.V., Gretskaia N.M., Bobrov M.I.u., Akimov M.G., FominaAgeeva E.V., Zinchenko G.N., Bairamashvili D.I., Miroshnikov A.I. (2009). [Modification of recombinant proteins by covalent polysialation illustrated with the example of human insulin]. Bioorg. Khim. 35 (2), 274–8 [+]

    Methods of selective and nonselective covalent immobilization of genetically engineered proteins on molecules of natural polysialic acid are described by the example of human insulin. Such modification increases insulin lifetime in vivo.

  11. Akimov M.G., Gretskaia N.M., Shevchenko K.V., Shevchenko V.P., Miasoedov N.F., Bobrov M.I.u., Bezuglov V.V. (2009). [New aspects of biosynthesis and metabolism of N-acyldopamines in rat tissues]. Bioorg. Khim. 33 (6), 648–52 [+]

    Possible biosynthetic pathways of N-acyldopamines in rat tissues were compared. It was shown that an insignificant amount of the conjugation products was formed during the incubation of arachidonic acid and dopamine, whereas the substitution of tyrosine for dopamine resulted in the productive biosynthesis of N-arachidonoyldopamine. The biosynthesis presumably involves several closely conjugated enzymatic stages, and free fatty acids rather than their CoA esters served as the starting substrates. The decarboxylation stage probably precedes the stage of catechol system formation, because N-acetyltyramine (a probable intermediate) was easily oxidized by monophenol monooxygenase to N-acyldopamine, whereas N-acyltyrosine is hydrolyzed under these conditions. Biosynthesis of N-acyldopamines in a cell-free medium was accompanied by their methylation. The possibility of oxidative metabolism of N-acyldopamines, which could serve as co-substrates or inhibitors of different oxidoreductases, was shown for the first time.

  12. Bezuglov V.V., Gretskaia N.M., Blazhenova A.V., Adrianova E.L., Akimov M.G., Bobrov M.I.u., Nazimov I.V., Kisel M.I., Sharko O.L., Novikov A.V., Krasnov N.V., Shevchenko V.P., Viunova T.V., Miasoedova N.F. (2009). [Arachidonoyl amino acids and arachidonoyl peptides: synthesis and properties]. Bioorg. Khim. 32 (3), 258–67 [+]

    N-Arachidonoyl (AA) derivatives of amino acids (glycine, phenylalanine, proline, valine, gamma-amino butyric acid (GABA), dihydroxyphenylalanine, tyrosine, tryptophan, and alanine) and peptides (Semax, MEHFPGP, and PGP) were synthesized in order to study the biological properties of acylamino acids. The mass spectra of all the compounds at atmospheric pressure electrospray ionization display the most intense peaks of protonated molecular ions; the detection limits for these compounds are 10 fmol per sample. AA-Gly showed the highest inhibitory activity toward fatty acid amide hydrolase from rat brain (IC50 6.5 microM) among all the acylamino acids studied. AA-Phe, AA-Tyr, and AA-GABA exhibited a weak but detectable inhibitory effect (IC50 55, 60, and 50 microM, respectively). The acylated amino acids themselves, except for AA-Gly, were stable to the hydrolysis by this enzyme. All the arachidonoylamino acids inhibited cabbage phospholipase D to various degrees; AA-GABA and AA-Phe proved to be the most active (IC50 20 and 27 microM, respectively). Attempts to detect the biosynthesis of AA-Tyr in homogenates of rat liver and nerve tissue showed no formation in vitro of either this acylamino acid or AA-dopamine and AA-Phe, the products of its metabolism. The highest contents of these metabolites were detected in liver homogenate and in the brain homogenate, respectively. Acylamino acids exert no cytotoxic effect toward the glioma C6 cells. It was shown that N-acylation of Semax with arachidonic acid results in enhancement of its hydrolytic stability and increases its affinity for the sites of specific binding in rat cerebellum membranes. The English version of the paper: Russian Journal of Bioorganic Chemistry, 2006, vol. 32, no. 3; see also

  13. Romanov V.P., Bezuglov V.V., Bobrov M.I.u., Kostromina T.I., Feofanov S.A., Miroshnikov A.I. (2009). [Isolation of expressed in E. coli human interferon beta1b (Ser17) by ion-exchange chromatography]. Bioorg. Khim. 37 (3), 327–33 [+]

    A method for isolation of interferon beta1b (Serl7) from inclusion bodies, comprising the steps of solution and reduction of protein from the inclusion bodies, refolding, chromatography on DEAE-Sepharose, chromatography on SP-Sepharose, concentrating, desalting and addition of stabilizers. The solution of reduced protein was diluted with pH 8.0 buffer of 50 mM Tris-HCl, 25 microM CuCl2 and 0.5% Twin 20 for refolding. We used gradient of pH (from 9.3 upto 11.3) for elution of interferon-beta from cation-exchange column. We concentrated of eluate and then desalted on the Sephadex G-50 column with 1 mM NaOH. Then the protein solution was neutralized with mannitol and Na-phosphate. Obtained preparation of interferon-beta was pure by gel-electrophoresis and by HPLC analysis, and had practically indentical level of antiproliferative activity with well-known preparation of Betaferone. Thus we show the possibility of isolation and obtaining of pure and active interferone-beta by ion-exchange chromatography in the presence of non-ion detergent Twin 20. We believe this method for interferon betalb preparation is perspective for scaling and using in the develop of industrial technology for production of this preparation.

  14. Grigorev V.V., Serkov I.V., Beznosko B.K., Ivanova T.A., Gretskaia N.M., Bezuglov V.V., Bachurin S.O. (2009). [Influence of derivatives of arachidonic and docosohexaenic acids on AMPA receptors in Purkinje neurons and cognitive functions in mice]. Izv. Akad. Nauk. Ser. Biol.  (3), 370–4 [+]

    The effect of derivatives of arachidonic and docosahexaenoic acids on AMPA receptors in Purkinje cells from the rat cerebellum was studied using the patch-clamp electrophysiological method. It was shown that derivatives of arachidonic acid-arachidonoyl dopamine and docosahexaenoic acid-docosahexaenoyl dopamine and ester of docosahexaenoic acid with ethylene glycol in nanomolar concentrations effectively potentiated the ionic currents caused by activation of AMPA receptors of kainic acid. Ester of docosahexaenoic acid with nitroethylene glycol blocked AMPA receptors, and anandamide (ethanolamide of arachidonic acid) was not effective. A behavioral test showed that docosahexaenoyl dopamine in doses of 0.1-20 mg/kg had no effect on the learning and memory abilities of the animals tested.

  15. Serkov I.V., Bezuglov V.V. (2009). [Endocannabinoid 2-arachidonoylglycerol: 1,3-dinitrates of cyclooxygenase metabolites, synthesis and properties]. Bioorg. Khim. 35 (2), 245–52 [+]

    Glycerol esters 1,3-dinitrates of the cyclooxygenase metabolites of natural prostaglandin 2-arachidonoylglycerol, an endogenous ligand of cannabinoid receptors, were synthesized for the first time. Four methods of synthesis of these esters were developed via the activation of a carboxyl group and their chemical and pharmacological properties were investigated. The esters exhibit a more selective pharmacological spectrum of activities in comparison with the corresponding natural prostaglandins: some types of myotropic activity were enhanced, while others were loosened. 1,3-dinitroglycerol esters act as vasodilators, whereas the majority of natural prostaglandins act as vasoconstrictors. The observed changes result from the introduction of an NO-releasing fragment into prostaglandin molecule.

  16. Serkov I.V., Bezuglov V.V. (2009). [Prostaglandin fluorides in synthesis of natural prostaglandin derivatives at a carboxyl group]. Bioorg. Khim. 35 (1), 122–8 [+]

    Methods of synthesis of prostaglandin fluorides were developed and their properties were investigated. These compounds were shown to be convenient synthetic precursors for obtaining esters and amides of natural prostaglandins and their fluorodeoxy analogues.

  17. Kurdiumov I.N., Ganshina T.S., Mirzoian N.R., Vasileva T.M., Makarov V.A., Petrukhina G.N., Bezuglov V.V., Gretskaia N.M., Zinchenko G.N., Mirzoian R.S. (2009). [Cerebrovascular and antiaggregative effects of GABA-docosahexaenoyldopamine conjugate]. Eksp Klin Farmakol 71 (4), 26–9 [+]

    The effects of GABA - docosahexaenoyldopamine (DHED) conjugate on the cerebral haemodynamics and thrombocyte aggregation were evaluated and compared to these of docosahexaenoyldopamine alone. The GABA - DHED conjugate was shown to significantly enhance the cerebral circulation in rats with a model of global transient cerebral ischemia, as compared to the intact animals. Administered alone, DHED increased the blood supply of both intact and ischemic brains to an equal extent. The GABA-DHED conjugate demonstrated the antiaggregative activity, but the effect was less expressed than that of DHED alone.

  18. Buznikov G.A., Nikitina L.A., Seidler F.J., Slotkin T.A., Bezuglov V.V., Milosević I., Lazarević L., Rogac L., Ruzdijić S., Rakić L.M. (2009). Amyloid precursor protein 96-110 and beta-amyloid 1-42 elicit developmental anomalies in sea urchin embryos and larvae that are alleviated by neurotransmitter analogs for acetylcholine, serotonin and cannabinoids. Neurotoxicol Teratol 30 (6), 503–9 [+]

    Amyloid precursor protein (APP) is overexpressed in the developing brain and portions of its extracellular domain, especially amino acid residues 96-110, play an important role in neurite outgrowth and neural cell differentiation. In the current study, we evaluated the developmental abnormalities caused by administration of exogenous APP(96-110) in sea urchin embryos and larvae, which, like the developing mammalian brain, utilize acetylcholine and other neurotransmitters as morphogens; effects were compared to those of beta-amyloid 1-42 (Abeta42), the neurotoxic APP fragment contained within neurodegenerative plaques in Alzheimer's Disease. Although both peptides elicited dysmorphogenesis, Abeta42 was far more potent; in addition, whereas Abeta42 produced abnormalities at developmental stages ranging from early cleavage divisions to the late pluteus, APP(96-110) effects were restricted to the intermediate, mid-blastula stage. For both agents, anomalies were prevented or reduced by addition of lipid-permeable analogs of acetylcholine, serotonin or cannabinoids; physostigmine, a carbamate-derived cholinesterase inhibitor, was also effective. In contrast, agents that act on NMDA receptors (memantine) or alpha-adrenergic receptors (nicergoline), and that are therapeutic in Alzheimer's Disease, were themselves embryotoxic, as was tacrine, a cholinesterase inhibitor from a different chemical class than physostigmine. Protection was also provided by agents acting downstream from receptor-mediated events: increasing cyclic AMP with caffeine or isobutylmethylxanthine, or administering the antioxidant, a-tocopherol, were all partially effective. Our findings reinforce a role for APP in development and point to specific interactions with neurotransmitter systems that act as morphogens in developing sea urchins as well as in the mammalian brain.

  19. Vunova T.V., Andreeva L.A., Shevchenko K.V., Shevchenko V.P., Bobrov M.Y., Bezuglov V.V., Myasoedov N.F. (2009). Binding of tripeptide Pro-Gly-Pro labeled at the C-terminal proline residue to plasma membranes of the rat forebrain. Dokl. Biol. Sci. 419, 95–6 ID:590
  20. Serkov I.V., Shevtsova E.F., Dubova L.G., Kireeva E.G., Vishnevskaya E.M., Gretskaya N.M., Bezuglov V.V., Bachurin S.O. (2009). Interaction of docosahexaenoic acid derivatives with mitochondria. Dokl. Biol. Sci. 414, 187–9 ID:594
  21. Serkov I.V., Grigorev V.V., Ivanova T.A., Gretskaya N.M., Bezuglov V.V., Bachurin S.O. (2009). Effect of derivatives of docosahexaenoic acid on AMPA receptors in Purkinje neurons. Dokl. Biol. Sci. 411, 434–5 ID:595
  22. Vyunova T.V., Shevchenko K.V., Shevchenko V.P., Bobrov M.Y., Bezuglov V.V., Myasoedov N.F. (2009). Specific binding of semax in different regions of the rat brain. Dokl. Biol. Sci. 410, 376–7 ID:598
  23. Serkov I.V., Bezuglov V.V. (2009). [O-Nitration of prostaglandins: synthesis of 15-O-nitrates of 11-deoxyprostaglandin E1 and its methyl ester]. Bioorg. Khim. 32 (1), 110–2 [+]

    O-Nitration of the allylic hydroxyl group in prostaglandins and the synthesis of 15-O-nitrates of 11-deoxyprostaglandin E1 and its methyl ester are described for the first time.

  24. Buznikov G.A., Peterson R.E., Nikitina L.A., Bezuglov V.V., Lauder J.M. (2009). The pre-nervous serotonergic system of developing sea urchin embryos and larvae: pharmacologic and immunocytochemical evidence. Neurochem. Res. 30 (6-7), 825–37 [+]

    Forty serotonin-related neurochemicals were tested on embryos and larvae of Lytechinus variegatus and other sea urchin species. Some of these substances (agonists of 5-HT1 receptors, antagonists of 5-HT2, 5-HT3 or 5-HT4 receptors, and inhibitors of the serotonin transporter, SERT) perturbed post-blastulation development, eliciting changes in embryonic/larval phenotypes typical for each class of receptor ligand. These developmental malformations were prevented completely or partially by serotonin (5-HT) or 5-HT analogs (5-HTQ, AA-5-HT), providing evidence for the putative localization of cellular targets. Immunoreactive 5-HT, 5-HT receptors and SERT were found in pre-nervous embryos and larvae of both L. variegatus and Strongylocentrotus droebachiensis. During gastrulation, these components of the serotonergic system were localized to the archenteron (primary gut), mesenchyme-like cells, and often the apical ectoderm. These results provide evidence that pre-nervous 5-HT may regulate early events of sea urchin embryogenesis, mediated by 5-HT receptors or the 5-HT transporter.

  25. Roshchina V.V., Bezuglov V.V., Markova L.N., Sakharova N.Y., Buznikov G.A., Karnaukhov V.N., Chailakhyan L.M. (2009). Interaction of living cells with fluorescent derivatives of biogenic amines. Dokl. Biochem. Biophys. 393, 346–9 ID:601
  26. Vasileva T.M., Petrukhina G.N., Makarov V.A., Serkov I.V., Gretskaia N.M., Bezuglov V.V. (2009). [Effects of the novel synthetic fatty acid dinitroglycerol esters on aggregation of the human platelets in vitro]. Eksp Klin Farmakol 66 (6), 36–41 [+]

    A series of six new synthetic dinitroglycerol esters of fatty acids on the human platelet aggregation was studied in vitro. Inclusion of the dinitroglycerol moiety into the molecule of arachidonic acid deprived this acid from pro-aggregant activity. All six compounds produced moderate (dose-dependent) inhibition of the platelet aggregation process induced by arachidonic acid (1 x 10(-3) M). Platelet aggregation was most significantly affected by dinitroglycerol esters of arachidonic and docosahexaenoic acids. This is probably explained by the influence of these esters on the oxidative metabolism of arachidonic acid to eicosanoids playing the role of proaggregants. In the presence of vessel wall (rat aorta fragments), dinitroglycerol esters of arachidonic and docosahexaenoic acids incubated with platelets (5 min, 37 degrees C) significantly reduced their aggregation induced by arachidonic acid (1 x 10(-3) M) or docosahexaenoic acid (1 x 10(-5) M) under the conditions of endothelial cyclooxygenase suppressed by acetylsalicylic acid (10 mg/ml). The pronounced antiaggregant effect of the synthetic dinitroglycerol esters studied is probably related to their ability to act as NO donors suppressing the activity of thrombocytes (provided that the NO production activity is present in the system).

  27. Vasileva T.M., Petrukhina G.N., Makarov V.A., Miftakhova N.T., Khaĭlov N.A., Ganshina T.S., Mirzoian R.S., Gretskaia N.M., Bobrov M.I.u., Bezuglov V.V. (2009). [Effect of dopamineamides of polyunsaturated fatty acids on blood coagulation system and cerebral circulation]. Eksp Klin Farmakol 65 (6), 41–5 [+]

    A series of original dopaminamides of polyunsaturated fatty acids were synthesized and characterized with respect to antiaggregant and cerebrovascular stimulant properties. It was established that dopaminamides of linolic, dimethyllinolic, docosapentaenoic, docosahexaenoic (DHEA) and stearidonic (C18:4 and C18:3) acids decrease ADP and arachidonic acid (AA) induced human thrombocyte aggregation in vitro. The most pronounced antiaggregant effect was observed for DHEA dopaminamide: in a dose of 10 mg/kg, this agent produced a significant decrease in the AA induced thrombocyte aggregation. DHEA per se in the same dose increases the activated partial thromboplastin time (APTT), while not affecting the prothrombin time. The synthesized dopaminamides of arachidonic, eicosapentaenoic, and docosahexaenoic acids stimulate local circulation in the cerebral cortex. The most pronounced cerebrovascular effect was also produced by DHEA dopaminamide.

  28. Bezuglov V.V., Zinchenko G.N., Nikitina L.A., Buznikov G.A. (2009). [Arachidonoylcholine and N,N-dimethylaminoethyl arachidonate are new cholinergic compounds]. Bioorg. Khim. 27 (3), 227–30 [+]

    Choline and N,N-dimethylaminoethyl esters of arachidonic and some other fatty acids were synthesized. Experiments on the embryos and larvae of sea urchins, sensitive to cholinergic compounds, showed that arachidonoylcholine exhibited cholinomimetic activity similar to that of nicotine whereas N,N-dimethylaminoethyl arachidonate acted as an acetylcholine antagonist. The corresponding esters of docosahexaenoic acid displayed similar biological properties.

  29. Willows A.O., Nikitina L.A., Bezuglov V.V., Gretskaia N.M., Buznikov G.A. (2009). [The possible functional interaction of serotonin and neuropeptides in embryogenic regulatory processes (experiments on embryos of the mollusk Tritona diomedea)]. Ontogenez 31 (2), 132–8 [+]

    Ritanserin and inmecarb hydrochloride, antagonists of serotonin, act cytostatically and teratogenically on early embryos of Tritonia diomedea, a nudibranch mollusk. On the basis of a pharmacological analysis and the type of developmental abnormalities observed, this action appears to be due to disturbances in the functional activity of endogenous serotonin and is associated with damage of to the cytoskeleton. The effects of ritanserin and inmecarb are prevented or attenuated by lipophilic serotonin analogs (serotoninamides of polyenoic fatty acids), as well as by polypeptides isolated from neurons Pd5 and Pd6 of the pedal ganglia of the adult Tritonia. In late embryos (stage of veligers), serotonin and to a lesser extent its lipophilic analogs strongly increase embryonic motility. This effect of serotonin is potentiated by some neuropeptides and inhibited by others. These results provide evidence for functional interaction between serotonin and neuropeptides in the control processes of embryogenesis.

  30. Markova L.N., Sakharova N.I.u., Bezuglov V.V. (2009). [The action of catecholamine-synthesis inhibitors and of spiperone on sea urchin and mouse embryos]. Ontogenez 31 (1), 32–9 [+]

    We studied the effects of three inhibitors of catecholamine synthesis on the development of sea urchins Sphaerechinus granularis and Paracentrotus lividus. These drugs affected the early embryogenesis, which was expressed in inhibition of the cleavage divisions, appearance of abnormal embryos, and developmental arrest. The addition of arachidonic acid amide and dopamine to the incubation medium weakened the effects of the inhibitors. Spiperone induced developmental defects in preimplantation mouse embryos and sea urchin embryos. Arachidonic acid amide with dopamine exerted a protective effect against spiperone when introduced to sea urchin embryos at the blastula or late gastrula stages, rather than after fertilization. In murine embryos, this amide induced developmental defects and arrest itself and its effect was reversible. Possible mechanisms underlying the effects of these drugs are discussed.

  31. Bezuglov V.V., Pashinnik V.E., Tovstenko V.I., Markovskiĭ L.N., Freĭmanis Ia.F., Serkov I.V. (2009). [Stereochemistry of substituting the allyl hydroxyl group with fluorine atom in prostaglandins. Synthesis of 15-fluoro-11,15-dideoxyprostaglandins E1]. Bioorg. Khim. 22 (10-11), 814–22 [+]

    (+/-)-15-Fluoro-11,15-dideoxyprostaglandin E1 and its methyl and ethyl esters were synthesized. Dehydroxyfluorination reaction (+/-)-11-deoxyprostaglandin E1 esters with various reagents based on SF4 was studied. Along with the target 15-fluorides (mixtures of alpha- and beta-epimers), products of allylic shift and dehydration in a ratio dependent on the fluorination agent were shown to be formed. With a morpholinotrifluorosulfuran-tris(morpholine)sulfonium trimethyldifluorosilicate mixture, the maximal excess (70%) of one of the 15-fluoro epimers was achieved. Possible mechanisms of dehydroxyfluorination of (+/-)-11-deoxyprostaglandin E1 esters with dialkylaminoflluorosulfurans were proposed. Methyl esters of 15-alpha-fluoro- and 15-beta-fluoro-11,15-dideoxyprostaglandin E1 exhibited moderate antiaggregation activity in rabbit platelet tests.

  32. Lakin K.M., Makarov V.A., Kovalev S.G., Petrukhina G.N., Serkov I.V., Golovanova N.K., Bezuglov V.V. (2009). [The effect of fluorinated prostaglandins on platelet aggregation]. Eksp Klin Farmakol 57 (2), 39–41 [+]

    The effects of fluorodeoxy prostanoids on platelet aggregability were studied. It was shown that introduction of fluorine into positions 9, 11 or 15 of prostaglandin F2 alpha led to enhanced proaggregation activity. The most active compound among fluorodeoxy analogs was 15-fluoro derivative; bisfluoro analog was moderately active, and 11-fluoro compound had the least activity. In the group of fluorodeoxy prostaglandins E2, a contrary effect was registered. Thus, the most active compound was 1-fluoride and the least, 15-fluoride. The incorporation of fluorine into position 15 of prostacyclin led to insignificantly lower antiaggregatory activity just as this modification of 6-keto-prostaglandin F1 alpha was accompanied by a dramatic increase in its ability to inhibit platelet aggregation.

  33. Nigamatov I.M., Makarov V.A., Serkov I.V., Gafurov R.G., Bezuglov V.V. (2009). [An 11-fluoroderivative of prostaglandin F2alpha--a stable thromboxano-mimetic]. Farmakol Toksikol 52 (4), 40–3 [+]

    In a search for stable thromboxanomimetics the action of 11-fluoro-11-deoxyprostaglandin F2 alpha (11-fluoro-PGF2 alpha) on the aggregatory properties of platelets and the tone of the vascular, respiratory and gastrointestinal smooth muscles was studied. It was found that 11-fluoro-PGF2 alpha, in contrast to PGF2 alpha, induces platelet aggregation and exhibits a high activity with respect to smooth muscles sensitive to thromboxane A2 alpha (TXA2) and a low activity for smooth muscles sensitive to PGF2 alpha 9-11-epoxyimino-PGH2 (a specific antagonist of TXA2 receptors) competitively blocked the action of 11-fluoro-PGF2 alpha on vascular smooth muscles. Thus, 11-fluoro-PGF2 exhibits properties of a thromboxanomimetic that makes it possible to propose it for modelling TXA2-induced biological effects.

  34. Buznikov G.A., Nikitina L.A., Bezuglov V.V., Milosević I., Lazarević L., Rogac L., Ruzdijić S., Slotkin T.A., Rakić L.M. (2008). Sea urchin embryonic development provides a model for evaluating therapies against beta-amyloid toxicity. Brain Res. Bull. 75 (1), 94–100 [+]

    Accumulation of beta-amyloid protein is an Alzheimer's disease hallmark but also may be mechanistically involved in neurodegeneration. One of its cleavage peptides, Abeta42, has been used to evaluate the mechanisms underlying amyloid-induced cytotoxicity and targeting of acetylcholine systems. We studied Sphaerechinus granularis sea urchin embryos which utilize acetylcholine and other neurotransmitters as morphogens. At a threshold concentration of 0.1 microM Abeta42, there was damage to the larval skeleton, accumulation of ectodermal cells in the blastocoele and underdevelopment of larval arms. Raising the Abeta42 concentration to 0.2-0.4 microM produced anomalies depending on the stage at which Abeta42 was introduced: at the first cleavage divisions, abnormalities appeared within 1-2 cell cycles; at the mid-blastula stage, the peak period of sensitivity to Abeta42, gastrulation was blocked; at later stages, there was progressive damage to the larval skeleton, digestive tract and larval spicules, as well as regression of larval arms. Each of these anomalies could be offset by the addition of lipid-permeable analogs of acetylcholine (arachidonoyl dimethylaminoethanol), serotonin (arachidonoyl serotonin) and cannabinoids (arachidonoyl vanillylamine), with the greatest activity exhibited by the acetylcholine analog. These results indicate that sea urchin embryos provide a model suitable to characterize the mechanisms underlying the cytotoxicity of Abeta42, as well as providing a system that enables the rapid screening of potential therapeutic interventions. The protection provided by neurotransmitter analogs, especially that for acetylcholine, points to unsuspected advantages of existing therapies that enhance cholinergic function, as well as indicating novel approaches that may prove protective in Alzheimer's disease.

  35. Lemak M.S., Bravarenko N.I., Bobrov M.Y., Bezuglov V.V., Ierusalimsky V.N., Storozhuk M.V., Malyshev A.Y., Balaban P.M. (2007). Cannabinoid regulation in identified synapse of terrestrial snail. Eur. J. Neurosci. 26 (11), 3207–14 [+]

    In the terrestrial snail a direct monosynaptic glutamatergic connection between the primary sensory neuron and a premotor interneuron involved in withdrawal behaviour can be functionally identified using electrophysiological techniques. We investigated the involvement of cannabinoids in regulation of this synaptic contact. The results demonstrate that the specific binding sites for agonists to mammalian type 1 cannabinoid receptors (CB1Rs) exist in the snail's nervous system. Application of a synthetic cannabinoid agonist anandamide selectively changed the efficacy of synaptic contacts between the identified neurons. A decrease in the long-term synaptic facilitation of the synaptic contact elicited by high-frequency nerve tetanization in the presence of cannabinoid agonist anandamide was observed, suggesting a possible role of endocannabinoids in regulation of plasticity at this synaptic site. The selective antagonist of CB1Rs [N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide] AM251 bath application was changing the efficacy of the synaptic contact only when the postsynaptic neuron had been intracellularly activated before its application. This observation implies an involvement of endocannabinoids in plasticity phenomena induced by activity in the postsynaptic target. Additional support of endocannabinoid involvement in synaptic function at this site was given by experiments in which AM251 blocked the short-term suppression of synaptic excitation evoked by low-frequency nerve tetanization, a phenomenon qualitatively similar to cannabinoid-dependent synaptically evoked suppression of excitation demonstrated in the mammalian nervous system. The results of the present study suggest an involvement of cannabinoids in the regulation of synaptic efficacy. Further, anandamide could be a candidate for an endogenous neuromessenger involved in plasticity processes.

  36. Buznikov G.A., Nikitina L.A., Rakić L.M., Milosević I., Bezuglov V.V., Lauder J.M., Slotkin T.A. (2007). The sea urchin embryo, an invertebrate model for mammalian developmental neurotoxicity, reveals multiple neurotransmitter mechanisms for effects of chlorpyrifos: therapeutic interventions and a comparison with the monoamine depleter, reserpine. Brain Res. Bull. 74 (4), 221–31 [+]

    Lower organisms show promise for the screening of neurotoxicants that might target mammalian brain development. Sea urchins use neurotransmitters as embryonic growth regulatory signals, so that adverse effects on neural substrates for mammalian brain development can be studied in this simple organism. We compared the effects of the organophosphate insecticide, chlorpyrifos in sea urchin embryos with those of the monoamine depleter, reserpine, so as to investigate multiple neurotransmitter mechanisms involved in developmental toxicity and to evaluate different therapeutic interventions corresponding to each neurotransmitter system. Whereas reserpine interfered with all stages of embryonic development, the effects of chlorpyrifos did not emerge until the mid-blastula stage. After that point, the effects of the two agents were similar. Treatment with membrane permeable analogs of the monoamine neurotransmitters, serotonin and dopamine, prevented the adverse effects of either chlorpyrifos or reserpine, despite the fact that chlorpyrifos works simultaneously through actions on acetylcholine, monoamines and other neurotransmitter pathways. This suggests that different neurotransmitters, converging on the same downstream signaling events, could work together or in parallel to offset the developmental disruption caused by exposure to disparate agents. We tested this hypothesis by evaluating membrane permeable analogs of acetylcholine and cannabinoids, both of which proved effective against chlorpyrifos- or reserpine-induced teratogenesis. Invertebrate test systems can provide both a screening procedure for mammalian neuroteratogenesis and may uncover novel mechanisms underlying developmental vulnerability as well as possible therapeutic approaches to prevent teratogenesis.

  37. Bobrov M.Y., Lyzhin A.A., Andrianova E.L., Gretskaya N.M., Zinchenko G.N., Frumkina L.E., Khaspekov L.G., Bezuglov V.V. (2006). Antioxidant and neuroprotective properties of N-docosahexaenoyl dopamine. Bull. Exp. Biol. Med. 142 (4), 425–7 [+]

    N-Docosahexaenoyl dopamine exhibited antioxidant activity in the test with a stable oxygen radical galvinoxyl. This compound produced a dose-dependent protective effect on cultured granular cells from rat cerebellum under conditions of oxidative stress. N-Docosahexaenoyl dopamine decelerated the development of symptoms of Parkinson's disease in mice receiving neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine.

  38. Kukushkin M.L., Igonkina S.I., Churyukanov M.V., Churyukanov V.V., Bobrov M.Y., Bezuglov V.V., Gretskaya N.V. (2006). Role of cannabinoid receptor agonists in mechanisms of suppression of central pain syndrome. Bull. Exp. Biol. Med. 142 (1), 39–42 [+]

    We studied the effect of cannabinoid receptor agonists anandamide and WIN 55,212-2 on the central pain syndrome induced by intraspinal injection of penicillin sodium salt in rats. Cannabinoids suppressed allodynia and spontaneous attacks in rats with the central pain syndrome. The analgesic effect was most pronounced after intrathecal injection of cannabinoid receptor agonist in a dose of 100 microg in 10 microl. After systemic treatment the analgesic effect was produced by only WIN 55,212-2 in a dose of 1 mg/kg. WIN 55,212-2 was superior to anandamide by the duration and intensity of the effect on allodynia and spontaneous attacks.

  39. Buznikov G.A., Nikitina L.A., Voronezhskaya E.E., Bezuglov V.V., DennisWillows A.O., Nezlin L.P. (2003). Localization of serotonin and its possible role in early embryos of Tritonia diomedea(Mollusca: Nudibranchia). Cell Tissue Res. 311 (2), 259–66 [+]

    A classical neurotransmitter serotonin (5-HT) was detected immunochemically using laser scanning microscopy at the early stages of Tritonia diomedea development. At the one- to eight-cell stages, immunolabeling suggested the presence of 5-HT in the cytoplasm close to the animal pole. At the morula and blastula stages, a group of micromeres at the animal pole showed immunoreactivity. At the gastrula stage no immunoreactive cells were detected, but they arose again at the early veliger stage. Antagonists of 5-HT(2) receptors, ritanserin and cyproheptadine, as well as lipophilic derivatives of dopamine blocked cleavage divisions or distorted their normal pattern. These effects were prevented by 5-HT and its highly lipophilic derivates, serotoninamides of polyenoic fatty acids, but not by the hydrophilic (quaternary) analog of 5-HT, 5-HTQ. The results confirm our earlier suggestion that endogenous 5-HT in pre-nervous embryos acts as a regulator of cleavage divisions in nudibranch molluscs.

  40. Buznikov G.A., Bezuglov V.V., Nikitina L.A., Slotkin T.A., Lauder J.M. (2001). [Cholinergic regulation of the sea urchin embryonic and larval development]. Ross Fiziol Zh Im I M Sechenova 87 (11), 1548–56 [+]

    Choline esters of polyenoic fatty acids block cleavage divisions of sea urchins and evoke the formation of one-cell multinuclear embryos. If the fatty acids AA-Ch or DHA-Ch are added at the mid or late blastula stage, many cells are extruded, forming extra-embryonic cell clusters near the animal pole of embryos or larvae. Both effects are prevented by dimethylaminoethyl esters of polyenoic fatty acids (AA-DMAE or DHA-DMAE) or their 5-hydroxytryptamides. Nicotinic acetylcholine receptor antagonists, imechine, d-tubocurarine or QX-222 provide partial protection against AA-Ch or DHA-Ch. The organophosphate pesticide, chlorpyrifos, or a combination of (-)-nicotine + phorbol 12-myristate 13-acetate, also evoke the mass extrusion of transformed embryonic cells at the animal pole of larvae. These effects are similarly antagonized by AA-DMAE, DHA-DMAE, or fatty acids 5-hydroxytryptamides. Taking together, these results suggest that AA-Ch and DHA-Ch act on sea urchin embryos and larvae as agonists of acetylcholine receptors, whereas AA-DMAE and DHA-DMAE act as antagonists. The ability of fatty acids 5-hydroxytryptamides to prevent the effects of AA-Ch or DHA-Ch may be due to restoration of the normal dynamic balance of cholinergic and serotonergic signaling during cleavage divisions and gastrulation.

  41. Buznikov G.A., Nikitina L.A., Bezuglov V.V., Lauder J.M., Padilla S., Slotkin T.A. (2001). An invertebrate model of the developmental neurotoxicity of insecticides: effects of chlorpyrifos and dieldrin in sea urchin embryos and larvae. Environ. Health Perspect. 109 (7), 651–61 [+]

    Chlorpyrifos targets mammalian brain development through a combination of effects directed at cholinergic receptors and intracellular signaling cascades that are involved in cell differentiation. We used sea urchin embryos as an invertebrate model system to explore the cellular mechanisms underlying the actions of chlorpyrifos and to delineate the critical period of developmental vulnerability. Sea urchin embryos and larvae were exposed to chlorpyrifos at different stages of development ranging from early cell cleavages through the prism stage. Although early cleavages were unaffected even at high chlorpyrifos concentrations, micromolar concentrations added at the mid-blastula stage evoked a prominent change in cell phenotype and overall larval structure, with appearance of pigmented cells followed by their accumulation in an extralarval cap that was extruded from the animal pole. At higher concentrations (20-40 microM), these abnormal cells constituted over 90% of the total cell number. Studies with cholinergic receptor blocking agents and protein kinase C inhibitors indicated two distinct types of effects, one mediated through stimulation of nicotinic cholinergic receptors and the other targeting intracellular signaling. The effects of chlorpyrifos were not mimicked by chlorpyrifos oxon, the active metabolite that inhibits cholinesterase, nor by nonorganophosphate cholinesterase inhibitors. Dieldrin, an organochlorine that targets GABA(A )receptors, was similarly ineffective. The effects of chlorpyrifos and its underlying cholinergic and signaling-related mechanisms parallel prior findings in mammalian embryonic central nervous system. Invertebrate test systems may thus provide both a screening procedure for potential neuroteratogenesis by organophosphate-related compounds, as well as a system with which to uncover novel mechanisms underlying developmental vulnerability.

  42. Bisogno T., Melck D., BobrovMYu , Gretskaya N.M., Bezuglov V.V., DePetrocellis L., DiMarzo V. (2000). N-acyl-dopamines: novel synthetic CB(1) cannabinoid-receptor ligands and inhibitors of anandamide inactivation with cannabimimetic activity in vitro and in vivo. Biochem. J. 351 Pt 3, 817–24 [+]

    We reported previously that synthetic amides of polyunsaturated fatty acids with bioactive amines can result in substances that interact with proteins of the endogenous cannabinoid system (ECS). Here we synthesized a series of N-acyl-dopamines (NADAs) and studied their effects on the anandamide membrane transporter, the anandamide amidohydrolase (fatty acid amide hydrolase, FAAH) and the two cannabinoid receptor subtypes, CB(1) and CB(2). NADAs competitively inhibited FAAH from N18TG2 cells (IC(50)=19-100 microM), as well as the binding of the selective CB(1) receptor ligand, [(3)H]SR141716A, to rat brain membranes (K(i)=250-3900 nM). The arachidonoyl (20:4 omega 6), eicosapentaenoyl (20:5 omega 3), docosapentaenoyl (22:5 omega 3), alpha-linolenoyl (18:3 omega 3) and pinolenoyl (5c,9c,12c 18:3 omega 6) homologues were also found to inhibit the anandamide membrane transporter in RBL-2H3 basophilic leukaemia and C6 glioma cells (IC(50)=17.5-33 microM). NADAs did not inhibit the binding of the CB(1)/CB(2) receptor ligand, [(3)H]WIN55,212-2, to rat spleen membranes (K(i)>10 microM). N-arachidonyl-dopamine (AA-DA) exhibited 40-fold selectivity for CB(1) (K(i)=250 nM) over CB(2) receptors, and N-docosapentaenoyl-dopamine exhibited 4-fold selectivity for the anandamide transporter over FAAH. AA-DA (0.1-10 microM) did not displace D1 and D2 dopamine-receptor high-affinity ligands from rat brain membranes, thus suggesting that this compound has little affinity for these receptors. AA-DA was more potent and efficacious than anandamide as a CB(1) agonist, as assessed by measuring the stimulatory effect on intracellular Ca(2+) mobilization in undifferentiated N18TG2 neuroblastoma cells. This effect of AA-DA was counteracted by the CB(1) antagonist SR141716A. AA-DA behaved as a CB(1) agonist in vivo by inducing hypothermia, hypo-locomotion, catalepsy and analgesia in mice (1-10 mg/kg). Finally, AA-DA potently inhibited (IC(50)=0.25 microM) the proliferation of human breast MCF-7 cancer cells, thus behaving like other CB(1) agonists. Also this effect was counteracted by SR141716A but not by the D2 antagonist haloperidol. We conclude that NADAs, and AA-DA in particular, may be novel and useful probes for the study of the ECS.

  43. Buznikov G.A., Bezuglov V.V. (2000). [Polyenic acid 5-hydroxytryptamides and 3-hydroxytyramides as tools for studying of the pre-nervous biogenic monoamine functions]. Ross Fiziol Zh Im I M Sechenova 86 (9), 1093–108 [+]

    Three main effects of the amides on embryos of opistobranch molluscs, sea urchins and starfish, were revealed. First, a rather independent and clear protective action of 5-HYDROXYTRYPTAMIDES AND 3-HYDROXYTYRAMIDES against cytostatic antagonists of serotonin and dopamine, resp. Second, prevention of developmental abnormalities induced by protein kinase C activators both by hydroxytryptamides and hydroxytyramides. Third, the cytostatic effect of 3-HYDROXYTRIPTAMIDES eliminated or prevented by 5-HYDROXYTRYPTAMIDES. These effects quantitatively depended on the structure of their fatty acids part. Some functionally active regulatory substances similar to 5-HYDROXYTRYPTAMIDES AND 3-HYDROXYTYRAMIDES may exist in the early embryos.

  44. Bezuglov V.V., Andreyuk G.M., Serkov I.V., Kisel M.A. (2000). Influence of dinitroglycerol and nitroethyleneglycol lipid derivatives on spectral parameters of human hemoglobin. Biochemistry Mosc. 65 (6), 685–9 [+]

    The interaction of organic nitrates (nitroethyleneglycol, dinitroglycerol, and their esters with arachidonic acid) with oxyhemoglobin and methemoglobin has been studied. Addition of nitroethyleneglycol and dinitroglycerol to oxyhemoglobin is accompanied by a modest but significant increase in oxidation rate of the heme protein to the high-spin ferri-form--methemoglobin. Arachidonoylglycerol dinitrate exerts a similar but more pronounced effect on hemoglobin: a molar excess of this dinitrate induces the transformation of a significant portion of oxyhemoglobin to methemoglobin, whereas arachidonoylnitroethyleneglycol is inactive. Arachidonoylglycerol dinitrate also induces changes in the spectral characteristics of methemoglobin; this may be due to disintegration of the methemoglobin with the loss of heme. The data demonstrate that some organic nitrates can interact with hemoglobin; this should be taken into account when using the oxyhemoglobin technique for measuring nitric oxide generation from these compounds.

  45. Bobrov M.Y., Shevchenko V.P., Yudushkin I.A., Rogov S.I., Remov M.N., FominaAgeeva E.V., Gretskaya N.M., Nagaev I.Y., Kuklev D.V., Bezuglov V.V. (2000). Hydrolysis of anandamide and eicosapentaenoic acid ethanolamide in mouse splenocytes. Biochemistry Mosc. 65 (5), 615–9 [+]

    The hydrolysis of anandamide has been studied in mouse splenocytes using tritiated anandamide analogs labeled in the acyl- or ethanolamide parts of the molecule. [3H]Anandamide undergoes rapid (t(1/2) = 2.5 min) uptake and hydrolysis, yielding ethanolamine and arachidonic acid. The anandamide hydrolysis in splenocytes is sensitive to inhibition by phenylmethylsulfonyl fluoride, and it is assumed that the observed activity is due to fatty acid amide hydrolase, which inactivates anandamide in central and peripheral tissues. Eicosapentaenoic acid ethanolamide and the 15-hydroxy-derivative of anandamide are shown to be amidohydrolase substrates as well. The fatty acids derived from hydrolytic cleavage of acylethanolamines undergo rapid oxidation by splenocyte lipoxygenase, yielding the corresponding 12-hydroxy-derivatives. Oxygenated ethanolamide derivatives were not found. The data suggest that polyenoic fatty acid ethanolamides are metabolic precursors of eicosanoids in splenocytes and that amide bond hydrolysis is the key point in switching of biological activity spectra between endocannabinoids and oxylipins.

  46. Bisogno T., Melck D., DePetrocellis L., BobrovMYu , Gretskaya N.M., Bezuglov V.V., Sitachitta N., Gerwick W.H., DiMarzo V. (1998). Arachidonoylserotonin and other novel inhibitors of fatty acid amide hydrolase. Biochem. Biophys. Res. Commun. 248 (3), 515–22 [+]

    Fatty acid amide hydrolase (FAAH) catalyzes the hydrolysis of bioactive fatty acid amides and esters such as the endogenous cannabinoid receptor ligands, anandamide (N-arachidonoyl-ethanolamine) and 2-arachidonoylglycerol, and the putative sleep inducing factor cis-9-octadecenoamide (oleamide). Most FAAH blockers developed to date also inhibit cytosolic phospholipase A2 (cPLA2) and/or bind to the CB1 cannabinoid receptor subtype. Here we report the finding of four novel FAAH inhibitors, two of which, malhamensilipin A and grenadadiene, were screened out of a series of thirty-two different algal natural products, and two others, arachidonoylethylene glycol (AEG) and arachidonoyl-serotonin (AA-5-HT) were selected out of five artificially functionalized polyunsaturated fatty acids. When using FAAH preparations from mouse neuroblastoma N18TG2 cells and [14C]anandamide as a substrate, the IC50s for these compounds ranged from 12.0 to 26 microM, the most active compound being AA-5-HT. This substance was also active on FAAH from rat basophilic leukaemia (RBL-2H3) cells (IC50 = 5.6 microM), and inhibited [14C]anandamide hydrolysis by both N18TG2 and RBL-2H3 intact cells without affecting [14C]anandamide uptake. While AEG behaved as a competitive inhibitor and was hydrolyzed to arachidonic acid (AA) by FAAH preparations, AA-5-HT was resistant to FAAH-catalyzed hydrolysis and behaved as a tight-binding, albeit non-covalent, mixed inhibitor. AA-5-HT did not interfere with cPLA2-mediated, ionomycin or antigen-induced release of [3H]AA from RBL-2H3 cells, nor with cPLA2 activity in cell-free experiments. Finally, AA-5-HT did not activate CB1 cannabinoid receptors since it acted as a very weak ligand in in vitro binding assays, and, at 10-15 mg/kg body weight, it was not active in the 'open field', 'hot plate' and rectal hypothermia tests carried out in mice. Conversely AEG behaved as a cannabimimetic substance in these tests as well as in the 'ring' immobility test where AA-5-HT was also active. AA-5-HT is the first FAAH inhibitor reported to date which is inactive both against cPLA2 and at CB1 receptors, whereas AEG represents a new type of cannabinoid receptor agonist.

  47. Bezuglov V.V., BobrovMYu , Archakov A.V. (1998). Bioactive amides of fatty acids. Biochemistry Mosc. 63 (1), 22–30 [+]

    Amides of fatty acids are lipid bioregulators formed from long chain saturated and unsaturated fatty acids via amidation by the corresponding amines. Ethanolamides of fatty acids are the most well-studied species of this group; an alternative pathway for their biosynthesis includes hydrolysis of N-acylated phosphatidylethanolamines by phospholipase D. Ethanolamides of fatty acids bind to the cannabinoid receptors of the central nervous system (CB1) or peripheral tissues (CB2) and can be considered as endogenous ligands of these receptors. Their pharmacological properties are similar to that of cannabimimetics. Simple amides of fatty acids are also endogenous bioregulators acting like sleep-inducing (oleamide) or angiogenic factors (erucamide). A new group of bioregulators comprise the amides of fatty acids and biologically active amines (vanillinamine, dopamine, and serotonin).

  48. Kogteva G.S., Bezuglov V.V. (1998). Unsaturated fatty acids as endogenous bioregulators. Biochemistry Mosc. 63 (1), 4–12 [+]

    Most biological functions of unsaturated fatty acids are due to their ability to act as second messengers or modulators of activities of functionally important proteins; these functions are not related to oxidative metabolism of unsaturated fatty acids. These acids regulate the activity of phospholipases, ion channels, ATPases, G-proteins, and protein kinases; they also modulate the phosphoinositide and sphingomyelin cycles, the transfer of hormonal information, and gene transcription. The great diversity of effects of unsaturated fatty acids and their presence at the earliest stages of evolution suggest for these bioregulators a system-forming role in the living body.

  49. Bezuglov V.V., Manevich E.M., Archakov A.V., Bobrov M.I.u., Kuklev D.V., Petrukhina G.N., Makarov V.A., Buznikov G.A. (1997). [Artificially functionalized polyenoic fatty acids--a new lipid bioregulators]. Bioorg. Khim. 23 (3), 211–20 [+]

    Dopamine, histamine, serotonin, and serotonin analogs were acylated with arachidonic and eicosapentaenoic acids, and the reaction products were named as artificially functionalized fatty acids (AFFA). The amides of arachidonic acid with serotonin, dopamine, and histamine were found to inhibit human platelet aggregation induced by ADP, arachidonic acid and adrenaline. Amides of arachidonic and eicosapentaeonic acids with serotonin and dopamine protect sea urchin early embryos against cytotoxic action of serotonin and histamine antagonists. These effects are not connected with the possible hydrolytic cleavage of AFFA to their constituent polyenoic fatty acids and amines. Arachidonic acid dopaminamide was shown to be a substrate of soybean 15-lipoxygenase, whereas the arachidonic acid amides with serotonin and its derivatives were resistant to this enzyme. Moreover, arachidonic acid serotoninamide turned out to be an irreversible lipoxygenase inhibitor. Considerable amount of hydroxyl radicals (fluorescent assay) were found for the first time to accompany lipoxygenase oxidation of linoleic acid; arachidonic acid serotoninamide blocked this process completely. Therefore, it was concluded that AFFA possess specific biological activity and can be considered as a novel group of lipid bioregulators.

  50. Bezuglov V.V., FominaAgeeva E.V., Tekieva E.A., Diatlovitskaia E.V. (1994). [Lipoxygenase oxidation of arachidonic acid in murine splenocytes and its modulation by the lactone ganglioside GM3]. Biokhimiia 59 (9), 1360–8 [+]

    The main arachidonic acid metabolites released into the medium by mouse splenocytes have been identified on the basis of chromatographic and spectral studies as well as by mass spectrometry of the derivatives. In the absence or presence of exogenous arachidonic acid mouse splenocytes produce mainly 12-hydroxy-5,8,10,14-eicosatetraenoic and 12,20-dihydroxy-5,8,10,14-eicosatetraenoic acids. Both products are constantly released by intact cells into surrounding media without stimulation by exogenous substrate or other modulators. For the first time it is shown that exogenously added ganglioside GM3 lactone as well as ganglioside GM3 itself can influence the arachidonic acid metabolism in splenocytes.

  51. Kuklev D.V., Rybin V.G., Imbs A.B., Bezuglov V.V. (1993). [Synthesis of monoepoxides of arachidonic, eicosapentaenoic, and docosahexaenoic acids]. Bioorg. Khim. 19 (11), 1122–7 [+]

    A practical synthesis of cis-monoepoxides from free arachidonic 20:4 (n-6), eicosapentaenoic 20:5 (n-3) and docosahexaenoic 22:6 (n-3) acids, their cytochrome P-450 epoxygenase metabolites, is described. The free polyunsaturated fatty acids (PUFA) were oxidized each by 1.25 eq. m-chloroperbenzoic acid in ethanol to give a mixture of PUFAs' mono-epoxy derivatives (45%) which was separated by HPLC and the individual isomers were characterised by mass spectrometry. All regioisomers of the free PUFAs' cis-monoepoxides were thus obtained.

  52. Gukovskaya A.S., Trepakova E.S., Zinchenko V.P., Korystov Y.N., Bezuglov V.V. (1992). Effect of the sulfhydryl reagent thimerosal on cytosolic free Ca2+ and membrane potential of thymocytes. Biochim. Biophys. Acta 1111 (1), 65–74 [+]

    The sulfhydryl reagent thimerosal at concentrations 5-100 microM has been found to induce a variety of changes in ion transport in rat thymocytes. In particular, [Ca2+]i increases about 10-fold from the basal level. The [Ca2+]i response to thimerosal displays a two-stage time course, with the main [Ca2+]i rise during the second stage. Evidence has been obtained for the depletion of intracellular Ca2+ pools in thimerosal-treated cells, however, Ca2+ mobilization from intracellular stores does not contribute significantly into [Ca2+]i rise. Thimerosal elicits permeability not only for Ca2+, but also for Mn2+ and Ni2+, which is Ca(2+)-dependent. We failed to get any evidence on thimerosal-induced inhibition of the plasma membrane Ca(2+)-ATPase. The induction of Ca2+ influx, rather than inhibition of Ca(2+)-ATPase, accounts for the disturbance of [Ca2+]i homeostasis in thimerosal-treated cells. Thimerosal also elicits changes in monovalent ion fluxes resulting in marked depolarization. The latter seems unrelated to the changes in [Ca2+]i and is suggested to be mediated both by increased permeability for Na+ and a decreased one for K+. Thimerosal significantly stimulates AA release from thymocytes. Evidence has been presented that AA metabolite(s), probably, LO product(s), may mediate the changes in the transport of mono- and divalent cations elicited by the sulfhydryl reagent. Prolonged treatment of thymocytes with thimerosal resulted in cell death.

  53. Sadovskaia V.L., Rakitin L.I.u., Golovanova N.K., Kogtev L.S., Bezuglov V.V. (1986). [Study of prostaglandins and thromboxanes B2 using mass-spectrometry of secondary ions]. Bioorg. Khim. 12 (7), 956–60 [+]

    Prostaglandins E, F, I2 and thromboxane B2 have been studied by secondary ion mass spectrometry. It is shown that the method is suitable for direct identification of these compounds either as free acids or as their sodium salts. The spectra of the former reveal their structural features, while with the latter information on the molecular weight can be obtained. The limit of detection (about 1 microgram) allows the analysis of prostaglandin solutions of 1 microgram/microliter concentrations used in pharmacological tests.


Vladimir Bezuglov

New fluorescent analogues of acyldopamines with the preserved the address moiety (2017-11-27)

New fluorescent analogues of natural acyldopamines mimicking residues of palmitic and oleic acids carrying the reporter BODIPY group at the distal end of molecule were synthesized. The specific uptake of mentioned compounds in rat pheochromocytoma PC12 cells was demonstrated and characteristics of this process were measured. Uptake rate of oleic analogue was more than three times as palmitic one. Synthesized compounds are ready to be used for investigation of acyldopamine transport in various cells of human or animal origin.

Modified peptides based on proglyprol structure with increased effectiveness (2016-03-29)

Hybrid compounds based on proglyprol (PGP) peptide carrying residues of docosahexaenoic acid and (or) dopamine were synthesized for the first time. Prepared compounds possess advanced neuroprotectory  and anti-inflammatory activity and may consider as prototypes of novel pharmaceuticals.


  1. Akimov M.G., Gretskaya N.M., Zinchenko G.N., Bezuglov V.V. (2015). Cytotoxicity of Endogenous Lipids N-acyl Dopamines and their Possible Metabolic Derivatives for Human Cancer Cell Lines of Different Histological Origin. Anticancer Res. 35 (5), 2657–61 [+]

    Dopamine amides of long chain fatty acids are a family of endogenous mammalian lipids with an unknown function; they are anti-proliferative for the C6 glioblastoma cell line. To assess their possible anti-cancer activity we evaluated their cytotoxicity for a set of cancer cell lines.