Science.gov

Sample records for dual receptor mechanisms

  1. Nuclear Receptor SHP Activates miR-206 Expression via a Cascade Dual Inhibitory Mechanism

    PubMed Central

    Song, Guisheng; Wang, Li

    2009-01-01

    MicroRNAs play a critical role in many essential cellular functions in the mammalian species. However, limited information is available regarding the regulation of miRNAs gene transcription. Microarray profiling and real-time PCR analysis revealed a marked down-regulation of miR-206 in nuclear receptor SHP−/− mice. To understand the regulatory function of SHP with regard to miR-206 gene expression, we determined the putative transcriptional initiation site of miR-206 and also its full length primary transcript using a database mining approach and RACE. We identified the transcription factor AP1 binding sites on the miR-206 promoter and further showed that AP1 (c-Jun and c-Fos) induced miR-206 promoter transactivity and expression which was repressed by YY1. ChIP analysis confirmed the physical association of AP1 (c-Jun) and YY1 with the endogenous miR-206 promoter. In addition, we also identified nuclear receptor ERRγ (NR3B3) binding site on the YY1 promoter and showed that YY1 promoter was transactivated by ERRγ, which was inhibited by SHP (NROB2). ChIP analysis confirmed the ERRγ binding to the YY1 promoter. Forced expression of SHP and AP1 induced miR-206 expression while overexpression of ERRγ and YY1 reduced its expression. The effects of AP1, ERRγ, and YY1 on miR-206 expression were reversed by siRNA knockdown of each gene, respectively. Thus, we propose a novel cascade “dual inhibitory” mechanism governing miR-206 gene transcription by SHP: SHP inhibition of ERRγ led to decreased YY1 expression and the de-repression of YY1 on AP1 activity, ultimately leading to the activation of miR-206. This is the first report to elucidate a cascade regulatory mechanism governing miRNAs gene transcription. PMID:19721712

  2. Molecular Docking and Prediction of Pharmacokinetic Properties of Dual Mechanism Drugs that Block MAO-B and Adenosine A(2A) Receptors for the Treatment of Parkinson's Disease.

    PubMed

    Azam, Faizul; Madi, Arwa M; Ali, Hamed I

    2012-07-01

    Monoamine oxidase B (MAO-B) inhibitory potential of adenosine A(2A) receptor (AA(2A)R) antagonists has raised the possibility of designing dual-target-directed drugs that may provide enhanced symptomatic relief and that may also slow the progression of Parkinson's disease (PD) by protecting against further neurodegeneration. To explain the dual inhibition of MAO-B and AA(2A)R at the molecular level, molecular docking technique was employed. Lamarckian genetic algorithm methodology was used for flexible ligand docking studies. A good correlation (R(2)= 0.524 and 0.627 for MAO-B and AA(2A)R, respectively) was established between docking predicted and experimental K(i) values, which confirms that the molecular docking approach is reliable to study the mechanism of dual interaction of caffeinyl analogs with MAO-B and AA(2A)R. Parameters for Lipinski's "Rule-of-Five" were also calculated to estimate the pharmacokinetic properties of dual-target-directed drugs where both MAO-B inhibition and AA(2A)R antagonism exhibited a positive correlation with calculated LogP having a correlation coefficient R(2) of 0.535 and 0.607, respectively. These results provide some beneficial clues in structural modification for designing new inhibitors as dual-target-directed drugs with desired pharmacokinetic properties for the treatment of PD.

  3. Biomembrane and receptor mechanisms

    SciTech Connect

    Chapman, D.; Bertoli, E.

    1987-01-01

    This book cover the reviews on biomembrane dynamics; recent spectroscopic studies. Topics covered are freeze fracture: Seeing and thinking biological membranes, membrane proteins and receptors: structure and organisation; techniques to determine the transbilayer distribution and mobility of phospholipids in biological membranes, transbilayer organisation of phospholipids in the plasma membranes of pro-erythroblasts and normal and abnormal red cells, aminophospholipid translocation in the erythroctye membrane is mediated by a specific AIP-dependent enzyme; membrane protein interactions, lipid-protein interactions: selectively and receptor binding, membrane fluidity in the regulation of membrane-linked enzymes, the lipid regulation of receptor functions, microheterogencity of biological membrane: structural and functional implications, fusion-fission reactions in biological membranes and in phospholpid bilayers, methods for studying the structure and function of the mitochondrial uncoupling protein, methods for studying metabolite transport in mitochondria, transport of metabolites in mitochondria, membrane gangliosides and allied glycosphingolipids: Biochemical features and physicochemical properties, the use of merocyanine 540 for monitoring aggregation properties of sialogangliosides in solution, hormone reception at the cell surface - an overview, double role for GIP in the stimulus secretion sequence of mast cells and neurophils, tumor promoters and hormone receptor coupling mechanisms in the anterior pituitary. The regulation of hormone-dependent adenylate cyclase in native membranes and systems reconstituted from purified components.- Immunological tools for the study of plasma membrane receptors.

  4. Control of gravitropic orientation. II. Dual receptor model for gravitropism

    NASA Technical Reports Server (NTRS)

    LaMotte, Clifford E.; Pickard, Barbara G.

    2004-01-01

    Gravitropism of vascular plants has been assumed to require a single gravity receptor mechanism. However, based on the evidence in Part I of this study, we propose that maize roots require two. The first mechanism is without a directional effect and, by itself, cannot give rise to tropism. Its role is quantitative facilitation of the second mechanism, which is directional like the gravitational force itself and provides the impetus for tropic curvature. How closely coupled the two mechanisms may be is, as yet, unclear. The evidence for dual receptors supports a general model for roots. When readiness for gravifacilitation, or gravifacilitation itself, is constitutive, orthogravitropic curvature can go to completion. If not constitutively enabled, gravifacilitation can be weak in the absence of light and water deficit or strong in the presence of light and water deficit. In either case, it can decay and permit roots to assume reproducible non-vertical orientations (plagiogravitropic or plagiotropic orientations) without using non-vertical setpoints. In this way roots are deployed in a large volume of soil. Gravitropic behaviours in shoots are more diverse than in roots, utilising oblique and horizontal as well as vertical setpoints. As a guide to future experiments, we assess how constitutive v. non-constitutive modes of gravifacilitation might contribute to behaviours based on each kind of setpoint.

  5. Tetrathiafulvalene diindolylquinoxaline: a dual signaling anion receptor with phosphate selectivity†

    PubMed Central

    Bejger, Christopher; Park, Jung Su; Silver, Eric S.; Sessler, Jonathan L.

    2011-01-01

    Incorporation of tetrathiafulvalene into the backbone of a known neutral phosphate receptor, diindolylquinoxaline, yields a dual optical-electrochemical chemosensor for dihydrogen phosphate that functions in dichloromethane. This system shows selectivity for dihydrogen phosphate over other small anions and can be used to detect the presence of this analyte via fluorescence quenching or cyclic voltammetry. PMID:20856940

  6. Quantum mechanics/molecular mechanics dual Hamiltonian free energy perturbation

    NASA Astrophysics Data System (ADS)

    Polyak, Iakov; Benighaus, Tobias; Boulanger, Eliot; Thiel, Walter

    2013-08-01

    The dual Hamiltonian free energy perturbation (DH-FEP) method is designed for accurate and efficient evaluation of the free energy profile of chemical reactions in quantum mechanical/molecular mechanical (QM/MM) calculations. In contrast to existing QM/MM FEP variants, the QM region is not kept frozen during sampling, but all degrees of freedom except for the reaction coordinate are sampled. In the DH-FEP scheme, the sampling is done by semiempirical QM/MM molecular dynamics (MD), while the perturbation energy differences are evaluated from high-level QM/MM single-point calculations at regular intervals, skipping a pre-defined number of MD sampling steps. After validating our method using an analytic model potential with an exactly known solution, we report a QM/MM DH-FEP study of the enzymatic reaction catalyzed by chorismate mutase. We suggest guidelines for QM/MM DH-FEP calculations and default values for the required computational parameters. In the case of chorismate mutase, we apply the DH-FEP approach in combination with a single one-dimensional reaction coordinate and with a two-dimensional collective coordinate (two individual distances), with superior results for the latter choice.

  7. Investigation of orexin-2 selective receptor antagonists: Structural modifications resulting in dual orexin receptor antagonists.

    PubMed

    Skudlarek, Jason W; DiMarco, Christina N; Babaoglu, Kerim; Roecker, Anthony J; Bruno, Joseph G; Pausch, Mark A; O'Brien, Julie A; Cabalu, Tamara D; Stevens, Joanne; Brunner, Joseph; Tannenbaum, Pamela L; Wuelfing, W Peter; Garson, Susan L; Fox, Steven V; Savitz, Alan T; Harrell, Charles M; Gotter, Anthony L; Winrow, Christopher J; Renger, John J; Kuduk, Scott D; Coleman, Paul J

    2017-03-15

    In an ongoing effort to explore the use of orexin receptor antagonists for the treatment of insomnia, dual orexin receptor antagonists (DORAs) were structurally modified, resulting in compounds selective for the OX2R subtype and culminating in the discovery of 23, a highly potent, OX2R-selective molecule that exhibited a promising in vivo profile. Further structural modification led to an unexpected restoration of OX1R antagonism. Herein, these changes are discussed and a rationale for selectivity based on computational modeling is proposed.

  8. Mechanism for the activation of glutamate receptors

    Cancer.gov

    Scientists at the NIH have used a technique called cryo-electron microscopy to determine a molecular mechanism for the activation and desensitization of ionotropic glutamate receptors, a prominent class of neurotransmitter receptors in the brain and spina

  9. Identification and mechanism of ABA receptor antagonism

    SciTech Connect

    Melcher, Karsten; Xu, Yong; Ng, Ley-Moy; Zhou, X. Edward; Soon, Fen-Fen; Chinnusamy, Viswanathan; Suino-Powell, Kelly M; Kovach, Amanda; Tham, Fook S.; Cutler, Sean R.; Li, Jun; Yong, Eu-Leong; Zhu, Jian-Kang; Xu, H. Eric

    2010-11-11

    The phytohormone abscisic acid (ABA) functions through a family of fourteen PYR/PYL receptors, which were identified by resistance to pyrabactin, a synthetic inhibitor of seed germination. ABA activates these receptors to inhibit type 2C protein phosphatases, such as ABI1, yet it remains unclear whether these receptors can be antagonized. Here we demonstrate that pyrabactin is an agonist of PYR1 and PYL1 but is unexpectedly an antagonist of PYL2. Crystal structures of the PYL2-pyrabactin and PYL1-pyrabactin-ABI1 complexes reveal the mechanism responsible for receptor-selective activation and inhibition, which enables us to design mutations that convert PYL1 to a pyrabactin-inhibited receptor and PYL2 to a pyrabactin-activated receptor and to identify new pyrabactin-based ABA receptor agonists. Together, our results establish a new concept of ABA receptor antagonism, illustrate its underlying mechanisms and provide a rational framework for discovering novel ABA receptor ligands.

  10. Smart dual-functional warhead for folate receptor-specific activatable imaging and photodynamic therapy.

    PubMed

    Kim, Jisu; Tung, Ching-Hsuan; Choi, Yongdoo

    2014-09-21

    A smart dual-targeted theranostic agent becomes highly fluorescent and phototoxic only when its linker is cleaved by tumor-associated lysosomal enzyme cathepsin B after internalization into folate receptor-positive cancer cells.

  11. Dual-action expanded-latch mechanism

    NASA Technical Reports Server (NTRS)

    Spencer, R. A.; Tewell, J. R.; Tobey, W. H.

    1978-01-01

    Single drive actuator operates novel mechanism that expands, attaches to object, and withdraws to latch object firmly to another part. Packaging is extremely simple and compact, and eliminates need for machined parts or close tolerances.

  12. Dual Modulators of GABA-A and Alpha7 Nicotinic Receptors for Treating Autism

    DTIC Science & Technology

    2014-08-01

    and Alpha7 Nicotinic Receptors for Treating Autism PRINCIPAL INVESTIGATOR: Kelvin W. Gee RECIPIENT: University of California Irvine...Aug 2014 4. TITLE AND SUBTITLE Dual Modulators of GABA-A and Alpha7 Nicotinic Receptors for Treating Autism 5a. CONTRACT NUMBER 5b. GRANT NUMBER...DISTRIBUTION / AVAILABILITY STATEMENT Approved for Public Release; Distribution Unlimited 13. SUPPLEMENTARY NOTES 14. ABSTRACT Autism

  13. Design and synthesis of dual 5-HT1A and 5-HT7 receptor ligands.

    PubMed

    Ofori, Edward; Zhu, Xue Y; Etukala, Jagan R; Peprah, Kwakye; Jordan, Kamanski R; Adkins, Adia A; Bricker, Barbara A; Kang, Hye J; Huang, Xi-Ping; Roth, Bryan L; Ablordeppey, Seth Y

    2016-08-15

    5-HT1A and 5-HT7 receptors have been at the center of discussions recently due in part to their major role in the etiology of major central nervous system diseases such as depression, sleep disorders, and schizophrenia. As part of our search to identify dual targeting ligands for these receptors, we have carried out a systematic modification of a selective 5HT7 receptor ligand culminating in the identification of several dual 5-HT1A and 5-HT7 receptor ligands. Compound 16, a butyrophenone derivative of tetrahydroisoquinoline (THIQ), was identified as the most potent agent with low nanomolar binding affinities to both receptors. Interestingly, compound 16 also displayed moderate affinity to other clinically relevant dopamine receptors. Thus, it is anticipated that compound 16 may serve as a lead for further exploitation in our quest to identify new ligands with the potential to treat diseases of CNS origin.

  14. Glucagon-Like Peptide 1/Glucagon Receptor Dual Agonism Reverses Obesity in Mice

    PubMed Central

    Pocai, Alessandro; Carrington, Paul E.; Adams, Jennifer R.; Wright, Michael; Eiermann, George; Zhu, Lan; Du, Xiaobing; Petrov, Aleksandr; Lassman, Michael E.; Jiang, Guoqiang; Liu, Franklin; Miller, Corey; Tota, Laurie M.; Zhou, Gaochao; Zhang, Xiaoping; Sountis, Michael M.; Santoprete, Alessia; Capito', Elena; Chicchi, Gary G.; Thornberry, Nancy; Bianchi, Elisabetta; Pessi, Antonello; Marsh, Donald J.; SinhaRoy, Ranabir

    2009-01-01

    OBJECTIVE Oxyntomodulin (OXM) is a glucagon-like peptide 1 (GLP-1) receptor (GLP1R)/glucagon receptor (GCGR) dual agonist peptide that reduces body weight in obese subjects through increased energy expenditure and decreased energy intake. The metabolic effects of OXM have been attributed primarily to GLP1R agonism. We examined whether a long acting GLP1R/GCGR dual agonist peptide exerts metabolic effects in diet-induced obese mice that are distinct from those obtained with a GLP1R-selective agonist. RESEARCH DESIGN AND METHODS We developed a protease-resistant dual GLP1R/GCGR agonist, DualAG, and a corresponding GLP1R-selective agonist, GLPAG, matched for GLP1R agonist potency and pharmacokinetics. The metabolic effects of these two peptides with respect to weight loss, caloric reduction, glucose control, and lipid lowering, were compared upon chronic dosing in diet-induced obese (DIO) mice. Acute studies in DIO mice revealed metabolic pathways that were modulated independent of weight loss. Studies in Glp1r−/− and Gcgr−/− mice enabled delineation of the contribution of GLP1R versus GCGR activation to the pharmacology of DualAG. RESULTS Peptide DualAG exhibits superior weight loss, lipid-lowering activity, and antihyperglycemic efficacy comparable to GLPAG. Improvements in plasma metabolic parameters including insulin, leptin, and adiponectin were more pronounced upon chronic treatment with DualAG than with GLPAG. Dual receptor agonism also increased fatty acid oxidation and reduced hepatic steatosis in DIO mice. The antiobesity effects of DualAG require activation of both GLP1R and GCGR. CONCLUSIONS Sustained GLP1R/GCGR dual agonism reverses obesity in DIO mice and is a novel therapeutic approach to the treatment of obesity. PMID:19602537

  15. Rational design of dual peptides targeting ghrelin and Y2 receptors to regulate food intake and body weight.

    PubMed

    Kilian, Tom-Marten; Klöting, Nora; Bergmann, Ralf; Els-Heindl, Sylvia; Babilon, Stefanie; Clément-Ziza, Mathieu; Zhang, Yixin; Beck-Sickinger, Annette G; Chollet, Constance

    2015-05-28

    Ghrelin and Y2 receptors play a central role in appetite regulation inducing opposite effects. The Y2 receptor induces satiety, while the ghrelin receptor promotes hunger and weight gain. However, the food regulating system is tightly controlled by interconnected pathways where redundancies can lead to poor efficacy and drug tolerance when addressing a single molecule. We developed a multitarget strategy to synthesize dual peptides simultaneously inhibiting the ghrelin receptor and stimulating the Y2 receptor. Dual peptides showed dual activity in vitro, and one compound induced a slight diminution of food intake in a rodent model of obesity. In addition, stability studies in rats revealed different behaviors between the dual peptide and its corresponding monomers. The Y2 receptor agonist was unstable in blood, while the dual peptide showed an intermediate stability compared to that of the highly stable ghrelin receptor inverse agonist.

  16. Directed Molecular Evolution of an Engineered Gammaretroviral Envelope Protein with Dual Receptor Use Shows Stable Maintenance of Both Receptor Specificities

    PubMed Central

    Friis, Kristina Pagh; Iturrioz, Xavier; Thomsen, Jonas; Alvear-Perez, Rodrigo; Bahrami, Shervin; Llorens-Cortes, Catherine

    2015-01-01

    ABSTRACT We have previously reported the construction of a murine leukemia virus-based replication-competent gammaretrovirus (SL3-AP) capable of utilizing the human G protein-coupled receptor APJ (hAPJ) as its entry receptor and its natural receptor, the murine Xpr1 receptor, with equal affinities. The apelin receptor has previously been shown to function as a coreceptor for HIV-1, and thus, adaptation of the viral vector to this receptor is of significant interest. Here, we report the molecular evolution of the SL3-AP envelope protein when the virus is cultured in cells harboring either the Xpr1 or the hAPJ receptor. Interestingly, the dual receptor affinity is maintained even after 10 passages in these cells. At the same time, the chimeric viral envelope protein evolves in a distinct pattern in the apelin cassette when passaged on D17 cells expressing hAPJ in three separate molecular evolution studies. This pattern reflects selection for reduced ligand-receptor interaction and is compatible with a model in which SL3-AP has evolved not to activate hAPJ receptor internalization. IMPORTANCE Few successful examples of engineered retargeting of a retroviral vector exist. The engineered SL3-AP envelope is capable of utilizing either the murine Xpr1 or the human APJ receptor for entry. In addition, SL3-AP is the first example of an engineered retrovirus retaining its dual tropism after several rounds of passaging on cells expressing only one of its receptors. We demonstrate that the virus evolves toward reduced ligand-receptor affinity, which sheds new light on virus adaptation. We provide indirect evidence that such reduced affinity leads to reduced receptor internalization and propose a novel model in which too rapid receptor internalization may decrease virus entry. PMID:26608314

  17. Dual mechanism of action of the atypical tetracycline chelocardin.

    PubMed

    Stepanek, Jennifer J; Lukežič, Tadeja; Teichert, Ines; Petković, Hrvoje; Bandow, Julia E

    2016-06-01

    Classical tetracyclines targeting the protein biosynthesis machinery are commonly applied in human and veterinary medicine. The development and spread of resistance seriously compromise the successful treatment of bacterial infections. The atypical tetracycline chelocardin holds promise as it retains activity against tetracycline-resistant strains. It has been suggested that chelocardin targets the bacterial membrane, thus differing in mode of action from that of classical tetracyclines. We investigated the mechanism of action of chelocardin using global proteome analysis. The proteome profiles after sublethal chelocardin stress were compared to a reference compendium containing antibiotic response profiles of Bacillus subtilis. This approach revealed a concentration-dependent dual mechanism of action. At low concentrations, like classical tetracyclines, chelocardin induces the proteomic signature for peptidyl transferase inhibition demonstrating that protein biosynthesis inhibition is the dominant physiological challenge. At higher concentrations B. subtilis mainly responds to membrane stress indicating that at clinically relevant concentrations the membrane is the main antibiotic target of chelocardin. Studying the effects on the membrane in more detail, we found that chelocardin causes membrane depolarization but does not lead to formation of large pores. We conclude that at growth inhibiting doses chelocardin not only targets protein biosynthesis but also corrupts the integrity of the bacterial membrane. This dual mechanism of action might prove beneficial in slowing the development of new resistance mechanisms against this atypical tetracycline.

  18. Behavioral analyses of GHB: receptor mechanisms.

    PubMed

    Carter, Lawrence P; Koek, Wouter; France, Charles P

    2009-01-01

    GHB is used therapeutically and recreationally, although the precise mechanism of action responsible for its different behavioral effects is not entirely clear. The purpose of this review is to summarize how behavioral procedures, especially drug discrimination procedures, have been used to study the mechanism of action of GHB. More specifically, we will review several different drug discrimination procedures and discuss how they have been used to qualitatively and quantitatively study different components of the complex mechanism of action of GHB. A growing number of studies have provided evidence that the behavioral effects of GHB are mediated predominantly by GABAB receptors. However, there is also evidence that the mechanisms mediating the effects of GHB and the prototypical GABAB receptor agonist baclofen are not identical, and that other mechanisms such as GHB receptors and subtypes of GABAA and GABAB receptors might contribute to the effects of GHB. These findings are consistent with the different behavioral profile, abuse liability, and therapeutic indications of GHB and baclofen. A better understanding of the similarities and differences between GHB and baclofen, as well as the pharmacological mechanisms of action underlying the recreational and therapeutic effects of GHB, could lead to more effective medications with fewer adverse effects.

  19. Behavioral Analyses of GHB: Receptor Mechanisms

    PubMed Central

    Carter, Lawrence P.; Koek, Wouter; France, Charles P.

    2009-01-01

    GHB is used therapeutically and recreationally, although the precise mechanism of action responsible for its different behavioral effects is not entirely clear. The purpose of this review is to summarize how behavioral procedures, especially drug discrimination procedures, have been used to study the mechanism of action of GHB. More specifically, we will review several different drug discrimination procedures and discuss how they have been used to qualitatively and quantitatively study different components of the complex mechanism of action of GHB. A growing number of studies have provided evidence that the behavioral effects of GHB are mediated predominantly by GABAB receptors. However, there is also evidence that the mechanisms mediating the effects of GHB and the prototypical GABAB receptor agonist baclofen are not identical, and that other mechanisms such as GHB receptors and subtypes of GABAA and GABAB receptors might contribute to the effects of GHB. These findings are consistent with the different behavioral profile, abuse liability, and therapeutic indications of GHB and baclofen. A better understanding of the similarities and differences between GHB and baclofen, as well as the pharmacological mechanisms of action underlying the recreational and therapeutic effects of GHB, could lead to more effective medications with fewer adverse effects. PMID:19010351

  20. Action mechanisms of Liver X Receptors

    SciTech Connect

    Gabbi, Chiara; Warner, Margaret; Gustafsson, Jan-Åke

    2014-04-11

    Highlights: • LXRα and LXRβ are ligand-activated nuclear receptors. • They share oxysterol ligands and the same heterodimerization partner, RXR. • LXRs regulate lipid and glucose metabolism, CNS and immune functions, and water transport. - Abstract: The two Liver X Receptors, LXRα and LXRβ, are nuclear receptors belonging to the superfamily of ligand-activated transcription factors. They share more than 78% homology in amino acid sequence, a common profile of oxysterol ligands and the same heterodimerization partner, Retinoid X Receptor. LXRs play crucial roles in several metabolic pathways: lipid metabolism, in particular in preventing cellular cholesterol accumulation; glucose homeostasis; inflammation; central nervous system functions and water transport. As with all nuclear receptors, the transcriptional activity of LXR is the result of an orchestration of numerous cellular factors including ligand bioavailability, presence of corepressors and coactivators and cellular context i.e., what other pathways are activated in the cell at the time the receptor recognizes its ligand. In this mini-review we summarize the factors regulating the transcriptional activity and the mechanisms of action of these two receptors.

  1. Primal-dual techniques for online algorithms and mechanisms

    NASA Astrophysics Data System (ADS)

    Liaghat, Vahid

    An offline algorithm is one that knows the entire input in advance. An online algorithm, however, processes its input in a serial fashion. In contrast to offline algorithms, an online algorithm works in a local fashion and has to make irrevocable decisions without having the entire input. Online algorithms are often not optimal since their irrevocable decisions may turn out to be inefficient after receiving the rest of the input. For a given online problem, the goal is to design algorithms which are competitive against the offline optimal solutions. In a classical offline scenario, it is often common to see a dual analysis of problems that can be formulated as a linear or convex program. Primal-dual and dual-fitting techniques have been successfully applied to many such problems. Unfortunately, the usual tricks come short in an online setting since an online algorithm should make decisions without knowing even the whole program. In this thesis, we study the competitive analysis of fundamental problems in the literature such as different variants of online matching and online Steiner connectivity, via online dual techniques. Although there are many generic tools for solving an optimization problem in the offline paradigm, in comparison, much less is known for tackling online problems. The main focus of this work is to design generic techniques for solving integral linear optimization problems where the solution space is restricted via a set of linear constraints. A general family of these problems are online packing/covering problems. Our work shows that for several seemingly unrelated problems, primal-dual techniques can be successfully applied as a unifying approach for analyzing these problems. We believe this leads to generic algorithmic frameworks for solving online problems. In the first part of the thesis, we show the effectiveness of our techniques in the stochastic settings and their applications in Bayesian mechanism design. In particular, we introduce new

  2. The dual orexin receptor antagonist TCS1102 does not affect reinstatement of nicotine-seeking

    PubMed Central

    McNally, Gavan P.; Clemens, Kelly J.

    2017-01-01

    The orexin/hypocretin system is important for appetitive motivation towards multiple drugs of abuse, including nicotine. Both OX1 and OX2 receptors individually have been shown to influence nicotine self-administration and reinstatement. Due to the increasing clinical use of dual orexin receptor antagonists in the treatment of disorders such as insomnia, we examined whether a dual orexin receptor antagonist may also be effective in reducing nicotine seeking. We tested the effect of intracerebroventricular (i.c.v.) administration of the potent and selective dual orexin receptor antagonist TCS1102 on orexin-A-induced food self-administration, nicotine self-administration and reinstatement of nicotine-seeking in rats. Our results show that 30 μg of TCS1102 i.c.v. abolishes orexin-A-induced increases in food self-administration but does not reduce nicotine self-administration. Neither i.c.v. 10 μg nor 30 μg of TCS1102 reduced compound reinstatement after short-term (15 days) self-administration nicotine, but 30 μg transiently reduced cue/nicotine compound reinstatement after chronic self-administration (29 days). These results indicate that TCS1102 has no substantial effect on motivation for nicotine seeking following chronic self-administration and no effect after shorter periods of intake. Orexin receptor antagonists may therefore have little clinical utility against nicotine addiction. PMID:28296947

  3. Kinetic properties of "dual" orexin receptor antagonists at OX1R and OX2R orexin receptors.

    PubMed

    Callander, Gabrielle E; Olorunda, Morenike; Monna, Dominique; Schuepbach, Edi; Langenegger, Daniel; Betschart, Claudia; Hintermann, Samuel; Behnke, Dirk; Cotesta, Simona; Fendt, Markus; Laue, Grit; Ofner, Silvio; Briard, Emmanuelle; Gee, Christine E; Jacobson, Laura H; Hoyer, Daniel

    2013-01-01

    Orexin receptor antagonists represent attractive targets for the development of drugs for the treatment of insomnia. Both efficacy and safety are crucial in clinical settings and thorough investigations of pharmacokinetics and pharmacodynamics can predict contributing factors such as duration of action and undesirable effects. To this end, we studied the interactions between various "dual" orexin receptor antagonists and the orexin receptors, OX1R and OX2R, over time using saturation and competition radioligand binding with [(3)H]-BBAC ((S)-N-([1,1'-biphenyl]-2-yl)-1-(2-((1-methyl-1H-benzo[d]imidazol-2-yl)thio)acetyl)pyrrolidine-2-carboxamide). In addition, the kinetics of these compounds were investigated in cells expressing human, mouse and rat OX1R and OX2R using FLIPR® assays for calcium accumulation. We demonstrate that almorexant reaches equilibrium very slowly at OX2R, whereas SB-649868, suvorexant, and filorexant may take hours to reach steady state at both orexin receptors. By contrast, compounds such as BBAC or the selective OX2R antagonist IPSU ((2-((1H-Indol-3-yl)methyl)-9-(4-methoxypyrimidin-2-yl)-2,9-diazaspiro[5.5]undecan-1-one) bind rapidly and reach equilibrium very quickly in binding and/or functional assays. Overall, the "dual" antagonists tested here tend to be rather unselective under non-equilibrium conditions and reach equilibrium very slowly. Once equilibrium is reached, each ligand demonstrates a selectivity profile that is however, distinct from the non-equilibrium condition. The slow kinetics of the "dual" antagonists tested suggest that in vitro receptor occupancy may be longer lasting than would be predicted. This raises questions as to whether pharmacokinetic studies measuring plasma or brain levels of these antagonists are accurate reflections of receptor occupancy in vivo.

  4. Mechanism of FGF receptor dimerization and activation

    NASA Astrophysics Data System (ADS)

    Sarabipour, Sarvenaz; Hristova, Kalina

    2016-01-01

    Fibroblast growth factors (fgfs) are widely believed to activate their receptors by mediating receptor dimerization. Here we show, however, that the FGF receptors form dimers in the absence of ligand, and that these unliganded dimers are phosphorylated. We further show that ligand binding triggers structural changes in the FGFR dimers, which increase FGFR phosphorylation. The observed effects due to the ligands fgf1 and fgf2 are very different. The fgf2-bound dimer structure ensures the smallest separation between the transmembrane (TM) domains and the highest possible phosphorylation, a conclusion that is supported by a strong correlation between TM helix separation in the dimer and kinase phosphorylation. The pathogenic A391E mutation in FGFR3 TM domain emulates the action of fgf2, trapping the FGFR3 dimer in its most active state. This study establishes the existence of multiple active ligand-bound states, and uncovers a novel molecular mechanism through which FGFR-linked pathologies can arise.

  5. Dual mechanisms regulate the nucleocytoplasmic localization of human DDX6.

    PubMed

    Huang, Jo-Hsi; Ku, Wei-Chi; Chen, Yen-Chun; Chang, Yi-Ling; Chu, Chia-Ying

    2017-02-20

    DDX6 is a conserved DEAD-box protein (DBP) that plays central roles in cytoplasmic RNA regulation, including processing body (P-body) assembly, mRNA decapping, and translational repression. Beyond its cytoplasmic functions, DDX6 may also have nuclear functions because its orthologues are known to localize to nuclei in several biological contexts. However, it is unclear whether DDX6 is generally present in human cell nuclei, and the molecular mechanism underlying DDX6 subcellular distribution remains elusive. In this study, we showed that DDX6 is commonly present in the nuclei of human-derived cells. Our structural and molecular analyses deviate from the current model that the shuttling of DDX6 is directly mediated by the canonical nuclear localization signal (NLS) and nuclear export signal (NES), which are recognized and transported by Importin-α/β and CRM1, respectively. Instead, we show that DDX6 can be transported by 4E-T in a piggyback manner. Furthermore, we provide evidence for a novel nuclear targeting mechanism in which DDX6 enters the newly formed nuclei by "hitch-hiking" on mitotic chromosomes with its C-terminal domain during M phase progression. Together, our results indicate that the nucleocytoplasmic localization of DDX6 is regulated by these dual mechanisms.

  6. Dual mechanisms regulate the nucleocytoplasmic localization of human DDX6

    PubMed Central

    Huang, Jo-Hsi; Ku, Wei-Chi; Chen, Yen-Chun; Chang, Yi-Ling; Chu, Chia-Ying

    2017-01-01

    DDX6 is a conserved DEAD-box protein (DBP) that plays central roles in cytoplasmic RNA regulation, including processing body (P-body) assembly, mRNA decapping, and translational repression. Beyond its cytoplasmic functions, DDX6 may also have nuclear functions because its orthologues are known to localize to nuclei in several biological contexts. However, it is unclear whether DDX6 is generally present in human cell nuclei, and the molecular mechanism underlying DDX6 subcellular distribution remains elusive. In this study, we showed that DDX6 is commonly present in the nuclei of human-derived cells. Our structural and molecular analyses deviate from the current model that the shuttling of DDX6 is directly mediated by the canonical nuclear localization signal (NLS) and nuclear export signal (NES), which are recognized and transported by Importin-α/β and CRM1, respectively. Instead, we show that DDX6 can be transported by 4E-T in a piggyback manner. Furthermore, we provide evidence for a novel nuclear targeting mechanism in which DDX6 enters the newly formed nuclei by “hitch-hiking” on mitotic chromosomes with its C-terminal domain during M phase progression. Together, our results indicate that the nucleocytoplasmic localization of DDX6 is regulated by these dual mechanisms. PMID:28216671

  7. A Dual Pathogenic Mechanism Links Tau Acetylation to Sporadic Tauopathy

    PubMed Central

    Trzeciakiewicz, Hanna; Tseng, Jui-Heng; Wander, Connor M.; Madden, Victoria; Tripathy, Ashutosh; Yuan, Chao-Xing; Cohen, Todd J.

    2017-01-01

    Tau acetylation has recently emerged as a dominant post-translational modification (PTM) in Alzheimer’s disease (AD) and related tauopathies. Mass spectrometry studies indicate that tau acetylation sites cluster within the microtubule (MT)-binding region (MTBR), suggesting acetylation could regulate both normal and pathological tau functions. Here, we combined biochemical and cell-based approaches to uncover a dual pathogenic mechanism mediated by tau acetylation. We show that acetylation specifically at residues K280/K281 impairs tau-mediated MT stabilization, and enhances the formation of fibrillar tau aggregates, highlighting both loss and gain of tau function. Full-length acetylation-mimic tau showed increased propensity to undergo seed-dependent aggregation, revealing a potential role for tau acetylation in the propagation of tau pathology. We also demonstrate that methylene blue, a reported tau aggregation inhibitor, modulates tau acetylation, a novel mechanism of action for this class of compounds. Our study identifies a potential “two-hit” mechanism in which tau acetylation disengages tau from MTs and also promotes tau aggregation. Thus, therapeutic approaches to limit tau K280/K281 acetylation could simultaneously restore MT stability and ameliorate tau pathology in AD and related tauopathies. PMID:28287136

  8. Mechanisms underlying dual effects of serotonin during development of Helisoma trivolvis (Mollusca)

    PubMed Central

    2014-01-01

    Background Serotonin (5-HT) is well known as widely distributed modulator of developmental processes in both vertebrates and invertebrates. It is also the earliest neurotransmitter to appear during neuronal development. In aquatic invertebrates, which have larvae in their life cycle, 5-HT is involved in regulation of stages transition including larval metamorphosis and settlement. However, molecular and cellular mechanisms underlying developmental transition in aquatic invertebrate species are yet poorly understood. Earlier we demonstrated that in larvae of freshwater molluscs and marine polychaetes, endogenous 5-HT released from the neurons of the apical sensory organ (ASO) in response to external stimuli retarded larval development at premetamorphic stages, and accelerated it at metamorphic stages. Here we used a freshwater snail Helisoma trivolvis to study molecular mechanisms underlying these dual developmental effects of 5-HT. Results Larval development of H. trivolvis includes transition from premetamorphic to metamorphic stages and shares the main features of metamorphosis with free-swimming aquatic larvae. Three types of 5-HT receptors (5-HT1-, 5-HT4- and 5-HT7-like) are functionally active at premetamorphic (trochophore, veliger) and metamorphic (veliconcha) stages, and expression patterns of these receptors and respective G proteins undergo coordinated changes during development. Stimulation of these receptors modulated cAMP-dependent regulation of cell divisions. Expression of 5-HT4- and 5-HT7-like receptors and their downstream Gs protein was down-regulated during the transition of pre- to metamorphic stage, while expression of 5-HT1 -like receptor and its downstream Gi protein was upregulated. In accordance with relative amount of these receptors, stimulation of 5-HTRs at premetamorphic stages induces developmental retardation, while their stimulation at metamorphic stages induces developmental acceleration. Conclusions We present a novel molecular

  9. Dual-Pitch Processing Mechanisms in Primate Auditory Cortex

    PubMed Central

    Bendor, Daniel; Osmanski, Michael S.

    2012-01-01

    Pitch, our perception of how high or low a sound is on a musical scale, is a fundamental perceptual attribute of sounds and is important for both music and speech. After more than a century of research, the exact mechanisms used by the auditory system to extract pitch are still being debated. Theoretically, pitch can be computed using either spectral or temporal acoustic features of a sound. We have investigated how cues derived from the temporal envelope and spectrum of an acoustic signal are used for pitch extraction in the common marmoset (Callithrix jacchus), a vocal primate species, by measuring pitch discrimination behaviorally and examining pitch-selective neuronal responses in auditory cortex. We find that pitch is extracted by marmosets using temporal envelope cues for lower pitch sounds composed of higher-order harmonics, whereas spectral cues are used for higher pitch sounds with lower-order harmonics. Our data support dual-pitch processing mechanisms, originally proposed by psychophysicists based on human studies, whereby pitch is extracted using a combination of temporal envelope and spectral cues. PMID:23152599

  10. Design and Synthesis of Norendoxifen Analogues with Dual Aromatase Inhibitory and Estrogen Receptor Modulatory Activities

    PubMed Central

    Lv, Wei; Liu, Jinzhong; Skaar, Todd C.; Flockhart, David A.; Cushman, Mark

    2015-01-01

    Both selective estrogen receptor modulators and aromatase inhibitors are widely used for the treatment of breast cancer. Compounds with both aromatase inhibitory and estrogen receptor modulatory activities could have special advantages for treatment of breast cancer. Our previous efforts led to the discovery of norendoxifen as the first compound with dual aromatase inhibitory and estrogen receptor binding activities. To optimize its efficacy and aromatase selectivity versus other cytochrome P450 enzymes, a series of structurally related norendoxifen analogues were designed and synthesized. The most potent compound, 4'-hydroxynorendoxifen (10), displayed elevated inhibitory potency against aromatase and enhanced affinity for estrogen receptors when compared to norendoxifen. The selectivity of 10 for aromatase versus other cytochrome P450 enzymes was also superior to norendoxifen. 4'-Hydroxynorendoxifen is therefore an interesting lead for further development to obtain new anticancer agents of potential value for the treatment of breast cancer. PMID:25751283

  11. Opioid receptor desensitization: mechanisms and its link to tolerance

    PubMed Central

    Allouche, Stéphane; Noble, Florence; Marie, Nicolas

    2014-01-01

    Opioid receptors (OR) are part of the class A of G-protein coupled receptors and the target of the opiates, the most powerful analgesic molecules used in clinic. During a protracted use, a tolerance to analgesic effect develops resulting in a reduction of the effectiveness. So understanding mechanisms of tolerance is a great challenge and may help to find new strategies to tackle this side effect. This review will summarize receptor-related mechanisms that could underlie tolerance especially receptor desensitization. We will focus on the latest data obtained on molecular mechanisms involved in opioid receptor desensitization: phosphorylation, receptor uncoupling, internalization, and post-endocytic fate of the receptor. PMID:25566076

  12. Quantum mechanics on SO(3) via noncommutative dual variables

    NASA Astrophysics Data System (ADS)

    Oriti, Daniele; Raasakka, Matti

    2011-07-01

    We formulate quantum mechanics on the group SO(3) using a noncommutative dual space representation for the quantum states, inspired by recent work in quantum gravity. The new noncommutative variables have a clear connection to the corresponding classical variables, and our analysis confirms them as the natural phase space variables, both mathematically and physically. In particular, we derive the first order (Hamiltonian) path integral in terms of the noncommutative variables, as a formulation of the transition amplitudes alternative to that based on harmonic analysis. We find that the nontrivial phase space structure gives naturally rise to quantum corrections to the action for which we find a closed expression. We then study both the semiclassical approximation of the first order path integral and the example of a free particle on SO(3). On the basis of these results, we comment on the relevance of similar structures and methods for more complicated theories with group-based configuration spaces, such as loop quantum gravity and spin foam models.

  13. The Dual Orexin Receptor Antagonist Almorexant Induces Sleep and Decreases Orexin-Induced Locomotion by Blocking Orexin 2 Receptors

    PubMed Central

    Mang, Géraldine M.; Dürst, Thomas; Bürki, Hugo; Imobersteg, Stefan; Abramowski, Dorothee; Schuepbach, Edi; Hoyer, Daniel; Fendt, Markus; Gee, Christine E.

    2012-01-01

    Study Objectives: Orexin peptides activate orexin 1 and orexin 2 receptors (OX1R and OX2R), regulate locomotion and sleep-wake. The dual OX1R/OX2R antagonist almorexant reduces activity and promotes sleep in multiple species, including man. The relative contributions of the two receptors in locomotion and sleep/wake regulation were investigated in mice. Design: Mice lacking orexin receptors were used to determine the contribution of OX1R and OX2R to orexin A-induced locomotion and to almorexant-induced sleep. Setting: N/A. Patients or Participants: C57BL/6J mice and OX1R+/+, OX1R-/-, OX2R+/+, OX2R-/- and OX1R-/-/OX2R-/- mice. Interventions: Intracerebroventricular orexin A; oral dosing of almorexant. Measurements and Results: Almorexant attenuated orexin A-induced locomotion. As in other species, almorexant dose-dependently increased rapid eye movement sleep (REM) and nonREM sleep in mice. Almorexant and orexin A were ineffective in OX1R-/-/OX2R-/- mice. Both orexin A-induced locomotion and sleep induction by almorexant were absent in OX2R-/- mice. Interestingly, almorexant did not induce cataplexy in wild-type mice under conditions where cataplexy was seen in mice lacking orexins and in OX1R-/-/OX2R-/- mice. Almorexant dissociates very slowly from OX2R as measured functionally and in radioligand binding. Under non equilibrium conditions in vitro, almorexant was a dual antagonist whereas at equilibrium, almorexant became OX2R selective. Conclusions: In vivo, almorexant specifically inhibits the actions of orexin A. The two known orexin receptors mediate sleep induction by almorexant and orexin A-induced locomotion. However, OX2R activation mediates locomotion induction by orexin A and antagonism of OX2R is sufficient to promote sleep in mice. Citation: Mang GM; Dürst T; Bürki H; Imobersteg S; Abramowski D; Schuepbach E; Hoyer D; Fendt M; Gee CE. The dual orexin receptor antagonist almorexant induces sleep and decreases orexin-induced locomotion by blocking orexin

  14. A catalytic mechanism for the dual-specific phosphatases.

    PubMed

    Denu, J M; Dixon, J E

    1995-06-20

    Dual-specific protein-tyrosine phosphatases have the common active-site sequence motif HCXXGXXRS(T). The role of the conserved hydroxyl was investigated by changing serine-131 to an alanine (S131A) in the dual-specific protein-tyrosine phosphatase VHR. The pH profile of the kcat/Km value for the S131A mutant is indistinguishable from that of the native enzyme. In contrast, the kcat value for S131A mutant is 100-fold lower than that for the native enzyme, and the shape of the pH profile was perturbed from bell-shaped in the native enzyme to a pH-independent curve over the pH range 4.5-9.0. This evidence, along with results from a previous study, suggests that the S131A mutation alters the rate-limiting step in the catalytic mechanism. Formation of a phosphoenzyme intermediate appears to be rate-limiting with the native enzyme, whereas in the S131A mutant breakdown of the intermediate is rate-limiting. This was confirmed by the appearance of a burst of p-nitrophenol formation when p-nitrophenyl phosphate rapidly reacted with the S131A enzyme in a stopped-flow spectrophotometer. Loss of this hydroxyl group at the active site dramatically diminished the ability of the enzyme to hydrolyze the thiol-phosphate intermediate without exerting any significant change in the steps leading to and including the formation of the intermediate. Consistent with rate-limiting intermediate formation in the native enzyme, the rate of burst in the S131A mutant was 1.5 s-1, which agrees well with the kcat value of 5 s-1 observed for native enzyme. The amplitude of the burst was stoichiometric with final enzyme concentration, and the slow linear rate (0.06 s-1) of p-nitrophenol formation after the burst was in agreement with the steady-state determined value of kcat (0.055 s-1).

  15. Dual Role of the Second Extracellular Loop of the Cannabinoid Receptor 1: Ligand Binding and Receptor Localization

    PubMed Central

    Ahn, Kwang H.; Bertalovitz, Alexander C.; Mierke, Dale F.

    2009-01-01

    are consistent with a dual role for EC2 in stabilizing receptor assembly and in ligand binding. PMID:19643997

  16. Dual signaling regulated by calcyon, a D1 dopamine receptor interacting protein.

    PubMed

    Lezcano, N; Mrzljak, L; Eubanks, S; Levenson, R; Goldman-Rakic, P; Bergson, C

    2000-03-03

    The synergistic response of cells to the stimulation of multiple receptors has been ascribed to receptor cross talk; however, the specific molecules that mediate the resultant signal amplification have not been defined. Here a 24-kilodalton single transmembrane protein, designated calcyon, we functionally characterize that interacts with the D1 dopamine receptor. Calcyon localizes to dendritic spines of D1 receptor-expressing pyramidal cells in prefrontal cortex. These studies delineate a mechanism of Gq- and Gs-coupled heterotrimeric GTP-binding protein-coupled receptor cross talk by which D1 receptors can shift effector coupling to stimulate robust intracellular calcium (Ca2+i) release as a result of interaction with calcyon. The role of calcyon in potentiating Ca2+-dependent signaling should provide insight into the D1 receptor-modulated cognitive functions of prefrontal cortex.

  17. Molecular mechanisms of glucocorticoid receptor signaling.

    PubMed

    Labeur, Marta; Holsboer, Florian

    2010-01-01

    This review highlights the most recent findings on the molecular mechanisms of the glucocorticoid receptor (GR). Most effects of glucocorticoids are mediated by the intracellular GR which is present in almost every tissue and controls transcriptional activation via direct and indirect mechanisms. Nevertheless the glu-cocorticoid responses are tissue -and gene- specific. GR associates selectively with corticosteroid ligands produced in the adrenal gland in response to changes of humoral homeostasis. Ligand interaction with GR promotes either GR binding to genomic glucocorticoid response elements, in turn modulating gene transcription, or interaction of GR monomers with other transcription factors activated by other signalling pathways leading to transrepression. The GR regulates a broad spectrum of physiological functions, including cell differentiation, metabolism and inflammatory responses. Thus, disruption or dysregulation of GR function will result in severe impairments in the maintenance of homeostasis and the control of adaptation to stress.

  18. Dual effects of anandamide on NMDA receptor-mediated responses and neurotransmission.

    PubMed

    Hampson, A J; Bornheim, L M; Scanziani, M; Yost, C S; Gray, A T; Hansen, B M; Leonoudakis, D J; Bickler, P E

    1998-02-01

    Anandamide is an endogenous ligand of cannabinoid receptors that induces pharmacological responses in animals similar to those of cannabinoids such as delta9-tetrahydrocannabinol (THC). Typical pharmacological effects of cannabinoids include disruption of pain, memory formation, and motor coordination, systems that all depend on NMDA receptor mediated neurotransmission. We investigated whether anandamide can influence NMDA receptor activity by examining NMDA-induced calcium flux (deltaCa2+NMDA) in rat brain slices. The presence of anandamide reduced deltaCa2+NMDA and the inhibition was disrupted by cannabinoid receptor antagonist, pertussis toxin treatment, and agatoxin (a calcium channel inhibitor). Whereas these treatments prevented anandamide inhibiting deltaCa2+NMDA, they also revealed another, underlying mechanism by which anandamide influences deltaCa2+NMDA. In the presence of cannabinoid receptor antagonist, anandamide potentiated deltaCa2+NMDA in cortical, cerebellar, and hippocampal slices. Anandamide (but not THC) also augmented NMDA-stimulated currents in Xenopus oocytes expressing cloned NMDA receptors, suggesting a capacity to directly modulate NMDA receptor activity. In a similar manner, anandamide enhanced neurotransmission across NMDA receptor-dependent synapses in hippocampus in a manner that was not mimicked by THC and was unaffected by cannabinoid receptor antagonist. These data demonstrate that anandamide can modulate NMDA receptor activity in addition to its role as a cannabinoid receptor ligand.

  19. A dual laminin/collagen receptor acts in peripheral nerve regeneration.

    PubMed Central

    Toyota, B; Carbonetto, S; David, S

    1990-01-01

    A regeneration chamber was created in vivo by suturing a synthetic tube sealed at its distal end onto the proximal stump of a severed rat sciatic nerve. Nerves regenerated into tubes coated with laminin at a rate of 0.33 mm/day after a lag of about 2 days. At 25 days, regenerating nerves had extended 23% farther into laminin-coated tubes as compared with uncoated ones. Monoclonal antibody 3A3, which functionally interferes with a dual laminin/collagen receptor, inhibited nerve regeneration into laminin-coated tubes by 32%. In contrast, monoclonal antibody JG22, which inhibits chicken matrix receptors, had no significant effect on regeneration. Immunohistochemical studies of teased adult rat sciatic nerves indicate that 3A3 bound to Schwann cells and possibly to axons. In other studies, the heterodimeric, laminin/collagen receptor recognized by 3A3 has been shown to be a member of the integrin superfamily of adhesive receptors. These data provide evidence that an integrin receptor functions in nerve regeneration in vivo. Images PMID:2154740

  20. Biaryls as potent, tunable dual neurokinin 1 receptor antagonists and serotonin transporter inhibitors.

    PubMed

    Degnan, Andrew P; Tora, George O; Han, Ying; Rajamani, Ramkumar; Bertekap, Robert; Krause, Rudolph; Davis, Carl D; Hu, Joanna; Morgan, Daniel; Taylor, Sarah J; Krause, Kelly; Li, Yu-Wen; Mattson, Gail; Cunningham, Melissa A; Taber, Matthew T; Lodge, Nicholas J; Bronson, Joanne J; Gillman, Kevin W; Macor, John E

    2015-08-01

    Depression is a serious illness that affects millions of patients. Current treatments are associated with a number of undesirable side effects. Neurokinin 1 receptor (NK1R) antagonists have recently been shown to potentiate the antidepressant effects of serotonin-selective reuptake inhibitors (SSRIs) in a number of animal models. Herein we describe the optimization of a biaryl chemotype to provide a series of potent dual NK1R antagonists/serotonin transporter (SERT) inhibitors. Through the choice of appropriate substituents, the SERT/NK1R ratio could be tuned to afford a range of target selectivity profiles. This effort culminated in the identification of an analog that demonstrated oral bioavailability, favorable brain uptake, and efficacy in the gerbil foot tap model. Ex vivo occupancy studies with compound 58 demonstrated the ability to maintain NK1 receptor saturation (>88% occupancy) while titrating the desired level of SERT occupancy (11-84%) via dose selection.

  1. Terminalia Chebula provides protection against dual modes of necroptotic and apoptotic cell death upon death receptor ligation

    PubMed Central

    Lee, Yoonjung; Byun, Hee Sun; Seok, Jeong Ho; Park, Kyeong Ah; Won, Minho; Seo, Wonhyoung; Lee, So-Ra; Kang, Kidong; Sohn, Kyung-Cheol; Lee, Ill Young; Kim, Hyeong-Geug; Son, Chang Gue; Shen, Han-Ming; Hur, Gang Min

    2016-01-01

    Death receptor (DR) ligation elicits two different modes of cell death (necroptosis and apoptosis) depending on the cellular context. By screening a plant extract library from cells undergoing necroptosis or apoptosis, we identified a water extract of Terminalia chebula (WETC) as a novel and potent dual inhibitor of DR-mediated cell death. Investigation of the underlying mechanisms of its anti-necroptotic and anti-apoptotic action revealed that WETC or its constituents (e.g., gallic acid) protected against tumor necrosis factor-induced necroptosis via the suppression of TNF-induced ROS without affecting the upstream signaling events. Surprisingly, WETC also provided protection against DR-mediated apoptosis by inhibition of the caspase cascade. Furthermore, it activated the autophagy pathway via suppression of mTOR. Of the WETC constituents, punicalagin and geraniin appeared to possess the most potent anti-apoptotic and autophagy activation effect. Importantly, blockage of autophagy with pharmacological inhibitors or genetic silencing of Atg5 selectively abolished the anti-apoptotic function of WETC. These results suggest that WETC protects against dual modes of cell death upon DR ligation. Therefore, WETC might serve as a potential treatment for diseases characterized by aberrantly sensitized apoptotic or non-apoptotic signaling cascades. PMID:27117478

  2. Targeting prostate cancer with compounds possessing dual activity as androgen receptor antagonists and HDAC6 inhibitors.

    PubMed

    Jadhavar, Pradeep S; Ramachandran, Sreekanth A; Riquelme, Eduardo; Gupta, Ashu; Quinn, Kevin P; Shivakumar, Devleena; Ray, Soumya; Zende, Dnyaneshwar; Nayak, Anjan K; Miglani, Sandeep K; Sathe, Balaji D; Raja, Mohd; Farias, Olivia; Alfaro, Ivan; Belmar, Sebastián; Guerrero, Javier; Bernales, Sebastián; Chakravarty, Sarvajit; Hung, David T; Lindquist, Jeffrey N; Rai, Roopa

    2016-11-01

    While enzalutamide and abiraterone are approved for treatment of metastatic castration-resistant prostate cancer (mCRPC), approximately 20-40% of patients have no response to these agents. It has been stipulated that the lack of response and the development of secondary resistance to these drugs may be due to the presence of AR splice variants. HDAC6 has a role in regulating the androgen receptor (AR) by modulating heat shock protein 90 (Hsp90) acetylation, which controls the nuclear localization and activation of the AR in androgen-dependent and independent scenarios. With dual-acting AR-HDAC6 inhibitors it should be possible to target patients who don't respond to enzalutamide. Herein, we describe the design, synthesis and biological evaluation of dual-acting compounds which target AR and are also specific towards HDAC6. Our efforts led to compound 10 which was found to have potent dual activity (HDAC6 IC50=0.0356μM and AR binding IC50=<0.03μM). Compound 10 was further evaluated for antagonist and other cell-based activities, in vitro stability and pharmacokinetics.

  3. Baclofen-induced antinociception and nicotinic receptor mechanism(s).

    PubMed

    Sabetkasai, M; Ahang, S; Shafaghi, B; Zarrindast, M R

    1999-11-01

    In this study, the influences of nicotinic receptor agents on baclofen-induced antinociception in the tail-flick test have been studied. Intraperitoneal administration of baclofen (2.5, 5 and 10 mg/kg) to mice induced a dose-dependent antinociception in the tail-flick test. Subcutaneous injection of nicotine (0.5-2.5 mg/kg) also caused a dose-dependent antinociceptive response. Intracerebral (10 and 20 microg/mouse) but not intraperitoneal administration of hexamethonium (5 and 10 mg/kg) to mice decreased the response of both nicotine and baclofen. However, administration of the GABA(B) antagonist CGP 35348 (100 and 200 mg/kg) decreased the response induced by baclofen but not by nicotine. It is concluded that at least part of the baclofen-induced antinociception may be mediated through a nicotinic mechanism.

  4. Characterization of U-97775 as a GABAA receptor ligand of dual functionality in cloned rat GABAA receptor subtypes.

    PubMed Central

    Im, H. K.; Im, W. B.; Pregenzer, J. F.; Carter, D. B.; Jacobsen, E. J.; Hamilton, B. J.

    1995-01-01

    1. U-97775 (tert-butyl 7-chloro-4,5-dihydro-5-[(1-(3,4,5-trimethyl)piperazino)carbonyl]- imidazo[1,5-a])quinoxaline-3-carboxylate) is a novel GABAA receptor ligand of dual functionality and was characterized for its interactions with cloned rat GABAA receptors expressed in human embryonic kidney cells. 2. The drug produced a bell-shaped dose-response profile in the alpha 1 beta 2 gamma 2 receptor subtype as monitored with GABA-induced Cl- currents in the whole cell patch-clamp technique. At low concentrations (< 0.5 microM), U-97775 enhanced the currents with a maximal increase of 120% as normalized to 5 microM GABA response (control). An agonist interaction of U-97775 with the benzodiazepine site is suggested, because Ro 15-1788 (an antagonist at the benzodiazepine site) abolished the current increase and [3H]-flunitrazepam binding was inhibited by U-97775 with a Ki of 1.2 nM. 3. The enhancement of GABA currents progressively disappeared as the U-97775 concentration was raised above 1 microM, and the current amplitude was reduced to 40% below the control at 10 microM U-97775. The current inhibition by U-97775 (10 microM) was not affected by Ro 15-1788. It appears that U-97775 interacts with a second site on GABA receptors, distinct from the benzodiazepine site, to reverse its agonistic activity on the benzodiazepine site and also to inhibit GABA currents. 4. U-97775 at low concentrations reduced and at high concentrations enhanced [35S]-TBPS binding. Ro 15-1788 selectively blocked the effect of U-97775 at low concentrations.(ABSTRACT TRUNCATED AT 250 WORDS) PMID:7647975

  5. Glycosylated SV2 and Gangliosides as Dual Receptors for Botulinum Neurotoxin Serotype F

    SciTech Connect

    Fu, Zhuji; Chen, Chen; Barbieri, Joseph T.; Kim, Jung-Ja P.; Baldwin, Michael R.

    2010-02-22

    Botulinum neurotoxin causes rapid flaccid paralysis through the inhibition of acetylcholine release at the neuromuscular junction. The seven BoNT serotypes (A-G) have been proposed to bind motor neurons via ganglioside-protein dual receptors. To date, the structure-function properties of BoNT/F host receptor interactions have not been resolved. Here, we report the crystal structures of the receptor binding domains (HCR) of BoNT/A and BoNT/F and the characterization of the dual receptors for BoNT/F. The overall polypeptide fold of HCR/A is essentially identical to the receptor binding domain of the BoNT/A holotoxin, and the structure of HCR/F is very similar to that of HCR/A, except for two regions implicated in neuronal binding. Solid phase array analysis identified two HCR/F binding glycans: ganglioside GD1a and oligosaccharides containing an N-acetyllactosamine core. Using affinity chromatography, HCR/F bound native synaptic vesicle glycoproteins as part of a protein complex. Deglycosylation of glycoproteins using {alpha}(1-3,4)-fucosidase, endo-{beta}-galactosidase, and PNGase F disrupted the interaction with HCR/F, while the binding of HCR/B to its cognate receptor, synaptotagmin I, was unaffected. These data indicate that the HCR/F binds synaptic vesicle glycoproteins through the keratan sulfate moiety of SV2. The interaction of HCR/F with gangliosides was also investigated. HCR/F bound specifically to gangliosides that contain {alpha}2,3-linked sialic acid on the terminal galactose of a neutral saccharide core (binding order GT1b = GD1a GM3; no binding to GD1b and GM1a). Mutations within the putative ganglioside binding pocket of HCR/F decreased binding to gangliosides, synaptic vesicle protein complexes, and primary rat hippocampal neurons. Thus, BoNT/F neuronal discrimination involves the recognition of ganglioside and protein (glycosylated SV2) carbohydrate moieties, providing a structural basis for the high affinity and specificity of BoNT/F for neurons.

  6. Mechanical Properties of a Superalloy Disk with a Dual Grain Structure

    NASA Technical Reports Server (NTRS)

    Gayda, John; Gabb, Timothy; Kantzos, Peter

    2003-01-01

    Mechanical properties from an advanced, nickel-base superalloy disk, with a dual grain structure consisting of a fine grain bore and coarse grain rim, were evaluated. The dual grain structure was produced using NASA's low cost Dual Microstructure Heat Treatment (DMHT) process. The results showed the DMHT disk to have a high strength, fatigue resistant bore comparable to a subsolvus (fine grain) heat treated disk, and a creep resistant rim comparable to a supersolvus (coarse grain) heat treated disk. Additional work on subsolvus solutioning before or after the DMHT conversion appears to be a viable avenue for further improvement in disk properties.

  7. Retinal Neuroprotective Effects of Flibanserin, an FDA-Approved Dual Serotonin Receptor Agonist-Antagonist

    PubMed Central

    Ryals, Renee C.; Ku, Cristy A.; Fischer, Cody M.; Patel, Rachel C.; Datta, Shreya; Yang, Paul; Wen, Yuquan; Hen, René; Pennesi, Mark E.

    2016-01-01

    Purpose To assess the neuroprotective effects of flibanserin (formerly BIMT-17), a dual 5-HT1A agonist and 5-HT2A antagonist, in a light-induced retinopathy model. Methods Albino BALB/c mice were injected intraperitoneally with either vehicle or increasing doses of flibanserin ranging from 0.75 to 15 mg/kg flibanserin. To assess 5-HT1A-mediated effects, BALB/c mice were injected with 10 mg/kg WAY 100635, a 5-HT1A antagonist, prior to 6 mg/kg flibanserin and 5-HT1A knockout mice were injected with 6 mg/kg flibanserin. Injections were administered once immediately prior to light exposure or over the course of five days. Light exposure lasted for one hour at an intensity of 10,000 lux. Retinal structure was assessed using spectral domain optical coherence tomography and retinal function was assessed using electroretinography. To investigate the mechanisms of flibanserin-mediated neuroprotection, gene expression, measured by RT-qPCR, was assessed following five days of daily 15 mg/kg flibanserin injections. Results A five-day treatment regimen of 3 to 15 mg/kg of flibanserin significantly preserved outer retinal structure and function in a dose-dependent manner. Additionally, a single-day treatment regimen of 6 to 15 mg/kg of flibanserin still provided significant protection. The action of flibanserin was hindered by the 5-HT1A antagonist, WAY 100635, and was not effective in 5-HT1A knockout mice. Creb, c-Jun, c-Fos, Bcl-2, Cast1, Nqo1, Sod1, and Cat were significantly increased in flibanserin-injected mice versus vehicle-injected mice. Conclusions Intraperitoneal delivery of flibanserin in a light-induced retinopathy mouse model provides retinal neuroprotection. Mechanistic data suggests that this effect is mediated through 5-HT1A receptors and that flibanserin augments the expression of genes capable of reducing mitochondrial dysfunction and oxidative stress. Since flibanserin is already FDA-approved for other indications, the potential to repurpose this drug for

  8. Kainate receptor trafficking: physiological roles and molecular mechanisms.

    PubMed

    Isaac, John T R; Mellor, Jack; Hurtado, David; Roche, Katherine W

    2004-12-01

    Recently, there has been intense interest in the mechanisms regulating the trafficking and synaptic targeting of kainate receptors in neurons. This topic is still in its infancy when compared with studies of trafficking of other ionotropic glutamate receptors; however, it is already clear that mechanisms exist for subunit- and splice variant-specific trafficking of kainate receptors. There is also enormous diversity of kainate receptor targeting, with the best-studied neurons in this regard being hippocampal CA3 pyramidal neurons and CA1 GABAergic interneurons. This review summarizes the current state of knowledge on this topic, focusing on the molecular mechanisms of kainate receptor trafficking and the potential for these mechanisms to regulate neuronal kainate receptor function.

  9. Piperazinyl carbamate fatty acid amide hydrolase inhibitors and transient receptor potential channel modulators as "dual-target" analgesics.

    PubMed

    Maione, Sabatino; Costa, Barbara; Piscitelli, Fabiana; Morera, Enrico; De Chiaro, Maria; Comelli, Francesca; Boccella, Serena; Guida, Francesca; Verde, Roberta; Ortar, Giorgio; Di Marzo, Vincenzo

    2013-10-01

    We showed previously that inhibiting fatty acid amide hydrolase (FAAH), an endocannabinoid degrading enzyme, and transient receptor potential vanilloid type-1 (TRPV1) channels with the same molecule, the naturally occurring N-arachidonoyl-serotonin (AA-5-HT), produces more efficacious anti-nociceptive and anti-hyperalgesic actions than the targeting of FAAH or TRPV1 alone. We also reported the synthesis of some piperazinyl carbamates as "dual" FAAH inhibitors and either antagonists at TRPV1 or agonists/desensitizers of the transient receptor potential ankyrin type-1 (TRPA1) cannel, another target for analgesic drugs. We investigated here if two such compounds, the FAAH/TRPV1 blocker OMDM198 and the FAAH inhibitor/TRPA1 agonist, OMDM202, exert anti-nociceptive actions in the formalin test of pain in mice, and through what mechanism. Both compounds inhibited the second phase of the response to formalin, the effect being maximal at 3 mg/kg, i.p. Antagonism of CB1 or CB2 receptors with AM251 or AM630 (1 mg/kg, i.p.), respectively, reversed this effect. A TRPV1 agonist, palvanil (0.1 mg/kg, i.p.), also reversed the analgesic effect of OMDM198. OMDM202 action was also antagonized by a per se inactive dose of the selective TRPA1 blocker, AP-18 (0.05 mg/kg, i.p.), but not by a TRPV1 antagonist. AP-18 at higher doses (0.1-0.2 mg/kg) inhibited both the first and second phase of the formalin response. The effects of OMDM198 and OMDM202 were accompanied by elevation of anandamide levels in the spinal cord. OMDM198 (0.1-5.0 mg/kg, i.p.) also reversed carrageenan-induced oedema and thermal hyperalgesia in mice with efficacy similar to that of AA-5-HT. These data suggest that "dual" fatty acid amide hydrolase and transient receptor potential channel modulators should be clinically evaluated as novel analgesics.

  10. Receptor recognition mechanisms of coronaviruses: a decade of structural studies.

    PubMed

    Li, Fang

    2015-02-01

    Receptor recognition by viruses is the first and essential step of viral infections of host cells. It is an important determinant of viral host range and cross-species infection and a primary target for antiviral intervention. Coronaviruses recognize a variety of host receptors, infect many hosts, and are health threats to humans and animals. The receptor-binding S1 subunit of coronavirus spike proteins contains two distinctive domains, the N-terminal domain (S1-NTD) and the C-terminal domain (S1-CTD), both of which can function as receptor-binding domains (RBDs). S1-NTDs and S1-CTDs from three major coronavirus genera recognize at least four protein receptors and three sugar receptors and demonstrate a complex receptor recognition pattern. For example, highly similar coronavirus S1-CTDs within the same genus can recognize different receptors, whereas very different coronavirus S1-CTDs from different genera can recognize the same receptor. Moreover, coronavirus S1-NTDs can recognize either protein or sugar receptors. Structural studies in the past decade have elucidated many of the puzzles associated with coronavirus-receptor interactions. This article reviews the latest knowledge on the receptor recognition mechanisms of coronaviruses and discusses how coronaviruses have evolved their complex receptor recognition pattern. It also summarizes important principles that govern receptor recognition by viruses in general.

  11. Identifying the receptor subtype selectivity of retinoid X and retinoic acid receptors via quantum mechanics.

    PubMed

    Tsuji, Motonori; Shudo, Koichi; Kagechika, Hiroyuki

    2017-03-01

    Understanding and identifying the receptor subtype selectivity of a ligand is an important issue in the field of drug discovery. Using a combination of classical molecular mechanics and quantum mechanical calculations, this report assesses the receptor subtype selectivity for the human retinoid X receptor (hRXR) and retinoic acid receptor (hRAR) ligand-binding domains (LBDs) complexed with retinoid ligands. The calculated energies show good correlation with the experimentally reported binding affinities. The technique proposed here is a promising method as it reveals the origin of the receptor subtype selectivity of selective ligands.

  12. The pesticin receptor of Yersinia enterocolitica: a novel virulence factor with dual function.

    PubMed

    Rakin, A; Saken, E; Harmsen, D; Heesemann, J

    1994-07-01

    The iron-repressible outer membrane protein FyuA of Yersinia enterocolitica operates as a receptor with dual function: (i) as a receptor for the Y. pestis bacteriocin pesticin, and (ii) as a receptor for yersiniabactin, a siderophore that is produced by mouse-virulent Y. enterocolitica strains of biogroup IB. Cloning of the FyuA-encoding gene was achieved by mobilization of a genomic cosmid library of the pesticin-sensitive and mouse-virulent Y. enterocolitica O:8 strain WA into the pesticin-resistant WA fyuA mutant and subsequent in vivo selection of transconjugants for the ability to survive and multiply in mice (phenotype mouse virulence). The reisolated transconjugants which survived in mice for 3 d harboured a unique cosmid and phenotypically were pesticin sensitive. From this cosmid a 2650 bp SalI-PstI fragment conferring pesticin sensitivity was subcloned. Sequencing of this DNA fragment revealed a single open reading frame of 2022 bp, which encodes a deduced polypeptide of 673 amino acids with a predicted molecular mass of 73,677 Da. Cleavage of a putative signal sequence composed of 22 amino acids should lead to a mature protein of 651 amino acids with a molecular mass of 71,368 Da. The open reading frame is preceded by a sequence which shares homology with the postulated consensus Fur iron-repressor protein-binding site. FyuA shows homology to other iron-regulated TonB-dependent outer membrane proteins with receptor functions (e.g. BtuB, CirA, FepA, IutA, FhuA, FoxA, FcuA). On the basis of multiple alignment of amino acid sequences of FyuA and other TonB-dependent receptors, a phylogenetic tree was constructed, demonstrating that FyuA probably belongs to the citrate subfamily or represents a new subfamily of TonB-dependent receptors. Moreover, by complementation of the WA fyuA mutant by the cloned fyuA gene, yersiniabactin uptake and mouse virulence were restored. These studies demonstrate that the cloned pesticin/yersiniabactin receptor FyuA of Y

  13. Synthesis of iboga-like isoquinuclidines: Dual opioid receptors agonists having antinociceptive properties.

    PubMed

    Banerjee, Tuhin Suvro; Paul, Sibasish; Sinha, Surajit; Das, Sumantra

    2014-11-01

    Some novel iboga-analogues consisting of benzofuran moiety and dehydroisoquinuclidine ring connected by -CH2-, (CH2)2 and (CH2)3 linkers have been synthesized with the view to develop potential antinociceptive drugs. The compounds 14 and 21 showed binding at the μ-opioid receptor (MOR), while the compound 11a exhibited dual affinities at both MOR and κ-opioid receptor (KOR). MAP kinase activation indicated all three compounds have opioid agonistic properties. The presence of a double bond and endo-methylcarboxylate group in the dehydroisoquinuclidine ring and the benzofuran and methylene spacer appeared to be essential for opioid receptor binding. Further studies demonstrated 11a caused significant antinociception in mice in the hot-plate test which was comparable to that produced by morphine. The compound 11a was also found to be nontremorigenic unlike various iboga congeners. This study identifies a new pharmacophore which may lead to the development of suitable substitute of morphine in the treatment of pain.

  14. Nuclear receptor Nurr1 agonists enhance its dual functions and improve behavioral deficits in an animal model of Parkinson's disease.

    PubMed

    Kim, Chun-Hyung; Han, Baek-Soo; Moon, Jisook; Kim, Deog-Joong; Shin, Joon; Rajan, Sreekanth; Nguyen, Quoc Toan; Sohn, Mijin; Kim, Won-Gon; Han, Minjoon; Jeong, Inhye; Kim, Kyoung-Shim; Lee, Eun-Hye; Tu, Yupeng; Naffin-Olivos, Jacqueline L; Park, Chang-Hwan; Ringe, Dagmar; Yoon, Ho Sup; Petsko, Gregory A; Kim, Kwang-Soo

    2015-07-14

    Parkinson's disease (PD), primarily caused by selective degeneration of midbrain dopamine (mDA) neurons, is the most prevalent movement disorder, affecting 1-2% of the global population over the age of 65. Currently available pharmacological treatments are largely symptomatic and lose their efficacy over time with accompanying severe side effects such as dyskinesia. Thus, there is an unmet clinical need to develop mechanism-based and/or disease-modifying treatments. Based on the unique dual role of the nuclear orphan receptor Nurr1 for development and maintenance of mDA neurons and their protection from inflammation-induced death, we hypothesize that Nurr1 can be a molecular target for neuroprotective therapeutic development for PD. Here we show successful identification of Nurr1 agonists sharing an identical chemical scaffold, 4-amino-7-chloroquinoline, suggesting a critical structure-activity relationship. In particular, we found that two antimalarial drugs, amodiaquine and chloroquine stimulate the transcriptional function of Nurr1 through physical interaction with its ligand binding domain (LBD). Remarkably, these compounds were able to enhance the contrasting dual functions of Nurr1 by further increasing transcriptional activation of mDA-specific genes and further enhancing transrepression of neurotoxic proinflammatory gene expression in microglia. Importantly, these compounds significantly improved behavioral deficits in 6-hydroxydopamine lesioned rat model of PD without any detectable signs of dyskinesia-like behavior. These findings offer proof of principle that small molecules targeting the Nurr1 LBD can be used as a mechanism-based and neuroprotective strategy for PD.

  15. Dual role of the receptor Tom20 in specificity and efficiency of protein import into mitochondria

    PubMed Central

    Yamamoto, Hayashi; Itoh, Nobuka; Kawano, Shin; Yatsukawa, Yoh-ichi; Momose, Takaki; Makio, Tadashi; Matsunaga, Mayumi; Yokota, Mihoko; Esaki, Masatoshi; Shodai, Toshihiro; Kohda, Daisuke; Aiken Hobbs, Alyson E.; Jensen, Robert E.; Endo, Toshiya

    2011-01-01

    Mitochondria import most of their resident proteins from the cytosol, and the import receptor Tom20 of the outer-membrane translocator TOM40 complex plays an essential role in specificity of mitochondrial protein import. Here we analyzed the effects of Tom20 binding on NMR spectra of a long mitochondrial presequence and found that it contains two distinct Tom20-binding elements. In vitro import and cross-linking experiments revealed that, although the N-terminal Tom20-binding element is essential for targeting to mitochondria, the C-terminal element increases efficiency of protein import in the step prior to translocation across the inner membrane. Therefore Tom20 has a dual role in protein import into mitochondria: recognition of the targeting signal in the presequence and tethering the presequence to the TOM40 complex to increase import efficiency. PMID:21173275

  16. Dual targeting of androgen receptor and mTORC1 by salinomycin in prostate cancer

    PubMed Central

    Jiang, Shoulei; Cropper, Jodie; Werner, Sherry L.; Song, Chung S.; Chatterjee, Bandana

    2016-01-01

    Androgen receptor (AR) and PI3K/AKT/mTORC1 are major survival signals that drive prostate cancer to a lethal disease. Reciprocal activation of these oncogenic pathways from negative cross talks contributes to low/limited success of pathway-selective inhibitors in curbing prostate cancer progression. We report that the antibiotic salinomycin, a cancer stem cell blocker, is a dual-acting AR and mTORC1 inhibitor, inhibiting PTEN-deficient castration-sensitive and castration-resistant prostate cancer in culture and xenograft tumors. AR expression, its transcriptional activity, and androgen biosynthesis regulating enzymes CYP17A1, HSD3β1 were reduced by sub-micro molar salinomycin. Estrogen receptor-α expression was unchanged. Loss of phosphorylated AR at serine-81, which is an index for nuclear AR activity, preceded total AR reduction. Rapamycin enhanced the AR protein level without altering phosphoAR-Ser81 and CYP17A1. Inactivation of mTORC1, evident from reduced phosphorylation of mTOR and downstream effectors, as well as AMPK activation led to robust autophagy induction. Apoptosis increased modestly, albeit significantly, by sub-micro molar salinomycin. Enhanced stimulatory TSC2 phosphorylation at Ser-1387 by AMPK, and reduced inhibitory TSC2 phosphorylation at Ser-939/Thr-1462 catalyzed by AKT augmented TSC2/TSC1 activity, which led to mTORC1 inhibition. AMPK-mediated raptor phosphorylation further reduced mTOR's kinase function and mTORC1 activity. Our novel finding on dual inhibition of AR and mTORC1 suggests that salinomycin is potentially active as monotherapy against advanced prostate cancer. PMID:27557496

  17. Quantitative electroencephalography within sleep/wake states differentiates GABAA modulators eszopiclone and zolpidem from dual orexin receptor antagonists in rats.

    PubMed

    Fox, Steven V; Gotter, Anthony L; Tye, Spencer J; Garson, Susan L; Savitz, Alan T; Uslaner, Jason M; Brunner, Joseph I; Tannenbaum, Pamela L; McDonald, Terrence P; Hodgson, Robert; Yao, Lihang; Bowlby, Mark R; Kuduk, Scott D; Coleman, Paul J; Hargreaves, Richard; Winrow, Christopher J; Renger, John J

    2013-11-01

    Dual orexin receptor antagonists (DORAs) induce sleep by blocking orexin 1 and orexin 2 receptor-mediated activities responsible for regulating wakefulness. DORAs represent a potential alternative mechanism to the current standard of care that includes the γ-aminobutyric acid (GABA)A receptor-positive allosteric modulators, eszopiclone and zolpidem. This work uses an innovative method to analyze electroencephalogram (EEG) spectral frequencies within sleep/wake states to differentiate the effects of GABAA modulators from DORA-22, an analog of the DORA MK-6096, in Sprague-Dawley rats. The effects of low, intermediate, and high doses of eszopiclone, zolpidem, and DORA-22 were examined after first defining each compound's ability to promote sleep during active-phase dosing. The EEG spectral frequency power within specific sleep stages was calculated in 1-Hz intervals from 1 to 100 Hz within each sleep/wake state for the first 4 h after the dose. Eszopiclone and zolpidem produced marked, dose-responsive disruptions in sleep stage-specific EEG spectral profiles compared with vehicle treatment. In marked contrast, DORA-22 exhibited marginal changes in the spectral profile, observed only during rapid eye movement sleep, and only at the highest dose tested. Moreover, while eszopiclone- and zolpidem-induced changes were evident in the inactive period, the EEG spectral responses to DORA-22 were absent during this phase. These results suggest that DORA-22 differs from eszopiclone and zolpidem whereby DORA-22 promotes somnolence without altering the neuronal network EEG activity observed during normal sleep.

  18. Effects of Strain Rates on Mechanical Properties and Fracture Mechanism of DP780 Dual Phase Steel

    NASA Astrophysics Data System (ADS)

    Li, Shengci; Kang, Yonglin; Zhu, Guoming; Kuang, Shuang

    2015-06-01

    The mechanical properties of DP780 dual phase steel were measured by quasi-static and high-speed tensile tests at strain rates between 0.001 and 1000 s-1 at room temperature. The deformation and fracture mechanisms were analyzed by observation of the tensile fracture and microstructure near the fracture. Dynamic factor and feret ratio quantitative methods were applied to study the effect of strain rate on the microstructure and properties of DP780 steel. The constitutive relation was described by a modified Johnson-Cook and Zerilli-Armstrong model. The results showed that the strain rate sensitivity of yield strength is bigger than that of ultimate tensile strength; as strain rate increased, the formation of microcracks and voids at the ferrite/martensite interface can be alleviated; the strain rate effect is unevenly distributed in the plastic deformation region. Moreover, both models can effectively describe the experimental results, while the modified Zerilli-Armstrong model is more accurate because the strain-hardening rate of this model is independent of strain rate.

  19. Mechanism of the estrogen receptor interaction with 4-hydroxytamoxifen

    SciTech Connect

    Sasson, S.; Notides, A.C.

    1988-04-01

    The binding mechanism of the estrogen receptor with 4-(/sup 3/H)hydroxytamoxifen was investigated. The equilibrium binding analysis with 4-(/sup 3/H)hydroxytamoxifen indicated a positive cooperative interaction: the Scatchard plot was convex and the Hill coefficient was 1.4-1.5. This binding appears similar to the positively cooperative interaction of the estrogen receptor with (/sup 3/H)estradiol. However, a competitive binding assay with a saturating concentration of (/sup 3/H) estradiol and variable concentrations of 4-hydroxytamoxifen produced nonparallel displacement curves indicating that the binding mechanism of the receptor with these two ligands is different. The competitive binding assay with (/sup 3/H)estradiol and 4-hydroxytamoxifen at constant molar ratios demonstrated that the receptor's affinity for estradiol was reduced and the receptor preferentially bound 4-hydroxytamoxifen. These data suggest that 4-hydroxytamoxifen interacts with the receptor differently than estradiol; it antagonizes the binding of estradiol when these two ligands are simultaneously present.

  20. Dual disease resistance mediated by the immune receptor Cf-2 in tomato requires a common virulence target of a fungus and a nematode.

    PubMed

    Lozano-Torres, Jose L; Wilbers, Ruud H P; Gawronski, Piotr; Boshoven, Jordi C; Finkers-Tomczak, Anna; Cordewener, Jan H G; America, Antoine H P; Overmars, Hein A; Van 't Klooster, John W; Baranowski, Lukasz; Sobczak, Miroslaw; Ilyas, Muhammad; van der Hoorn, Renier A L; Schots, Arjen; de Wit, Pierre J G M; Bakker, Jaap; Goverse, Aska; Smant, Geert

    2012-06-19

    Plants lack the seemingly unlimited receptor diversity of a somatic adaptive immune system as found in vertebrates and rely on only a relatively small set of innate immune receptors to resist a myriad of pathogens. Here, we show that disease-resistant tomato plants use an efficient mechanism to leverage the limited nonself recognition capacity of their innate immune system. We found that the extracellular plant immune receptor protein Cf-2 of the red currant tomato (Solanum pimpinellifolium) has acquired dual resistance specificity by sensing perturbations in a common virulence target of two independently evolved effectors of a fungus and a nematode. The Cf-2 protein, originally identified as a monospecific immune receptor for the leaf mold fungus Cladosporium fulvum, also mediates disease resistance to the root parasitic nematode Globodera rostochiensis pathotype Ro1-Mierenbos. The Cf-2-mediated dual resistance is triggered by effector-induced perturbations of the apoplastic Rcr3(pim) protein of S. pimpinellifolium. Binding of the venom allergen-like effector protein Gr-VAP1 of G. rostochiensis to Rcr3(pim) perturbs the active site of this papain-like cysteine protease. In the absence of the Cf-2 receptor, Rcr3(pim) increases the susceptibility of tomato plants to G. rostochiensis, thus showing its role as a virulence target of these nematodes. Furthermore, both nematode infection and transient expression of Gr-VAP1 in tomato plants harboring Cf-2 and Rcr3(pim) trigger a defense-related programmed cell death in plant cells. Our data demonstrate that monitoring host proteins targeted by multiple pathogens broadens the spectrum of disease resistances mediated by single plant immune receptors.

  1. Intact learning and memory in rats following treatment with the dual orexin receptor antagonist almorexant

    PubMed Central

    Jenck, François

    2010-01-01

    Rationale Orexins play a key role in the maintenance of alertness and are implicated in the modulation of diverse physiological processes, including cognitive function. Almorexant, a dual orexin receptor antagonist, transiently and reversibly blocks the action of orexin peptides at both OX1 and OX2 receptors and increases time spent in rapid eye movement (REM) and non-REM sleep. Objectives We explored the direct effects on learning and memory of single and repeated administration of almorexant in rats. Methods Following administration of high doses of almorexant (300 mg/kg, p.o.), scopolamine (0.8 mg/kg, i.p.), combination almorexant-scopolamine, or vehicle alone, rats were trained on a Morris water maze spatial navigation task, or on a passive avoidance task. Results Rats treated with almorexant learned the spatial navigation task with similar efficacy as vehicle-treated animals. After 4 days, almorexant—but not vehicle-treated rats had established spatial memory; after 8 days, spatial memory had been established in both vehicle—and almorexant-treated rats. Scopolamine-treated rats failed to learn the spatial task. Both vehicle—and almorexant—but not scopolamine-treated rats demonstrated passive avoidance learning. Almorexant did not ameliorate scopolamine-induced impairment of learning in either task. Conclusions Rats treated with almorexant are fully capable of spatial and avoidance learning. PMID:20631993

  2. Modeling biofilms with dual extracellular electron transfer mechanisms

    SciTech Connect

    Renslow, Ryan S.; Babauta, Jerome T.; Kuprat, Andrew P.; Schenk, Jim; Ivory, Cornelius; Fredrickson, Jim K.; Beyenal, Haluk

    2013-11-28

    Electrochemically active biofilms have a unique form of respiration in which they utilize solid external materials as their terminal electron acceptor for metabolism. Currently, two primary mechanisms have been identified for long-range extracellular electron transfer (EET): a diffusion- and a conduction-based mechanism. Evidence in the literature suggests that some biofilms, particularly Shewanella oneidensis, produce components requisite for both mechanisms. In this study, a generic model is presented that incorporates both diffusion- and conduction-based mechanisms and allows electrochemically active biofilms to utilize both simultaneously. The model was applied to Shewanella oneidensis and Geobacter sulfurreducens biofilms using experimentally generated data found the literature. Our simulation results showed that 1) biofilms having both mechanisms available, especially if they can interact, may have metabolic advantage over biofilms that can use only a single mechanism; 2) the thickness of Geobacter sulfurreducens biofilms is likely not limited by conductivity; 3) accurate intrabiofilm diffusion coefficient values are critical for current generation predictions; and 4) the local biofilm potential and redox potential are two distinct measurements and cannot be assumed to have identical values. Finally, we determined that cyclic and squarewave voltammetry are currently not good tools to determine the specific percentage of extracellular electron transfer mechanisms used by biofilms. The developed model will be a critical tool in designing experiments to explain EET mechanisms.

  3. Modeling biofilms with dual extracellular electron transfer mechanisms

    PubMed Central

    Renslow, Ryan; Babauta, Jerome; Kuprat, Andrew; Schenk, Jim; Ivory, Cornelius; Fredrickson, Jim; Beyenal, Haluk

    2013-01-01

    Electrochemically active biofilms have a unique form of respiration in which they utilize solid external materials as terminal electron acceptors for their metabolism. Currently, two primary mechanisms have been identified for long-range extracellular electron transfer (EET): a diffusion- and a conduction-based mechanism. Evidence in the literature suggests that some biofilms, particularly Shewanella oneidensis, produce the requisite components for both mechanisms. In this study, a generic model is presented that incorporates the diffusion- and the conduction-based mechanisms and allows electrochemically active biofilms to utilize both simultaneously. The model was applied to S. oneidensis and Geobacter sulfurreducens biofilms using experimentally generated data found in the literature. Our simulation results show that 1) biofilms having both mechanisms available, especially if they can interact, may have a metabolic advantage over biofilms that can use only a single mechanism; 2) the thickness of G. sulfurreducens biofilms is likely not limited by conductivity; 3) accurate intrabiofilm diffusion coefficient values are critical for current generation predictions; and 4) the local biofilm potential and redox potential are two distinct parameters and cannot be assumed to have identical values. Finally, we determined that simulated cyclic and squarewave voltammetry based on our model are currently not capable of determining the specific percentages of extracellular electron transfer mechanisms in a biofilm. The developed model will be a critical tool for designing experiments to explain EET mechanisms. PMID:24113651

  4. Systematic review of SGLT2 receptor inhibitors in dual or triple therapy in type 2 diabetes

    PubMed Central

    Clar, Christine; Gill, James Alexander; Court, Rachel; Waugh, Norman

    2012-01-01

    Background Despite the number of medications for type 2 diabetes, many people with the condition do not achieve good glycaemic control. Some existing glucose-lowering agents have adverse effects such as weight gain or hypoglycaemia. Type 2 diabetes tends to be a progressive disease, and most patients require treatment with combinations of glucose-lowering agents. The sodium glucose co-transporter 2 (SGLT2) receptor inhibitors are a new class of glucose-lowering agents. Objective To assess the clinical effectiveness and safety of the SGLT2 receptor inhibitors in dual or triple therapy in type 2 diabetes. Data sources MEDLINE, Embase, Cochrane Library (all sections); Science Citation Index; trial registries; conference abstracts; drug regulatory authorities; bibliographies of retrieved papers. Inclusion criteria Randomised controlled trials of SGLT2 receptor inhibitors compared with placebo or active comparator in type 2 diabetes in dual or combination therapy. Methods Systematic review. Quality assessment used the Cochrane risk of bias score. Results Seven trials, published in full, assessed dapagliflozin and one assessed canagliflozin. Trial quality appeared good. Dapagliflozin 10 mg reduced HbA1c by −0.54% (weighted mean differences (WMD), 95% CI −0.67 to −0.40) compared to placebo, but there was no difference compared to glipizide. Canagliflozin reduced HbA1c slightly more than sitagliptin (up to −0.21% vs sitagliptin). Both dapagliflozin and canagliflozin led to weight loss (dapagliflozin WMD −1.81 kg (95% CI −2.04 to −1.57), canagliflozin up to −2.3 kg compared to placebo). Limitations Long-term trial extensions suggested that effects were maintained over time. Data on canagliflozin are currently available from only one paper. Costs of the drugs are not known so cost-effectiveness cannot be assessed. More data on safety are needed, with the Food and Drug Administration having concerns about breast and bladder cancers. Conclusions

  5. Dual GPCR and GAG mimicry by the M3 chemokine decoy receptor

    SciTech Connect

    Alexander-Brett, Jennifer M.; Fremont, Daved H.

    2008-09-23

    Viruses have evolved a myriad of evasion strategies focused on undermining chemokine-mediated immune surveillance, exemplified by the mouse {gamma}-herpesvirus 68 M3 decoy receptor. Crystal structures of M3 in complex with C chemokine ligand 1/lymphotactin and CC chemokine ligand 2/monocyte chemoattractant protein 1 reveal that invariant chemokine features associated with G protein-coupled receptor binding are primarily recognized by the decoy C-terminal domain, whereas the N-terminal domain (NTD) reconfigures to engage divergent basic residue clusters on the surface of chemokines. Favorable electrostatic forces dramatically enhance the association kinetics of chemokine binding by M3, with a primary role ascribed to acidic NTD regions that effectively mimic glycosaminoglycan interactions. Thus, M3 employs two distinct mechanisms of chemical imitation to potently sequester chemokines, thereby inhibiting chemokine receptor binding events as well as the formation of chemotactic gradients necessary for directed leukocyte trafficking.

  6. A Dual Role for P2X7 Receptor during Porphyromonas gingivalis Infection

    PubMed Central

    Ramos-Junior, E.S.; Morandini, A.C.; Almeida-da-Silva, C.L.C.; Franco, E.J.; Potempa, J.; Nguyen, K.A.; Oliveira, A.C.; Zamboni, D.S.; Ojcius, D.M.; Scharfstein, J.

    2015-01-01

    Emerging evidence suggests a role for purinergic signaling in the activation of multiprotein intracellular complexes called inflammasomes, which control the release of potent inflammatory cytokines, such as interleukin (IL) -1β and -18. Porphyromonas gingivalis is intimately associated with periodontitis and is currently considered one of the pathogens that can subvert the immune system by limiting the activation of the NLRP3 inflammasome. We recently showed that P. gingivalis can dampen eATP-induced IL-1β secretion by means of its fimbriae in a purinergic P2X7 receptor–dependent manner. Here, we further explore the role of this purinergic receptor during eATP-induced IL-1β processing and secretion by P. gingivalis–infected macrophages. We found that NLRP3 was necessary for eATP-induced IL-1β secretion as well as for caspase 1 activation irrespective of P. gingivalis fimbriae. Additionally, although the secretion of IL-1β from P. gingivalis–infected macrophages was dependent on NLRP3, its adaptor protein ASC, or caspase 1, the cleavage of intracellular pro-IL-1β to the mature form was found to occur independently of NLRP3, its adaptor protein ASC, or caspase 1. Our in vitro findings revealed that P2X7 receptor has a dual role, being critical not only for eATP-induced IL-1β secretion but also for intracellular pro-IL-1β processing. These results were relevant in vivo since P2X7 receptor expression was upregulated in a P. gingivalis oral infection model, and reduced IFN-γ and IL-17 were detected in draining lymph node cells from P2rx7-/- mice. Furthermore, we demonstrated that P2X7 receptor and NLRP3 transcription were modulated in human chronic periodontitis. Overall, we conclude that the P2X7 receptor has a role in periodontal immunopathogenesis and suggest that targeting of the P2X7/NLRP3 pathway should be considered in future therapeutic interventions in periodontitis. PMID:26152185

  7. Presynaptic glutamate receptors: physiological functions and mechanisms of action.

    PubMed

    Pinheiro, Paulo S; Mulle, Christophe

    2008-06-01

    Glutamate acts on postsynaptic glutamate receptors to mediate excitatory communication between neurons. The discovery that additional presynaptic glutamate receptors can modulate neurotransmitter release has added complexity to the way we view glutamatergic synaptic transmission. Here we review evidence of a physiological role for presynaptic glutamate receptors in neurotransmitter release. We compare the physiological roles of ionotropic and metabotropic glutamate receptors in short- and long-term regulation of synaptic transmission. Furthermore, we discuss the physiological conditions that are necessary for their activation, the source of the glutamate that activates them, their mechanisms of action and their involvement in higher brain function.

  8. Turnover of Acetylcholine Receptors: Mechanisms of Regulation

    DTIC Science & Technology

    1988-12-01

    ME, Whittingham S, and Duane DD (1976) Antibody to acetylcholine receptor in myasthenia gravis : prevalance, clinical correlates and diagnostic value...transferred to nitorcellulose. Proc Natl Acad Sci 77:5201-5205. Weinberg CB and Hall ZW (1979) Antibodies from patients with myasthenia gravis recognize

  9. Dual role of dopamine D(2)-like receptors in the mediation of conditioned and unconditioned fear.

    PubMed

    Brandão, Marcus Lira; de Oliveira, Amanda Ribeiro; Muthuraju, Sangu; Colombo, Ana Caroline; Saito, Viviane Mitsuko; Talbot, Teddy

    2015-11-14

    A reduction of dopamine release or D2 receptor blockade in the terminal fields of the mesolimbic system, particularly the amygdala, clearly reduces conditioned fear. Similar D2 receptor antagonism in the neural substrates of fear in the midbrain tectum attenuates the processing of unconditioned aversive information. However, the implications of the interplay between opposing actions of dopamine in the rostral and caudal segments of the dopaminergic system are still unclear. Previous studies from this laboratory have reported the effects of dopaminergic drugs on behavior in rats in the elevated plus maze, auditory-evoked potentials (AEPs) recorded from the midbrain tectum, fear-potentiated startle, and conditioned freezing. These findings led to an interesting framework on the functional roles of dopamine in both anxiety and fear states. Dopamine D2 receptor inhibition in the terminal fields of the mesolimbic dopamine system generally causes anxiolytic-like effects, whereas the activity of midbrain substrates of unconditioned fear are enhanced by D2 receptor antagonists, suggesting that D2 receptor-mediated mechanisms play opposing roles in fear/anxiety processes, depending on the brain region under study. Dopamine appears to mediate conditioned fear by acting at rostral levels of the brain and regulate unconditioned fear at the midbrain level, likely by reducing the sensorimotor gating of aversive events.

  10. Discovery of Dual ETA/ETB Receptor Antagonists from Traditional Chinese Herbs through in Silico and in Vitro Screening

    PubMed Central

    Wang, Xing; Zhang, Yuxin; Liu, Qing; Ai, Zhixin; Zhang, Yanling; Xiang, Yuhong; Qiao, Yanjiang

    2016-01-01

    Endothelin-1 receptors (ETAR and ETBR) act as a pivotal regulator in the biological effects of ET-1 and represent a potential drug target for the treatment of multiple cardiovascular diseases. The purpose of the study is to discover dual ETA/ETB receptor antagonists from traditional Chinese herbs. Ligand- and structure-based virtual screening was performed to screen an in-house database of traditional Chinese herbs, followed by a series of in vitro bioassay evaluation. Aristolochic acid A (AAA) was first confirmed to be a dual ETA/ETB receptor antagonist based intracellular calcium influx assay and impedance-based assay. Dose-response curves showed that AAA can block both ETAR and ETBR with IC50 of 7.91 and 7.40 μM, respectively. Target specificity and cytotoxicity bioassay proved that AAA is a selective dual ETA/ETB receptor antagonist and has no significant cytotoxicity on HEK293/ETAR and HEK293/ETBR cells within 24 h. It is a feasible and effective approach to discover bioactive compounds from traditional Chinese herbs using in silico screening combined with in vitro bioassay evaluation. The structural characteristic of AAA for its activity was especially interpreted, which could provide valuable reference for the further structural modification of AAA. PMID:26999111

  11. Dual structural color mechanisms in a scarab beetle.

    PubMed

    Xu, Man; Seago, Ainsley E; Sutherland, Tara D; Weisman, Sarah

    2010-11-01

    The cuticle of a Mycterophallus cetoniine scarab species displays both red iridescence due to a multilayer reflector mechanism and rainbow iridescence due to a superimposed diffraction grating mechanism. This is the first reported example of an animal possessing two independent classes of structural colors arising from interference at the wavelengths of visible light. In this work, the Mycterophallus cuticle is characterized by light microscopy, spectrophotometry, scanning electron microscopy, and transmission electron microscopy. We compare the cuticle of the Mycterophallus species to two closely related Lomaptera scarab species, one with only a multilayer reflector and the second with only a diffraction grating. We calculate the correspondence between the nanostructural parameters and the optical properties of the Mycterophallus cuticle to determine the relative optical contributions of the two color mechanisms and the interactions between them.

  12. Dual actions of (-)-stepholidine on the dopamine receptor-mediated adenylate cyclase activity in rat corpus striatum.

    PubMed

    Dong, Z J; Guo, X; Chen, L J; Han, Y F; Jin, G Z

    1997-01-01

    (-)-Stepholidine (SPD) is an antagonist of normosensitive dopamine (DA) receptors, but it exhibits D1 agonistic action on rotational behaviour in rats with unilateral 6-hydroxydopamine (6-OHDA) lesions of the substantia nigra pars compacta (SNC). In the present study, agonistic and antagonistic effects of SPD on the DA receptor-mediated synaptosomal adenylate cyclase (AC) activity in rat striatum were investigated. After blockade of D2 receptors, SPD augmented AC activity dose-dependently. The EC50 value was 41.1 +/- 8.6 micromol/L. At the concentration of 10 micromol/L, SPD increased cAMP formation from a basal level (50.8 +/- 10.3 pmol/mg protein/min) to 133.7 +/- 31.8 pmol/mg protein/min. The SPD-induced stimulation of AC activity was almost completely reversed by 10 micromol/L Sch23390. These results indicate that SPD possesses an agonistic action on the D1 receptor. Forskolin-stimulated adenylate cyclase (FSAC) activity was used as a model to elucidate the effect of SPD on D2 receptors. The results indicate that DA inhibited FSAC activity dose-dependently, while SPD partially restored FSAC activity. Taken together, these results support the conclusion that SPD has dual actions on DA receptors that mediate AC activity, i.e., an agonistic action on D1 receptors and an antagonistic action on D2 receptors.

  13. Operating mechanism for dual valves in an internal combustion engine

    SciTech Connect

    Nagahiro, K.; Ishida, A.; Kajiwara, S.

    1987-04-14

    A valve operating mechanism is described for an internal combustion engine having a camshaft, a pair of intake or exhaust valves for each engine cylinder and a rocker shaft, comprising: first and second rocker arms pivotally mounted on the rocker shaft in adjacent relationship and engaging the pair of valves. The first rocker arm engages the camshaft; and piston means in the rocker arms selectively shiftable between positions connecting the rocker arms for pivotal movement in unison and disconnecting the rocker arms for independent movement. The piston means includes two pistons slidably mounted in the first rocker arm with one piston slidable into the second rocker arm for connecting the first and second rocker arms.

  14. Dual allosteric activation mechanisms in monomeric human glucokinase.

    PubMed

    Whittington, A Carl; Larion, Mioara; Bowler, Joseph M; Ramsey, Kristen M; Brüschweiler, Rafael; Miller, Brian G

    2015-09-15

    Cooperativity in human glucokinase (GCK), the body's primary glucose sensor and a major determinant of glucose homeostatic diseases, is fundamentally different from textbook models of allostery because GCK is monomeric and contains only one glucose-binding site. Prior work has demonstrated that millisecond timescale order-disorder transitions within the enzyme's small domain govern cooperativity. Here, using limited proteolysis, we map the site of disorder in unliganded GCK to a 30-residue active-site loop that closes upon glucose binding. Positional randomization of the loop, coupled with genetic selection in a glucokinase-deficient bacterium, uncovers a hyperactive GCK variant with substantially reduced cooperativity. Biochemical and structural analysis of this loop variant and GCK variants associated with hyperinsulinemic hypoglycemia reveal two distinct mechanisms of enzyme activation. In α-type activation, glucose affinity is increased, the proteolytic susceptibility of the active site loop is suppressed and the (1)H-(13)C heteronuclear multiple quantum coherence (HMQC) spectrum of (13)C-Ile-labeled enzyme resembles the glucose-bound state. In β-type activation, glucose affinity is largely unchanged, proteolytic susceptibility of the loop is enhanced, and the (1)H-(13)C HMQC spectrum reveals no perturbation in ensemble structure. Leveraging both activation mechanisms, we engineer a fully noncooperative GCK variant, whose functional properties are indistinguishable from other hexokinase isozymes, and which displays a 100-fold increase in catalytic efficiency over wild-type GCK. This work elucidates specific structural features responsible for generating allostery in a monomeric enzyme and suggests a general strategy for engineering cooperativity into proteins that lack the structural framework typical of traditional allosteric systems.

  15. Dynamic Regulation of the GABAA Receptor Function by Redox Mechanisms.

    PubMed

    Calvo, Daniel J; Beltrán González, Andrea N

    2016-09-01

    Oxidizing and reducing agents, which are currently involved in cell metabolism and signaling pathways, can regulate fast inhibitory neurotransmission mediated by GABA receptors in the nervous system. A number of in vitro studies have shown that diverse redox compounds, including redox metabolites and reactive oxygen and nitrogen species, modulate phasic and tonic responses mediated by neuronal GABAA receptors through both presynaptic and postsynaptic mechanisms. We review experimental data showing that many redox agents, which are normally present in neurons and glia or are endogenously generated in these cells under physiologic states or during oxidative stress (e.g., hydrogen peroxide, superoxide and hydroxyl radicals, nitric oxide, ascorbic acid, and glutathione), induce potentiating or inhibiting actions on different native and recombinant GABAA receptor subtypes. Based on these results, it is thought that redox signaling might represent a homeostatic mechanism that regulates the function of synaptic and extrasynaptic GABAA receptors in physiologic and pathologic conditions.

  16. Bilingualism modulates dual mechanisms of cognitive control: Evidence from ERPs.

    PubMed

    Morales, Julia; Yudes, Carolina; Gómez-Ariza, Carlos J; Bajo, M Teresa

    2015-01-01

    Recent behavioral findings with the AX-Continous Performance Task (AX-CPT; Morales et al., 2013) show that bilinguals only outperform monolinguals under conditions that require the highest adjustment between monitoring (proactive) and inhibitory (reactive) control, which supports the idea that bilingualism modulates the coordination of different control mechanisms. In an ERP experiment we aimed to further investigate the role that bilingualism plays in the dynamic combination of proactive and reactive control in the AX-CPT. Our results strongly indicate that bilingualism facilitates an effective adjustment between both components of cognitive control. First, we replicated previous behavioral results. Second, ERP components indicated that bilingualism influences the conflict monitoring, response inhibition and error monitoring components of control (as indexed by the N2 and P3a elicited by the probe and the error-related negativity following incorrect responses, respectively). Thus, bilinguals exerted higher reactive control than monolinguals but only when they needed to overcome the competing cue-information. These findings join others in suggesting that a better understanding of the cognitive benefits of bilingualism may require consideration of a multi-component perspective.

  17. A mechanical cooler for dual-temperature applications

    NASA Astrophysics Data System (ADS)

    Gully, W.; Carrington, H.; Kiehl, W.; Byrne, Kevin

    1998-01-01

    Ball Aerospace has been developing Stirling cycle mechanical cryocoolers specifically for space applications. These coolers are special in that they are designed from the beginning for power efficiency, high reliability, and compatibility with sensitive instruments. We have delivered several of these coolers to NASA Goddard Space Flight Center, and are currently assembling one for the High Resolution Dynamics Limb Sounder (HIRDLS) program. In our current research effort, funded by the Ballistic Missile Defense Organization (BMDO), we are tailoring our basic design to new requirements from the Air Force Research Laboratory and its customers. We describe our success in optimizing a cooler to efficiently provide refrigeration at two different temperatures simultaneously. This two-temperature application requires 0.4 W of cooling at 35 K, and 0.6 W of cooling at 60 K. We have met these requirements with an input power of approximately 70 W from a dc source with a breadboard version of the cooler. We expect to deliver the protoflight version of this cooler to the Air Force Research Laboratory in January 1998.

  18. Mechanisms of Regulation of the Chemokine-Receptor Network

    PubMed Central

    Stone, Martin J.; Hayward, Jenni A.; Huang, Cheng; E. Huma, Zil; Sanchez, Julie

    2017-01-01

    The interactions of chemokines with their G protein-coupled receptors promote the migration of leukocytes during normal immune function and as a key aspect of the inflammatory response to tissue injury or infection. This review summarizes the major cellular and biochemical mechanisms by which the interactions of chemokines with chemokine receptors are regulated, including: selective and competitive binding interactions; genetic polymorphisms; mRNA splice variation; variation of expression, degradation and localization; down-regulation by atypical (decoy) receptors; interactions with cell-surface glycosaminoglycans; post-translational modifications; oligomerization; alternative signaling responses; and binding to natural or pharmacological inhibitors. PMID:28178200

  19. Mechanical design and force calibration of dual-axis micromechanical probe for friction force microscopy

    SciTech Connect

    Fukuzawa, Kenji; Terada, Satoshi; Shikida, Mitsuhiro; Amakawa, Hiroaki; Zhang, Hedong; Mitsuya, Yasunaga

    2007-02-01

    A dual-axis micromechanical probe that combines a double cantilever and torsion beams is presented. This probe can reduce the mechanical cross-talk between the lateral and vertical force detections. In addition, dual-axis forces can be detected by measuring the dual-axis displacement of the probe end using the optical lever-based method used in conventional friction force microscopes (FFMs). In this paper, the mechanical design of the probe, the details of the fabrication method, FFM performance, and calibration of the friction force are discussed. The mechanical design and the microfabrication method for probes that can provide a force resolution of the order of 1 nN without mechanical cross-talk are presented. Calibration of the lateral force signal is possible by using the relationship between the lateral force and the piezodisplacement at the onset of the probe scanning. The micromechanical probe enables simultaneous and independent detection of atomic and friction forces. This leads to accurate investigation of nanotribological phenomena and visualization of the distribution of the friction properties, which helps the identification of the material properties.

  20. Analysis method and principle of dual-mode electro-mechanical variable transmission program

    NASA Astrophysics Data System (ADS)

    Li, Hongcai; Yan, Qingdong; Xiang, Changle; Wang, Weida

    2012-05-01

    Automotive industry, as an important pillar of the national economy, has been rapidly developing in recent years. But proplems such as energy comsumption and environmental pollution are posed at the same time. Electro-mechanical variable transmission system is considered one of avilable workarounds. It is brought forward a kind of design methods of dual-mode electro-mechanical variable transmission system rotational speed characteristics and dual-mode drive diagrams. With the motor operating behavior of running in four quadrants and the speed characteristics of the simple internal and external meshing single planetary gear train, four kinds of dual-mode electro-mechanical transmission system scheme are designed. And the velocity, torque and power characteristics of one of the programs are analyzed. The magnitude of the electric split-flow power is an important factor which influences the system performance, so in the parameters matching design, it needs to reduce the power needs under the first mode of the motor. The motor, output rotational speed range and the position of the mode switching point have relationships with the characteristics design of the planetary gear set. The analysis method is to provide a reference for hybrid vehicles' design. As the involved rotational speed and torque relationships are the natural contact of every part of transmission system, a theory basis of system program and performance analysis is provided.

  1. M2 Muscarinic Receptors Induce Airway Smooth Muscle Activation via a Dual, Gβγ-mediated Inhibition of Large Conductance Ca2+-activated K+ Channel Activity*

    PubMed Central

    Zhou, Xiao-Bo; Wulfsen, Iris; Lutz, Susanne; Utku, Emine; Sausbier, Ulrike; Ruth, Peter; Wieland, Thomas; Korth, Michael

    2008-01-01

    Airway smooth muscle is richly endowed with muscarinic receptors of the M2 and M3 subtype. Stimulation of these receptors inhibits large conductance calcium-activated K+ (BK) channels, a negative feed back regulator, in a pertussis toxinsensitive manner and thus facilitates contraction. The underlying mechanism, however, is unknown. We therefore studied the activity of bovine trachea BK channels in HEK293 cells expressing the M2 or M3 receptor (M2RorM3R). In M2R- but not M3R-expressing cells, maximal effective concentrations of carbamoylcholine (CCh) inhibited whole cell BK currents by 53%. This M2R-induced inhibition was abolished by pertussis toxin treatment or overexpression of the Gβγ scavenger transducin-α. In inside-out patches, direct application of 300 nm purified Gβγ decreased channel open probability by 55%. The physical interaction of Gβγ with BK channels was confirmed by co-immunoprecipitation. Interestingly, inhibition of phospholipase C as well as protein kinase C activities also reversed the CCh effect but to a smaller (∼20%) extent. Mouse tracheal cells responded similarly to CCh, purified Gβγ and phospholipase C/protein kinase C inhibition as M2R-expressing HEK293 cells. Our results demonstrate that airway M2Rs inhibit BK channels by a dual, Gβγ-mediated mechanism, a direct membrane-delimited interaction, and the activation of the phospholipase C/protein kinase C pathway. PMID:18524769

  2. Mechanism of Positive Allosteric Modulators Acting on AMPA Receptors

    SciTech Connect

    Jin,R.; Clark, S.; Weeks, A.; Dudman, J.; Gouaux, E.; Partin, K.

    2005-01-01

    Ligand-gated ion channels involved in the modulation of synaptic strength are the AMPA, kainate, and NMDA glutamate receptors. Small molecules that potentiate AMPA receptor currents relieve cognitive deficits caused by neurodegenerative diseases such as Alzheimer's disease and show promise in the treatment of depression. Previously, there has been limited understanding of the molecular mechanism of action for AMPA receptor potentiators. Here we present cocrystal structures of the glutamate receptor GluR2 S1S2 ligand-binding domain in complex with aniracetam [1-(4-methoxybenzoyl)-2-pyrrolidinone] or CX614 (pyrrolidino-1, 3-oxazino benzo-1, 4-dioxan-10-one), two AMPA receptor potentiators that preferentially slow AMPA receptor deactivation. Both potentiators bind within the dimer interface of the nondesensitized receptor at a common site located on the twofold axis of molecular symmetry. Importantly, the potentiator binding site is adjacent to the 'hinge' in the ligand-binding core 'clamshell' that undergoes conformational rearrangement after glutamate binding. Using rapid solution exchange, patch-clamp electrophysiology experiments, we show that point mutations of residues that interact with potentiators in the cocrystal disrupt potentiator function. We suggest that the potentiators slow deactivation by stabilizing the clamshell in its closed-cleft, glutamate-bound conformation.

  3. Pharmacokinetic and pharmacodynamic interactions between almorexant, a dual orexin receptor antagonist, and desipramine.

    PubMed

    Cruz, Hans G; Hay, Justin L; Hoever, Petra; Alessi, Federica; te Beek, Erik T; van Gerven, Joop M A; Dingemanse, Jasper

    2014-08-01

    Almorexant is a dual orexin receptor antagonist (DORA) with sleep-enabling effects in humans. Insomnia is often associated with mental health problems, including depression. Hence, potential interactions with antidepressants deserve attention. Desipramine was selected as a model drug because it is mainly metabolized by CYP2D6, which is inhibited by almorexant in vitro. A single-center, randomized, placebo-controlled, two-way crossover study in 20 healthy male subjects was conducted to evaluate the pharmacokinetic and pharmacodynamic interactions between almorexant and desipramine. Almorexant 200mg or matching placebo (double-blind) was administered orally once daily in the morning for 10 days, and a single oral dose of 50mg desipramine (open-label) was administered on Day 5. Almorexant increased the exposure to desipramine 3.7-fold, suggesting that almorexant is a moderate inhibitor of desipramine metabolism through inhibition of CYP2D6. Conversely, desipramine showed no relevant effects on the pharmacokinetics of almorexant. Pharmacodynamic evaluations indicated that almorexant alone reduced visuomotor coordination, postural stability, and alertness, and slightly increased calmness. Desipramine induced a reduction in subjective alertness and an increase in pupil/iris ratio. Despite the increase in exposure to desipramine, almorexant and desipramine in combination showed the same pharmacodynamic profile as almorexant alone, except for prolonging reduced alertness and preventing the miotic effect of almorexant. Co-administration also prolonged the mydriatic effect of desipramine. Overall, repeated administration of almorexant alone or with single-dose desipramine was well tolerated. The lack of a relevant interaction with antidepressants, if confirmed for other DORAs, would be a key feature for a safer class of hypnotics.

  4. Mechanical properties and dual atmosphere tolerance of Ag-Al based braze

    SciTech Connect

    Kim, Jin Yong; Choi, Jung-Pyung; Weil, K. Scott

    2008-03-14

    Reactive air brazing (RAB) based on the silver-copper oxide system was recently developed for use in sealing high-temperature electrochemical devices such as solid oxide fuel cells. One of the concerns regarding the viability of this joining technique is the long-term stability of silver-based alloys under a high-temperature, dual oxidizing/reducing gas environment. One possible solution to improve the dual atmosphere tolerance of the silver-based system is the addition of elements which can preferentially react with oxygen over hydrogen and minimize the pore formation caused by the reaction of oxygen with hydrogen in the silver matrix. In this paper, the effects of aluminum addition into silver-based air braze filler materials on microstructure, mechanical properties, and high temperature dual atmosphere tolerance were investigated using foils and pastes of aluminum-added braze filler materials. Joints brazed with binary Ag-Al braze foils containing more than 2 at% of Al retained a metallic form of aluminum in the metallic braze filler matrix after brazing at 1000°C in air. The flexural strength of joints prepared with binary Ag-Al braze foils decreased with increase in Al content due to the formation of interfacial aluminum oxide. The existence of metallic aluminum in the braze filler matrix, however, enhanced the high temperature dual atmosphere tolerance of the silver-based braze filler, showing smaller size of porosity after dual reducing/oxidizing atmosphere tests at 800°C for 1000 hrs. The Binary and ternary braze pastes based on the Ag-Al(-Cu) system were also tried as a sealant. Alumina joints brazed with these pastes showed increase in flexural strength with Cu content. However, a braze filler containing 5 at% Al and 8 at% Cu possessed nearly no metallic aluminum in the braze filler matrix after brazing, while the as-brazed sample prepared using a binary braze filler with 5 at% Al kept some metallic Al in the braze matrix. Thus, the addition of copper

  5. Dual agonist Surrobody™ simultaneously activates death receptors DR4 and DR5 to induce cancer cell death

    PubMed Central

    Milutinovic, Snezana; Kashyap, Arun K.; Yanagi, Teruki; Wimer, Carina; Zhou, Sihong; O' Neil, Ryann; Kurtzman, Aaron L.; Faynboym, Alexsandr; Xu, Li; Hannum, Charles H.; Diaz, Paul W.; Matsuzawa, Shu-ichi; Horowitz, Michael; Horowitz, Lawrence; Bhatt, Ramesh R.; Reed, John C.

    2015-01-01

    Death receptors of the Tumor Necrosis Factor (TNF) family are found on surface of most cancer cells and their activation typically kills cancer cells through the stimulation of the extrinsic apoptotic pathway. The endogenous ligand for death receptors-4 and -5 (DR4 and DR5) is Tumor Necrosis Factor-Related Apoptosis-Inducing Ligand, TRAIL (Apo2L). Since most untransformed cells are not susceptible to TRAIL-induced apoptosis, death receptor activators have emerged as promising cancer therapeutic agents. One strategy to stimulate death receptors in cancer patients is to use soluble human recombinant TRAIL protein, but this agent has limitations of a short half-life and decoy receptor sequestration. Another strategy that attempted to evade decoy receptor sequestration and to provide improved pharmacokinetic properties was to generate DR4 or DR5 agonist antibodies. The resulting monoclonal agonist antibodies overcame the limitations of short half-life and avoided decoy receptor sequestration, but are limited by activating only one of the two death receptors. Here, we describe a DR4 and DR5 dual agonist produced using Surrobody™ technology that activates both DR4 and DR5 to induce apoptotic death of cancer cells in vitro and in vivo and also avoids decoy receptor sequestration. This fully human anti-DR4/DR5 Surrobody displays superior potency to DR4- and DR5-specific antibodies, even when combined with TRAIL-sensitizing pro-apoptotic agents. Moreover, cancer cells were less likely to acquire resistance to Surrobody than either anti-DR4 or anti-DR5 mono-specific antibodies. Taken together, Surrobody shows promising preclinical pro-apoptotic activity against cancer cells, meriting further exploration of its potential as a novel cancer therapeutic agent. PMID:26516157

  6. Adrenergic receptor control mechanism for growth hormone secretion.

    PubMed

    Blackard, W G; Heidingsfelder, S A

    1968-06-01

    The influence of catecholamines on growth hormone secretion has been difficult to establish previously, possibly because of the suppressive effect of the induced hyperglycemia on growth hormone concentrations. In this study, an adrenergic receptor control mechanism for human growth hormone (HGH) secretion was uncovered by studying the effects of alpha and beta receptor blockade on insulin-induced growth hormone elevations in volunteer subjects. Alpha adrenergic blockade with phentolamine during insulin hypoglycemia, 0.1 U/kg, inhibited growth hormon elevations to 30-50% of values in the same subjects during insulin hypoglycemia without adrenergic blockade. More complete inhibition by phentolamine could not be demonstrated at a lower dose of insulin (0.05 U/kg). Beta adrenergic blockade with propranolol during insulin hypoglycemia significantly enhanced HGH concentrations in paired experiments. The inhibiting effect of alpha adrenergic receptor blockade on HGH concentrations could not be attributed to differences in blood glucose or free fatty acid values; however, more prolonged hypoglycemia and lower plasma free fatty acid values may have been a factor in the greater HGH concentrations observed during beta blockade. In the absence of insulin induced hypoglycemia, neither alpha nor beta adrenergic receptor blockade had a detectable effect on HGH concentrations. Theophylline, an inhibitor of cyclic 3'5'-AMP phosphodiesterase activity, also failed to alter plasma HGH concentrations. These studies demonstrate a stimulatory effect of alpha receptors and a possible inhibitory effect of beta receptors on growth hormone secretion.

  7. Structural Basis for Iloprost as a Dual Peroxisome Proliferator-activated Receptor [alpha/delta] Agonist

    SciTech Connect

    Jin, Lihua; Lin, Shengchen; Rong, Hui; Zheng, Songyang; Jin, Shikan; Wang, Rui; Li, Yong

    2012-03-15

    Iloprost is a prostacyclin analog that has been used to treat many vascular conditions. Peroxisome proliferator-activated receptors (PPARs) are ligand-regulated transcription factors with various important biological effects such as metabolic and cardiovascular physiology. Here, we report the crystal structures of the PPAR{alpha} ligand-binding domain and PPAR{delta} ligand-binding domain bound to iloprost, thus providing unambiguous evidence for the direct interaction between iloprost and PPARs and a structural basis for the recognition of PPAR{alpha}/{delta} by this prostacyclin analog. In addition to conserved contacts for all PPAR{alpha} ligands, iloprost also initiates several specific interactions with PPARs using its unique structural groups. Structural and functional studies of receptor-ligand interactions reveal strong functional correlations of the iloprost-PPAR{alpha}/{delta} interactions as well as the molecular basis of PPAR subtype selectivity toward iloprost ligand. As such, the structural mechanism may provide a more rational template for designing novel compounds targeting PPARs with more favorable pharmacologic impact based on existing iloprost drugs.

  8. Structural basis for iloprost as a dual peroxisome proliferator-activated receptor alpha/delta agonist.

    PubMed

    Jin, Lihua; Lin, Shengchen; Rong, Hui; Zheng, Songyang; Jin, Shikan; Wang, Rui; Li, Yong

    2011-09-09

    Iloprost is a prostacyclin analog that has been used to treat many vascular conditions. Peroxisome proliferator-activated receptors (PPARs) are ligand-regulated transcription factors with various important biological effects such as metabolic and cardiovascular physiology. Here, we report the crystal structures of the PPARα ligand-binding domain and PPARδ ligand-binding domain bound to iloprost, thus providing unambiguous evidence for the direct interaction between iloprost and PPARs and a structural basis for the recognition of PPARα/δ by this prostacyclin analog. In addition to conserved contacts for all PPARα ligands, iloprost also initiates several specific interactions with PPARs using its unique structural groups. Structural and functional studies of receptor-ligand interactions reveal strong functional correlations of the iloprost-PPARα/δ interactions as well as the molecular basis of PPAR subtype selectivity toward iloprost ligand. As such, the structural mechanism may provide a more rational template for designing novel compounds targeting PPARs with more favorable pharmacologic impact based on existing iloprost drugs.

  9. Exploring dual inhibitors for STAT1 and STAT5 receptors utilizing virtual screening and dynamics simulation validation.

    PubMed

    Raj, Utkarsh; Kumar, Himansu; Gupta, Saurabh; Varadwaj, Pritish Kumar

    2016-10-01

    Signal transducer and activator of transcription (STAT) proteins are latent cytoplasmic transcription factors that transduce signals from cytokines and growth factors to the nucleus and thereby regulate the expression of a variety of target genes. Although mutations of STATs have not been reported in human tumors but the activity of several members of the family, such as STAT1 and STAT5, is deregulated in a variety of human carcinoma. STAT1 and STAT5 share a structural similarity with a highly conserved SH2 domain which is responsible for the activation of STAT proteins on interaction with phosphotyrosine motifs for specific STAT-receptor contacts and STAT dimerization. The purpose of this study is to identify domain-specific dual inhibitors for both STAT1 and STAT5 proteins from a database of natural products and natural product-like compounds comprising of over 90,000 compounds. Virtual screening-based molecular docking was performed in order to find novel natural dual inhibitors. Further, the study was supported by the 50-ns molecular dynamics simulation for receptor-ligand complexes (STAT1-STOCK-1N-69677 and STAT5-STOCK-1N-69677). Analysis of molecular interactions in the SH2 domains of both STAT1 and STAT5 proteins with the ligand revealed few conserved amino acid residues which are responsible to stabilize the ligands within the binding pocket through bonded and non-bonded interactions. This study suggested that compound STOCK-1N-69677 might putatively act as a dual inhibitor of STAT1 and STAT5 receptors, through its binding to the SH2 domain.

  10. An NMDA Receptor-Dependent Mechanism Underlies Inhibitory Synapse Development.

    PubMed

    Gu, Xinglong; Zhou, Liang; Lu, Wei

    2016-01-26

    In the mammalian brain, GABAergic synaptic transmission provides inhibitory balance to glutamatergic excitatory drive and controls neuronal output. The molecular mechanisms underlying the development of GABAergic synapses remain largely unclear. Here, we report that NMDA-type ionotropic glutamate receptors (NMDARs) in individual immature neurons are the upstream signaling molecules essential for GABAergic synapse development, which requires signaling via Calmodulin binding motif in the C0 domain of the NMDAR GluN1 subunit. Interestingly, in neurons lacking NMDARs, whereas GABAergic synaptic transmission is strongly reduced, the tonic inhibition mediated by extrasynaptic GABAA receptors is increased, suggesting a compensatory mechanism for the lack of synaptic inhibition. These results demonstrate a crucial role for NMDARs in specifying the development of inhibitory synapses, and suggest an important mechanism for controlling the establishment of the balance between synaptic excitation and inhibition in the developing brain.

  11. Dual and pan-peroxisome proliferator-activated receptors (PPAR) co-agonism: the bezafibrate lessons

    PubMed Central

    Tenenbaum, Alexander; Motro, Michael; Fisman, Enrique Z

    2005-01-01

    There are three peroxisome proliferator-activated receptors (PPARs) subtypes which are commonly designated PPAR alpha, PPAR gamma and PPAR beta/delta. PPAR alpha activation increases high density lipoprotein (HDL) cholesterol synthesis, stimulates "reverse" cholesterol transport and reduces triglycerides. PPAR gamma activation results in insulin sensitization and antidiabetic action. Until recently, the biological role of PPAR beta/delta remained unclear. However, treatment of obese animals by specific PPAR delta agonists results in normalization of metabolic parameters and reduction of adiposity. Combined treatments with PPAR gamma and alpha agonists may potentially improve insulin resistance and alleviate atherogenic dyslipidemia, whereas PPAR delta properties may prevent the development of overweight which typically accompanies "pure" PPAR gamma ligands. The new generation of dual-action PPARs – the glitazars, which target PPAR-gamma and PPAR-alpha (like muraglitazar and tesaglitazar) are on deck in late-stage clinical trials and may be effective in reducing cardiovascular risk, but their long-term clinical effects are still unknown. A number of glitazars have presented problems at a late stage of clinical trials because of serious side-effects (including ragaglitazar and farglitazar). The old and well known lipid-lowering fibric acid derivative bezafibrate is the first clinically tested pan – (alpha, beta/delta, gamma) PPAR activator. It is the only pan-PPAR activator with more than a quarter of a century of therapeutic experience with a good safety profile. Therefore, bezafibrate could be considered (indeed, as a "post hoc" understanding) as an "archetype" of a clinically tested pan-PPAR ligand. Bezafibrate leads to considerable raising of HDL cholesterol and reduces triglycerides, improves insulin sensitivity and reduces blood glucose level, significantly lowering the incidence of cardiovascular events and new diabetes in patients with features of

  12. Mechanical Coupling Error Suppression Technology for an Improved Decoupled Dual-Mass Micro-Gyroscope.

    PubMed

    Yang, Bo; Wang, Xingjun; Deng, Yunpeng; Hu, Di

    2016-04-08

    This paper presents technology for the suppression of the mechanical coupling errors for an improved decoupled dual-mass micro-gyroscope (DDMG). The improved micro-gyroscope structure decreases the moment arm of the drive decoupled torque, which benefits the suppression of the non-ideal decoupled error. Quadrature correction electrodes are added to eliminate the residual quadrature error. The structure principle and the quadrature error suppression means of the DDMG are described in detail. ANSYS software is used to simulate the micro-gyroscope structure to verify the mechanical coupling error suppression effect. Compared with the former structure, simulation results demonstrate that the rotational displacements of the sense frame in the improved structure are substantially suppressed in the drive mode. The improved DDMG structure chip is fabricated by the deep dry silicon on glass (DDSOG) process. The feedback control circuits with quadrature control loops are designed to suppress the residual mechanical coupling error. Finally, the system performance of the DDMG prototype is tested. Compared with the former DDMG, the quadrature error in the improved dual-mass micro-gyroscope is decreased 9.66-fold, and the offset error is decreased 6.36-fold. Compared with the open loop sense, the feedback control circuits with quadrature control loop decrease the bias drift by 20.59-fold and the scale factor non-linearity by 2.81-fold in the ±400°/s range.

  13. A dual analysis for recycled particulate composites: linking micro- and macro-mechanics

    SciTech Connect

    Avila, Antonio F.; Rodrigues, Paulo C.M.; Santos, Dagoberto B.; Faria, Ana C.A

    2003-06-15

    The large amount of disposable bottles produced nowadays makes imperative the search for alternative procedures for recycling them since they are not biodegradable. This paper takes into consideration the thermomechanical recycling of post-consumed plastic bottles, especially the ones made of polyethylene terephthalate (PET) and high-density polyethylene (HDPE), and their use as composite materials for engineering applications. As changes on the composite's microstructure can have an influence on macroscopic behavior, a new type of analysis is needed. To be able to evaluate the composite performance, a dual analysis procedure was developed. It consists of a micro-mechanical analysis where the microstructure is observed by optical microscopy, and variations in morphology are related to composite overall mechanical behavior. The macro-mechanical analysis is performed by ASTM D 3039/3039 M tensile tests. By doing this, the composite effective moduli can be determined. The new composite seems to be encouraging, i.e., an HDPE/PET composite with 40:60 ratio, in weight, experiments a stiffness recovery from the third to the fourth recycle. Moreover, the dual analysis was able to capture this variation.

  14. Mechanical Coupling Error Suppression Technology for an Improved Decoupled Dual-Mass Micro-Gyroscope

    PubMed Central

    Yang, Bo; Wang, Xingjun; Deng, Yunpeng; Hu, Di

    2016-01-01

    This paper presents technology for the suppression of the mechanical coupling errors for an improved decoupled dual-mass micro-gyroscope (DDMG). The improved micro-gyroscope structure decreases the moment arm of the drive decoupled torque, which benefits the suppression of the non-ideal decoupled error. Quadrature correction electrodes are added to eliminate the residual quadrature error. The structure principle and the quadrature error suppression means of the DDMG are described in detail. ANSYS software is used to simulate the micro-gyroscope structure to verify the mechanical coupling error suppression effect. Compared with the former structure, simulation results demonstrate that the rotational displacements of the sense frame in the improved structure are substantially suppressed in the drive mode. The improved DDMG structure chip is fabricated by the deep dry silicon on glass (DDSOG) process. The feedback control circuits with quadrature control loops are designed to suppress the residual mechanical coupling error. Finally, the system performance of the DDMG prototype is tested. Compared with the former DDMG, the quadrature error in the improved dual-mass micro-gyroscope is decreased 9.66-fold, and the offset error is decreased 6.36-fold. Compared with the open loop sense, the feedback control circuits with quadrature control loop decrease the bias drift by 20.59-fold and the scale factor non-linearity by 2.81-fold in the ±400°/s range. PMID:27070616

  15. Qualification of a High Accuracy Dual-Axis Antenna Deployment and Trimming Mechanism

    NASA Technical Reports Server (NTRS)

    Gossant, Alain; Morichon, Francois

    2010-01-01

    The Antenna Deployment and Trimming Mechanism Mark 2 (ADTM Mk2) has been developed to answer today's need for a generic antenna deployment and high accuracy pointing mechanism, allowing RF sensing applications and easier dual deployments configurations. This paper presents the design and evolution from its predecessor, the experience of the design team from kick off to qualification and batch manufacture, as well as some lessons learned from ramping up "mass-production" capabilities while implementing customer driven changes. Astrium has manufactured and flown ADTM units for the past 20 years, from an initial deployment-only mechanism developed for the Orion program to today's Eurostar E3000 ADTM family. The Antenna ADTM Mk2 is an evolution of the original ADTM Mk1. Although it uses Mk1 building blocks to minimize risks associated with the development of a new product, it incorporates major evolutions and is the new baseline for Astrium latest generation of Eurostar E3000 telecom satellites.

  16. Unidirectional, dual-comb lasing under multiple pulse formation mechanisms in a passively mode-locked fiber ring laser.

    PubMed

    Liu, Ya; Zhao, Xin; Hu, Guoqing; Li, Cui; Zhao, Bofeng; Zheng, Zheng

    2016-09-19

    Dual-comb lasers simultaneously generating asynchronous ultrashort pulses could be an intriguing alternative to the current dual-laser comb source. When generated through a common light path, the low common-mode noises and good coherence between the pulse trains could be realized. Here we demonstrate the completely common-path, unidirectional dual-comb lasing using a carbon nanotube saturable absorber with additional pulse narrowing and broadening mechanisms. The interactions between multiple soliton formation mechanisms result in bifurcation into unusual two-pulse states with pulses of four-fold bandwidth difference and tens-of-Hz repetition rate difference. Coherence between the pulses is verified by the asynchronous cross-sampling and dual-comb spectroscopy measurements.

  17. Synthesis and pharmacological evaluation of dual acting ligands targeting the adenosine A2A and dopamine D2 receptors for the potential treatment of Parkinson's disease.

    PubMed

    Jörg, Manuela; May, Lauren T; Mak, Frankie S; Lee, Kiew Ching K; Miller, Neil D; Scammells, Peter J; Capuano, Ben

    2015-01-22

    A relatively new strategy in drug discovery is the development of dual acting ligands. These molecules are potentially able to interact at two orthosteric binding sites of a heterodimer simultaneously, possibly resulting in enhanced subtype selectivity, higher affinity, enhanced or modified physiological response, and reduced reliance on multiple drug administration regimens. In this study, we have successfully synthesized a series of classical heterobivalent ligands as well as a series of more integrated and "drug-like" dual acting molecules, incorporating ropinirole as a dopamine D2 receptor agonist and ZM 241385 as an adenosine A2A receptor antagonist. The best compounds of our series maintained the potency of the original pharmacophores at both receptors (adenosine A2A and dopamine D2). In addition, the integrated dual acting ligands also showed promising results in preliminary blood-brain barrier permeability tests, whereas the classical heterobivalent ligands are potentially more suited as pharmacological tools.

  18. Differences in Gene Regulation by Dual Ligands of Nuclear Receptors Constitutive Androstane Receptor (CAR) and Pregnane X Receptor (PXR) in HepG2 Cells Stably Expressing CAR/PXR.

    PubMed

    Kanno, Yuichiro; Tanuma, Nobuaki; Yazawa, Saki; Zhao, Shuai; Inaba, Miki; Nakamura, Satoshi; Nemoto, Kiyomitsu; Inouye, Yoshio

    2016-08-01

    The constitutive androstane receptor (CAR) and pregnane X receptor (PXR) regulate various genes involved in xenobiotics and drug metabolism. In many cases, CAR/PXR share ligands termed dual ligands of CAR/PXR. It is difficult to investigate the effect of CAR/PXR dual ligands in cell lines because CAR and PXR expression is scarcely detected in cultured cell lines. Here, we established a tetracycline-inducible human CAR and stably human PXR-overexpressing HepG2 cell line (HepTR/hCAR/hPXR) to examine CAR/PXR dual ligands. In the present study, we investigated the regulation of CYP2B6, CYP2C9, CYP3A4, and UDP-glucuronosyl transferase, which are target genes of CAR/PXR, by dual ligands of CAR/PXR in two transfectants. Activation of CAR and PXR in cells treated with a high dose of CITCO [6-(4-chlorophenyl)-imidazo(2,1-b)thiazole-5-carbaldehyde] or cotreated with rifampicin and tetracycline resulted in synergistic enhancement of CYP3A4, but not CYP2B6, CYP2C9, or UGT1A1, mRNA expression in HepTR/hCAR/hPXR cells. In contrast, this synergistic effect was not observed in HepTR/hCAR cells. These observations were also demonstrated in human primary hepatocytes. Taken together, our results suggest that dual ligands of CAR/PXR show distinct gene regulation patterns by cross-talk between CAR and PXR. Furthermore, the two newly established cell lines are useful tools to investigate dual ligands of CAR/PXR.

  19. Dual effect on the RET receptor of MEN 2 mutations affecting specific extracytoplasmic cysteines.

    PubMed

    Chappuis-Flament, S; Pasini, A; De Vita, G; Ségouffin-Cariou, C; Fusco, A; Attié, T; Lenoir, G M; Santoro, M; Billaud, M

    1998-12-03

    The RET gene encodes a receptor tyrosine kinase whose function is essential during the development of kidney and the intestinal nervous system. Germline mutations affecting one of five cysteines (Cys609, 611, 618, 620 and 634) located in the juxtamembrane domain of the RET receptor are responsible for the vast majority of two cancer-prone disorders, multiple endocrine neoplasia type 2A (MEN 2A) and familial medullary thyroid carcinoma (FMTC). These mutations lead to the replacement of a cysteine by an alternate amino acid. Mutations of the RET gene are also the underlying genetic cause of Hirschsprung disease (HSCR), a congenital aganglionosis of the hindgut. In a fraction of kindreds, MEN 2A cosegregate with HSCR and affected individuals carry a single mutation at codons 609, 618 or 620. To examine the consequences of cysteine substitution on RET function, we have introduced a Cys to Arg mutation into the wild-type RET at either codons 609, 618, 620, 630 or 634. We now report that each mutation induces a constitutive catalytic activity due to the aberrant disulfide homodimerization of RET. However, mutations 630 and 634 activate RET more strongly than mutations 609, 618 or 620 as demonstrated by quantitative assays in rodent fibroblasts and pheochromocytoma PC12 cells. Biochemical analysis revealed that mutations 618 and 620, and to a lesser extent mutation 609, result in a marked reduction of the level of RET at the cell surface and as a consequence decrease the amount of RET covalent dimer. These findings provide a molecular basis explaining the range of phenotype engendered by alterations of RET cysteines and suggest a novel mechanism whereby mutations of cysteines 609, 618 and 620 exert both activating and inactivating effects.

  20. Mechanisms of oestrogen receptor (ER) gene regulation in breast cancer

    PubMed Central

    2016-01-01

    Most breast cancers are driven by a transcription factor called oestrogen receptor (ER). Understanding the mechanisms of ER activity in breast cancer has been a major research interest and recent genomic advances have revealed extraordinary insights into how ER mediates gene transcription and what occurs during endocrine resistance. This review discusses our current understanding on ER activity, with an emphasis on several evolving, but important areas of ER biology. PMID:26884552

  1. Piperazine-2,3-dicarboxylic acid Derivatives as Dual Antagonists of NMDA and GluK1-Containing Kainate Receptors

    PubMed Central

    Irvine, Mark W.; Costa, Blaise M.; Dlaboga, Daniel; Culley, Georgia; Hulse, Richard; Scholefield, Caroline L.; Atlason, Palmi; Fang, Guangyu; Eaves, Richard; Morley, Richard; Mayo-Martin, Maria B.; Amici, Mascia; Bortolotto, Zuner A.; Donaldson, Lucy; Collingridge, Graham L.; Molnár, Elek; Monaghan, Daniel T.; Jane, David E.

    2011-01-01

    Competitive N-methyl-D-aspartate receptor (NMDAR) antagonists bind to the GluN2 subunit, of which there are four types (GluN2A-D). We report that some N1-substituted derivatives of cis-piperazine-2,3-dicarboxylic acid display improved relative affinity for GluN2C and GluN2D versus GluN2A and GluN2B. These derivatives also display subtype-selectivity among the more distantly related kainate receptor family. Compounds 18i and (−)-4 were the most potent kainate receptor antagonists and 18i was selective for GluK1 versus GluK2, GluK3 and AMPA receptors. Modeling studies revealed structural features required for activity at GluK1 subunits and suggested that S674 was vital for antagonist activity. Consistent with this hypothesis, replacing the equivalent residue in GluK3 (alanine) with a serine imparts 18i antagonist activity. Antagonists with dual GluN2D and GluK1 antagonist activity may have beneficial effects in various neurological disorders. Consistent with this idea, antagonist 18i (30 mg/Kg i.p.) showed antinociceptive effects in an animal model of mild nerve injury. PMID:22111545

  2. Genomic and non-genomic mechanisms of oxytocin receptor regulation.

    PubMed

    Zingg, H H; Grazzini, E; Breton, C; Larcher, A; Rozen, F; Russo, C; Guillon, G; Mouillac, B

    1998-01-01

    Our recent studies have shown that regulation of uterine oxytocin (OT) binding involves at least two different mechanism: Estradiol (E2)-induced upregulation is accompanied by an increase in OT receptor (OTR) mRNA accumulation, implying that the E2 effect is mediated via increased OTR gene transcription and/or OTR mRNA stabilization. In contrast, P (P)-induced OTR down-regulation occurs via a novel non-genomic mechanism, involving a direct interaction of P with the OTR at the level of the cell membrane. We found that P specifically binds to the OTR and inhibits its ligand binding and signalling functions. Physiological levels of P repress in vitro the ligand binding capacity (Bmax) of the OTR by > 50%. When expressed in CHO cells, the OTR provides a high affinity (Kd: 20nM) membrane binding site for P. OT-induced inositol phosphate production and intracellular calcium mobilization is inhibited 85% and 90%, respectively, by P. These effects are specific as signalling and binding functions of the closely related V1a vasopressin receptor remain unaffected by P, and as other, related steroids are devoid of any effect on OTR binding or signalling functions. The present observation of a specific interaction of a steroid with a G-protein-linked receptor defines a new mechanism of non-genomic steroid action and uncovers a novel level of crosstalk between steroid and peptide hormone action.

  3. Bisphenol A affects androgen receptor function via multiple mechanisms.

    PubMed

    Teng, Christina; Goodwin, Bonnie; Shockley, Keith; Xia, Menghang; Huang, Ruili; Norris, John; Merrick, B Alex; Jetten, Anton M; Austin, Christopher P; Tice, Raymond R

    2013-05-25

    Bisphenol A (BPA), is a well-known endocrine disruptor compound (EDC) that affects the normal development and function of the female and male reproductive system, however the mechanisms of action remain unclear. To investigate the molecular mechanisms of how BPA may affect ten different nuclear receptors, stable cell lines containing individual nuclear receptor ligand binding domain (LBD)-linked to the β-Gal reporter were examined by a quantitative high throughput screening (qHTS) format in the Tox21 Screening Program of the NIH. The results showed that two receptors, estrogen receptor alpha (ERα) and androgen receptor (AR), are affected by BPA in opposite direction. To confirm the observed effects of BPA on ERα and AR, we performed transient transfection experiments with full-length receptors and their corresponding response elements linked to luciferase reporters. We also included in this study two BPA analogs, bisphenol AF (BPAF) and bisphenol S (BPS). As seen in African green monkey kidney CV1 cells, the present study confirmed that BPA and BPAF act as ERα agonists (half maximal effective concentration EC50 of 10-100 nM) and as AR antagonists (half maximal inhibitory concentration IC50 of 1-2 μM). Both BPA and BPAF antagonized AR function via competitive inhibition of the action of synthetic androgen R1881. BPS with lower estrogenic activity (EC50 of 2.2 μM), did not compete with R1881 for AR binding, when tested at 30 μM. Finally, the effects of BPA were also evaluated in a nuclear translocation assays using EGPF-tagged receptors. Similar to 17β-estradiol (E2) which was used as control, BPA was able to enhance ERα nuclear foci formation but at a 100-fold higher concentration. Although BPA was able to bind AR, the nuclear translocation was reduced. Furthermore, BPA was unable to induce functional foci in the nuclei and is consistent with the transient transfection study that BPA is unable to activate AR.

  4. Design and Functional Validation of a Mechanism for Dual-Spinning CubeSats

    NASA Technical Reports Server (NTRS)

    Peters, Eric; Dave, Pratik; Kingsbury, Ryan; Marinan, Anne; Wise, Evan; Pong, Chris; Prinkey, Meghan; Cahoy, Kerri; Miller, David W.; Sklair, Devon

    2014-01-01

    The mission of the Micro-sized Microwave Atmospheric Satellite (MicroMAS) is to collect useful atmospheric images using a miniature passive microwave radiometer payload hosted on a low-cost CubeSat platform. In order to collect this data, the microwave radiometer payload must rotate to scan the ground-track perpendicular to the satellite's direction of travel. A custom motor assembly was developed to facilitate the rotation of the payload while allowing the spacecraft bus to remained fixed in the local-vertical, local-horizontal (LVLH) frame for increased pointing accuracy. This paper describes the mechanism used to enable this dual-spinning operation for CubeSats, and the lessons learned during the design, fabrication, integration, and testing phases of the mechanism's development lifecycle.

  5. Mechanism of TGFbeta receptor inhibition by FKBP12.

    PubMed Central

    Chen, Y G; Liu, F; Massague, J

    1997-01-01

    Transforming growth factor-beta (TGFbeta) signaling requires phosphorylation of the type I receptor TbetaR-I by TbetaR-II. Although TGFbeta promotes the association of TbetaR-I with TbetaR-II, these receptor components have affinity for each other which can lead to their ligand-independent activation. The immunophilin FKBP12 binds to TbetaR-I and inhibits its signaling function. We investigated the mechanism and functional significance of this effect. FKBP12 binding to TbetaR-I involves the rapamycin/Leu-Pro binding pocket of FKBP12 and a Leu-Pro sequence located next to the activating phosphorylation sites in TbetaR-I. Mutations in the binding sites of FKBP12 or TbetaR-I abolish the interaction between these proteins, leading to receptor activation in the absence of added ligand. FKBP12 does not inhibit TbetaR-I association with TbetaR-II, but inhibits TbetaR-I phosphorylation by TbetaR-II. Rapamycin, which blocks FKBP12 binding to TbetaR-I, reverses the inhibitory effect of FKBP12 on TbetaR-I phosphorylation. By impeding the activation of TGFbeta receptor complexes formed in the absence of ligand, FKBP12 may provide a safeguard against leaky signaling resulting from the innate tendency of TbetaR-I and TbetaR-II to interact with each other. PMID:9233797

  6. Envelope determinants for dual-receptor specificity in feline leukemia virus subgroup A and T variants.

    PubMed

    Cheng, Heather H; Anderson, Maria M; Hankenson, F Claire; Johnston, Lily; Kotwaliwale, Chitra V; Overbaugh, Julie

    2006-02-01

    Gammaretroviruses, including the subgroups A, B, and C of feline leukemia virus (FeLV), use a multiple-membrane-spanning transport protein as a receptor. In some cases, such as FeLV-T, a nonclassical receptor that includes both a transport protein (Pit1) and a soluble cofactor (FeLIX) is required for entry. To define which regions confer specificity to classical versus nonclassical receptor pathways, we engineered mutations found in either FeLV-A/T or FeLV-T, individually and in combination, into the backbone of the transmissible form of the virus, FeLV-A. The receptor specificities of these viruses were tested by measuring infection and binding to cells expressing the FeLV-A receptor or the FeLV-T receptors. FeLV-A receptor specificity was maintained when changes at amino acid position 6, 7, or 8 of the mature envelope glycoprotein were introduced, although differences in infection efficiency were observed. When these N-terminal mutations were introduced together with a C-terminal 4-amino-acid insertion and an adjacent amino acid change, the resulting viruses acquired FeLV-T receptor specificity. Additionally, a W-->L change at amino acid position 378, although not required, enhanced infectivity for some viruses. Thus, we have found that determinants in the N and C termini of the envelope surface unit can direct entry via the nonclassical FeLV-T receptor pathway. The region that has been defined as the receptor binding domain of gammaretroviral envelope proteins determined entry via the FeLV-A receptor independently of the presence of the N- and C-terminal FeLV-T receptor determinants.

  7. Sodium Absorption by Barley Roots: Role of the Dual Mechanisms of Alkali Cation Transport 1

    PubMed Central

    Rains, D. W.; Epstein, Emanuel

    1967-01-01

    Radioactively labeled Na+ absorbed by barley roots was sequestered in an intracellular compartment or compartments (“inner” spaces) in which it was only very slowly exchangeable with exogenous Na+. Absorption of this fraction proceeded at a constant rate for at least 1 hour. When the rate of Na+ absorption was examined over the range of concentrations, 0.005 to 50 mm, the isotherm depicting the relation showed dual kinetics as follows. Over the range, 0.005 to 0.2 mm, a single Michaelis-Menten term describes the relation between the concentration of Na+ and the rate of its absorption. The mechanism of Na+ absorption operating over this range of concentrations, mechanism 1 of alkali cation transport, is severely inhibited in the presence of Ca2+ and virtually rendered inoperative for Na+ transport by the combined presence of Ca2+ and K+. The mechanism is equally effective in Na+ transport whether Cl− or F− is the anion, but is somewhat inhibited when the anion is SO42−. Over the high range of concentrations, 0.5 to 50 mm Na+, a second, low-affinity mechanism of Na+ absorption comes into play. In the presence of Ca2+ and K+, this mechanism 2 is the only one to transport Na+ effectively, since Na+ absorption via mechanism 1 is virtually abolished under these conditions. Anaerobic conditions, low temperature, and the uncoupler, 2,4-dinitrophenol, inhibit Na+ absorption both at low and high Na+ concentrations. PMID:16656509

  8. Dual Receptor Recognizing Cell Penetrating Peptide for Selective Targeting, Efficient Intratumoral Diffusion and Synthesized Anti-Glioma Therapy

    PubMed Central

    Liu, Yayuan; Mei, Ling; Xu, Chaoqun; Yu, Qianwen; Shi, Kairong; Zhang, Li; Wang, Yang; Zhang, Qianyu; Gao, Huile; Zhang, Zhirong; He, Qin

    2016-01-01

    Cell penetrating peptides (CPPs) were widely used for drug delivery to tumor. However, the nonselective in vivo penetration greatly limited the application of CPPs-mediated drug delivery systems. And the treatment of malignant tumors is usually followed by poor prognosis and relapse due to the existence of extravascular core regions of tumor. Thus it is important to endue selective targeting and stronger intratumoral diffusion abilities to CPPs. In this study, an RGD reverse sequence dGR was conjugated to a CPP octa-arginine to form a CendR (R/KXXR/K) motif contained tandem peptide R8-dGR (RRRRRRRRdGR) which could bind to both integrin αvβ3 and neuropilin-1 receptors. The dual receptor recognizing peptide R8-dGR displayed increased cellular uptake and efficient penetration ability into glioma spheroids in vitro. The following in vivo studies indicated the active targeting and intratumoral diffusion capabilities of R8-dGR modified liposomes. When paclitaxel was loaded in the liposomes, PTX-R8-dGR-Lip induced the strongest anti-proliferation effect on both tumor cells and cancer stem cells, and inhibited the formation of vasculogenic mimicry channels in vitro. Finally, the R8-dGR liposomal drug delivery system prolonged the medium survival time of intracranial C6 bearing mice by 2.1-fold compared to the untreated group, and achieved an exhaustive anti-glioma therapy including anti-tumor cells, anti-vasculogenic mimicry and anti-brain cancer stem cells. To sum up, all the results demonstrated that R8-dGR was an ideal dual receptor recognizing CPP with selective glioma targeting and efficient intratumoral diffusion, which could be further used to equip drug delivery system for effective glioma therapy. PMID:26877777

  9. Freud-2/CC2D1B mediates dual repression of the serotonin-1A receptor gene.

    PubMed

    Hadjighassem, Mahmoud R; Galaraga, Kimberly; Albert, Paul R

    2011-01-01

    The serotonin-1A (5-HT1A) receptor functions as a pre-synaptic autoreceptor in serotonin neurons that regulates their activity, and is also widely expressed on non-serotonergic neurons as a post-synaptic heteroreceptor to mediate serotonin action. The 5-HT1A receptor gene is strongly repressed by a dual repressor element (DRE), which is recognized by two proteins: Freud-1/CC2D1A and another unknown protein. Here we identify mouse Freud-2/CC2D1B as the second repressor of the 5-HT1A-DRE. Freud-2 shares 50% amino acid identity with Freud-1, and contains conserved structural domains. Mouse Freud-2 bound specifically to the rat 5-HT1A-DRE adjacent to, and partially overlapping, the Freud-1 binding site. By supershift assay using nuclear extracts from L6 myoblasts, Freud-2-DRE complexes were distinguished from Freud-1-DRE complexes. Freud-2 mRNA and protein were detected throughout mouse brain and peripheral tissues. Freud-2 repressed 5-HT1A promoter-reporter constructs in a DRE-dependent manner in non-neuronal (L6) or 5-HT1A-expressing neuronal (NG108-15, RN46A) cell models. In NG108-15 cells, knockdown of Freud-2 using a specific short-interfering RNA reduced endogenous Freud-2 protein levels and decreased Freud-2 bound to the 5-HT1A-DRE as detected by chromatin immunoprecipitation assay, but increased 5-HT1A promoter activity and 5-HT1A protein levels. Taken together, these data show that Freud-2 is the second component that, with Freud-1, mediates dual repression of the 5-HT1A receptor gene at the DRE.

  10. Lysine Specific Demethylase 1 has Dual Functions as a Major Regulator of Androgen Receptor Transcriptional Activity

    PubMed Central

    Cai, Changmeng; He, Housheng Hansen; Gao, Shuai; Chen, Sen; Yu, Ziyang; Gao, Yanfei; Chen, Shaoyong; Chen, Mei Wei; Zhang, Jesse; Ahmed, Musaddeque; Wang, Yang; Metzger, Eric; Schüle, Roland; Liu, X. Shirley; Brown, Myles; Balk, Steven P.

    2014-01-01

    SUMMARY Lysine Specific Demethylase 1 (LSD1, KDM1A) functions as a transcriptional corepressor through demethylation of histone 3 lysine 4 (H3K4), but has coactivator function on some genes through unclear mechanisms. We show that LSD1, interacting with CoREST, associates with and coactivates androgen receptor (AR) on a large fraction of androgen-stimulated genes. A subset of these AR/LSD1-associated enhancer sites have histone 3 threonine 6 phosphorylation (H3T6ph), and these sites are further enriched for androgen-stimulated genes. Significantly, despite its coactivator activity, LSD1 still mediates H3K4me2 demethylation at these androgen-stimulated enhancers. FOXA1 is also associated with LSD1 at AR regulated enhancer sites, and a FOXA1 interaction with LSD1 enhances binding of both proteins at these sites. These findings show LSD1 functions broadly as a regulator of AR function, that it maintains a transcriptional repression function at AR-regulated enhancers through H3K4 demethylation, and has a distinct AR-linked coactivator function mediated by demethylation of other substrates. PMID:25482560

  11. Dual Effect of Adenosine A1 Receptor Activation on Renal O2 Consumption.

    PubMed

    Babich, Victor; Vadnagara, Komal; Di Sole, Francesca

    2015-12-01

    The high requirement of O2 in the renal proximal tubule stems from a high rate of Na(+) transport. Adenosine A1 receptor (A1R) activation regulates Na(+) transport in this nephron segment. Thus, the effect of the acute activation and the mechanisms of A1R on the rate of O2 consumption were evaluated. The A1R-antagonist, 8-cyclopentyl-1,3-dipropylxanthine (CPX) and adenosine deaminase (ADA), which metabolize endogenous adenosine, reduced O2 consumption (40-50%). Replacing Na(+) in the buffer reversed the ADA- or CPX-mediated reduction of O2 consumption. Blocking the Na/H-exchanger activity, which decreases O2 usage per se, did not enhance the ADA- or CPX-induced inhibition of O2 consumption. These data indicate that endogenous adenosine increases O2 usage via the activation of Na(+) transport. In the presence of endogenous adenosine, A1R was further activated by the A1R-agonist N(6)-cyclopentyladenosine (CPA); CPA inhibited O2 usage (30%) and this effect also depended on Na(+) transport. Moreover, a low concentration of CPA activated O2 usage in tissue pretreated with ADA, whereas a high concentration of CPA inhibited O2 usage; both effects depended on Na(+). Protein kinase C signaling mediated the inhibitory effect of A1R, while adenylyl cyclase mediated its stimulatory effect on O2 consumption. In summary, increasing the local concentrations of adenosine can either activate or inhibit O2 consumption via A1R, and this mechanism depends on Na(+) transport. The inhibition of O2 usage by A1R activation might restore the compromised balance between energy supply and demand under pathophysiological conditions, such as renal ischemia, which results in high adenosine production.

  12. The nicotinic acetylcholine receptor CHRNA5/A3/B4 gene cluster: Dual role in nicotine addiction and lung cancer

    PubMed Central

    Improgo, Ma. Reina D.; Scofield, Michael D.; Tapper, Andrew R.; Gardner, Paul D.

    2010-01-01

    More than 1 billion people around the world smoke, with 10 million cigarettes sold every minute. Cigarettes contain thousands of harmful chemicals including the psychoactive compound, nicotine. Nicotine addiction is initiated by the binding of nicotine to nicotinic acetylcholine receptors, ligand-gated cation channels activated by the endogenous neurotransmitter, acetylcholine. These receptors serve as prototypes for all ligand-gated ion channels and have been extensively studied in an attempt to elucidate their role in nicotine addiction. Many of these studies have focused on heteromeric nicotinic acetylcholine receptors containing α4 and β2 subunits and homomeric nicotinic acetylcholine receptors containing the α7 subunit, two of the most abundant subtypes expressed in the brain. Recently however, a series of linkage analyses, candidate-gene analyses and genome-wide association studies have brought attention to three other members of the nicotinic acetylcholine receptor family: the α5, α3 and β4 subunits. The genes encoding these subunits lie in a genomic cluster that contains variants associated with increased risk for several diseases including nicotine dependence and lung cancer. The underlying mechanisms for these associations have not yet been elucidated but decades of research on the nicotinic receptor gene family as well as emerging data provide insight on how these receptors may function in pathological states. Here, we review this body of work, focusing on the clustered nicotinic acetylcholine receptor genes and evaluating their role in nicotine addiction and lung cancer. PMID:20685379

  13. Molecular Mechanisms of Antiseizure Drug Activity at GABAA Receptors

    PubMed Central

    Greenfield, L. John

    2013-01-01

    The GABAA receptor (GABAAR) is a major target of antiseizure drugs (ASDs). A variety of agents that act at GABAARs s are used to terminate or prevent seizures. Many act at distinct receptor sites determined by the subunit composition of the holoreceptor. For the benzodiazepines, barbiturates, and loreclezole, actions at the GABAAR are the primary or only known mechanism of antiseizure action. For topiramate, felbamate, retigabine, losigamone and stiripentol, GABAAR modulation is one of several possible antiseizure mechanisms. Allopregnanolone, a progesterone metabolite that enhances GABAAR function, led to the development of ganaxolone. Other agents modulate GABAergic “tone” by regulating the synthesis, transport or breakdown of GABA. GABAAR efficacy is also affected by the transmembrane chloride gradient, which changes during development and in chronic epilepsy. This may provide an additional target for “GABAergic” ASDs. GABAAR subunit changes occur both acutely during status epilepticus and in chronic epilepsy, which alter both intrinsic GABAAR function and the response to GABAAR-acting ASDs. Manipulation of subunit expression patterns or novel ASDs targeting the altered receptors may provide a novel approach for seizure prevention. PMID:23683707

  14. [Forgetfulness and amnesia: receptor mechanisms and brain mapping].

    PubMed

    Ilíuchenok, R Iu; Dubrovina, N I; Podgornaia, O V; Galkina, O V

    1994-01-01

    Inability to remember and amnesia have been shown to be active neurochemical processes. The coupled processes (blockade of the triggering DA stimulating system and activation of the inhibitory GABA-ergic system with the predominant value of postsynaptic D-2 receptors) are a neurochemical basis for development of amnesia. The mechanisms of spontaneous forgetting is provided by a decrease in the activity of the dopaminergic system along with the enhancement of benzodiazepine-GABA-ergic interferentional inhibition. The observed changes in dopamine metabolism, para-tyramine appearance, as well as restructure of D-2 receptors provide the activity of dopamine increasing mechanism which determines the retention of memory traces. A computer model of the spatial interaction of the dopamine membrane-receptor complex was constructed by scanning the samples of synaptic membranes after learning and amnesia. A new method of inducing psychogenic amnesia in human beings has been elaborated. Amnesia is characterized by the absence of increases in the number of cortical connections reflecting the emotional factor of information.

  15. Benzothiazoles as probes for the 5HT1A receptor and the serotonin transporter (SERT): a search for new dual-acting agents as potential antidepressants.

    PubMed

    Zhu, Xue Y; Etukala, Jagan R; Eyunni, Suresh V K; Setola, Vincent; Roth, Bryan L; Ablordeppey, Seth Y

    2012-07-01

    The synthesis and evaluation of several benzothiazole-based compounds are described in an attempt to identify novel dual-acting 5HT(1A) receptor and SERT inhibitors as new antidepressants. Binding affinities at the 5HT(1A) receptor and the serotonin transporter do not appear to be congruent and other areas of the binding sites would need to be explored in order to improve binding simultaneously at both sites. Compounds 20 and 23 show moderate binding affinity at the 5HT(1A) receptor and the SERT site and thus, have the potential to be further explored as dual-acting agents. In addition, compound 20 binds with low affinity to the dopamine transporter (DAT), the norepinephrine transporter (NET) and 5HT(2C) receptor, which are desirable properties as selectivity for SERT (and not DAT or NET) is associated with an absence of cardiovascular side effects.

  16. First implementation of burrowing motions in dual-reciprocating drilling using an integrated actuation mechanism

    NASA Astrophysics Data System (ADS)

    Pitcher, Craig; Gao, Yang

    2017-03-01

    The dual-reciprocating drill (DRD) is a biologically-inspired low-mass alternative to traditional drilling techniques, using backwards-facing teethed halves to grip the surrounding substrate, generating a traction force that reduces the required overhead penetration force. Previous experiments using a proof-of-concept test bench have provided evidence as to the significant role of sideways movements and lateral forces in improving drilling performance. The system is also progressing to a first system prototype concept, in which an actuation mechanism is integrated within the drill heads. To experimentally determine the effect of lateral motions, a new internal actuation mechanism was developed to allow the inclusion of controlled sideways movements, resulting in the creation of the circular and diagonal burrowing motions. This paper presents an investigation into the performance of the reciprocation and burrowing motions by testing them in a planetary regolith simulant. Analysis of force sensor measurements has shown a relationship between the penetration and traction forces and the internal friction of the mechanism and depth achieved. These tests have also experimentally demonstrated the benefit of lateral motions in drilling performance, with both the burrowing mechanisms and drilling tests performed at an angle able to penetrate further than traditional vertical reciprocation, leading to the proposition of new burrowing and diagonal drilling mechanics. From this, a new fully integrated system prototype can be developed which incorporates lateral motions that can optimise the drilling performance.

  17. Receptor- and reactive intermediate-mediated mechanisms of teratogenesis.

    PubMed

    Wells, Peter G; Lee, Crystal J J; McCallum, Gordon P; Perstin, Julia; Harper, Patricia A

    2010-01-01

    Drugs and environmental chemicals can adversely alter the development of the fetus at critical periods during pregnancy, resulting in death, or in structural and functional birth defects in the surviving offspring. This process of teratogenesis may not be evident until a decade or more after birth. Postnatal functional abnormalities include deficits in brain function, a variety of metabolic diseases, and cancer. Due to the high degree of fetal cellular division and differentiation, and to differences from the adult in many biochemical pathways, the fetus is highly susceptible to teratogens, typically at low exposure levels that do not harm the mother. Insights into the mechanisms of teratogenesis come primarily from animal models and in vitro systems, and involve either receptor-mediated or reactive intermediate-mediated processes. Receptor-mediated mechanisms involving the reversible binding of xenobiotic substrates to a specific receptor are exemplified herein by the interaction of the environmental chemical 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD or "dioxin") with the cytosolic aryl hydrocarbon receptor (AHR), which translocates to the nucleus and, in association with other proteins, binds to AH-responsive elements (AHREs) in numerous genes, initiating changes in gene transcription that can perturb development. Alternatively, many xenobiotics are bioactivated by fetal enzymes like the cytochromes P450 (CYPs) and prostaglandin H synthases (PHSs) to highly unstable electrophilic or free radical reactive intermediates. Electrophilic reactive intermediates can covalently (irreversibly) bind to and alter the function of essential cellular macromolecules (proteins, DNA), causing developmental anomalies. Free radical reactive intermediates can enhance the formation of reactive oxygen species (ROS), resulting in oxidative damage to cellular macromolecules and/or altered signal transduction. The teratogenicity of reactive intermediates is determined to a large extent

  18. Microstructure evolution and mechanical behavior of a high strength dual-phase steel under monotonic loading

    SciTech Connect

    Nesterova, E.V.; Bouvier, S.; Bacroix, B.

    2015-02-15

    Transmission electron microscopy (TEM) microstructures of a high-strength dual-phase steel DP800 have been examined after moderate plastic deformations in simple shear and uniaxial tension. Special attention has been paid to the effect of the intergranular hard phase (martensite) on the microstructure evolution in the near-grain boundary regions. Quantitative parameters of dislocation patterning have been determined and compared with the similar characteristics of previously examined single-phase steels. The dislocation patterning in the interiors of the ferrite grains in DP800 steel is found to be similar to that already observed in the single-phase IF (Interstitial Free) steel whereas the martensite-affected zones present a delay in patterning and display very high gradients of continuous (gradual) disorientations associated with local internal stresses. The above stresses are shown to control the work-hardening of dual-phase materials at moderate strains for monotonic loading and are assumed to influence their microstructure evolution and mechanical behavior under strain-path changes. - Highlights: • The microstructure evolution has been studied by TEM in a DP800 steel. • It is influenced by both martensite and dislocations in the initial state. • The DP800 steel presents a high work-hardening rate due to internal stresses.

  19. Ammonium carbamates as highly active transdermal permeation enhancers with a dual mechanism of action.

    PubMed

    Novotný, Michal; Klimentová, Jana; Janůšová, Barbora; Palát, Karel; Hrabálek, Alexandr; Vávrová, Kateřina

    2011-03-10

    Transdermal permeation enhancers are compounds that temporarily increase drug flux through the skin by interacting with constituents of the stratum corneum. Transkarbam 12 (T12) is a highly active, broad-spectrum, biodegradable enhancer with low toxicity and low dermal irritation. We show here that T12 acts by a dual mechanism of action. The first part of this activity is associated with its ammonium carbamate polar head as shown by its pH-dependent effects on the permeation of two model drugs. Once this ammonium carbamate penetrates into the stratum corneum intercellular lipids, it rapidly decomposes releasing two molecules of protonated dodecyl 6-aminohexanoate (DDEAC) and carbon dioxide. This was observed by thermogravimetric analysis and infrared spectroscopy. This step of T12 action influences drug permeation through lipidic pathways, not through the aqueous pores (polar pathway) as shown by its effects on various model drugs and electrical impedance. Consequently, protonated DDEAC released in the stratum corneum is also an active enhancer. It broadens the scope of T12 action since it is also able to increase permeation of hydrophilic drugs that prefer the pore pathway. Thus, this dual effect of T12 is likely responsible for its favorable properties, which make it a good candidate for prospective clinical use.

  20. Synthesis and characterization of a dual kappa-delta opioid receptor agonist analgesic blocking cocaine reward behavior.

    PubMed

    Váradi, András; Marrone, Gina F; Eans, Shainnel O; Ganno, Michelle L; Subrath, Joan J; Le Rouzic, Valerie; Hunkele, Amanda; Pasternak, Gavril W; McLaughlin, Jay P; Majumdar, Susruta

    2015-11-18

    3-Iodobenzoyl naltrexamine (IBNtxA) is a potent analgesic belonging to the pharmacologically diverse 6β-amidoepoxymorphinan group of opioids. We present the synthesis and pharmacological evaluation of five analogs of IBNtxA. The scaffold of IBNtxA was modified by removing the 14-hydroxy group, incorporating a 7,8 double bond and various N-17 alkyl substituents. The structural modifications resulted in analogs with picomolar affinities for opioid receptors. The lead compound (MP1104) was found to exhibit approximately 15-fold greater antinociceptive potency (ED50 = 0.33 mg/kg) compared with morphine, mediated through the activation of kappa- and delta-opioid receptors. Despite its kappa agonism, this lead derivative did not cause place aversion or preference in mice in a place-conditioning assay, even at doses 3 times the analgesic ED50. However, pretreatment with the lead compound prevented the reward behavior associated with cocaine in a conditioned place preference assay. Together, these results suggest the promise of dual acting kappa- and delta-opioid receptor agonists as analgesics and treatments for cocaine addiction.

  1. Gutenberg-Richter law for deep earthquakes revisited: A dual-mechanism hypothesis

    NASA Astrophysics Data System (ADS)

    Zhan, Zhongwen

    2017-03-01

    Deep earthquake b values appear to vary with slab thermal state and earthquake magnitude. The physical reason for the variations and the relation with deep rupture mechanisms are still unclear. Here I confirm the spatial variations of b value and the dependence on slab temperature using about 40 yr of data from the Global Centroid Moment Tensor catalog. A new bimodal pattern is observed for the 500-700 km depth range: b is close to 1 in the cold Tonga slab, while in warmer slabs (e.g., South America, Japan-Kuril, Izu-Bonin-Mariana), b is close to 0.5 for intermediate magnitudes (Mw5.3-6.5) and increases to ∼1 for large magnitudes (Mw > 6.5). To explain these observations, I propose a dual-mechanism hypothesis in which deep earthquakes nucleate only within the metastable olivine wedge (MOW), but can rupture outside MOW by a different mechanism. The fractal dimension of earthquake size distribution changes from 2 to 1 as the thermally controlled MOW thickness decreases, and back to 2 as the mechanism outside MOW dominates.

  2. Antibacterial activity of phenyllactic acid against Listeria monocytogenes and Escherichia coli by dual mechanisms.

    PubMed

    Ning, Yawei; Yan, Aihong; Yang, Kun; Wang, Zhixin; Li, Xingfeng; Jia, Yingmin

    2017-08-01

    Phenyllactic acid (PLA), a phenolic acid phytochemical, is considered to be a promising candidate for use as a chemical preservative due to its broad antimicrobial activity. The antibacterial target of PLA has rarely been reported, thus investigations were performed to elucidate the antibacterial mechanism of PLA against Listeria monocytogenes and Escherichia coli. Flow cytometry analysis stained with propidium iodide (PI) demonstrated that PLA could damage the membrane integrity of L. monocytogenes, while it could not disrupt that of E. coli. The uptake of 1-N-phenylnaphthylamine (NPN) indicated that PLA interrupted the outer membrane permeability of E. coli. Scanning electron microscopy (SEM) observation visualized the damage caused by PLA as morphological changes in L. monocytogenes and E. coli. Fluorescence assays demonstrated that PLA could interact with bacterial genomic DNA in the manner of intercalation. This finding suggested dual antibacterial targets of PLA, namely membrane and genomic DNA.

  3. Inhibitory Mechanism of an Allosteric Antibody Targeting the Glucagon Receptor*

    PubMed Central

    Mukund, Susmith; Shang, Yonglei; Clarke, Holly J.; Madjidi, Azadeh; Corn, Jacob E.; Kates, Lance; Kolumam, Ganesh; Chiang, Vicky; Luis, Elizabeth; Murray, Jeremy; Zhang, Yingnan; Hötzel, Isidro; Koth, Christopher M.; Allan, Bernard B.

    2013-01-01

    Elevated glucagon levels and increased hepatic glucagon receptor (GCGR) signaling contribute to hyperglycemia in type 2 diabetes. We have identified a monoclonal antibody that inhibits GCGR, a class B G-protein coupled receptor (GPCR), through a unique allosteric mechanism. Receptor inhibition is mediated by the binding of this antibody to two distinct sites that lie outside of the glucagon binding cleft. One site consists of a patch of residues that are surface-exposed on the face of the extracellular domain (ECD) opposite the ligand-binding cleft, whereas the second binding site consists of residues in the αA helix of the ECD. A docking model suggests that the antibody does not occlude the ligand-binding cleft. We solved the crystal structure of GCGR ECD containing a naturally occurring G40S mutation and found a shift in the register of the αA helix that prevents antibody binding. We also found that alterations in the αA helix impact the normal function of GCGR. We present a model for the allosteric inhibition of GCGR by a monoclonal antibody that may form the basis for the development of allosteric modulators for the treatment of diabetes and other class B GPCR-related diseases. PMID:24189067

  4. Mechanism of dimerization of the human melanocortin 1 receptor

    SciTech Connect

    Zanna, Paola T.; Sanchez-Laorden, Berta L.; Perez-Oliva, Ana B.; Turpin, Maria C.; Herraiz, Cecilia; Jimenez-Cervantes, Celia; Garcia-Borron, Jose C.

    2008-04-04

    The melanocortin 1 receptor (MC1R) is a dimeric G protein-coupled receptor expressed in melanocytes, where it regulates the amount and type of melanins produced and determines the tanning response to ultraviolet radiation. We have studied the mechanisms of MC1R dimerization. Normal dimerization of a deleted mutant lacking the seventh transmembrane fragment and the C-terminal cytosolic extension excluded coiled-coil interactions as the basis of dimerization. Conversely, the electrophoretic pattern of wild type receptor and several Cys {yields} Ala mutants showed that four disulfide bonds are established between the monomers. Disruption of any of these bonds abolished MC1R function, but only the one involving Cys35 was essential for traffic to the plasma membrane. A quadruple Cys35-267-273-275Ala mutant migrating as a monomer in SDS-PAGE in the absence of reducing agents was able to dimerize with WT, suggesting that in addition to disulfide bond formation, dimerization involves non-covalent interactions, likely of domain swap type.

  5. Inhibitory mechanism of an allosteric antibody targeting the glucagon receptor.

    PubMed

    Mukund, Susmith; Shang, Yonglei; Clarke, Holly J; Madjidi, Azadeh; Corn, Jacob E; Kates, Lance; Kolumam, Ganesh; Chiang, Vicky; Luis, Elizabeth; Murray, Jeremy; Zhang, Yingnan; Hötzel, Isidro; Koth, Christopher M; Allan, Bernard B

    2013-12-13

    Elevated glucagon levels and increased hepatic glucagon receptor (GCGR) signaling contribute to hyperglycemia in type 2 diabetes. We have identified a monoclonal antibody that inhibits GCGR, a class B G-protein coupled receptor (GPCR), through a unique allosteric mechanism. Receptor inhibition is mediated by the binding of this antibody to two distinct sites that lie outside of the glucagon binding cleft. One site consists of a patch of residues that are surface-exposed on the face of the extracellular domain (ECD) opposite the ligand-binding cleft, whereas the second binding site consists of residues in the αA helix of the ECD. A docking model suggests that the antibody does not occlude the ligand-binding cleft. We solved the crystal structure of GCGR ECD containing a naturally occurring G40S mutation and found a shift in the register of the αA helix that prevents antibody binding. We also found that alterations in the αA helix impact the normal function of GCGR. We present a model for the allosteric inhibition of GCGR by a monoclonal antibody that may form the basis for the development of allosteric modulators for the treatment of diabetes and other class B GPCR-related diseases.

  6. Mechanical design of SST-GATE, a dual-mirror telescope for the Cherenkov Telescope Array

    NASA Astrophysics Data System (ADS)

    Dournaux, Jean-Laurent; Huet, Jean-Michel; Amans, Jean-Philippe; Dumas, Delphine; Laporte, Philippe; Sol, Hélène; Blake, Simon

    2014-07-01

    The Cherenkov Telescope Array (CTA) project aims to create the next generation Very High Energy (VHE) gamma-ray telescope array. It will be devoted to the observation of gamma rays over a wide band of energy, from a few tens of GeV to more than 100 TeV. Two sites are foreseen to view the whole sky where about 100 telescopes, composed of three different classes, related to the specific energy region to be investigated, will be installed. Among these, the Small Size class of Telescopes, SSTs, are devoted to the highest energy region, to beyond 100 TeV. Due to the large number of SSTs, their unit cost is an important parameter. At the Observatoire de Paris, we have designed a prototype of a Small Size Telescope named SST-GATE, based on the dual-mirror Schwarzschild-Couder optical formula, which has never before been implemented in the design of a telescope. Over the last two years, we developed a mechanical design for SST-GATE from the optical and preliminary mechanical designs made by the University of Durham. The integration of this telescope is currently in progress. Since the early stages of mechanical design of SST-GATE, finite element method has been used employing shape and topology optimization techniques to help design several elements of the telescope. This allowed optimization of the mechanical stiffness/mass ratio, leading to a lightweight and less expensive mechanical structure. These techniques and the resulting mechanical design are detailed in this paper. We will also describe the finite element analyses carried out to calculate the mechanical deformations and the stresses in the structure under observing and survival conditions.

  7. Strain rate effects on the mechanical behavior of two Dual Phase steels in tension

    NASA Astrophysics Data System (ADS)

    Cadoni, E.; Singh, N. K.; Forni, D.; Singha, M. K.; Gupta, N. K.

    2016-05-01

    This paper presents an experimental investigation on the strain rate sensitivity of Dual Phase steel 1200 (DP1200) and Dual Phase steel 1400 (DP1400) under uni-axial tensile loads in the strain rate range from 0.001 s-1 to 600 s-1. These materials are advanced high strength steels (AHSS) having high strength, high capacity to dissipate crash energy and high formability. Flat sheet specimens of the materials having gauge length 10 mm, width 4 mm and thickness 2 mm (DP1200) and 1.25 mm (DP1400), are tested at room temperature (20∘C) on electromechanical universal testing machine to obtain their stress-strain relation under quasi-static condition (0.001 s-1), and on Hydro-Pneumatic machine and modified Hopkinson bar to study their mechanical behavior at medium (3 s-1, and 18 s-1) and high strain rates (200 s-1, 400 s-1, and 600 s-1) respectively. Tests under quasi-static condition are performed at high temperature (200∘C) also, and found that tensile flow stress is a increasing function of temperature. The stress-strain data has been analysed to determine the material parameters of the Cowper-Symonds and the Johnson-Cook models. A simple modification of the Johnson-Cook model has been proposed in order to obtain a better fit of tests at high temperatures. Finally, the fractographs of the broken specimens are taken by scanning electron microscope (SEM) to understand the fracture mechanism of these advanced high strength steels at different strain rates.

  8. CD44 Receptor Targeting and Endosomal pH-Sensitive Dual Functional Hyaluronic Acid Micelles for Intracellular Paclitaxel Delivery.

    PubMed

    Liu, Yanhua; Zhou, Chengming; Wang, Wenping; Yang, Jianhong; Wang, Hao; Hong, Wei; Huang, Yu

    2016-12-05

    A novel CD44 receptor targeting and endosome pH-sensitive dual functional hyaluronic acid-deoxycholic acid-histidine (HA-DOCA-His) micellar system was designed for intracellular paclitaxel (PTX) delivery. The HA-DOCA-His micelles exhibited desirable endosome pH (5.0-6.0)-induced aggregation and deformation behavior verified by size distribution, critical micellar concentration, and zeta potential changes. The HA-DOCA-His micelles presented excellent encapsulation efficiency and loading capacity of 90.0% and 18.9% for PTX, respectively. The PTX release from HA-DOCA-His micelles was pH-dependent, with more rapid PTX release at pH 6.0 and 5.0 than those at pH 7.4 and 6.5. The cellular uptake performance of HA-DOCA-His micelles was enhanced comparing with pH-insensitive HA-DOCA micelles by qualitative and quantitative measurements. HA-DOCA-His micelles could be taken up via CD44-receptor mediated endocytosis, transported into endosomes, and triggered drug release to cytoplasm. In vitro cytotoxicity study exhibited PTX-loaded HA-DOCA-His micelles were more active in tumor cell growth inhibition in MCF-7 cells at pH 5.8 than those at pH 6.8 and pH 7.4. A superior antitumor efficacy was demonstrated with HA-DOCA-His micelles in a MCF-7 breast tumor model. These indicated that the dual functional HA-DOCA-His micelles combined targeted intracellular delivery and endosomal release strategies could be developed as a promising nanocarrier for anticancer efficacy improvement of PTX.

  9. Potent neutralization of hepatitis A virus reveals a receptor mimic mechanism and the receptor recognition site

    PubMed Central

    Wang, Xiangxi; Zhu, Ling; Dang, Minghao; Hu, Zhongyu; Gao, Qiang; Yuan, Shuai; Sun, Yao; Zhang, Bo; Ren, Jingshan; Kotecha, Abhay; Walter, Thomas S.; Wang, Junzhi; Fry, Elizabeth E.; Stuart, David I.; Rao, Zihe

    2017-01-01

    Hepatitis A virus (HAV) infects ∼1.4 million people annually and, although there is a vaccine, there are no licensed therapeutic drugs. HAV is unusually stable (making disinfection problematic) and little is known of how it enters cells and releases its RNA. Here we report a potent HAV-specific monoclonal antibody, R10, which neutralizes HAV infection by blocking attachment to the host cell. High-resolution cryo-EM structures of HAV full and empty particles and of the complex of HAV with R10 Fab reveal the atomic details of antibody binding and point to a receptor recognition site at the pentamer interface. These results, together with our observation that the R10 Fab destabilizes the capsid, suggest the use of a receptor mimic mechanism to neutralize virus infection, providing new opportunities for therapeutic intervention. PMID:28074040

  10. Potent neutralization of hepatitis A virus reveals a receptor mimic mechanism and the receptor recognition site.

    PubMed

    Wang, Xiangxi; Zhu, Ling; Dang, Minghao; Hu, Zhongyu; Gao, Qiang; Yuan, Shuai; Sun, Yao; Zhang, Bo; Ren, Jingshan; Kotecha, Abhay; Walter, Thomas S; Wang, Junzhi; Fry, Elizabeth E; Stuart, David I; Rao, Zihe

    2017-01-24

    Hepatitis A virus (HAV) infects ∼1.4 million people annually and, although there is a vaccine, there are no licensed therapeutic drugs. HAV is unusually stable (making disinfection problematic) and little is known of how it enters cells and releases its RNA. Here we report a potent HAV-specific monoclonal antibody, R10, which neutralizes HAV infection by blocking attachment to the host cell. High-resolution cryo-EM structures of HAV full and empty particles and of the complex of HAV with R10 Fab reveal the atomic details of antibody binding and point to a receptor recognition site at the pentamer interface. These results, together with our observation that the R10 Fab destabilizes the capsid, suggest the use of a receptor mimic mechanism to neutralize virus infection, providing new opportunities for therapeutic intervention.

  11. Allosteric mechanisms of nuclear receptors: insights from computational simulations.

    PubMed

    Mackinnon, Jonathan A G; Gallastegui, Nerea; Osguthorpe, David J; Hagler, Arnold T; Estébanez-Perpiñá, Eva

    2014-08-05

    The traditional structural view of allostery defines this key regulatory mechanism as the ability of one conformational event (allosteric site) to initiate another in a separate location (active site). In recent years computational simulations conducted to understand how this phenomenon occurs in nuclear receptors (NRs) has gained significant traction. These results have yield insights into allosteric changes and communication mechanisms that underpin ligand binding, coactivator binding site formation, post-translational modifications, and oncogenic mutations. Moreover, substantial efforts have been made in understanding the dynamic processes involved in ligand binding and coregulator recruitment to different NR conformations in order to predict cell/tissue-selective pharmacological outcomes of drugs. They also have improved the accuracy of in silico screening protocols so that nowadays they are becoming part of optimisation protocols for novel therapeutics. Here we summarise the important contributions that computational simulations have made towards understanding the structure/function relationships of NRs and how these can be exploited for rational drug design.

  12. GABAA receptor inhibition triggers a nicotinic neuroprotective mechanism

    PubMed Central

    Ferchmin, P. A; Pérez, Dinely; Alvarez, William Castro; Penzo, Mario A.; Maldonado, Héctor M.; Eterovic, Vesna A.

    2014-01-01

    Nicotinic acetylcholine receptor (nAChR)-mediated neuroprotection has been implicated in the treatment of neurodegenerative disorders such as Alzheimer’s, Parkinson’s and hypoxic ischemic events, as well as other diseases hallmarked by excitotoxic and apoptotic neuronal death. Several modalities of nicotinic neuroprotection have been reported. However, although this process generally involves α4β2 and α7 subtypes, the underlying mechanisms are largely unknown. Interestingly, both activation and inhibition of α7 nAChRs have been reported to be neuroprotective. We have shown that inhibition of α7 nAChRs protects the function of acute hippocampal slices against excitotoxicity in a α4β2-dependent manner. Neuroprotection was assessed as the prevention of the NMDA-dependent loss of the area of population spikes (PSs) in the CA1 area of acute hippocampal slices. Our results support a model in which α7 AChRs control the release of GABA. Blocking either α7 or GABAA receptors reduces the inhibitory tone on cholinergic terminals, thereby promoting α4β2 activation, which in turn mediates neuroprotection. These results shed light on how α7 nAChR inhibition can be neuroprotective through a mechanism mediated by activation of α4β2 nAChRs. PMID:23280428

  13. A smart pinless ejection mechanism using dual-resonance excitation Langevin piezoelectric transducers

    NASA Astrophysics Data System (ADS)

    Wang, Yu-Jen; Fu, Kuo-Chieh; Wang, Chun-Chieh

    2016-01-01

    This study investigated a smart pinless ejection mechanism comprising two dual-resonance excitation Langevin piezoelectric transducers (DRELPTs) for keeping the injection parts intact and protecting their top and bottom surfaces from scarring during plastic injection molding. The dimensions of each DRELPT were determined using longitudinal vibration models, and an optimization method was used to set the frequency ratio of the first to the second longitudinal mode to 1:2. This concept enables the driving of DRELPT in its two longitudinal modes consistent with the ejection direction in resonant-type smooth impact drive mechanisms. During the ejection process, DRELPT provides an ejection force, which is applied on the sidewalls of the injection parts to protect their top and bottom surfaces from scarring. Considering individual differences in the resonance frequencies of DRELPTs, a resonance frequency tracking circuit based on a phase-locked loop was designed to keep DRELPT actuating in resonance. The ejection velocity of the injection part was estimated using the kinetic models derived from the dynamic behavior of the mold cavity and injection parameters. A characteristic number S was defined to evaluate the average velocity of the injection part during ejection. Proof-of-concept experimental results of the pinless ejection mechanism are presented. The ejection time, that is, the time from triggering the composite wave to the full departure of the injection part from the mold cavity, was 72 ms.

  14. The receptor kinase CERK1 has dual functions in symbiosis and immunity signalling.

    PubMed

    Zhang, Xiaowei; Dong, Wentao; Sun, Jongho; Feng, Feng; Deng, Yiwen; He, Zuhua; Oldroyd, Giles E D; Wang, Ertao

    2015-01-01

    The establishment of symbiotic interactions between mycorrhizal fungi, rhizobial bacteria and their legume hosts involves a common symbiosis signalling pathway. This signalling pathway is activated by Nod factors produced by rhizobia and these are recognised by the Nod factor receptors NFR1/LYK3 and NFR5/NFP. Mycorrhizal fungi produce lipochitooligosaccharides (LCOs) similar to Nod factors, as well as short-chain chitin oligomers (CO4/5), implying commonalities in signalling during mycorrhizal and rhizobial associations. Here we show that NFR1/LYK3, but not NFR5/NFP, is required for the establishment of the mycorrhizal interaction in legumes. NFR1/LYK3 is necessary for the recognition of mycorrhizal fungi and the activation of the symbiosis signalling pathway leading to induction of calcium oscillations and gene expression. Chitin oligosaccharides also act as microbe associated molecular patterns that promote plant immunity via similar LysM receptor-like kinases. CERK1 in rice has the highest homology to NFR1 and we show that this gene is also necessary for the establishment of the mycorrhizal interaction as well as for resistance to the rice blast fungus. Our results demonstrate that NFR1/LYK3/OsCERK1 represents a common receptor for chitooligosaccharide-based signals produced by mycorrhizal fungi, rhizobial bacteria (in legumes) and fungal pathogens. It would appear that mycorrhizal recognition has been conserved in multiple receptors across plant species, but additional diversification in certain plant species has defined other signals that this class of receptors can perceive.

  15. Peripheral Receptor Mechanisms Underlying Orofacial Muscle Pain and Hyperalgesia

    NASA Astrophysics Data System (ADS)

    Saloman, Jami L.

    Musculoskeletal pain conditions, particularly those associated with temporomandibular joint and muscle disorders (TMD) are severely debilitating and affect approximately 12% of the population. Identifying peripheral nociceptive mechanisms underlying mechanical hyperalgesia, a prominent feature of persistent muscle pain, could contribute to the development of new treatment strategies for the management of TMD and other muscle pain conditions. This study provides evidence of functional interactions between ligand-gated channels, P2X3 and TRPV1/TRPA1, in trigeminal sensory neurons, and proposes that these interactions underlie the development of mechanical hyperalgesia. In the masseter muscle, direct P2X3 activation, via the selective agonist αβmeATP, induced a dose- and time-dependent hyperalgesia. Importantly, the αβmeATP-induced hyperalgesia was prevented by pretreatment of the muscle with a TRPV1 antagonist, AMG9810, or the TRPA1 antagonist, AP18. P2X3 was co-expressed with both TRPV1 and TRPA1 in masseter muscle afferents confirming the possibility for intracellular interactions. Moreover, in a subpopulation of P2X3 /TRPV1 positive neurons, capsaicin-induced Ca2+ transients were significantly potentiated following P2X3 activation. Inhibition of Ca2+-dependent kinases, PKC and CaMKII, prevented P2X3-mechanical hyperalgesia whereas blockade of Ca2+-independent PKA did not. Finally, activation of P2X3 induced phosphorylation of serine, but not threonine, residues in TRPV1 in trigeminal sensory neurons. Significant phosphorylation was observed at 15 minutes, the time point at which behavioral hyperalgesia was prominent. Similar data were obtained regarding another nonselective cation channel, the NMDA receptor (NMDAR). Our data propose P2X3 and NMDARs interact with TRPV1 in a facilitatory manner, which could contribute to the peripheral sensitization underlying masseter hyperalgesia. This study offers novel mechanisms by which individual pro-nociceptive ligand

  16. Concentration-Dependent Dual Mode of Zn Action at Serotonin 5-HT1A Receptors: In Vitro and In Vivo Studies.

    PubMed

    Satała, Grzegorz; Duszyńska, Beata; Stachowicz, Katarzyna; Rafalo, Anna; Pochwat, Bartlomiej; Luckhart, Christine; Albert, Paul R; Daigle, Mireille; Tanaka, Kenji F; Hen, René; Lenda, Tomasz; Nowak, Gabriel; Bojarski, Andrzej J; Szewczyk, Bernadeta

    2016-12-01

    Recent data has indicated that Zn can modulate serotonergic function through the 5-HT1A receptor (5-HT1AR); however, the exact mechanisms are unknown. In the present studies, radioligand binding assays and behavioural approaches were used to characterize the pharmacological profile of Zn at 5-HT1ARs in more detail. The influence of Zn on agonist binding to 5-HT1ARs stably expressed in HEK293 cells was investigated by in vitro radioligand binding methods using the agonist [(3)H]-8-OH-DPAT. The in vivo effects of Zn were compared with those of 8-OH-DPAT in hypothermia, lower lip retraction (LLR), 5-HT behavioural syndrome and the forced swim (FST) tests. In the in vitro studies, biphasic effects, which involved allosteric potentiation of agonist binding at sub-micromolar Zn concentrations and inhibition at sub-millimolar Zn concentrations, were found. The in vivo studies showed that Zn did not induce LLR or elements of 5-HT behavioural syndrome but blocked such effects induced by 8-OH-DPAT. Zn decreased body temperature in rats and mice; however, Zn failed to induce hypothermia in the 5-HT1A autoreceptor knockout mice. In the FST, Zn potentiated the effect of 8-OH-DPAT. However, in the FST performed with the 5-HT1A autoreceptor knockout mice, the anti-immobility effect of Zn was partially blocked. Both the binding and behavioural studies suggest a concentration-dependent dual mechanism of Zn action at 5-HT1ARs, with potentiation at low dose and inhibition at high dose. Moreover, the in vivo studies indicate that Zn can modulate both presynaptic and postsynaptic 5-HT1ARs; however, Zn's effects at presynaptic receptors seem to be more potent.

  17. On the mechanism of extractive electrospray ionization (EESI) in the dual-spray configuration.

    PubMed

    Wang, Rui; Gröhn, Arto Juhani; Zhu, Liang; Dietiker, Rolf; Wegner, Karsten; Günther, Detlef; Zenobi, Renato

    2012-03-01

    Dual-spray extractive electrospray ionization (EESI) mass spectrometry as a versatile analytical technique has attracted much interest due to its advantages over conventional electrospray ionization (ESI). The crucial difference between EESI and ESI is that in the EESI process, the analytes are introduced in nebulized form via a neutral spray and ionized by collisions with the charged droplets from an ESI source formed by spraying pure solvent. However, the mechanism of the droplet-droplet interactions in the EESI process is still not well understood. For example, it is unclear which type of droplet-droplet interaction is dominant: bounce, coalescence, disruption, or fragmentation? In this work, droplet-droplet interaction was investigated in detail based on a theoretical model. Phase Doppler anemometry (PDA) was employed to investigate the droplet behavior in the EESI plume and provide the experimental data (droplet size and velocity) necessary for theoretical analysis. Furthermore, numerical simulations were performed to clarify the influence of the sheath gas flow on the EESI process. No coalescence between the droplets in the ESI spray and the droplets in the sample spray was observed using various geometries and sample flow rates. Theoretical analysis, together with the PDA results, suggests that droplet fragmentation may be the dominant type of droplet-droplet interaction in the EESI. The interaction time between the ESI droplet and the sample droplet was estimated to be <5 μs. This work gives a clear picture of droplet-droplet interactions in the dual-spray EESI process and detailed information for the optimization of this method for future applications that require higher sensitivity.

  18. Dual temporal encoding mechanisms in human auditory cortex: Evidence from MEG and EEG.

    PubMed

    Tang, Huizhen; Crain, Stephen; Johnson, Blake W

    2016-03-01

    Current hypotheses about language processing advocate an integral relationship between encoding of temporal information and linguistic processing in the brain. All such explanations must accommodate the evident ability of the perceptual system to process both slow and fast time scales in speech. However most cortical neurons are limited in their capability to precisely synchronise to temporal modulations at rates faster than about 50Hz. Hence, a central question in auditory neurophysiology concerns how the full range of perceptually relevant modulation rates might be encoded in the cerebral cortex. Here we show with concurrent noninvasive magnetoencephalography (MEG) and electroencephalography (EEG) measurements that the human auditory cortex transitions between a phase-locked (PL) mode of responding to modulation rates below about 50Hz, and a non-phase-locked (NPL) mode at higher rates. Precisely such dual response modes are predictable from the behaviours of single neurons in auditory cortices of non-human primates. Our data point to a common mechanistic explanation for the single neuron and MEG/EEG results and support the hypothesis that two distinct types of neuronal encoding mechanisms are employed by the auditory cortex to represent a wide range of temporal modulation rates. This dual encoding model allows slow and fast modulations in speech to be processed in parallel and is therefore consistent with theoretical frameworks in which slow temporal modulations (such as rhythm or syllabic structure) are akin to the contours or edges of visual objects, whereas faster modulations (such as periodicity pitch or phonemic structure) are more like visual texture.

  19. Synergistic growth inhibitory effects of the dual endothelin-1 receptor antagonist bosentan on pancreatic stellate and cancer cells.

    PubMed

    Fitzner, Brit; Brock, Peter; Holzhüter, Stephanie-Anna; Nizze, Horst; Sparmann, Gisela; Emmrich, Jörg; Liebe, Stefan; Jaster, Robert

    2009-02-01

    Pancreatic stellate cells (PSC) play a key role in pancreatic fibrosis. Activation of PSC occurs in response to pro-fibrogenic stimuli and is maintained by autocrine loops of mediators, such as endothelin (ET)-1. Here, we have evaluated effects of the dual ET receptor antagonist bosentan in models of pancreatic fibrogenesis and cancer. Cell culture studies revealed that PSC and DSL6A pancreatic cancer cells expressed both ET-1 and ET receptors. Bosentan efficiently inhibited proliferation of both cell types and collagen synthesis in PSC. Expression of the myofibroblastic marker alpha-smooth muscle actin, connective tissue growth factor, and ET-1 itself in PSC was reduced, while expression of matrix metalloproteinase-9 was enhanced. Like PSC, DSL6A cells secrete less ET-1 when cultured with bosentan. In a rat model of pancreatic fibrosis, chronic pancreatitis induced by dibutyltin dichloride, a tendency towards a diminished disease progression was observed in a subgroup of rats with less severe disease. Together, our results indicate that bosentan exerts antifibrotic and antitumor effects in vitro. Its efficiency in vivo warrants further investigation.

  20. A Dual Target-directed Agent against Interleukin-6 Receptor and Tumor Necrosis Factor α ameliorates experimental arthritis

    PubMed Central

    Kim, Youngkyun; Yi, Hyoju; Jung, Hyerin; Rim, Yeri Alice; Park, Narae; Kim, Juryun; Jung, Seung Min; Park, Sung-Hwan; Park, Young Woo; Ju, Ji Hyeon

    2016-01-01

    A considerable proportion of patients with rheumatoid arthritis (RA) do not respond to monospecific agents. The purpose of our study was to generate a hybrid form of biologics, targeting tumor-necrosis factor alpha (TNFα) and interleukin-6 receptor (IL-6R), and determine its anti-arthritic properties in vitro and in vivo. A novel dual target-directed agent (DTA(A7/sTNFR2)) was generated by conjugating soluble TNF receptor 2 (sTNFR2) to the Fc region of A7, a new anti-IL-6R antibody obtained by screening the phage display human antibody library. DTA(A7/sTNFR2) inhibited the proliferation and migration of fibroblast-like synoviocytes from patients with RA (RA-FLS) more efficiently than single target-directed agents. DTA(A7/sTNFR2) also blocked osteoclastogenesis from bone marrow cells. The arthritis severity scores of the experimental arthritis mice with DTA(A7/sTNFR2) tended to be lower than those of mice with IgG, A7, or sTNFR2. Histological data suggested that DTA(A7/sTNFR2) is more efficient than single-target drugs in preventing joint destruction and bone loss. These results were confirmed in vivo using the minicircle system. Taken together, the results show that DTA(A7/sTNFR2) may be a promising therapeutic agent for the treatment of RA. PMID:26841833

  1. Dual Endothelin Receptor Blockade Abrogates Right Ventricular Remodeling and Biventricular Fibrosis in Isolated Elevated Right Ventricular Afterload

    PubMed Central

    Nielsen, Eva Amalie; Sun, Mei; Honjo, Osami; Hjortdal, Vibeke E.; Redington, Andrew N.; Friedberg, Mark K.

    2016-01-01

    Background Pulmonary arterial hypertension is usually fatal due to right ventricular failure and is frequently associated with co-existing left ventricular dysfunction. Endothelin-1 is a powerful pro-fibrotic mediator and vasoconstrictor that is elevated in pulmonary arterial hypertension. Endothelin receptor blockers are commonly used as pulmonary vasodilators, however their effect on biventricular injury, remodeling and function, despite elevated isolated right ventricular afterload is unknown. Methods Elevated right ventricular afterload was induced by progressive pulmonary artery banding. Seven rabbits underwent pulmonary artery banding without macitentan; 13 received pulmonary artery banding + macitentan; and 5 did not undergo inflation of the pulmonary artery band (sham-operated controls). Results: Right and left ventricular collagen content was increased with pulmonary artery banding compared to sham-operated controls and ameliorated by macitentan. Right ventricular fibrosis signaling (connective tissue growth factor and endothelin-1 protein levels); extra-cellular matrix remodeling (matrix-metalloproteinases 2 and 9), apoptosis and apoptosis-related peptides (caspases 3 and 8) were increased with pulmonary artery banding compared with sham-operated controls and decreased with macitentan. Conclusion Isolated right ventricular afterload causes biventricular fibrosis, right ventricular apoptosis and extra cellular matrix remodeling, mediated by up-regulation of endothelin-1 and connective tissue growth factor signaling. These pathological changes are ameliorated by dual endothelin receptor blockade despite persistent elevated right ventricular afterload. PMID:26765263

  2. A dual inhibition mechanism of herpesviral ICP47 arresting a conformationally thermostable TAP complex

    PubMed Central

    Herbring, Valentina; Bäucker, Anja; Trowitzsch, Simon; Tampé, Robert

    2016-01-01

    As a centerpiece of antigen processing, the ATP-binding cassette transporter associated with antigen processing (TAP) became a main target for viral immune evasion. The herpesviral ICP47 inhibits TAP function, thereby suppressing an adaptive immune response. Here, we report on a thermostable ICP47-TAP complex, generated by fusion of different ICP47 fragments. These fusion complexes allowed us to determine the direction and positioning in the central cavity of TAP. ICP47-TAP fusion complexes are arrested in a stable conformation, as demonstrated by MHC I surface expression, melting temperature, and the mutual exclusion of herpesviral TAP inhibitors. We unveiled a conserved region next to the active domain of ICP47 as essential for the complete stabilization of the TAP complex. Binding of the active domain of ICP47 arrests TAP in an open inward facing conformation rendering the complex inaccessible for other viral factors. Based on our findings, we propose a dual interaction mechanism for ICP47. A per se destabilizing active domain inhibits the function of TAP, whereas a conserved C-terminal region additionally stabilizes the transporter. These new insights into the ICP47 inhibition mechanism can be applied for future structural analyses of the TAP complex. PMID:27845362

  3. A plausible mechanism of biosorption in dual symbioses by vesicular-arbuscular mycorrhizal in plants.

    PubMed

    Azmat, Rafia; Hamid, Neelofer

    2015-03-01

    Dual symbioses of vesicular-arbuscular mycorrhizal (VAM) fungi with growth of Momordica charantia were elucidated in terms of plausible mechanism of biosorption in this article. The experiment was conducted in green house and mixed inoculum of the VAM fungi was used in the three replicates. Results demonstrated that the starch contents were the main source of C for the VAM to builds their hyphae. The increased plant height and leaves surface area were explained in relation with an increase in the photosynthetic rates to produce rapid sugar contents for the survival of plants. A decreased in protein, and amino acid contents and increased proline and protease activity in VAM plants suggested that these contents were the main bio-indicators of the plants under biotic stress. The decline in protein may be due to the degradation of these contents, which later on converted into dextrose where it can easily be absorbed by for the period of symbioses. A mechanism of C chemisorption in relation with physiology and morphology of plant was discussed.

  4. Two separate, but interacting, neural systems for familiarity and novelty detection: a dual-route mechanism.

    PubMed

    Kafkas, Alexandros; Montaldi, Daniela

    2014-05-01

    It has long been assumed that familiarity- and novelty-related processes fall on a single continuum drawing on the same cognitive and neural mechanisms. The possibility that familiarity and novelty processing involve distinct neural networks was explored in a functional magnetic resonance imaging study (fMRI), in which familiarity and novelty judgments were made in contexts emphasizing either familiarity or novelty decisions. Parametrically modulated BOLD responses to familiarity and novelty strength were isolated in two separate, nonoverlapping brain networks. The novelty system involved brain regions along the ventral visual stream, the hippocampus, and the perirhinal and parahippocampal cortices. The familiarity system, on the other hand, involved the dorsomedial thalamic nucleus, and regions within the medial prefrontal cortex and the medial and lateral parietal cortex. Convergence of the two networks, treating familiarity and novelty as a single continuum was only found in a fronto-parietal network. Finally, the orbitomedial prefrontal cortex was found to be sensitive to reported strength/confidence, irrespective of stimulus' familiarity or novelty. This pattern of results suggests a dual-route mechanism supported by the existence of two distinct but interacting functional systems for familiarity and novelty. Overall, these findings challenge current assumptions regarding the neural systems that support the processing of novel and familiar information, and have important implications for research into the neural bases of recognition memory.

  5. Investigating enhanced mechanical properties in dual-phase Fe-Ga-Tb alloys.

    PubMed

    Meng, Chongzheng; Wang, Hui; Wu, Yuye; Liu, Jinghua; Jiang, Chengbao

    2016-10-03

    Dual-phase (Fe83Ga17)100-xTbx alloys with 0 ≤ x ≤ 1 were synthesized by arc melting and homogenization treatment. The microstructures and the corresponding mechanical properties were systematically investigated. The chemical composition of the body centered cubic matrix is Fe83Ga17. The monoclinic second phase was composed of meltable precipitates with approximate composition Fe57Ga33Tb10. The nano-hardness of matrix and precipitates were 2.55 ± 0.17 GPa and 6.81 ± 1.03 GPa, respectively. Both the ultimate tensile strength (UTS) and fracture strain (ε) of the alloys were improved by the precipitates for x ≤ 0.2 alloys, but the strain decreases significantly at higher values of x. As potential structural-functional materials, the best mechanical properties obtained were a UTS of 595 ± 10 MPa and an ε of 3.5 ± 0.1%, four-fold and seven-fold improvements compared with the un-doped alloy. The mechanism for these anomalous changes of mechanical properties was attributed to the dispersed precipitates and semi-coherent interfaces, which serve as strong obstacles to dislocation motion and reduce the stress concentration at the grain boundaries. A sizeable improvement of magnetostriction induced by the precipitates in the range 0 ≤ x ≤ 0.2 was discovered and an optimal value of 150 ± 5 ppm is found, over three times higher than that of the un-doped alloy.

  6. Investigating enhanced mechanical properties in dual-phase Fe-Ga-Tb alloys

    PubMed Central

    Meng, Chongzheng; Wang, Hui; Wu, Yuye; Liu, Jinghua; Jiang, Chengbao

    2016-01-01

    Dual-phase (Fe83Ga17)100−xTbx alloys with 0 ≤ x ≤ 1 were synthesized by arc melting and homogenization treatment. The microstructures and the corresponding mechanical properties were systematically investigated. The chemical composition of the body centered cubic matrix is Fe83Ga17. The monoclinic second phase was composed of meltable precipitates with approximate composition Fe57Ga33Tb10. The nano-hardness of matrix and precipitates were 2.55 ± 0.17 GPa and 6.81 ± 1.03 GPa, respectively. Both the ultimate tensile strength (UTS) and fracture strain (ε) of the alloys were improved by the precipitates for x ≤ 0.2 alloys, but the strain decreases significantly at higher values of x. As potential structural-functional materials, the best mechanical properties obtained were a UTS of 595 ± 10 MPa and an ε of 3.5 ± 0.1%, four-fold and seven-fold improvements compared with the un-doped alloy. The mechanism for these anomalous changes of mechanical properties was attributed to the dispersed precipitates and semi-coherent interfaces, which serve as strong obstacles to dislocation motion and reduce the stress concentration at the grain boundaries. A sizeable improvement of magnetostriction induced by the precipitates in the range 0 ≤ x ≤ 0.2 was discovered and an optimal value of 150 ± 5 ppm is found, over three times higher than that of the un-doped alloy. PMID:27694839

  7. Investigating enhanced mechanical properties in dual-phase Fe-Ga-Tb alloys

    NASA Astrophysics Data System (ADS)

    Meng, Chongzheng; Wang, Hui; Wu, Yuye; Liu, Jinghua; Jiang, Chengbao

    2016-10-01

    Dual-phase (Fe83Ga17)100‑xTbx alloys with 0 ≤ x ≤ 1 were synthesized by arc melting and homogenization treatment. The microstructures and the corresponding mechanical properties were systematically investigated. The chemical composition of the body centered cubic matrix is Fe83Ga17. The monoclinic second phase was composed of meltable precipitates with approximate composition Fe57Ga33Tb10. The nano-hardness of matrix and precipitates were 2.55 ± 0.17 GPa and 6.81 ± 1.03 GPa, respectively. Both the ultimate tensile strength (UTS) and fracture strain (ε) of the alloys were improved by the precipitates for x ≤ 0.2 alloys, but the strain decreases significantly at higher values of x. As potential structural-functional materials, the best mechanical properties obtained were a UTS of 595 ± 10 MPa and an ε of 3.5 ± 0.1%, four-fold and seven-fold improvements compared with the un-doped alloy. The mechanism for these anomalous changes of mechanical properties was attributed to the dispersed precipitates and semi-coherent interfaces, which serve as strong obstacles to dislocation motion and reduce the stress concentration at the grain boundaries. A sizeable improvement of magnetostriction induced by the precipitates in the range 0 ≤ x ≤ 0.2 was discovered and an optimal value of 150 ± 5 ppm is found, over three times higher than that of the un-doped alloy.

  8. Methylphenidate Enhances NMDA-Receptor Response in Medial Prefrontal Cortex via Sigma-1 Receptor: A Novel Mechanism for Methylphenidate Action

    PubMed Central

    Liu, Yue; Ji, Xiao-Hua; Peng, Ji-Yun; Zhang, Xue-Han; Zhen, Xue-Chu; Li, Bao-Ming

    2012-01-01

    Methylphenidate (MPH), commercially called Ritalin or Concerta, has been widely used as a drug for Attention Deficit Hyperactivity Disorder (ADHD). Noteworthily, growing numbers of young people using prescribed MPH improperly for pleasurable enhancement, take high risk of addiction. Thus, understanding the mechanism underlying high level of MPH action in the brain becomes an important goal nowadays. As a blocker of catecholamine transporters, its therapeutic effect is explained as being due to proper modulation of D1 and α2A receptor. Here we showed that higher dose of MPH facilitates NMDA-receptor mediated synaptic transmission via a catecholamine-independent mechanism, in layer V∼VI pyramidal cells of the rat medial prefrontal cortex (PFC). To indicate its postsynaptic action, we next found that MPH facilitates NMDA-induced current and such facilitation could be blocked by σ1 but not D1/5 and α2 receptor antagonists. And this MPH eliciting enhancement of NMDA-receptor activity involves PLC, PKC and IP3 receptor mediated intracellular Ca2+ increase, but does not require PKA and extracellular Ca2+ influx. Our additional pharmacological studies confirmed that higher dose of MPH increases locomotor activity via interacting with σ1 receptor. Together, the present study demonstrates for the first time that MPH facilitates NMDA-receptor mediated synaptic transmission via σ1 receptor, and such facilitation requires PLC/IP3/PKC signaling pathway. This novel mechanism possibly explains the underlying mechanism for MPH induced addictive potential and other psychiatric side effects. PMID:23284812

  9. Structure and mechanism of activity-based inhibition of the EGF-Receptor by Mig6

    PubMed Central

    Ficarro, Scott B.; Zhang, Yi; Lee, Byung Il; Cho, Ahye; Kim, Kihong; Park, Angela K.J.; Park, Woong-Yang; Murray, Bradley; Meyerson, Matthew; Beroukhim, Rameen; Marto, Jarrod A.; Cho, Jeonghee; Eck, Michael J.

    2016-01-01

    Mig6 is a feedback inhibitor that directly binds, inhibits and drives internalization of ErbB-family receptors. Mig6 selectivity targets activated receptors. Here we find that the EGF receptor phosphorylates Mig6 on Tyr394, and that this phosphorylation is primed by prior phosphorylation of an adjacent residue, Tyr395, by Src. Crystal structures of human EGFR–Mig6 complexes reveal the structural basis for enhanced phosphorylation of primed Mig6 and show how Mig6 rearranges after phosphorylation by EGFR to effectively irreversibly inhibit the same receptor that catalyzed its phosphorylation. This dual phosphorylation site allows Mig6 to inactivate EGFR in a manner that requires activation of the target receptor and can be modulated by Src. Loss of Mig6 is a driving event in human cancer; analysis of 1057 gliomas reveals frequent focal deletions of ERRFI, the gene that encodes Mig6, in EGFR-amplified glioblastomas. PMID:26280531

  10. Mechanical stress activates NMDA receptors in the absence of agonists.

    PubMed

    Maneshi, Mohammad Mehdi; Maki, Bruce; Gnanasambandam, Radhakrishnan; Belin, Sophie; Popescu, Gabriela K; Sachs, Frederick; Hua, Susan Z

    2017-01-03

    While studying the physiological response of primary rat astrocytes to fluid shear stress in a model of traumatic brain injury (TBI), we found that shear stress induced Ca(2+) entry. The influx was inhibited by MK-801, a specific pore blocker of N-Methyl-D-aspartic acid receptor (NMDAR) channels, and this occurred in the absence of agonists. Other NMDA open channel blockers ketamine and memantine showed a similar effect. The competitive glutamate antagonists AP5 and GluN2B-selective inhibitor ifenprodil reduced NMDA-activated currents, but had no effect on the mechanically induced Ca(2+) influx. Extracellular Mg(2+) at 2 mM did not significantly affect the shear induced Ca(2+) influx, but at 10 mM it produced significant inhibition. Patch clamp experiments showed mechanical activation of NMDAR and inhibition by MK-801. The mechanical sensitivity of NMDARs may play a role in the normal physiology of fluid flow in the glymphatic system and it has obvious relevance to TBI.

  11. Mechanical stress activates NMDA receptors in the absence of agonists

    PubMed Central

    Maneshi, Mohammad Mehdi; Maki, Bruce; Gnanasambandam, Radhakrishnan; Belin, Sophie; Popescu, Gabriela K.; Sachs, Frederick; Hua, Susan Z.

    2017-01-01

    While studying the physiological response of primary rat astrocytes to fluid shear stress in a model of traumatic brain injury (TBI), we found that shear stress induced Ca2+ entry. The influx was inhibited by MK-801, a specific pore blocker of N-Methyl-D-aspartic acid receptor (NMDAR) channels, and this occurred in the absence of agonists. Other NMDA open channel blockers ketamine and memantine showed a similar effect. The competitive glutamate antagonists AP5 and GluN2B-selective inhibitor ifenprodil reduced NMDA-activated currents, but had no effect on the mechanically induced Ca2+ influx. Extracellular Mg2+ at 2 mM did not significantly affect the shear induced Ca2+ influx, but at 10 mM it produced significant inhibition. Patch clamp experiments showed mechanical activation of NMDAR and inhibition by MK-801. The mechanical sensitivity of NMDARs may play a role in the normal physiology of fluid flow in the glymphatic system and it has obvious relevance to TBI. PMID:28045032

  12. Leaky ryanodine receptors contribute to diaphragmatic weakness during mechanical ventilation

    PubMed Central

    Matecki, Stefan; Dridi, Haikel; Jung, Boris; Saint, Nathalie; Reiken, Steven R.; Scheuermann, Valérie; Mrozek, Ségolène; Umanskaya, Alisa; Petrof, Basil J.; Jaber, Samir; Marks, Andrew R.; Lacampagne, Alain

    2016-01-01

    Ventilator-induced diaphragmatic dysfunction (VIDD) refers to the diaphragm muscle weakness that occurs following prolonged controlled mechanical ventilation (MV). The presence of VIDD impedes recovery from respiratory failure. However, the pathophysiological mechanisms accounting for VIDD are still not fully understood. Here, we show in human subjects and a mouse model of VIDD that MV is associated with rapid remodeling of the sarcoplasmic reticulum (SR) Ca2+ release channel/ryanodine receptor (RyR1) in the diaphragm. The RyR1 macromolecular complex was oxidized, S-nitrosylated, Ser-2844 phosphorylated, and depleted of the stabilizing subunit calstabin1, following MV. These posttranslational modifications of RyR1 were mediated by both oxidative stress mediated by MV and stimulation of adrenergic signaling resulting from the anesthesia. We demonstrate in the murine model that such abnormal resting SR Ca2+ leak resulted in reduced contractile function and muscle fiber atrophy for longer duration of MV. Treatment with β-adrenergic antagonists or with S107, a small molecule drug that stabilizes the RyR1–calstabin1 interaction, prevented VIDD. Diaphragmatic dysfunction is common in MV patients and is a major cause of failure to wean patients from ventilator support. This study provides the first evidence to our knowledge of RyR1 alterations as a proximal mechanism underlying VIDD (i.e., loss of function, muscle atrophy) and identifies RyR1 as a potential target for therapeutic intervention. PMID:27457930

  13. Theoretical studies of the activation mechanism of histamine H sub 2 -receptors: Dimaprit and the receptor model

    SciTech Connect

    Pardo, L.; Mazurek, A.P.; Osman, R.; Weinstein, H. )

    1989-01-01

    Ab initio quantum mechanical calculations are used to explore the interaction of dimaprit, a histamine H,-receptor agonist, with a molecular complex designed to model the specific recognition of histamine, and the H{sub 2}-receptor activation mechanism triggered by the binding of the ligand. The stabilization of several isomeric forms of the isothiourea moiety of dimaprit in the receptor model is considered, including models for the monocationic and dicationic forms that are likely to exist under physiological conditions. The energetics of proton transfer from a receptor site to the ligand are evaluated in the presence and absence of models for other sites in the receptor. The energetic contribution of ligand desolvation to the various steps in the receptor binding and activation mechanism is estimated from calculations of the enthalpy of solvation in water represented as a continuum dielectric. The results indicate that the most likely manner in which dimaprit mimics the binding of histamine to the proposed proton donor site in the H{sub 2}-receptor model requires the sulfur in the isothiourea moiety of dimaprit to act as the proton acceptor in the activation mechanism. The simulation of this mechanism reveals its feasibility and indicates that the monocation form of dimaprit, rather than the dication, is likely to be the physiologically active species.

  14. Sugar and pH dual-responsive mesoporous silica nanocontainers based on competitive binding mechanisms

    NASA Astrophysics Data System (ADS)

    Yilmaz, M. Deniz; Xue, Min; Ambrogio, Michael W.; Buyukcakir, Onur; Wu, Yilei; Frasconi, Marco; Chen, Xinqi; Nassar, Majed S.; Stoddart, J. Fraser; Zink, Jeffrey I.

    2014-12-01

    A sugar and pH dual-responsive controlled release system, which is highly specific towards molecular stimuli, has been developed based on the binding between catechol and boronic acid on a platform of mesoporous silica nanoparticles (MSNs). By grafting phenylboronic acid stalks onto the silica surface, catechol-containing β-cyclodextrins can be attached to the orifices of the MSNs' nanopores through formation of boronate esters which block access to the nanopores. These esters are stable enough to prevent cargo molecules from escaping. The boronate esters disassociate in the presence of sugars, enabling the molecule-specific controlled-release feature of this hybrid system. The rate of release has been found to be tunable by varying both the structures and the concentrations of sugars, as a result of the competitive binding nature associated with the mechanism of its operation. Acidification also induces the release of cargo molecules. Further investigations show that the presence of both a low pH and sugar molecules provides cooperative effects which together control the rate of release.A sugar and pH dual-responsive controlled release system, which is highly specific towards molecular stimuli, has been developed based on the binding between catechol and boronic acid on a platform of mesoporous silica nanoparticles (MSNs). By grafting phenylboronic acid stalks onto the silica surface, catechol-containing β-cyclodextrins can be attached to the orifices of the MSNs' nanopores through formation of boronate esters which block access to the nanopores. These esters are stable enough to prevent cargo molecules from escaping. The boronate esters disassociate in the presence of sugars, enabling the molecule-specific controlled-release feature of this hybrid system. The rate of release has been found to be tunable by varying both the structures and the concentrations of sugars, as a result of the competitive binding nature associated with the mechanism of its operation

  15. Dual signal transduction pathways activated by TSH receptors in rat primary tanycyte cultures.

    PubMed

    Bolborea, Matei; Helfer, Gisela; Ebling, Francis J P; Barrett, Perry

    2015-06-01

    Tanycytes play multiple roles in hypothalamic functions, including sensing peripheral nutrients and metabolic hormones, regulating neurosecretion and mediating seasonal cycles of reproduction and metabolic physiology. This last function reflects the expression of TSH receptors in tanycytes, which detect photoperiod-regulated changes in TSH secretion from the neighbouring pars tuberalis. The present overall aim was to determine the signal transduction pathway by which TSH signals in tanycytes. Expression of the TSH receptor in tanycytes of 10-day-old Sprague Dawley rats was observed by in situ hybridisation. Primary ependymal cell cultures prepared from 10-day-old rats were found by immunohistochemistry to express vimentin but not GFAP and by PCR to express mRNA for Dio2, Gpr50, Darpp-32 and Tsh receptors that are characteristic of tanycytes. Treatment of primary tanycyte/ependymal cultures with TSH (100  IU/l) increased cAMP as assessed by ELISA and induced a cAMP-independent increase in the phosphorylation of ERK1/2 as assessed by western blot analysis. Furthermore, TSH (100  IU/l) stimulated a 2.17-fold increase in Dio2 mRNA expression. We conclude that TSH signal transduction in cultured tanycytes signals via Gαs to increase cAMP and via an alternative G protein to increase phosphorylation of ERK1/2.

  16. A new mechanism for growth hormone receptor activation of JAK2, and implications for related cytokine receptors

    PubMed Central

    Waters, Michael J; Brooks, Andrew J; Chhabra, Yash

    2014-01-01

    The growth hormone receptor was the first cytokine receptor to be cloned and crystallized, and provides a valuable exemplar for activation of its cognate kinase, JAK2. We review progress in understanding its activation mechanism, in particular the molecular movements made by this constitutively dimerized receptor in response to ligand binding, and how these lead to a separation of JAK-binding Box1 motifs. Such a separation leads to removal of the pseudokinase inhibitory domain from the kinase domain of a partner JAK2 bound to the receptor, and vice versa, leading to apposition of the kinase domains and transactivation. This may be a general mechanism for class I cytokine receptor action. PMID:25101218

  17. Dual strain mechanisms in a lead-free morphotropic phase boundary ferroelectric.

    PubMed

    Walker, Julian; Simons, Hugh; Alikin, Denis O; Turygin, Anton P; Shur, Vladimir Y; Kholkin, Andrei L; Ursic, Hana; Bencan, Andreja; Malic, Barbara; Nagarajan, Valanoor; Rojac, Tadej

    2016-01-21

    Electromechanical properties such as d33 and strain are significantly enhanced at morphotropic phase boundaries (MPBs) between two or more different crystal structures. Many actuators, sensors and MEMS devices are therefore systems with MPBs, usually between polar phases in lead (Pb)-based ferroelectric ceramics. In the search for Pb-free alternatives, systems with MPBs between polar and non-polar phases have recently been theorized as having great promise. While such an MPB was identified in rare-earth (RE) modified bismuth ferrite (BFO) thin films, synthesis challenges have prevented its realization in ceramics. Overcoming these, we demonstrate a comparable electromechanical response to Pb-based materials at the polar-to-non-polar MPB in Sm modified BFO. This arises from 'dual' strain mechanisms: ferroelectric/ferroelastic switching and a previously unreported electric-field induced transition of an anti-polar intermediate phase. We show that intermediate phases play an important role in the macroscopic strain response, and may have potential to enhance electromechanical properties at polar-to-non-polar MPBs.

  18. Photoluminescence and doping mechanism of theranostic Eu3+/Fe3+ dual-doped hydroxyapatite nanoparticles

    PubMed Central

    Chen, Min-Hua; Yoshioka, Tomohiko; Ikoma, Toshiyuki; Hanagata, Nobutaka; Lin, Feng-Huei; Tanaka, Junzo

    2014-01-01

    Theranostic nanoparticles currently have been regarded as an emerging concept of ‘personalized medicine’ with diagnostic and therapeutic dual-functions. Eu3+ doped hydroxyapatite (HAp) has been regarded as a promising fluorescent probe for in vivo imaging applications. Additionally, substitution of Ca2+ with Fe3+ in HAp crystal may endow the capability of producing heat upon exposure to a magnetic field. Here we report a preliminary study of doping mechanism and photoluminescence of Eu3+ and Fe3+ doped HAp nanoparticles (Eu/Fe:HAp). HAp with varied concentration of Eu3+ and Fe3+ doping are presented as Eu(10 mol%):HAp, Eu(7 mol%)-Fe(3 mol%):HAp, Eu(5 mol%)-Fe(5 mol%):HAp, Eu(3 mol%)-Fe(7 mol%):HAp, and Fe(10 mol%):HAp in the study. The results showed that the HAp particles, in nano-size with rod-like morphology, were successfully doped with Eu3+ and Fe3+, and the particles can be well suspended in cell culture medium. Photoluminescence analysis revealed that particles have prominent emissions at 536 nm, 590 nm, 615 nm, 650 nm and 695 nm upon excitation at a wavelength of 397 nm. Moreover, these Eu/Fe:HAp nanoparticles belonged to B-type carbonated HAp, which has been considered an effective biodegradable and biocompatible drug/gene carrier in biological applications. PMID:27877717

  19. First experimental investigation of dual-reciprocating drilling in planetary regoliths: Proposition of penetration mechanics

    NASA Astrophysics Data System (ADS)

    Gouache, Thibault P.; Gao, Yang; Coste, Pierre; Gourinat, Yves

    2011-10-01

    The search for life in the solar system requires sub-surface exploration capabilities of extra-terrestrial bodies like the Moon and Mars. To do so different techniques are being developed: from the classical rotary drilling techniques widely used on Earth to more original techniques like ultrasonic drilling. Dual-reciprocating drilling (DRD) is a bio-mimetic drilling principle inspired by the manner wood-wasps drill into wood to lay its eggs. It was proposed as an efficient extra-terrestrial drilling technique requiring low over-head force. To deepen the understanding of this novel drilling technique, DRD has been tested for the first time in planetary regolith simulants. These experiments are reported here. To do so a new test bench was built and is presented. The soil forces on the drill bit are analysed and the final depth reached by the DRD system is compared to the final depth reached by static penetration. The experiments have shown very high levels of slippage (defined here specifically for DRD). The observations of the surface deformations and the importance of slippage lead to the proposal of DRD penetration mechanics in regoliths. Finally a re-evaluation of previous DRD experiments conducted on low compressive strength rocks also show the high levels of slippage during DRD.

  20. Modeling of dual emission laser induced fluorescence for slurry thickness measurements in chemical mechanical polishing

    NASA Astrophysics Data System (ADS)

    Gray, Caprice; Rogers, Chris B.; Manno, Vincent P.; White, Robert D.

    2011-07-01

    Dual emission laser induced fluorescence (DELIF) is a technique for measuring the instantaneous thin fluid film thickness in dynamic systems. Two fluorophores within the system produce laser induced emissions that are filtered and captured by two cameras. The ratio of the images from these cameras is used to cancel the effect of the laser beam profile on the image intensity. The resultant intensity ratio can be calibrated to a fluid film thickness. The utilization of a 2-dye system when applied to Chemical Mechanical Polishing (CMP) is complicated by the fluorescence of the polymeric polishing pad and the light scattering particles in the polishing slurry. We have developed a model of DELIF for CMP with 1-dye employing the polishing pad as the second fluorophore. While scattering particles in the slurry decrease the overall intensity of the individual images, the contrast in the image ratio increases. Using the 1-dye DELIF system to measure thin slurry films, our model results indicate that a cubic calibration may be needed. However, experimental results suggest a linear calibration is achieved for slurry films between 0 and 133 μm thick with scattering coefficients as high as 8.66 mm-1 at a wavelength equal to 410 nm.

  1. Crystal structures and enzyme mechanisms of a dual fucose mutarotase/ribose pyranase.

    PubMed

    Lee, Kwang-Hoon; Ryu, Kyoung-Seok; Kim, Min-Sung; Suh, Hye-Young; Ku, Bonsu; Song, Young-Lan; Ko, Sunggeon; Lee, Weontae; Oh, Byung-Ha

    2009-08-07

    Escherichia coli FucU (Fucose Unknown) is a dual fucose mutarotase and ribose pyranase, which shares 44% sequence identity with its human counterpart. Herein, we report the structures of E. coli FucU and mouse FucU bound to L-fucose and delineate the catalytic mechanisms underlying the interconversion between stereoisomers of fucose and ribose. E. coli FucU forms a decameric toroid with each active site formed by two adjacent subunits. While one subunit provides most of the fucose-interacting residues including a catalytic tyrosine residue, the other subunit provides a catalytic His-Asp dyad. This active-site feature is critical not only for the mutarotase activity toward L-fucose but also for the pyranase activity toward D-ribose. Structural and biochemical analyses pointed that mouse FucU assembles into four different oligomeric forms, among which the smallest homodimeric form is most abundant and would be the predominant species under physiological conditions. This homodimer has two fucose-binding sites that are devoid of the His-Asp dyad and catalytically inactive, indicating that the mutarotase and the pyranase activities appear dispensable in vertebrates. The defective assembly of the mouse FucU homodimer into the decameric form is due to an insertion of two residues at the N-terminal extreme, which is a common aspect of all the known vertebrate FucU proteins. Therefore, vertebrate FucU appears to serve for as yet unknown function through the quaternary structural alteration.

  2. Dual Mechanisms of Ion Uptake in Relation to Vacuolation in Corn Roots

    PubMed Central

    Torii, Kenji; Laties, George G.

    1966-01-01

    Absorption isotherms for chloride and rubidium ions have been determined through a wide concentration range for nonvacuolate root tips, and for vacuolate subapical sections of corn root. In the range 0 to 0.5 mm, chloride absorption is hyperbolic with concentration in both tips and proximal sections. At high concentrations, 1 to 50 mm, a second multiple-hyperbolic isotherm for chloride is noted in vacuolate tissue, while the isotherm for nonvacuolate tips rises exponentially. A linear to exponentially rising isotherm is taken to signify diffusive permeation. The same distinction between tip and subapical tissue characterizes Rb absorption. Rb uptake is indifferent to the nature of the counterion at all concentrations in the tip, while the counterion exerts a predictable influence on Rb absorption in proximal tissue. The effect of a poorly absorbable anion on Rb uptake is greater in the high concentration range. Evidence is presented for the metabolic nature of ion transport into nonvacuolate root tips. Verification is offered that ion uptake is mediated by dual mechanisms, and the thesis is developed that the high-affinity (low Ks) system mediates ion passage through the plasma membrane while the low-affinity (high Ks) system implements transport through the tonoplast. PMID:16656332

  3. Dual mechanisms of ion uptake in relation to vacuolation in corn roots.

    PubMed

    Torii, K; Laties, G G

    1966-05-01

    Absorption isotherms for chloride and rubidium ions have been determined through a wide concentration range for nonvacuolate root tips, and for vacuolate subapical sections of corn root. In the range 0 to 0.5 mm, chloride absorption is hyperbolic with concentration in both tips and proximal sections. At high concentrations, 1 to 50 mm, a second multiple-hyperbolic isotherm for chloride is noted in vacuolate tissue, while the isotherm for nonvacuolate tips rises exponentially. A linear to exponentially rising isotherm is taken to signify diffusive permeation.The same distinction between tip and subapical tissue characterizes Rb absorption. Rb uptake is indifferent to the nature of the counterion at all concentrations in the tip, while the counterion exerts a predictable influence on Rb absorption in proximal tissue. The effect of a poorly absorbable anion on Rb uptake is greater in the high concentration range. Evidence is presented for the metabolic nature of ion transport into nonvacuolate root tips. Verification is offered that ion uptake is mediated by dual mechanisms, and the thesis is developed that the high-affinity (low K(s)) system mediates ion passage through the plasma membrane while the low-affinity (high K(s)) system implements transport through the tonoplast.

  4. Dual and tetraelectrode QCMs using imprinted polymers as receptors for ions and neutral analytes.

    PubMed

    Latif, Usman; Mujahid, Adnan; Afzal, Adeel; Sikorski, Renatus; Lieberzeit, Peter A; Dickert, Franz L

    2011-06-01

    Polymers as coating materials were combined with quartz crystal microbalances (QCMs) to design sensor devices for the detection of both ionic and neutral analytes in liquid phase. The design and geometry of dual and tetraelectrode QCMs have been optimized to reduce electric field interferences. An unusual Sauerbrey effect was observed while exposing potassium salt solution to 10- and 20-MHz QCMs, i.e. increase in the frequency shifts by a factor of seven, which is attributed to electro-acoustic phenomena. Non-functionalized sol-gel materials were synthesized by templating with hydrophobic salt such as tetraethyl ammonium picrate. Imprinting with these ions of low charge density leads to sensitive layers, and UV-Vis spectroscopy was used to check re-inclusion of this analyte. In the next strategy, functionalized polyurethane for potassium ions and sol-gel materials with aminopropyl group as ligand were generated to tune selectivity and sensitivity towards Ni(2+) and Cu(2+). Methacrylic acid polymers were optimized for the detection of atrazine by hydrogen bonding; double molecular imprinted polyurethane approach was followed for pyrene recognition. Finally, these imprinted polymers were combined with tetraelectrode QCM to develop sensor platform.

  5. Dual Targeting of the Chemokine Receptors CXCR4 and ACKR3 with Novel Engineered Chemokines*

    PubMed Central

    Hanes, Melinda S.; Salanga, Catherina L.; Chowdry, Arnab B.; Comerford, Iain; McColl, Shaun R.; Kufareva, Irina; Handel, Tracy M.

    2015-01-01

    The chemokine CXCL12 and its G protein-coupled receptors CXCR4 and ACKR3 are implicated in cancer and inflammatory and autoimmune disorders and are targets of numerous antagonist discovery efforts. Here, we describe a series of novel, high affinity CXCL12-based modulators of CXCR4 and ACKR3 generated by selection of N-terminal CXCL12 phage libraries on live cells expressing the receptors. Twelve of 13 characterized CXCL12 variants are full CXCR4 antagonists, and four have Kd values <5 nm. The new variants also showed high affinity for ACKR3. The variant with the highest affinity for CXCR4, LGGG-CXCL12, showed efficacy in a murine model for multiple sclerosis, demonstrating translational potential. Molecular modeling was used to elucidate the structural basis of binding and antagonism of selected variants and to guide future designs. Together, this work represents an important step toward the development of therapeutics targeting CXCR4 and ACKR3. PMID:26216880

  6. Discovery of dual positive allosteric modulators (PAMs) of the metabotropic glutamate 2 receptor and CysLT1 antagonists for treating migraine headache.

    PubMed

    Blanco, Maria-Jesus; Benesh, Dana R; Knobelsdorf, James A; Khilevich, Albert; Cortez, Guillermo S; Mokube, Fese; Aicher, Thomas D; Groendyke, Todd M; Marmsater, Fredrik P; Tang, Tony P; Johnson, Kirk W; Clemens-Smith, Amy; Muhlhauser, Mark A; Swanson, Steven; Catlow, John; Emkey, Renee; Johnson, Michael P; Schkeryantz, Jeffrey M

    2017-01-15

    Pyridylmethylsulfonamide series were the first reported example of positive allosteric modulators (PAM) of the mGlu2 receptor. The hydroxyacetophenone scaffold is a second series of mGlu2 PAMs we have identified. This series of molecules are potent mGlu2 potentiators and possess significant CysLT1 (cysteinyl leukotriene receptor 1) antagonist activity, showing in vivo efficacy in a dural plasma protein extravasation (PPE) model of migraine. In this paper, we describe the dual SAR, pharmacokinetics and preclinical in vivo efficacy data for a tetrazole containing hydroxyacetophenone scaffold.

  7. Pharmacokinetics and preliminary safety data of a single oral dose of bosentan, a dual endothelin receptor antagonist, in cats.

    PubMed

    Puza, N; Papich, M G; Reinero, C; Chang, C H; Yu, D-H; Sharp, C; DeClue, A

    2014-04-01

    The objective of this study was to evaluate the pharmacokinetic properties and adverse effect profile of single-dose oral bosentan, a dual endothelin receptor antagonist, in healthy cats. Pharmacokinetic parameters were determined following a single mean ± SD oral dose of 3.2 ± 0.6 mg/kg of bosentan in 6 adult cats. Blood was collected for quantification of bosentan via high-performance liquid chromatography with ultraviolet detection. Blood and urine were evaluated for CBC, plasma biochemical profile, and urinalysis, and repeat physical examinations were performed to evaluate for adverse effects. The mean terminal half-life of bosentan was 20.4 ± 17.2 h. The mean peak plasma concentration was 0.49 ± 0.24 g/mL, and the mean time to maximum plasma concentration was 6.8 ± 8.6 h. The area under the curve was 5.14 ± 3.81 h·μg/mL. Oral bosentan tablets were absorbed in cats, and no clinically important adverse events were noted. Further evaluation of repeat dosing, investigation into the in vivo efficacy of decreasing endothelin-1 concentrations in cats, as well as safety in conjunction with other medications is warranted.

  8. A novel peroxisome proliferator-activated receptor alpha/gamma dual agonist demonstrates favorable effects on lipid homeostasis.

    PubMed

    Guo, Qiu; Sahoo, Soumya P; Wang, Pei-Ran; Milot, Denise P; Ippolito, Marc C; Wu, Margaret S; Baffic, Joanne; Biswas, Chhabi; Hernandez, Melba; Lam, My-Hanh; Sharma, Neelam; Han, Wei; Kelly, Linda J; MacNaul, Karen L; Zhou, Gaochao; Desai, Ranjit; Heck, James V; Doebber, Thomas W; Berger, Joel P; Moller, David E; Sparrow, Carl P; Chao, Yu-Sheng; Wright, Samuel D

    2004-04-01

    Patients with type 2 diabetes mellitus exhibit hyperglycemia and dyslipidemia as well as a markedly increased incidence of atherosclerotic cardiovascular disease. Here we report the characterization of a novel arylthiazolidinedione capable of lowering both glucose and lipid levels in animal models. This compound, designated TZD18, is a potent agonist with dual human peroxisome proliferator-activated receptor (PPAR)-alpha/gamma activities. In keeping with its PPARgamma activity, TZD18 caused complete normalization of the elevated glucose in db/db mice and Zucker diabetic fatty rats. TZD18 lowered both cholesterol and triglycerides in hamsters and dogs. TZD18 inhibited cholesterol biosynthesis at steps before mevalonate and reduced hepatic levels of 3-hydroxy-3-methylglutaryl coenzyme A reductase activity. Moreover, TZD18 significantly suppressed gene expression of fatty acid synthesis and induced expression of genes for fatty acid degradation and triglyceride clearance. Studies on 17 additional PPARalpha or PPARalpha/gamma agonists showed that lipid lowering in hamsters correlated with the magnitude of hepatic gene expression changes. Importantly, the presence of PPARgamma agonism did not affect the relationship between hepatic gene expression and lipid lowering. Taken together, these data suggest that PPARalpha/gamma agonists, such as TZD18, affect lipid homeostasis, leading to an antiatherogenic plasma lipid profile. Agents with these properties may provide favorable means for treatment of type 2 diabetes and dyslipidemia and the prevention of atherosclerotic cardiovascular disease.

  9. Neural mechanisms of dual-task interference and cognitive capacity limitation in the prefrontal cortex.

    PubMed

    Watanabe, Kei; Funahashi, Shintaro

    2014-04-01

    Simultaneous performance of two tasks often leads to performance deficits in the component tasks. This effect, known as dual-task interference, is thought to be a proof of capacity limitation in cognition, and the lateral prefrontal cortex (LPFC) has been highlighted as its putative neural substrate. Here we recorded single-neuron activities in LPFC while monkeys performed dual tasks that required the simultaneous performance of a varying-load spatial attention task and a spatial memory task. We found that the performance of the monkeys exhibited dual-task interference, and prefrontal neuron activities showed a decreased ability to represent task-relevant information to a degree proportional to the increased demand of the concurrent counterpart task. The locus of the interference was shown to originate in the simultaneous, overloaded recruitment of the same LPFC neural population by the two tasks. These results provide direct neurophysiological evidence for, and constraints to, psychological models of dual-task interference and capacity limitation.

  10. Dual mechanism of vascular endothelial growth factor upregulation by hypoxia in human hepatocellular carcinoma

    PubMed Central

    von Marschall, Z; Cramer, T; Hocker, M; Finkenzeller, G; Wiedenmann, B; Rosewicz, S

    2001-01-01

    BACKGROUND/AIMS—Vascular endothelial growth factor (VEGF) plays a key role in regulation of tumour associated angiogenesis. In the current study we analysed expression of VEGF and its receptors in human hepatocellular carcinoma (HCC) and investigated the molecular mechanisms of VEGF regulation by hypoxia.
METHODS—VEGF, kinase domain region (KDR)/fetal liver kinase 1 (flk-1), and flt-1 expression were examined by immunohistochemistry and in situ hybridisation in 15 human HCC tissues. Expression of VEGF and regulation by hypoxia were assessed in three human HCC cell lines using a quantitative competitive reverse transcription-polymerase chain reaction, ELISA, and a series of 5' deletion reporter gene constructs of the human VEGF promoter in transient transfection assays.
RESULTS—We observed over expression of VEGF mRNA and protein in HCC compared with cirrhosis or normal liver. Expression of VEGF in tumour cells was strongly increased in areas directly adjacent to necrotic/hypoxic regions. Both VEGF receptors were detected in vascular endothelia of HCC while only KDR/flk-1 receptors were detected in endothelial cells of cirrhotic livers. Expression of VEGF was observed in all human HCC cell lines examined. Hypoxia (1% oxygen) resulted in profound upregulation of VEGF mRNA and protein levels. Furthermore, hypoxia treatment resulted in a doubling of VEGF mRNA stability. Deletion analysis of the human VEGF 5' flanking region −2018 and +50 demonstrated induction of VEGF promoter activity under hypoxic conditions which was significantly decreased following deletion of the region −1286 and −789 suggesting a substantial contribution of the −975 putative hypoxia inducible factor 1 binding site to hypoxia mediated transcriptional activation of the VEGF gene.
CONCLUSION—These data suggest hypoxia as a central stimulus of angiogenesis in human HCC through upregulation of VEGF gene expression by at least two distinct molecular mechanisms: activation of

  11. Lipid-based nanoformulation of irinotecan: dual mechanism of action allows for combination chemo/angiogenic therapy.

    PubMed

    Waterhouse, Dawn N; Yapp, Donald; Verreault, Maite; Anantha, Malathi; Sutherland, Brent; Bally, Marcel B

    2011-11-01

    A number of studies have outlined the antiangiogenic effects of cytotoxic agents when administered frequently at low doses. These studies suggest that the effect of the cytotoxic agent is on the vasculature within the tumor and it is assumed that there is little or negligible cytotoxicity. Liposomal drug delivery systems have the ability to provide a dual mechanism of activity where tumor accumulation can deliver high local concentrations of the drug at the site of action with concomitant slow release of the drug from carriers in the blood compartment that results in antivascular effects, similar to that achieved when dosing frequently at low levels. Although this dual mechanism of activity may be linked to other lipid nanoparticle formulations of anticancer drugs, this article summarizes the evidence supporting direct (cytotoxic) and indirect (antivascular) actions of a liposomal formulation of irinotecan.

  12. Synthesis and dual histamine H₁ and H₂ receptor antagonist activity of cyanoguanidine derivatives.

    PubMed

    Sadek, Bassem; Alisch, Rudi; Buschauer, Armin; Elz, Sigurd

    2013-11-15

    Premedication with a combination of histamine H₁ receptor (H₁R) and H₂ receptor (H₂R) antagonists has been suggested as a prophylactic principle, for instance, in anaesthesia and surgery. Aiming at pharmacological hybrids combining H₁R and H₂R antagonistic activity, a series of cyanoguanidines 14-35 was synthesized by linking mepyramine-type H₁R antagonist substructures with roxatidine-, tiotidine-, or ranitidine-type H₂R antagonist moieties. N-desmethylmepyramine was connected via a poly-methylene spacer to a cyanoguanidine group as the "urea equivalent" of the H₂R antagonist moiety. The title compounds were screened for histamine antagonistic activity at the isolated ileum (H₁R) and the isolated spontaneously beating right atrium (H₂R) of the guinea pig. The results indicate that, depending on the nature of the H₂R antagonist partial structure, the highest H₁R antagonist potency resided in roxatidine-type compounds with spacers of six methylene groups in length (compound 21), and tiotidine-type compounds irrespective of the alkyl chain length (compounds 28, 32, 33), N-cyano-N'-[2-[[(2-guanidino-4-thiazolyl)methyl]thio]ethyl]-N″-[2-[N-[2-[N-(4-methoxybenzyl)-N-(pyridyl)-amino] ethyl]-N-methylamino]ethyl] guanidine (25, pKB values: 8.05 (H₁R, ileum) and 7.73 (H₂R, atrium) and the homologue with the mepyramine moiety connected by a six-membered chain to the tiotidine-like partial structure (compound 32, pKB values: 8.61 (H₁R) and 6.61 (H₂R) were among the most potent hybrid compounds. With respect to the development of a potential pharmacotherapeutic agent, structural optimization seems possible through selection of other H₁R and H₂R pharmacophoric moieties with mutually affinity-enhancing properties.

  13. Design and Investigation of a [(18)F]-Labeled Benzamide Derivative as a High Affinity Dual Sigma Receptor Subtype Radioligand for Prostate Tumor Imaging.

    PubMed

    Yang, Dongzhi; Comeau, Anthony; Bowen, Wayne D; Mach, Robert H; Ross, Brian D; Hong, Hao; Van Dort, Marcian E

    2017-03-06

    High overexpression of sigma (σ) receptors (σ1 and σ2 subtypes) in a variety of human solid tumors has prompted the development of σ receptor-targeting radioligands, as imaging agents for tumor detection. A majority of these radioligands to date target the σ2 receptor, a potential marker of tumor proliferative status. The identification of approximately equal proportions of both σ receptor subtypes in prostate tumors suggests that a high affinity, dual σ receptor-targeting radioligand could potentially provide enhanced tumor targeting efficacy in prostate cancer. To accomplish this goal, we designed a series of ligands which bind to both σ receptor subtypes with high affinity. Ligand 3a in this series, displaying optimal dual σ receptor subtype affinity (σ1, 6.3 nM; σ2, 10.2 nM) was radiolabeled with fluorine-18 ((18)F) to give [(18)F]3a and evaluated as a σ receptor-targeting radioligand in the mouse PC-3 prostate tumor model. Cellular assays with PC-3 cells demonstrated that a major proportion of [(18)F]3a was localized to cell surface σ receptors, while ∼10% of [(18)F]3a was internalized within cells after incubation for 3.5 h. Serial PET imaging in mice bearing PC-3 tumors revealed that uptake of [(18)F]3a was 1.6 ± 0.8, 4.4 ± 0.3, and 3.6 ± 0.6% ID/g (% injection dose per gram) in σ receptor-positive prostate tumors at 15 min, 1.5 h, and 3.5 h postinjection, respectively (n = 3) resulting in clear tumor visualization. Blocking studies conducted with haloperidol (a nonselective inhibitor for both σ receptor subtypes) confirmed that the uptake of [(18)F]3a was σ receptor-mediated. Histology analysis confirmed similar expression of σ1 and σ2 in PC-3 tumors which was significantly greater than its expression in normal organs/tissues such as liver, kidney, and muscle. Metabolite studies revealed that >50% of radioactivity in PC-3 tumors at 30 min postinjection represented intact [(18)F]3a. Prominent σ receptor-specific uptake of [(18)F]3a in

  14. 5-HT7 receptor activation inhibits mechanical hypersensitivity secondary to capsaicin sensitization in mice.

    PubMed

    Brenchat, Alex; Romero, Luz; García, Mónica; Pujol, Marta; Burgueño, Javier; Torrens, Antoni; Hamon, Michel; Baeyens, José Manuel; Buschmann, Helmut; Zamanillo, Daniel; Vela, José Miguel

    2009-02-01

    This work aimed to evaluate the potential role of the 5-HT(7) receptor in nociception secondary to a sensitizing stimulus in mice. For this purpose, the effects of relevant ligands (5-HT(7) receptor agonists: AS-19, MSD-5a, E-55888; 5-HT(7) receptor antagonists: SB-258719, SB-269970; 5-HT(1A) receptor agonist: F-13640; 5-HT(1A) receptor antagonist: WAY-100635) were assessed on capsaicin-induced mechanical hypersensitivity, a pain behavior involving hypersensitivity of dorsal horn neurons (central sensitization). For the 5-HT(7) receptor agonists used, binding profile and intrinsic efficacy to stimulate cAMP formation in HEK-293F cells expressing the human 5-HT(7) receptor were also evaluated. AS-19 and E-55888 were selective for 5-HT(7) receptors. E-55888 was a full agonist whereas AS-19 and MSD-5a behaved as partial agonists, with maximal effects corresponding to 77% and 61%, respectively, of the cAMP response evoked by the full agonist 5-HT. Our in vivo results revealed that systemic administration of 5-HT(7) receptor agonists exerted a clear-cut dose-dependent antinociceptive effect that was prevented by 5-HT(7) receptor antagonists, but not by the 5-HT(1A) receptor antagonist. The order of efficacy (E-55888>AS-19>MSD-5a) matched their in vitro efficacy as 5-HT(7) receptor agonists. Contrary to agonists, a dose-dependent promotion of mechanical hypersensitivity was observed after administration of 5-HT(7) receptor antagonists, substantiating the involvement of the 5-HT(7) receptor in the control of capsaicin-induced mechanical hypersensitivity. These findings suggest that serotonin exerts an inhibitory role in the control of nociception through activation of 5-HT(7) receptors, and point to a new potential therapeutic use of 5-HT(7) receptor agonists in the field of analgesia.

  15. Intrarenal dopamine D1-like receptor stimulation induces natriuresis via an angiotensin type-2 receptor mechanism.

    PubMed

    Salomone, Leslie J; Howell, Nancy L; McGrath, Helen E; Kemp, Brandon A; Keller, Susanna R; Gildea, John J; Felder, Robin A; Carey, Robert M

    2007-01-01

    We explored the effects of direct renal interstitial stimulation of dopamine D(1)-like receptors with fenoldopam, a selective D(1)-like receptor agonist, on renal sodium excretion and angiotensin type-2 (AT(2)) receptor expression and cellular distribution in rats on a high-sodium intake. In contrast to vehicle-infused rats, sodium excretion increased in fenoldopam-infused rats during each of three 1-hour experimental periods (<0.001). Blood pressure was unaffected by vehicle or fenoldopam. In plasma membranes of renal cortical cells, fenoldopam increased D(1) receptor expression by 38% (P<0.05) and AT(2) receptor expression by 69% (P<0.01). In plasma membranes of renal proximal tubule cells, fenoldopam increased AT(2) receptor expression by 108% (P<0.01). In outer apical membranes of proximal tubule cells, fenoldopam increased AT(2) receptor expression by 59% (P<0.01). No significant change in total AT(2) receptor protein expression was detectable in response to fenoldopam. Fenoldopam-induced natriuresis was abolished when either PD-123319, a specific AT(2) receptor antagonist, or SCH-23390, a potent D(1)-like receptor antagonist, was coinfused with F (P<0.001). In summary, direct renal D(1)-like receptor activation increased urinary sodium excretion and the plasma membrane expression of AT(2) receptors in renal cortical and proximal tubule cells. D(1)-like receptor-induced natriuresis was abolished by intrarenal AT(2) receptor inhibition. These findings suggest that dopaminergic regulation of sodium excretion involves recruitment of AT(2) receptors to the outer plasma membranes of renal proximal tubule cells and that dopamine-induced natriuresis requires AT(2) receptor activation.

  16. Mechanical Stimulation of Piezo1 Receptors Depends on Extracellular Matrix Proteins and Directionality of Force.

    PubMed

    Gaub, Benjamin M; Müller, Daniel J

    2017-02-08

    Piezo receptors convert mechanical forces into electrical signals. In mammals, they play important roles in basic physiological functions including proprioception, sensation of touch, and vascular development. However, basic receptor properties like the gating mechanism, the interaction with extracellular matrix (ECM) proteins, and the response to mechanical stimulation, remain poorly understood. Here, we establish an atomic force microscopy (AFM)-based assay to mechanically stimulate Piezo1 receptors in living animal cells, while monitoring receptor activation in real-time using functional calcium imaging. Our experiments show that in the absence of ECM proteins Piezo1 receptors are relatively insensitive to mechanical forces pushing the cellular membrane, whereas they can hardly be activated by mechanically pulling the membrane. Yet, if conjugated with Matrigel, a mix of ECM proteins, the receptors become sensitized. Thereby, forces pulling the cellular membrane activate the receptor much more efficiently compared to pushing forces. Finally, we found that collagen IV, a component of the basal lamina, which forms a cohesive network and mechanical connection between cells, sensitizes Piezo1 receptors to mechanical pulling.

  17. Architecture and conformational switch mechanism of the ryanodine receptor.

    PubMed

    Efremov, Rouslan G; Leitner, Alexander; Aebersold, Ruedi; Raunser, Stefan

    2015-01-01

    Muscle contraction is initiated by the release of calcium (Ca(2+)) from the sarcoplasmic reticulum into the cytoplasm of myocytes through ryanodine receptors (RyRs). RyRs are homotetrameric channels with a molecular mass of more than 2.2 megadaltons that are regulated by several factors, including ions, small molecules and proteins. Numerous mutations in RyRs have been associated with human diseases. The molecular mechanism underlying the complex regulation of RyRs is poorly understood. Using electron cryomicroscopy, here we determine the architecture of rabbit RyR1 at a resolution of 6.1 Å. We show that the cytoplasmic moiety of RyR1 contains two large α-solenoid domains and several smaller domains, with folds suggestive of participation in protein-protein interactions. The transmembrane domain represents a chimaera of voltage-gated sodium and pH-activated ion channels. We identify the calcium-binding EF-hand domain and show that it functions as a conformational switch allosterically gating the channel.

  18. Dual role of serotonin in the pathogenesis of indomethacin-induced small intestinal ulceration: pro-ulcerogenic action via 5-HT3 receptors and anti-ulcerogenic action via 5-HT4 receptors.

    PubMed

    Kato, Shinichi; Matsuda, Narumi; Matsumoto, Kenjiro; Wada, Mai; Onimaru, Naoki; Yasuda, Masashi; Amagase, Kikuko; Horie, Syunji; Takeuchi, Koji

    2012-09-01

    Serotonin (5-HT) exerts multiple physiological functions not only in the central and peripheral nervous systems but also in the gastrointestinal tract, and these multiple functions are accounted for by a variety of 5-HT receptor subtypes. We investigated the role of 5-HT in the pathogenesis of indomethacin-induced intestinal lesions in mice, in relation to 5-HT receptor subtypes. A single oral administration of indomethacin (10 mg/kg) provoked damage in the small intestine of mice 24 h later, and this response was prevented by pretreatment with p-chlorophenylalanine (a 5-HT synthesis inhibitor). The administration of 5-HT3 receptor antagonists, such as ondansetron and ramosetron, dose-dependently reduced the severity of the intestinal lesions, whereas a high dose of GR113808 (a 5-HT4 receptor antagonist) significantly aggravated these lesions. In contrast, NAN-190 (a 5-HT1 receptor antagonist), ketanserin (a 5-HT2 receptor antagonist), and SB269970 (a 5-HT7 receptor antagonist) had no effect on these lesions. Mosapride (a 5-HT4 receptor agonist) significantly reduced the severity of indomethacin-induced intestinal lesions, and this protective effect was totally prevented by either GR113808 or methyllycaconitine (an α7-nicotinic acetylcholine receptor antagonist). Indomethacin increased the activity of myeloperoxidase and the expression of inducible nitric oxide synthase, inflammatory cytokines, and chemokines in the small intestine; these responses were significantly attenuated by ondansetron and mosapride. These findings suggest that endogenous 5-HT exerts a dual role in the pathogenesis of indomethacin-induced intestinal lesions: pro-ulcerogenic action via 5-HT3 receptors and anti-ulcerogenic action via 5-HT4 receptors, and the latter effect via 5-HT4 receptors may be mediated by activation of α7-nicotinic acetylcholine receptors.

  19. Dual pH-responsive and CD44 receptor targeted multifunctional nanoparticles for anticancer intracellular delivery

    NASA Astrophysics Data System (ADS)

    Chen, Daquan; Sun, Jingfang; Lian, Shengnan; Liu, Zongliang; Sun, Kaoxiang; Liu, Wanhui; Wu, Zimei; Zhang, Qiang

    2014-11-01

    In this article, we prepared a multifunctional oligosaccharides of hyaluronan (oHA) conjugates, oHA-histidine-menthone 1,2-glycerol ketal (oHM). The oHM conjugates possess pH-sensitive menthone 1,2-glycerol ketal (MGK) as hydrophobic moieties and oHA as the target of CD44 receptor. The polymeric mPEG-Chitosan-Ketal (PCK) carrying pH-sensitive ketal group as hydrophobic moieties and PEG group as hydrophilic moieties were synthesized. The two pH-sensitive ketal derivatives were employed to fabricate nanoparticles for anti-tumor drug delivery. The oHM-PCK nanoparticles (oHPN) can spontaneously self-assemble into mixed micellar structure with nano-sized spherical shape of 100-200 nm at pH 7.4 PBS conditions. The oHPN could release encapsulated curcumin with 92.6 % at pH 5.0 compared with 55.3 % at pH 7.4. The results of cytotoxicity assay indicated that encapsulated curcumin in oHPN (Cur-oHPN) have less toxicity compared to curcumin suspension. The anti-tumor efficacy in vivo suggested that Cur-oHPN suppressed tumor growth most efficiently. These results present the promising potential of oHPN as an effective nano-sized pH-sensitive drug delivery system for intracellular delivery.

  20. Species dependent dual modulation of the benzodiazepine/GABA receptor chloride channel by dihydroergosine

    SciTech Connect

    Pericic, D.; Tvrdeic, A. )

    1990-01-01

    Dihydroergosine enhanced the incidence of bicuculline induced convulsions in female rats, while 100 mg/kg of dihydroergosine given to female mice made 45% convulsive dose of bicuculline to be subconvulsive. The same dose of dihydroergosine enhanced in mice the latency of bicuculline-induced convulsions. Although, in in vitro experiments dihydroergosine showed very weak ability to prevent the binding of {sup 3}H-muscimol, the drug was able to diminish and to augment the IC{sub 50} of bicuculline and GABA when added to crude synaptosomal pellet of the rat and mouse brain respectively. Lower concentrations of dihydroergosine stimulated and higher inhibited {sup 3}H-TBOB binding to the crude synaptosomal pellet of the rat brain. In the preparation of mouse brain dihydroergosine produced only inhibition of {sup 3}H-TBOB binding. Only slight quantitative differences were observed in bicuculline-induced stimulation and in GABA- and diazepam-induced inhibition of {sup 3}H-TBOB binding between the two species. The results suggest that the opposite species-dependent effects of dihydroergosine on bicuculline-induced convulsions are due to the ability of this drug to modulate species-dependently the benzodiazepine/GABA receptor chloride channel complex.

  1. Adenosine A2A receptor antagonism and neuroprotection: mechanisms, lights, and shadows.

    PubMed

    Popoli, Patrizia; Minghetti, Luisa; Tebano, Maria Teresa; Pintor, Annita; Domenici, Maria Rosaria; Massotti, Marino

    2004-01-01

    Adenosine A2A receptor antagonists are regarded as potential neuroprotective drugs, although the mechanisms underlying their effects remain to be elucidated. In this review, quinolinic acid (QA)-induced striatal toxicity was used as a tool to investigate the mechanisms of the neuroprotective effects of A2A receptor antagonists. After having examined the effects of selective A2A receptor antagonists toward different mechanisms of QA toxicity, we conclude that (1) the effect elicited by A2A receptor blockade on QA-induced glutamate outflow may be one of the mechanisms of the neuroprotective activity of A2A receptor antagonists; (2) A2A receptor antagonists have a potentially worsening influence on QA-dependent NMDA receptor activation; and (3) the ability of A2A receptor antagonists to prevent QA-induced lipid peroxidation does not correlate with the neuroprotective effects. These results suggest that A2A receptor antagonists may have either potentially beneficial or detrimental influence in models of neurodegeneration that are mainly due to increased glutamate levels or enhanced sensitivity of NMDA receptors, respectively.

  2. Pharmacological activation of 5-HT7 receptors reduces nerve injury-induced mechanical and thermal hypersensitivity.

    PubMed

    Brenchat, Alex; Nadal, Xavier; Romero, Luz; Ovalle, Sergio; Muro, Asunción; Sánchez-Arroyos, Ricard; Portillo-Salido, Enrique; Pujol, Marta; Montero, Ana; Codony, Xavier; Burgueño, Javier; Zamanillo, Daniel; Hamon, Michel; Maldonado, Rafael; Vela, José Miguel

    2010-06-01

    The involvement of the 5-HT(7) receptor in nociception and pain, particularly chronic pain (i.e., neuropathic pain), has been poorly investigated. In the present study, we examined whether the 5-HT(7) receptor participates in some modulatory control of nerve injury-evoked mechanical hypersensitivity and thermal (heat) hyperalgesia in mice. Activation of 5-HT(7) receptors by systemic administration of the selective 5-HT(7) receptor agonist AS-19 (1 and 10mg/kg) exerted a clear-cut reduction of mechanical and thermal hypersensitivities that were reversed by co-administering the selective 5-HT(7) receptor antagonist SB-258719. Interestingly, blocking of 5-HT(7) receptors with SB-258719 (2.5 and 10mg/kg) enhanced mechanical (but not thermal) hypersensitivity in nerve-injured mice and induced mechanical hypersensitivity in sham-operated mice. Effectiveness of the treatment with a 5-HT(7) receptor agonist was maintained after repeated systemic administration: no tolerance to the antiallodynic and antihyperalgesic effects was developed following treatment with the selective 5-HT(7) receptor agonist E-57431 (10mg/kg) twice daily for 11 days. The 5-HT(7) receptor co-localized with GABAergic cells in the dorsal horn of the spinal cord, suggesting that the activation of spinal inhibitory GABAergic interneurons could contribute to the analgesic effects of 5-HT(7) receptor agonists. In addition, a significant increase of 5-HT(7) receptors was found by immunohistochemistry in the ipsilateral dorsal horn of the spinal cord after nerve injury, suggesting a "pain"-triggered regulation of receptor expression. These results support the idea that the 5-HT(7) receptor subtype is involved in the control of pain and point to a new potential use of 5-HT(7) receptor agonists for the treatment of neuropathic pain.

  3. A new method for ligand docking to flexible receptors by dual alanine scanning and refinement (SCARE)

    NASA Astrophysics Data System (ADS)

    Bottegoni, Giovanni; Kufareva, Irina; Totrov, Maxim; Abagyan, Ruben

    2008-05-01

    Protein binding sites undergo ligand specific conformational changes upon ligand binding. However, most docking protocols rely on a fixed conformation of the receptor, or on the prior knowledge of multiple conformations representing the variation of the pocket, or on a known bounding box for the ligand. Here we described a general induced fit docking protocol that requires only one initial pocket conformation and identifies most of the correct ligand positions as the lowest score. We expanded a previously used diverse "cross-docking" benchmark to thirty ligand-protein pairs extracted from different crystal structures. The algorithm systematically scans pairs of neighbouring side chains, replaces them by alanines, and docks the ligand to each `gapped' version of the pocket. All docked positions are scored, refined with original side chains and flexible backbone and re-scored. In the optimal version of the protocol pairs of residues were replaced by alanines and only one best scoring conformation was selected from each `gapped' pocket for refinement. The optimal SCARE (SCan Alanines and REfine) protocol identifies a near native conformation (under 2 Å RMSD) as the lowest rank for 80% of pairs if the docking bounding box is defined by the predicted pocket envelope, and for as many as 90% of the pairs if the bounding box is derived from the known answer with ˜5 Å margin as used in most previous publications. The presented fully automated algorithm takes about 2 h per pose of a single processor time, requires only one pocket structure and no prior knowledge about the binding site location. Furthermore, the results for conformationally conserved pockets do not deteriorate due to substantial increase of the pocket variability.

  4. Dual estrogenic regulation of the nuclear progestin receptor and spermatogonial renewal during gilthead seabream (Sparus aurata) spermatogenesis.

    PubMed

    Chauvigné, François; Parhi, Janmejay; Ollé, Judith; Cerdà, Joan

    2017-04-01

    Studies in teleosts suggest that progestins have crucial functions during early spermatogenesis. However, the role of the different progestin receptors in these mechanisms is poorly understood. In this work, we investigated the expression pattern and hormonal regulation of the classical nuclear progestin receptor (Pgr) in the gilthead seabream at three different stages of spermatogenesis: the resting (postspawning) phase, onset of spermatogenesis, and spermiation. Immunolocalization experiments using a seabream specific Pgr antibody revealed that the receptor was expressed in Sertoli and Leydig cells, and also in a subset of spermatogonia type A, throughout spermatogenesis. Short-term treatment of testis explants with 17β-estradiol (E2) increased pgr mRNA expression at all stages, while the progestin 17α,20β-dihydroxy-4-pregnen-3-one (17,20βP) had the opposite effect. At the resting stage, Sertoli cell Pgr expression was positively correlated with the occurrence of proliferating spermatogonia type A in the tubules, and both processes were incremented in vitro by E2 likely through the estrogen receptor alpha (Era) expressed in Sertoli and Leydig cells. In contrast, treatment with 17,20βP downregulated Pgr expression in somatic cells. The androgen 11-ketotestosterone (11-KT) upregulated pgr expression in Leydig cells and promoted the proliferation of mostly spermatogonia type B, but only during spermiation. No relationship between the changes in the cell type-specific expression of the Pgr with the entry into meiosis of germ cells was found. These data suggest a differential steroid regulation of Pgr expression during seabream spermatogenesis and the potential interplay of the E2/Era and 17,20βP/Pgr pathways for the maintenance of spermatogonial renewal rather than entry into meiosis.

  5. A novel smooth impact drive mechanism actuation method with dual-slider for a compact zoom lens system.

    PubMed

    Lee, Jonghyun; Kwon, Won Sik; Kim, Kyung-Soo; Kim, Soohyun

    2011-08-01

    In this paper, a novel actuation method for a smooth impact drive mechanism that positions dual-slider by a single piezo-element is introduced and applied to a compact zoom lens system. A mode chart that determines the state of the slider at the expansion or shrinkage periods of the piezo-element is presented, and the design guide of a driving input profile is proposed. The motion of dual-slider holding lenses is analyzed at each mode, and proper modes for zoom functions are selected for the purpose of positioning two lenses. Because the proposed actuation method allows independent movement of two lenses by a single piezo-element, the zoom lens system can be designed to be compact. For a feasibility test, a lens system composed of an afocal zoom system and a focusing lens was developed, and the passive auto-focus method was implemented.

  6. Synthesis and structure-activity relationships of new carbonyl guanidine derivatives as novel dual 5-HT2B and 5-HT7 receptor antagonists.

    PubMed

    Moritomo, Ayako; Yamada, Hiroyoshi; Watanabe, Toshihiro; Itahana, Hirotsune; Akuzawa, Shinobu; Okada, Minoru; Ohta, Mitsuaki

    2013-12-15

    To identify potent dual 5-HT2B and 5-HT7 receptor antagonists, we synthesized a series of novel carbonyl guanidine derivatives and examined their structure-activity relationships. Among these compounds, N-(9-hydroxy-9H-fluorene-2-carbonyl)guanidine (10) had a good in vitro profile, that is, potent affinity for human 5-HT2B and 5-HT7 receptor subtypes (Ki=1.8 nM and Ki=17.6 nM, respectively) and high selectivity over 5-HT2A, 5-HT2C, α1, D2 and M1 receptors. Compound 10 also showed a suppressing effect on 5-HT-induced dural protein extravasation in guinea pigs when orally administered.

  7. A novel glucagon-like peptide 1/glucagon receptor dual agonist improves steatohepatitis and liver regeneration in mice.

    PubMed

    Valdecantos, M Pilar; Pardo, Virginia; Ruiz, Laura; Castro-Sánchez, Luis; Lanzón, Borja; Fernández-Millán, Elisa; García-Monzón, Carmelo; Arroba, Ana I; González-Rodríguez, Águeda; Escrivá, Fernando; Álvarez, Carmen; Rupérez, Francisco J; Barbas, Coral; Konkar, Anish; Naylor, Jacqui; Hornigold, David; Santos, Ana Dos; Bednarek, Maria; Grimsby, Joseph; Rondinone, Cristina M; Valverde, Ángela M

    2017-03-01

    Because nonalcoholic steatohepatitis (NASH) is associated with impaired liver regeneration, we investigated the effects of G49, a dual glucagon-like peptide-1/glucagon receptor agonist, on NASH and hepatic regeneration. C57Bl/6 mice fed chow or a methionine and choline-deficient (MCD) diet for 1 week were divided into 4 groups: control (chow diet), MCD diet, chow diet plus G49, and M+G49 (MCD diet plus G49). Mice fed a high-fat diet (HFD) for 10 weeks were divided into groups: HFD and H+G49 (HFD plus G49). Following 2 (MCD groups) or 3 (HFD groups) weeks of treatment with G49, partial hepatectomy (PH) was performed, and all mice were maintained on the same treatment schedule for 2 additional weeks. Analysis of liver function, hepatic regeneration, and comprehensive genomic and metabolic profiling were conducted. NASH was ameliorated in the M+G49 group, manifested by reduced inflammation, steatosis, oxidative stress, and apoptosis and increased mitochondrial biogenesis. G49 treatment was also associated with replenishment of intrahepatic glucose due to enhanced gluconeogenesis and reduced glucose use through the pentose phosphate cycle and oxidative metabolism. Following PH, G49 treatment increased survival, restored the cytokine-mediated priming phase, and enhanced the proliferative capacity and hepatic regeneration ratio in mice on the MCD diet. NASH markers remained decreased in M+G49 mice after PH, and glucose use was shifted to the pentose phosphate cycle and oxidative metabolism. G49 administered immediately after PH was also effective at alleviating the pathological changes induced by the MCD diet. Benefits in terms of liver regeneration were also found in mice fed HFD and treated with G49.

  8. The Dual Syk/JAK Inhibitor Cerdulatinib Antagonizes B-cell Receptor and Microenvironmental Signaling in Chronic Lymphocytic Leukemia.

    PubMed

    Blunt, Matthew D; Koehrer, Stefan; Dobson, Rachel C; Larrayoz, Marta; Wilmore, Sarah; Hayman, Alice; Parnell, Jack; Smith, Lindsay D; Davies, Andrew; Johnson, Peter W M; Conley, Pamela B; Pandey, Anjali; Strefford, Jonathan C; Stevenson, Freda K; Packham, Graham; Forconi, Francesco; Coffey, Greg P; Burger, Jan A; Steele, Andrew J

    2016-10-03

    Purpose: B-cell receptor (BCR)-associated kinase inhibitors, such as ibrutinib, have revolutionized the treatment of chronic lymphocytic leukemia (CLL). However, these agents are not curative, and resistance is already emerging in a proportion of patients. IL4, expressed in CLL lymph nodes, can augment BCR signaling and reduce the effectiveness of BCR kinase inhibitors. Therefore, simultaneous targeting of the IL4- and BCR signaling pathways by cerdulatinib, a novel dual Syk/JAK inhibitor currently in clinical trials (NCT01994382), may improve treatment responses in patients.Experimental Design: PBMCs from patients with CLL were treated in vitro with cerdulatinib alone or in combination with venetoclax. Cell death, chemokine, and cell signaling assay were performed and analyzed by flow cytometry, immunoblotting, q-PCR, and ELISA as indicated.Results: At concentrations achievable in patients, cerdulatinib inhibited BCR- and IL4-induced downstream signaling in CLL cells using multiple readouts and prevented anti-IgM- and nurse-like cell (NLC)-mediated CCL3/CCL4 production. Cerdulatinib induced apoptosis of CLL cells, in a time- and concentration-dependent manner, and particularly in IGHV-unmutated samples with greater BCR signaling capacity and response to IL4, or samples expressing higher levels of sIgM, CD49d(+), or ZAP70(+) Cerdulatinib overcame anti-IgM, IL4/CD40L, or NLC-mediated protection by preventing upregulation of MCL-1 and BCL-XL; however, BCL-2 expression was unaffected. Furthermore, in samples treated with IL4/CD40L, cerdulatinib synergized with venetoclax in vitro to induce greater apoptosis than either drug alone.Conclusion: Cerdulatinib is a promising therapeutic for the treatment of CLL either alone or in combination with venetoclax, with the potential to target critical survival pathways in this currently incurable disease. Clin Cancer Res; 1-12. ©2016 AACR.

  9. Biased agonism as a mechanism for differential signaling by chemokine receptors.

    PubMed

    Rajagopal, Sudarshan; Bassoni, Daniel L; Campbell, James J; Gerard, Norma P; Gerard, Craig; Wehrman, Tom S

    2013-12-06

    Chemokines display considerable promiscuity with multiple ligands and receptors shared in common, a phenomenon that is thought to underlie their biochemical "redundancy." Their receptors are part of a larger seven-transmembrane receptor superfamily, commonly referred to as G protein-coupled receptors, which have been demonstrated to be able to signal with different efficacies to their multiple downstream signaling pathways, a phenomenon referred to as biased agonism. Biased agonism has been primarily reported as a phenomenon of synthetic ligands, and the biologic prevalence and importance of such signaling are unclear. Here, to assess the presence of biased agonism that may underlie differential signaling by chemokines targeting the same receptor, we performed a detailed pharmacologic analysis of a set of chemokine receptors with multiple endogenous ligands using assays for G protein signaling, β-arrestin recruitment, and receptor internalization. We found that chemokines targeting the same receptor can display marked differences in their efficacies for G protein- or β-arrestin-mediated signaling or receptor internalization. This ligand bias correlates with changes in leukocyte migration, consistent with different mechanisms underlying the signaling downstream of these receptors induced by their ligands. These findings demonstrate that biased agonism is a common and likely evolutionarily conserved biological mechanism for generating qualitatively distinct patterns of signaling via the same receptor in response to different endogenous ligands.

  10. Mechanism of GABAB receptor-induced BDNF secretion and promotion of GABAA receptor membrane expression.

    PubMed

    Kuczewski, Nicola; Fuchs, Celine; Ferrand, Nadine; Jovanovic, Jasmina N; Gaiarsa, Jean-Luc; Porcher, Christophe

    2011-08-01

    Recent studies have shown that GABA(B) receptors play more than a classical inhibitory role and can function as an important synaptic maturation signal early in life. In a previous study, we reported that GABA(B) receptor activation triggers secretion of brain-derived neurotrophic factor (BDNF) and promotes the functional maturation of GABAergic synapses in the developing rat hippocampus. To identify the signalling pathway linking GABA(B) receptor activation to BDNF secretion in these cells, we have now used the phosphorylated form of the cAMP response element-binding protein as a biological sensor for endogenous BDNF release. In the present study, we show that GABA(B) receptor-induced secretion of BDNF relies on the activation of phospholipase C, followed by the formation of diacylglycerol, activation of protein kinase C, and the opening of L-type voltage-dependent Ca(2+) channels. We further show that once released by GABA(B) receptor activation, BDNF increases the membrane expression of β(2/3) -containing GABA(A) receptors in neuronal cultures. These results reveal a novel function of GABA(B) receptors in regulating the expression of GABA(A) receptor through BDNF-tropomyosin-related kinase B receptor dependent signalling pathway.

  11. Coupling characteristics and control of dual mechanical port machine with spoke type permanent magnet arrangement

    NASA Astrophysics Data System (ADS)

    Zhuang, Xingming; Song, Qiang; Wen, Xuhui; Zhao, Feng; Fan, Tao

    2014-11-01

    Dual mechanical port machine(DMPM), as a novel electromechanical energy conversion device, has attracted widespread attention. DMPM with spoke type permanent magnet arrangements(STPM-DMPM), which is one of several types of DMPM, has been of interest recently. The unique coupling characteristics of STPM-DMPM are beneficial to improving system performance, but these same characteristics increase the difficulties of control. Now there has been little research about the control of STPM-DMPM, and this has hindered its practical application. Based on a mathematical model of STPM-DMPM, the coupling characteristics and the merits and demerits of such devices are analyzed as applied to a hybrid system. The control strategies for improving the disadvantages and for utilizing the advantage of coupling are researched. In order to weaken the interaction effect of torque outputs in the inner motor and the outer motor that results from coupling in STPM-DMPM, a decoupling control method based on equivalent current control is proposed, and independent torque control for the inner motor and outer motor is achieved. In order to solve address the problem of adequately utilization of coupling, minimizing the overall copper loss of the inner motor and the outer motor of STPM-DMPM is taken as the optimization objective for optimal control, and the purpose of utilizing the coupling adequately and reasonably is achieved. The verification tests of the proposed decoupling control and optimal control strategies are carried out on a prototype STPM-DMPM, and the experimental results show that the interaction effect of torque outputs in the inner motor and the outer motor can be markedly weakened through use of the control method. The overall copper loss of the inner motor and the outer motor can be markedly reduced through use of the optimal control method, while the power output remains unchanged. A breakthrough in the control problem of STPM-DMPM is accomplished by combining the control

  12. Isolating the Neural Mechanisms of Interference During Continuous Multisensory Dual-task Performance

    DTIC Science & Technology

    2014-01-01

    sion accuracy in each task, the signal detection theory sensitivity measure of d0 was used (Macmillan & Creelman , 1991; Green & Swets, 1966). First...J. (1998). Sources of dual-task interference: Evidence from human electrophysiology. Psychological Science, 9, 223–227. Macmillan, N. A., & Creelman

  13. Cellular mechanisms of the 5-HT7 receptor-mediated signaling

    PubMed Central

    Guseva, Daria; Wirth, Alexander; Ponimaskin, Evgeni

    2014-01-01

    Serotonin (5-hydroxytryptamine or 5-HT) is an important neurotransmitter regulating a wide range of physiological and pathological functions via activation of heterogeneously expressed 5-HT receptors. The 5-HT7 receptor is one of the most recently described members of the 5-HT receptor family. Functionally, 5-HT7 receptor is associated with a number of physiological and pathological responses, including serotonin-induced phase shifting of the circadian rhythm, control of memory as well as locomotor and exploratory activity. A large body of evidence indicates involvement of the 5-HT7 receptor in anxiety and depression, and recent studies suggest that 5-HT7 receptor can be highly relevant for the treatment of major depressive disorders. The 5-HT7 receptor is coupled to the stimulatory Gs-protein, and receptor stimulation results in activation of adenylyl cyclase (AC) leading to a rise of cAMP concentration. In addition, this receptor is coupled to the G12-protein to activate small GTPases of the Rho family. This review focuses on molecular mechanisms responsible for the 5-HT7 receptor-mediated signaling. We provide detailed overview of signaling cascades controlled and regulated by the 5-HT7 receptor and discuss the functional impact of 5-HT7 receptor for the regulation of different cellular and subcellular processes. PMID:25324743

  14. Cellular mechanisms of the 5-HT7 receptor-mediated signaling.

    PubMed

    Guseva, Daria; Wirth, Alexander; Ponimaskin, Evgeni

    2014-01-01

    Serotonin (5-hydroxytryptamine or 5-HT) is an important neurotransmitter regulating a wide range of physiological and pathological functions via activation of heterogeneously expressed 5-HT receptors. The 5-HT7 receptor is one of the most recently described members of the 5-HT receptor family. Functionally, 5-HT7 receptor is associated with a number of physiological and pathological responses, including serotonin-induced phase shifting of the circadian rhythm, control of memory as well as locomotor and exploratory activity. A large body of evidence indicates involvement of the 5-HT7 receptor in anxiety and depression, and recent studies suggest that 5-HT7 receptor can be highly relevant for the treatment of major depressive disorders. The 5-HT7 receptor is coupled to the stimulatory Gs-protein, and receptor stimulation results in activation of adenylyl cyclase (AC) leading to a rise of cAMP concentration. In addition, this receptor is coupled to the G12-protein to activate small GTPases of the Rho family. This review focuses on molecular mechanisms responsible for the 5-HT7 receptor-mediated signaling. We provide detailed overview of signaling cascades controlled and regulated by the 5-HT7 receptor and discuss the functional impact of 5-HT7 receptor for the regulation of different cellular and subcellular processes.

  15. Heterodimeric interaction between retinoid X receptor alpha and orphan nuclear receptor OR1 reveals dimerization-induced activation as a novel mechanism of nuclear receptor activation.

    PubMed Central

    Wiebel, F F; Gustafsson, J A

    1997-01-01

    OR1 is a member of the steroid/thyroid hormone nuclear receptor superfamily which has been described to mediate transcriptional responses to retinoids and oxysterols. On a DR4 response element, an OR1 heterodimer with the nuclear receptor retinoid X receptor alpha (RXR alpha) has been described to convey transcriptional activation in both the absence and presence of the RXR ligand 9-cis retinoic acid, the mechanisms of which have remained unclear. Here, we dissect the effects of RXR alpha and OR1 ligand-binding domain interaction on transcriptional regulation and the role of the respective carboxy-terminal activation domains (AF-2s) in the absence and presence of the RXR ligand, employing chimeras of the nuclear receptors containing the heterologous GAL4 DNA-binding domain as well as natural receptors. The results show that the interaction of the RXR and OR1 ligand-binding domains unleashes a transcription activation potential that is mainly dependent on the AF-2 of OR1, indicating that interaction with RXR activates OR1. This defines dimerization-induced activation as a novel function of heterodimeric interaction and mechanism of receptor activation not previously described for nuclear receptors. Moreover, we present evidence that activation of OR1 occurs by a conformational change induced upon heterodimerization with RXR. PMID:9199332

  16. Mechanisms of acquired resistance to androgen receptor targeting drugs in castration resistant prostate cancer

    PubMed Central

    Chism, David D.; De Silva, Dinuka; Whang, Young E.

    2014-01-01

    After initial response to androgen receptor targeting drugs abiraterone or enzalutamide, most patients develop progressive disease and therefore, castration resistant prostate cancer (CRPC) remains a terminal disease. Multiple mechanisms underlying acquired resistance have been postulated. Intratumoral androgen synthesis may resume after abiraterone treatment. A point mutation in the ligand binding domain of androgen receptor may confer resistance to enzalutamide. Emergence of androgen receptor splice variants lacking the ligand binding domain may mediate resistance to abiraterone and enzalutamide. Steroid receptors such as glucocorticoid receptor may substitute for androgen receptor. Drugs with novel mechanisms of action or combination therapy, along with biomarkers for patient selection, may be needed to improve the therapy of CRPC. PMID:24927631

  17. Distinct mechanisms of endocrine disruption of DDT-related pesticides toward estrogen receptor α and estrogen-related receptor γ.

    PubMed

    Zhuang, Shulin; Zhang, Jing; Wen, Yuezhong; Zhang, Chunlong; Liu, Weiping

    2012-11-01

    Dichlorodiphenyltrichloroethane (DDT) is ubiquitous in the environment, and the exposure to DDT and its related pesticides has long been linked to endocrine disruption. The mechanism of endocrine disruption toward targeted receptors, however, remains unclear. Probing the molecular recognition of DDT analogs by targeted receptors at the atomic level is critical for deciphering this mechanism. Molecular dynamics (MD) simulations were applied to probe the molecular recognition process of DDT and its five analogs, including dichlordiphenyldichloroethylene (DDE), dichlorodiphenyldichloroethane (DDD), methoxychlor (MXC), p,p'-hydroxy-DDT (HPTE), and dicofol by human estrogen receptor (ER) α and human ER-related receptor (ERR) γ. Van der Waals interactions mainly drive the interactions of DDT analogs with ERα ligand-binding domain (LBD) and ERRγ LBD. Minor structural changes of DDT analogs in the number and position of chlorine and phenolic hydroxyl moiety cause differences in binding modes through aromatic stacking and hydrogen bonding and thus affect differently conformational changes of ERα LBD and ERRγ LBD. The binding of DDT analogs affects the helix 12 orientation of ERα LBD but causes no rearrangement of helix 12 of ERRγ LBD. These results extend our understanding of how DDT analogs exert their estrogen-disrupting effects toward different receptors via multiple mechanisms.

  18. Molecular mechanism for opioid dichotomy: bidirectional effect of μ-opioid receptors on P2X₃ receptor currents in rat sensory neurones.

    PubMed

    Chizhmakov, Igor; Kulyk, Vyacheslav; Khasabova, Iryna; Khasabov, Sergey; Simone, Donald; Bakalkin, Georgy; Gordienko, Dmitri; Verkhratsky, Alexei; Krishtal, Oleg

    2015-06-01

    Here, we describe a molecular switch associated with opioid receptors-linked signalling cascades that provides a dual opioid control over P2X3 purinoceptor in sensory neurones. Leu-enkephalin inhibited P2X3-mediated currents with IC50 ~10 nM in ~25% of small nociceptive rat dorsal root ganglion (DRG) neurones. In contrast, in neurones pretreated with pertussis toxin leu-enkephalin produced stable and significant increase of P2X3 currents. All effects of opioid were abolished by selective μ-opioid receptor antagonist D-Phe-Cys-Tyr-D-Trp-Orn-Thr-Pen-Thr-NH2 (CTOP), nonselective inhibitor naloxone, and by PLC inhibitor U73122. Thus, we discovered a dual link between purinoceptors and μ-opioid receptors: the latter exert both inhibitory (pertussis toxin-sensitive) and stimulatory (pertussis toxin-insensitive) actions on P2X3 receptors through phospholipase C (PLC)-dependent pathways. This dual opioid control of P2X3 receptors may provide a molecular explanation for dichotomy of opioid therapy. Pharmacological control of this newly identified facilitation/inhibition switch may open new perspectives for the adequate medical use of opioids, the most powerful pain-killing agents known today.

  19. Mechanism of kinase activation in the receptor for colony-stimulating factor 1.

    PubMed Central

    Lee, A W; Nienhuis, A W

    1990-01-01

    Receptor tyrosine kinases remain dormant until activated by ligand binding to the extracellular domain. Two mechanisms have been proposed for kinase activation: (i) ligand binding to the external domain of a receptor monomer may induce a conformational change that is transmitted across the cell membrane (intramolecular model) or (ii) the ligand may facilitate oligomerization, thereby allowing interactions between the juxtaposed kinase domains (intermolecular model). The receptor for colony-stimulating factor 1 was used to test these models. Large insertions at the junction between the external and transmembrane domains of the receptor, introduced by site-directed mutagenesis of the cDNA, were positioned to isolate the external domain and prevent transmembrane conformational propagation while allowing for receptor oligomerization. Such mutant receptors were expressed on the cell surface, bound ligand with high affinity, exhibited ligand-stimulated autophosphorylation, and signaled mitogenesis and cellular proliferation in the presence of ligand. A second experimental strategy directly tested the intermolecular model of ligand activation. A hybrid receptor composed of the external domain of human glycophorin A and the transmembrane and cytoplasmic domains of the colony-stimulating factor 1 receptor exhibited anti-glycophorin antibody-induced kinase activity that supported mitogenesis. Our data strongly support a mechanism of receptor activation based on ligand-induced receptor oligomerization. Images PMID:2169623

  20. Differences in the binding mechanism of RU486 and progesterone to the progesterone receptor

    SciTech Connect

    Skafar, D.F. )

    1991-11-12

    The binding mechanism of the antagonist RU486 to the progesterone receptor was compared with that of the agonists progesterone and R5020. Both progesterone and RU486 bound to the receptor with a Hill coefficient of 1.2, indicating the binding of each ligand is positive cooperative. However, when each ligand was used to compete with ({sup 3}H)progesterone for binding to the receptor at receptor concentrations near 8 nM, at which the receptor is likely a dimer, the competition curve for RU486 was significantly steeper than the curves for progesterone and R5020. This indicated that a difference in the binding mechanism of RU486 and progesterone can be detected when both ligands are present. In contrast, at receptor concentrations near 1 nM, at which the receptor is likely a monomer, the competition curves for all three ligands were indistinguishable. These results indicate that RU486 and agonists have different binding mechanisms for the receptor and further suggest that this difference may be related to site-site interactions within the receptor.

  1. Antidepressant effects of ketamine and the roles of AMPA glutamate receptors and other mechanisms beyond NMDA receptor antagonism.

    PubMed

    Aleksandrova, Lily R; Phillips, Anthony G; Wang, Yu Tian

    2017-01-31

    The molecular mechanisms underlying major depressive disorder remain poorly understood, and current antidepressant treatments have many shortcomings. The recent discovery that a single intravenous infusion of ketamine at a subanesthetic dose had robust, rapid and sustained antidepressant effects in individuals with treatment-resistant depression inspired tremendous interest in investigating the molecular mechanisms mediating ketamine's clinical efficacy as well as increased efforts to identify new targets for antidepressant action. We review the clinical utility of ketamine and recent insights into its mechanism of action as an antidepressant, including the roles of N-methyl-D-aspartate receptor inhibition, α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid receptor upregulation, activation of downstream synaptogenic signalling pathways and the production of an active ketamine metabolite, hydroxynorketamine. Emerging knowledge of the molecular mechanisms underlying both ketamine's positive therapeutic and detrimental side effects will aid the development of a new generation of much-needed superior antidepressant agents.

  2. The transient receptor potential ankyrin-1 mediates mechanical hyperalgesia induced by the activation of B1 receptor in mice.

    PubMed

    Meotti, Flavia Carla; Figueiredo, Cláudia Pinto; Manjavachi, Marianne; Calixto, João B

    2017-02-01

    The kinin receptor B1 and the transient receptor potential ankyrin 1 (TRPA1) work as initiators and gatekeepers of nociception and inflammation. This study reports that the nociceptive transmission induced by activation of B1 receptor is dependent on TRPA1 ion channel. The mechanical hyperalgesia was induced by intrathecal (i.t.) injection of B1 agonist des-Arginine(9)-bradykinin (DABK) or TRPA1 agonist cinnamaldehyde and was evaluated by the withdrawal response after von Frey Hair application in the hind paw. After behavioral experiments, lumbar spinal cord and dorsal root ganglia (DRG) were harvested to assess protein expression and mRNA by immunohistochemistry and real time-PCR, respectively. The pharmacological antagonism (HC030031) or the down-regulation of TRPA1 greatly inhibited the mechanical hyperalgesia induced by DABK. Intrathecal injection of DABK up regulated the ionized calcium binding adaptor molecule (Iba-1) in lumbar spinal cord (L5-L6); TRPA1 protein and mRNA in lumbar spinal cord; and B1 receptor mRNA in both lumbar spinal cord and DRG. The knockdown of TRPA1 prevented microglia activation induced by DABK. Furthermore, the mechanical hyperalgesia induced by either DABK or by cinnamaldehyde was significantly reduced by inhibition of cyclooxygenase (COX), protein kinase C (PKC) or phospholipase C (PLC). In summary, this study revealed that TRPA1 positively modulates the mechanical hyperalgesia induced by B1 receptor activation in the spinal cord and that the classical GPCR downstream molecules PLC, diacylglycerol (DAG), 3,4,5-inositide phosphate (IP3) and PKC are involved in the nociceptive transmission triggered by these two receptors.

  3. Influence of Thermal Aging on the Microstructure and Mechanical Behavior of Dual Phase Precipitation Hardened Powder Metallurgy Stainless Steels

    NASA Astrophysics Data System (ADS)

    Stewart, Jennifer

    2011-12-01

    Increasing demand for high strength powder metallurgy (PM) steels has resulted in the development of dual phase PM steels. In this work, the effects of thermal aging on the microstructure and mechanical behavior of dual phase precipitation hardened powder metallurgy (PM) stainless steels of varying ferrite-martensite content were examined. Quantitative analyses of the inherent porosity and phase fractions were conducted on the steels and no significant differences were noted with respect to aging temperature. Tensile strength, yield strength, and elongation to fracture all increased with increasing aging temperature reaching maxima at 538°C in most cases. Increased strength and decreased ductility were observed in steels of higher martensite content. Nanoindentation of the individual microconstituents was employed to obtain a fundamental understanding of the strengthening contributions. Both the ferrite and martensite hardness values increased with aging temperature and exhibited similar maxima to the bulk tensile properties. Due to the complex non-uniform stresses and strains associated with conventional nanoindentation, micropillar compression has become an attractive method to probe local mechanical behavior while limiting strain gradients and contributions from surrounding features. In this study, micropillars of ferrite and martensite were fabricated by focused ion beam (FIB) milling of dual phase precipitation hardened powder metallurgy (PM) stainless steels. Compression testing was conducted using a nanoindenter equipped with a flat punch indenter. The stress-strain curves of the individual microconstituents were calculated from the load-displacement curves less the extraneous displacements of the system. Using a rule of mixtures approach in conjunction with porosity corrections, the mechanical properties of ferrite and martensite were combined for comparison to tensile tests of the bulk material, and reasonable agreement was found for the ultimate tensile

  4. A novel dual-glucagon-like peptide-1 and glucose-dependent insulinotropic polypeptide receptor agonist is neuroprotective in transient focal cerebral ischemia in the rat.

    PubMed

    Han, Ling; Hölscher, Christian; Xue, Guo-Fang; Li, Guanglai; Li, Dongfang

    2016-01-06

    Glucagon-like peptide-1 (GLP-1) and glucose-dependent insulinotropic polypeptide (GIP) receptor agonists have been shown to be neuroprotective in previous studies in animal models of Alzheimer's or Parkinson's disease. Recently, novel dual-GLP-1/GIP receptor agonists that activate both receptors (DA) were developed to treat diabetes. We tested the protective effects of a novel potent DA against middle cerebral artery occlusion injury in rats and compared it with a potent GLP-1 analog, Val(8)-GLP-1(glu-PAL). Animals were evaluated for neurologic deficit score, infarct volume, and immunohistochemical analyses of the brain at several time points after ischemia. The Val(8)-GLP-1(glu-PAL)-treated and DA-treated groups showed significantly reduced scores of neurological dysfunction, cerebral infarction size, and percentage of TUNEL-positive apoptotic neurons. Furthermore, the expression of the apoptosis marker Bax, the inflammation marker iNOS, and the survival marker Bcl-2 was significantly increased. The DA-treated group was better protected against neurodegeneration than the Val(8)-GLP-1(glu-PAL) group, and the scores of neurological dysfunction, cerebral infarction size, and expression of Bcl-2 were higher, whereas the percentage of TUNEL-positive neurons and the levels of Bax and iNOS were lower in the DA group. DA treatment reduced the infarct volume and improved the functional deficit. It also suppressed the inflammatory response and cell apoptosis after reperfusion. In conclusion, the novel GIP and GLP-1 dual-receptor agonist is more neuroprotective than a GLP-1 receptor agonist in key biomarkers of neuronal degeneration.

  5. Effect of curing mode on the micro-mechanical properties of dual-cured self-adhesive resin cements.

    PubMed

    Ilie, Nicoleta; Simon, Alexander

    2012-04-01

    Light supplying to luting resin cements is impeded in several clinical situations, causing us to question whether materials can properly be cured to achieve adequately (or adequate) mechanical properties. The aim of this study was therefore to analyse the effect of light on the micro-mechanical properties of eight popular dual-cured self-adhesive resin cements by comparing them with two conventional, also dual-cured, resin cements. Four different curing procedures were applied: auto-polymerisation (dark curing) and light curing (LED unit, Freelight 2, 20 s) by applying the unit directly on the samples' surface, at a distance of 5 and 10 mm. Twenty minutes after curing, the samples were stored for 1 week at 37°C in a water-saturated atmosphere. The micro-mechanical properties-Vickers hardness, modulus of elasticity, creep and elastic/plastic deformation-were measured. Data were analysed with multivariate ANOVA followed by Tukey's test and partial eta-squared statistics (p < 0.05). A very strong influence of the material as well as filler volume and weight on the micro-mechanical properties was measured, whereas the influence of the curing procedure and type of cement-conventional or self-adhesive-was generally low. The influence of light on the polymerisation process was material dependent, with four different behaviour patterns to be distinguished. As a material category, significantly higher micro-mechanical properties were measured for the conventional compared to the self-adhesive resin cements, although this difference was low. Within the self-adhesive resin cements group, the variation in micro-mechanical properties was high. The selection of suitable resin cements should be done by considering, besides its adhesive properties, its micro-mechanical properties and curing behaviour also.

  6. Signal transduction of receptor-mediated antiproliferative action of melatonin on human prostate epithelial cells involves dual activation of Gα(s) and Gα(q) proteins.

    PubMed

    Shiu, Stephen Y W; Pang, Bo; Tam, Chun W; Yao, Kwok-Ming

    2010-10-01

    Melatonin has been shown to inhibit the proliferation of malignant and transformed human prostate epithelial cells by transcriptional up-regulation of p27(Kip1) expression via MTNR1A receptor-mediated activation of protein kinase A (PKA) and protein kinase C (PKC) in parallel. Given that melatonin MTNR1A receptor is a G protein-coupled receptor, this study was conducted to identify the specific G proteins that mediate the antiproliferative action of melatonin on human prostate epithelial cells. In 22Rv1 and RWPE-1 cells, knockdown of either Gα(s) or Gα(q) , but not Gα(i2) expression by RNA interference, abrogated the effects of melatonin on p27(Kip1) and cell proliferation. Conversely, cellular overexpression of activated mutants of Gα(s) and Gα(q) in 22Rv1 and RWPE-1 cells mimicked the effects of melatonin on prostate epithelial cell antiproliferation by increasing p27(Kip1) expression through downstream activation of PKA and PKC in parallel. Moreover, melatonin or 2-iodomelatonin induced elevation of adenosine-3',5'-cyclic monophosphate (cAMP) in 22Rv1 and RWPE-1 cells. The effects of 2-iodomelatonin on cAMP were blocked by the nonselective MTNR1A/MTNR1B receptor antagonist luzindole but were not affected by the selective MTNR1B receptor antagonist 4-phenyl-2-propionamidotetraline (4-P-PDOT). Furthermore, knockdown of Gα(s) mitigated the stimulatory effects of 2-iodomelatonin on cAMP. Collectively, the data demonstrated, for the first time, functional coupling of MTNR1A receptor to Gα(s) in cancerous or transformed human cells expressing endogenous melatonin receptors. Our results also showed that dual activation of Gα(s) and Gα(q) proteins is involved in the signal transduction of MTNR1A receptor-mediated antiproliferative action of melatonin on human prostate epithelial cells.

  7. How does a protein with dual mitotic spindle and extracellular matrix receptor functions affect tumor susceptibility and progression?

    PubMed Central

    Tolg, Cornelia; McCarthy, James B

    2011-01-01

    The mechanisms responsible for the oncogenic effects of the hyaluronan (HA) receptor and mitotic spindle binding protein, RHAMM, are poorly understood. On one hand, extracellular RHAMM interacts with HA and cellsurface receptors such as CD44 to coordinately activate the MAPK/ERK1,2 pathway, thus contributing to the spread and proliferation of tumor cells. On the other hand, intracellular RHAMM decorates mitotic spindles and is necessary for spindle formation and progression through G2/M and overexpression or loss of RHAMM can result in multipole spindles and chromosome missegregation. The deregulation of these intracellular functions could lead to genomic instability and fuel tumor progression. This suggests that both extracellular and intracellular RHAMM can promote tumor progression. Intracellular RHAMM can bind directly to ERK1 to form complexes with ERK2, MEK1 and ERK1,2 substrates, and we present a model whereby RHAMM's function is as a scaffold protein, controlling activation and targeting of ERK1,2 to specific substrates. PMID:21655434

  8. Mechanism of cooperative catalysis in a Lewis acid promoted nickel-catalyzed dual C-H activation reaction.

    PubMed

    Anand, Megha; Sunoj, Raghavan B

    2012-09-07

    The mechanism of cooperativity offered by AlMe(3) in a Ni-catalyzed dehydrogenative cycloaddition between substituted formamides and an alkyne is investigated by using DFT(SMD(toluene)/M06/6-31G**) methods. The preferred pathway is identified to involve dual C-H activation, with first a higher barrier formyl C(sp(2))-H oxidative insertion followed by benzylic methyl C(sp(3))-H activation. The cooperativity is traced to be of kinetic origin as evidenced by stabilized transition states when AlMe(3) is bound to the formyl group, particularly in the oxidative insertion step.

  9. [1+1 or 2x1? Another form of dual antidepressive mechanism of action].

    PubMed

    Lakatos, László; Rihmer, Zoltán

    2005-09-01

    The primary aim of this review is to summarize the result of clinical trials comparing tricyclic antidepressants with first generation SSRIs, dual action (serotonergic and noradrenergic) agents like venlafaxine or mirtazapine with SSRIs and escitalopram with other SSRIs and venlafaxine. The review analyzes the relationship between the clinical efficacy and tolerability data and the mode of action. The clinical data suggest that multiple mode of action can substantially improve efficacy, however the relative risk of side-effects also increases if a drug affects multiple neurotransmitters. A new opportunity of development is the molecule of escitalopram introduced to the market some years ago. Its unique dual action on serotonergic transmission offers improved efficacy without compromising the favourable side-effect profile due to selectivity. Based on clinical trials escitalopram shows improved efficacy, faster onset of action and higher remission rates compared to first generation SSRIs. Remission rates with escitalopram are at least as high as with venlafaxine and escitalopram shows faster onset of action, earlier sustained remission and better tolerability. The good clinical efficacy data of escitalopram may be explained by its unique dual serotonergic effect on serotonin transporter protein.

  10. Mechanisms of Action of Anticholinesterases and Oximes on Acetylcholine Receptors

    DTIC Science & Technology

    1988-07-23

    J.F. and D.B. Sanders. The management of patients with myasthenia gravis , in Myasthenia Gravis (E.X. Albuquerque and A.T. Eldefrawi, eds.), Chapman...Eldefrawi. Affinity of myasthenia drugs to acetylcholinesterase and acetylcholine receptor. Biochem. Med. 10:258-265 (1974). 9. Carpenter, D.O., L.A

  11. Mechanism of partial agonism in AMPA-type glutamate receptors

    PubMed Central

    Salazar, Hector; Eibl, Clarissa; Chebli, Miriam; Plested, Andrew

    2017-01-01

    Neurotransmitters trigger synaptic currents by activating ligand-gated ion channel receptors. Whereas most neurotransmitters are efficacious agonists, molecules that activate receptors more weakly—partial agonists—also exist. Whether these partial agonists have weak activity because they stabilize less active forms, sustain active states for a lesser fraction of the time or both, remains an open question. Here we describe the crystal structure of an α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate receptor (AMPAR) ligand binding domain (LBD) tetramer in complex with the partial agonist 5-fluorowillardiine (FW). We validate this structure, and others of different geometry, using engineered intersubunit bridges. We establish an inverse relation between the efficacy of an agonist and its promiscuity to drive the LBD layer into different conformations. These results suggest that partial agonists of the AMPAR are weak activators of the receptor because they stabilize multiple non-conducting conformations, indicating that agonism is a function of both the space and time domains. PMID:28211453

  12. Role of angiotensin AT(2) receptors in natriuresis: Intrarenal mechanisms and therapeutic potential.

    PubMed

    Carey, Robert M; Padia, Shetal H

    2013-08-01

    The renin-angiotensin system is a coordinated hormonal cascade critical for the regulation of blood pressure (BP) and kidney function. Angiotensin (Ang) II, the major angiotensin effector peptide, binds to two major receptors, namely AT1 and AT2 receptors. The AT1 receptors engender antinatriuresis and raise BP, whereas AT2 receptors oppose these effects, inducing natriuresis and reducing BP. There is high AT2 receptor expression in the adult kidney, especially in the proximal tubule. In AT2 receptor-null mice, long-term AngII infusion results in pressor and antinatriuretic hypersensivivity compared with responses in wild-type mice. The major endogenous receptor ligand for AT2 receptor-mediated natriuretic responses appears to be des-aspartyl(1) -AngII (AngIII) instead of AngII. Recent studies have demonstrated that AngII requires metabolism to AngIII by aminopeptidase A to induce natriuresis and that inhibition of aminopeptidase N increases intrarenal AngIII and augments AngIII-induced natriuresis. The renal dopaminergic system is another important natriuretic pathway. Renal proximal tubule the D1 and D5 receptor subtypes (D1 -like receptors (D1LIKE R)) control approximately 50% of basal sodium excretion. Recently, we have found that natriuresis induced by proximal tubule D1LIKE R requires AT2 receptor activation and that D1LIKE R stimulation induces recruitment of AT2 receptors to the apical plasma membrane via a cAMP-dependent mechanism. Initial studies using the potent AT2 receptor non-peptide agonist Compound 21 demonstrate natriuresis in both the presence and absence of AT1 receptor blockade, indicating the therapeutic potential of this compound in fluid-retaining states and hypertension.

  13. Growth factor control of epidermal growth factor receptor kinase activity via an intramolecular mechanism.

    PubMed

    Koland, J G; Cerione, R A

    1988-02-15

    The mechanism by which the protein kinase activity of the epidermal growth factor (EGF) receptor is activated by binding of growth factor was investigated. Detergent-solubilized receptor in monomeric form was isolated by sucrose density gradient centrifugation and both its kinase and autophosphorylation activities monitored. In a low ionic strength medium and with MnCl2 as an activator, the activity of the monomeric receptor was EGF-independent. However, with 0.25 M ammonium sulfate present, the MnCl2-stimulated kinase activity was strikingly EGF-dependent. In contrast, the kinase activity expressed in the presence of MgCl2 showed growth factor control in the absence of added salt. Under the conditions of these experiments there was apparently little tendency for growth factor to induce aggregation of the receptor, indicating that the allosteric activation of the receptor kinase by EGF occurred via an intramolecular mechanism. Whereas detergent-solubilized receptor was the subject of these studies, the kinase activity of cell surface receptors might also be controlled by an intramolecular mechanism. These results indicate that an individual receptor molecule has the potential to function as a transmembrane signal transducer.

  14. New low-density lipoprotein receptor upregulators acting via a novel mechanism.

    PubMed

    Ashton, M J; Brown, T J; Fenton, G; Halley, F; Harper, M F; Lockey, P M; Porter, B; Roach, A G; Stuttle, K A; Vicker, N; Walsh, R J

    1996-08-16

    The synthesis and biological activity of a new series of benzamides and related compounds that upregulate the expression of the low-density lipoprotein (LDL) receptor in human hepatocytes (HepG2 cells) by a novel mechanism are described. The lead compound, N-[5-[(3-cyclohexylpropionyl)amino]-2-methylphenyl]-4-hydroxybe nzamide (1, RPR102359), increased the expression of the LDL receptors in HepG2 cells by 80% when tested at a concentration of 3 microM. Mevinolin (lovastatin) was found to increase the LDL receptor expression by 70% at the same concentration. In contrast to mevinolin, 1 was found to have no effect on cholesterol biosynthesis in liver homogenates or in HepG2 cells at doses where substantial upregulation of the LDL receptor was observed and thus stimulated LDL receptor expression by a novel mechanism.

  15. ERBB receptors: from oncogene discovery to basic science to mechanism-based cancer therapeutics.

    PubMed

    Arteaga, Carlos L; Engelman, Jeffrey A

    2014-03-17

    ERBB receptors were linked to human cancer pathogenesis approximately three decades ago. Biomedical investigators have since developed substantial understanding of the biology underlying the dependence of cancers on aberrant ERBB receptor signaling. An array of cancer-associated genetic alterations in ERBB receptors has also been identified. These findings have led to the discovery and development of mechanism-based therapies targeting ERBB receptors that have improved outcome for many cancer patients. In this Perspective, we discuss current paradigms of targeting ERBB receptors with cancer therapeutics and our understanding of mechanisms of action and resistance to these drugs. As current strategies still have limitations, we also discuss challenges and opportunities that lie ahead as basic scientists and clinical investigators work toward more breakthroughs.

  16. Histaminergic receptors of medial septum and conditioned place preference: D1 dopamine receptor mechanism.

    PubMed

    Zarrindast, Mohammad-Reza; Moghimi, Maryam; Rostami, Parvin; Rezayof, Ameneh

    2006-09-13

    In the present study, the effects of intra-medial septum injections of histamine and/or the histamine H1 or H2 receptor antagonists on the acquisition of conditioned place preference (CPP) in male Wistar rats have been investigated. Our data showed that the conditioning treatments with intra-medial septum injection of different doses of histamine (0.5-15 microg/rat) induced a significant CPP for the drug-associated place. Using a 3-day schedule of conditioning, it was found that the histamine H1 receptor antagonist, pyrilamine (10 and 15 microg/rat, intra-medial septum) also induced a significant place preference. In addition, pyrilamine inhibited the histamine (7.5 microg/rat)-induced place preference. Intra-medial septum administration of the histamine H2 receptor antagonist, ranitidine (5-15 microg/rat) alone or in combination with histamine did not produce a significant place preference or place aversion. On the other hand, intra-medial septum administration of the dopamine D1 receptor antagonist, SCH 233390 (0.5, 0.75 and 1 microg/rat) inhibited the histamine (7.5 microg/rat) or pyrilamine (15 microg/rat)-induced place preference in a dose-dependent manner, but no effect was observed for the dopamine D2 receptor antagonist, sulpiride on the histamine or pyrilamine response. The administration of histamine (2.5-15 microg/rat) or pyrilamine (10 and 15 microg/rat) during acquisition increased locomotor activity of the animals on the testing days. The results suggest that histaminergic receptors of the medial septum may be involved in CPP and thus it is postulated that dopamine D1 receptors may play an important role in this effect.

  17. Common molecular mechanisms in field- and agrin-induced acetylcholine receptor clustering.

    PubMed

    Sabrina, F; Stollberg, J

    1997-04-01

    1. The aggregation of acetylcholine receptors at the developing neuromuscular junction is critical to the development and function of this synapse. In vitro studies have shown that receptor aggregation can be induced by the finding of agrin to the muscle cell surface and by the electric field-induced concentration of a (nonreceptor) molecule at the cathodal cell pole. 2. We report here on the interaction between agrin binding and electric fields with respect to the distribution of receptors and agrin binding sites. 3. (a) Pretreatment of cells with agrin completely blocks the development of field-induced receptor clusters. (b) Field-induced aggregation of receptors precedes the field-induced aggregation of agrin binding sites by approximately 30 min. (c) Electric fields prevent agrin-induced receptor clustering despite the presence of agrin binding sites and freely diffusing receptors. 4. These results indicate that another membrane component-but not the agrin binding site and not the receptor-is required for agrin-induced receptor clustering. They also suggest that electric fields and agrin cause receptor clustering via common molecular mechanisms.

  18. Glycyrrhizic Acid Reduces Heart Rate and Blood Pressure by a Dual Mechanism.

    PubMed

    Singh, Kailash; Zaw, Aung Moe; Sekar, Revathi; Palak, Ahuja; Allam, Ahmed A; Ajarem, Jamaan; Chow, Billy K C

    2016-09-27

    Beta adrenergic receptors are crucial for their role in rhythmic contraction of heart along with their role in the pathological conditions such as tachycardia and high risk of heart failure. Studies report that the levels of beta-1 adrenergic receptor tend to decrease by 50%, whereas, the levels of beta-2 adrenergic receptor remains constant during the risk of heart failure. Beta blockers-the antagonistic molecules for beta-adrenergic receptors, function by slowing the heart rate, which thereby allows the left ventricle to fill completely during tachycardia incidents and hence helps in blood pumping capacity of heart and reducing the risk of heart failure. In the present study, we investigate the potential of glycyrrhizic acid (GA) as a possible principal drug molecule for cardiac arrhythmias owing to its ability to induce reduction in the heart rate and blood pressure. We use in vitro and in silico approach to study GA's effect on beta adrenergic receptor along with an in vivo study to examine its effect on heart rate and blood pressure. Additionally, we explore GA's proficiency in eliciting an increase in the plasma levels of vasoactive intestinal peptide, which by dilating the blood vessel consequently, can be a crucial aid during the occurrence of a potential heart attack. Therefore, we propose GA as a potential principal drug molecule via its potential in modulating heart rate and blood pressure.

  19. Molecular Mechanism of AMPA Receptor Modulation by TARP/Stargazin.

    PubMed

    Ben-Yaacov, Anat; Gillor, Moshe; Haham, Tomer; Parsai, Alon; Qneibi, Mohammad; Stern-Bach, Yael

    2017-03-08

    AMPA receptors (AMPARs) mediate the majority of fast excitatory transmission in the brain and critically contribute to synaptic plasticity and pathology. AMPAR trafficking and gating are tightly controlled by auxiliary transmembrane AMPAR regulatory proteins (TARPs). Here, using systematic domain swaps with the TARP-insensitive kainate receptor GluK2, we show that AMPAR interaction with the prototypical TARP stargazin/γ2 primarily involves the AMPAR membrane domains M1 and M4 of neighboring subunits, initiated or stabilized by the AMPAR C-tail, and that these interactions are sufficient to enable full receptor modulation. Moreover, employing TARP chimeras disclosed a key role in this process also for the TARP transmembrane domains TM3 and TM4 and extracellular loop 2. Mechanistically, our data support a two-step action in which binding of TARP to the AMPAR membrane domains destabilizes the channel closed state, thereby enabling an efficient opening upon agonist binding, which then stabilizes the open state via subsequent interactions.

  20. Glucocorticoid hormone resistance during primate evolution: receptor-mediated mechanisms.

    PubMed

    Chrousos, G P; Renquist, D; Brandon, D; Eil, C; Pugeat, M; Vigersky, R; Cutler, G B; Loriaux, D L; Lipsett, M B

    1982-03-01

    The concentrations of total and protein-unbound plasma cortisol of New World monkeys are higher than those of Old World primates and prosimians. The urinary free-cortisol excretion also is increased markedly. However, there is no physiologic evidence of increased cortisol effect. These findings suggest end-organ resistance to glucocorticoids. This was confirmed by showing that the hypothalamic-pituitary adrenal axis is resistant to suppression by dexamethasone. To study this phenomenon, glucocorticoid receptors were examined in circulating mononuclear leukocytes and cultured skin fibroblasts from both New and Old World species. The receptor content is the same in all species, but the New World monkeys have a markedly decreased binding affinity for dexamethasone. Thus, the resistance of these species to the action of cortisol is due to the decreased binding affinity of the glucocorticoid receptor. This presumed mutation must have occurred after the bifurcation of Old and New World primates (approximately 60 x 10(6) yr ago) and before the diversion of the New World primates from each other (approximately 15 x 10(6) yr ago).

  1. Structural Mapping and Functional Characterization of Zebrafish Class B G-Protein Coupled Receptor (GPCR) with Dual Ligand Selectivity towards GLP-1 and Glucagon

    PubMed Central

    Oren, Deena A.; Wei, Yang; Skrabanek, Luce; Chow, Billy K. C.; Mommsen, Thomas; Mojsov, Svetlana

    2016-01-01

    GLP-1 and glucagon regulate glucose metabolism through a network of metabolic pathways initiated upon binding to their specific receptors that belong to class B G-protein coupled receptors (GPCRs). The therapeutic potential of glucagon is currently being evaluated, while GLP-1 is already used in the treatment of type 2 diabetes and obesity. Development of a second generation of GLP-1 based therapeutics depends on a molecular and structural understanding of the interactions between the GLP-1 receptor (GLP-1R) and its ligand GLP-1. There is considerable sequence conservation between GLP-1 and glucagon and between the hGLP-1R and human glucagon receptor (hGCGR), yet each receptor recognizes only its own specific ligand. Glucagon receptors in fish and frogs also exhibit ligand selectivity only towards glucagon and not GLP-1. Based on competitive binding experiments and assays of increase in intracellular cAMP, we demonstrate here that a GPCR in zebrafish (Danio rerio) exhibits dual ligand selectivity towards GLP-1 and glucagon, a characteristic not found in mammals. Further, many structural features found in hGLP-1R and hGCGR are also found in this zebrafish GPCR (zfGPCR). We show this by mapping of its sequence and structural features onto the hGLP-1R and hGCGR based on their partial and complementary crystal structures. Thus, we propose that zfGPCR represents a dual GLP-1R/GCGR. The main differences between the three receptors are in their stalk regions that connect their N-terminal extracellular domains (NECDs) with their transmembrane domains and the absence of loop 3 in the NECD in zfGLP-1R/GCGR. These observations suggest that the interactions between GLP-1 and glucagon with loop 3 and the stalk regions may induce different conformational changes in hGLP-1R and hGCGR upon ligand binding and activation that lead to selective recognition of their native ligands. PMID:27930690

  2. The mechansims by which solute nitrogen affects phase transformations and mechanical properties of automotive dual-phase sheet steel

    NASA Astrophysics Data System (ADS)

    Brown, Tyson W.

    Dual-phase steels have seen increased use in automotive applications in recent years, in order to meet the goals of weight reduction and occupant safety. Variations in nitrogen content that may be encountered in steel sourced from a basic oxygen furnace process compared to an electric arc furnace process require that dual-phase steel producers understand the ways that nitrogen affects processing and properties. In the current work, the distribution of nitrogen was investigated in a dual-phase steel with a base chemistry of 0.1 C, 2.0 Mn, 0.2 Cr, 0.2 Mo (wt pct) across a range of nitrogen contents (30-159 ppm) with Al (0.2 and 0.08 wt pct), and Ti (0.02 wt pct) additions used for precipitation control of nitrogen amounts. The distribution of nitrogen amongst trapping sites, including precipitates, grain boundaries, dislocations, and interstitial sites (away from other types of defects) was determined from a combination of electrolytic dissolution, internal friction, and three-dimensional atom probe tomography experiments. Various mechanisms by which different amounts and locations of nitrogen affect phase transformations and mechanical properties were identified from quantitative metallography, dilatometric measurement of phase transformations, tensile testing, and nanoindentation hardness testing. Results indicate nitrogen that is not precipitated with Ti or Al (free nitrogen) partitions to austenite (and thus martensite) during typical intercritical annealing treatments, and is mostly contained in Cottrell atmospheres in martensite. Due to the austenite stabilizing effect of nitrogen, the presence of free nitrogen during intercritical annealing leads to a higher austenite fraction in certain conditions. Thus, the presence of free nitrogen in a dual-phase microstructure will lead to an increase in tensile and yield strengths from both an increase in martensite fraction, and an increase in martensite hardness due to solid solution strengthening. Despite the presence

  3. Hybrid Electrochemical Mechanical Planarization Process for Cu Dual-Damascene Through-Silicon Via Using Noncontact Electrode Pad

    NASA Astrophysics Data System (ADS)

    Shigeru Tominaga,; Daisuke Abe,; Taro Enomoto,; Seiichi Kondo,; Hideki Kitada,; Takayuki Ohba,

    2010-05-01

    A hybrid electrochemical mechanical planarization and chemical mechanical planarization (e-CMP/CMP) was applied to the Cu dual-damascene through-silicon via (TSV) process for wafer-level three-dimensional integrated circuit (3D-IC) stacking. In this process, an electrochemically deposited Cu film was removed by e-CMP at a removal rate of 3.5 μm/min until the voltage endpoint was detected. Then, residual Cu film was polished off in the CMP mode using the same e-CMP pad. A fine Cu damascene structure was successfully fabricated with a dishing depth of less than 200 nm in a metal pad of 200× 200 μm2 area. The criterion of dishing without failure in the adhesive coat for 3D-IC stacking is discussed.

  4. Hybrid Electrochemical Mechanical Planarization Process for Cu Dual-Damascene Through-Silicon Via Using Noncontact Electrode Pad

    NASA Astrophysics Data System (ADS)

    Tominaga, Shigeru; Abe, Daisuke; Enomoto, Taro; Kondo, Seiichi; Kitada, Hideki; Ohba, Takayuki

    2010-05-01

    A hybrid electrochemical mechanical planarization and chemical mechanical planarization (e-CMP/CMP) was applied to the Cu dual-damascene through-silicon via (TSV) process for wafer-level three-dimensional integrated circuit (3D-IC) stacking. In this process, an electrochemically deposited Cu film was removed by e-CMP at a removal rate of 3.5 µm/min until the voltage endpoint was detected. Then, residual Cu film was polished off in the CMP mode using the same e-CMP pad. A fine Cu damascene structure was successfully fabricated with a dishing depth of less than 200 nm in a metal pad of 200×200 µm2 area. The criterion of dishing without failure in the adhesive coat for 3D-IC stacking is discussed.

  5. ``Lock and key mechanism'' for ligand binding with adrenergic receptors and the arising mechanical effects on the cell membrane

    NASA Astrophysics Data System (ADS)

    Lunghi, Laura; Deseri, Luca

    2013-03-01

    Chemicals hitting the surface of cell aggregates are known to give arise to cyclic Adenosine Mono Phosphate (cAMP), a second messenger that transduces inside the cell the effects of species that cannot get through the cell membrane. Ligands bind to a specific receptor following the so called ``lock and key mechanism'' (beta)-adrenergic receptors are proteins embedded in the lipid bilayer characterized by seven transmembrane helices. Thinning and thickening in cell membranes may be initiated by conformational changes of some of three of the seven domains above. The cell response is linked to the coupling of chemical, conformational and mechanical effects. Part of the cAMP remains intracellular, whereas the remaining fractions migrates outside the cell due to membrane transporters. A new Helmholtz free energy, accounting for receptor and transporter densities, receptor conformation field and membrane elasticity is investigated. It is shown how the density of active receptors is directly related to the conformation field and it enters the resulting balance equation for the membrane stress. Balance laws for fluxes of transporters and receptors, coupled with the former because of the outgoing cAMP flux caused by the transporters, as well as for the diffusive powers must be supplied. The Center for Nonlinear Analysis through the NSF Grant No. DMS-0635983 is gratefully acknowledged.

  6. Development and Acceptance Testing of the Dual Wheel Mechanism for the Tunable Filter Imager Cryogenic Instrument on the JWST

    NASA Technical Reports Server (NTRS)

    Leckie, Martin; Ahmad, Zakir

    2010-01-01

    The James Webb Space Telescope (JWST) will carry four scientific instruments, one of which is the Tunable Filter Imager (TFI), which is an instrument within the Fine Guidance Sensor. The Dual Wheel (DW) mechanism is being designed, built and tested by COM DEV Ltd. under contract from the Canadian Space Agency. The DW mechanism includes a pupil wheel (PW) holding seven coronagraphic masks and two calibration elements and a filter wheel (FW) holding nine blocking filters. The DW mechanism must operate at both room temperature and at 35K. Successful operation at 35K comprises positioning each optical element with the required repeatability, for several thousand occasions over the five year mission. The paper discusses the results of testing geared motors and bearings at the cryogenic temperature. In particular bearing retainer design and PGM-HT material, the effects of temperature gradients across bearings and the problems associated with cooling mechanisms down to cryogenic temperatures. The results of additional bearing tests are described that were employed to investigate an abnormally high initial torque experienced at cryogenic temperatures. The findings of these tests, was that the bearing retainer and the ball/race system could be adversely affected by the large temperature change from room temperature to cryogenic temperature and also the temperature gradient across the bearing. The DW mechanism is now performing successfully at both room temperature and at cryogenic temperature. The life testing of the mechanism is expected to be completed in the first quarter of 2010.

  7. Ombuin-3-O-β-D-glucopyranoside from Gynostemma pentaphyllum is a dual agonistic ligand of peroxisome proliferator-activated receptors α and δ/β

    SciTech Connect

    Malek, Mastura Abd; Hoang, Minh-Hien; Jia, Yaoyao; Lee, Ji Hae; Jun, Hee Jin; Lee, Dong-Ho; Lee, Hak Ju; Lee, Chul; Lee, Myung Koo; Hwang, Bang Yeon; Lee, Sung-Joon

    2013-01-25

    Highlights: ► Ombuin-3-O-β-D-glucopyranoside is a dual ligand for PPARα and δ/β. ► Ombuin-3-O-β-D-glucopyranoside reduces cellular lipid levels in multiple cell types. ► Cells stimulated with ombuine up-regulated target genes in cholesterol efflux. ► Cells stimulated with ombuine regulated target fatty acid β-oxidation and synthesis. ► Ombuin-3-O-β-D-glucopyranoside could ameliorate hyperlipidemia and hepatic steatosis. -- Abstract: We demonstrated that ombuin-3-O-β-D-glucopyranoside (ombuine), a flavonoid from Gynostemma pentaphyllum, is a dual agonist for peroxisome proliferator-activated receptors (PPARs) α and δ/β. Using surface plasmon resonance (SPR), time-resolved fluorescence resonance energy transfer (FRET) analyses, and reporter gene assays, we showed that ombuine bound directly to PPARα and δ/β but not to PPARγ or liver X receptors (LXRs). Cultured HepG2 hepatocytes stimulated with ombuine significantly reduced intracellular concentrations of triglyceride and cholesterol and downregulated the expression of lipogenic genes, including sterol regulatory element binding protein-1c (SREBP1c) and stearoyl-CoA desaturase-1 (SCD-1), with activation of PPARα and δ/β. Activation of LXRs by ombuine was confirmed by reporter gene assays, however, SPR and cell-based FRET assays showed no direct binding of ombuine to either of the LXRs suggesting LXR activation by ombuine may be operated via PPARα stimulation. Ombuine-stimulated macrophages showed significantly induced transcription of ATP binding cassette cholesterol transporter A1 (ABCA1) and G1 (ABCG1), the key genes in reverse cholesterol transport, which led to reduced cellular cholesterol concentrations. These results suggest that ombuine is a dual PPAR ligand for PPARα and δ/β with the ability to decrease lipid concentrations by reducing lipogenic gene expression in hepatocytes and inducing genes involved in cholesterol efflux in macrophages.

  8. Allosteric Modulators of GABAB Receptors: Mechanism of Action and Therapeutic Perspective

    PubMed Central

    Pin, Jean-Philippe; Prézeau, Laurent

    2007-01-01

    γ-aminobutyric acid (GABA) plays important roles in the central nervous system, acting as a neurotransmitter on both ionotropic ligand-gated Cl--channels, and metabotropic G-protein coupled receptors (GPCRs). These two types of receptors called GABAA (and C) and GABAB are the targets of major therapeutic drugs such as the anxiolytic benzodiazepines, and antispastic drug baclofen (lioresal®), respectively. Although the multiplicity of GABAA receptors offer a number of possibilities to discover new and more selective drugs, the molecular characterization of the GABAB receptor revealed a unique, though complex, heterodimeric GPCR. High throughput screening strategies carried out in pharmaceutical industries, helped identifying new compounds positively modulating the activity of the GABAB receptor. These molecules, almost devoid of apparent activity when applied alone, greatly enhance both the potency and efficacy of GABAB agonists. As such, in contrast to baclofen that constantly activates the receptor everywhere in the brain, these positive allosteric modulators induce a large increase in GABAB-mediated responses only WHERE and WHEN physiologically needed. Such compounds are then well adapted to help GABA to activate its GABAB receptors, like benzodiazepines favor GABAA receptor activation. In this review, the way of action of these molecules will be presented in light of our actual knowledge of the activation mechanism of the GABAB receptor. We will then show that, as expected, these molecules have more pronounced in vivo responses and less side effects than pure agonists, offering new potential therapeutic applications for this new class of GABAB ligands. PMID:19305802

  9. Discovery of a Series of Imidazo[4,5-b]pyridines with Dual Activity at Angiotensin II Type 1 Receptor and Peroxisome Proliferator-Activated Receptor-[gamma

    SciTech Connect

    Casimiro-Garcia, Agustin; Filzen, Gary F.; Flynn, Declan; Bigge, Christopher F.; Chen, Jing; Davis, Jo Ann; Dudley, Danette A.; Edmunds, Jeremy J.; Esmaeil, Nadia; Geyer, Andrew; Heemstra, Ronald J.; Jalaie, Mehran; Ohren, Jeffrey F.; Ostroski, Robert; Ellis, Teresa; Schaum, Robert P.; Stoner, Chad

    2013-03-07

    Mining of an in-house collection of angiotensin II type 1 receptor antagonists to identify compounds with activity at the peroxisome proliferator-activated receptor-{gamma} (PPAR{gamma}) revealed a new series of imidazo[4,5-b]pyridines 2 possessing activity at these two receptors. Early availability of the crystal structure of the lead compound 2a bound to the ligand binding domain of human PPAR{gamma} confirmed the mode of interaction of this scaffold to the nuclear receptor and assisted in the optimization of PPAR{gamma} activity. Among the new compounds, (S)-3-(5-(2-(1H-tetrazol-5-yl)phenyl)-2,3-dihydro-1H-inden-1-yl)-2-ethyl-5-isobutyl-7-methyl-3H-imidazo[4,5-b]pyridine (2l) was identified as a potent angiotensin II type I receptor blocker (IC{sub 50} = 1.6 nM) with partial PPAR{gamma} agonism (EC{sub 50} = 212 nM, 31% max) and oral bioavailability in rat. The dual pharmacology of 2l was demonstrated in animal models of hypertension (SHR) and insulin resistance (ZDF rat). In the SHR, 2l was highly efficacious in lowering blood pressure, while robust lowering of glucose and triglycerides was observed in the male ZDF rat.

  10. Discovery of a series of imidazo[4,5-b]pyridines with dual activity at angiotensin II type 1 receptor and peroxisome proliferator-activated receptor-γ.

    PubMed

    Casimiro-Garcia, Agustin; Filzen, Gary F; Flynn, Declan; Bigge, Christopher F; Chen, Jing; Davis, Jo Ann; Dudley, Danette A; Edmunds, Jeremy J; Esmaeil, Nadia; Geyer, Andrew; Heemstra, Ronald J; Jalaie, Mehran; Ohren, Jeffrey F; Ostroski, Robert; Ellis, Teresa; Schaum, Robert P; Stoner, Chad

    2011-06-23

    Mining of an in-house collection of angiotensin II type 1 receptor antagonists to identify compounds with activity at the peroxisome proliferator-activated receptor-γ (PPARγ) revealed a new series of imidazo[4,5-b]pyridines 2 possessing activity at these two receptors. Early availability of the crystal structure of the lead compound 2a bound to the ligand binding domain of human PPARγ confirmed the mode of interaction of this scaffold to the nuclear receptor and assisted in the optimization of PPARγ activity. Among the new compounds, (S)-3-(5-(2-(1H-tetrazol-5-yl)phenyl)-2,3-dihydro-1H-inden-1-yl)-2-ethyl-5-isobutyl-7-methyl-3H-imidazo[4,5-b]pyridine (2l) was identified as a potent angiotensin II type I receptor blocker (IC(50) = 1.6 nM) with partial PPARγ agonism (EC(50) = 212 nM, 31% max) and oral bioavailability in rat. The dual pharmacology of 2l was demonstrated in animal models of hypertension (SHR) and insulin resistance (ZDF rat). In the SHR, 2l was highly efficacious in lowering blood pressure, while robust lowering of glucose and triglycerides was observed in the male ZDF rat.

  11. Extracellular matrix hyaluronan signals via its CD44 receptor in the increased responsiveness to mechanical stimulation.

    PubMed

    Ferrari, L F; Araldi, D; Bogen, O; Levine, J D

    2016-06-02

    We propose that the extracellular matrix (ECM) signals CD44, a hyaluronan receptor, to increase the responsiveness to mechanical stimulation in the rat hind paw. We report that intradermal injection of hyaluronidase induces mechanical hyperalgesia, that is inhibited by co-administration of a CD44 receptor antagonist, A5G27. The intradermal injection of low (LMWH) but not high (HMWH) molecular weight hyaluronan also induces mechanical hyperalgesia, an effect that was attenuated by pretreatment with HMWH or A5G27. Pretreatment with HMWH also attenuated the hyperalgesia induced by hyaluronidase. Similarly, intradermal injection of A6, a CD44 receptor agonist, produced hyperalgesia that was inhibited by HMWH and A5G27. Inhibitors of protein kinase A (PKA) and Src, but not protein kinase C (PKC), significantly attenuated the hyperalgesia induced by both A6 and LMWH. Finally, to determine if CD44 receptor signaling is involved in a preclinical model of inflammatory pain, we evaluated the effect of A5G27 and HMWH on the mechanical hyperalgesia associated with the inflammation induced by carrageenan. Both A5G27 and HMWH attenuated carrageenan-induced mechanical hyperalgesia. Thus, while LMWH acts at its cognate receptor, CD44, to induce mechanical hyperalgesia, HMWH acts at the same receptor as an antagonist. That the local administration of HMWH or A5G27 inhibits carrageenan-induced hyperalgesia supports the suggestion that carrageenan produces changes in the ECM that contributes to inflammatory pain. These studies define a clinically relevant role for signaling by the hyaluronan receptor, CD44, in increased responsiveness to mechanical stimulation.

  12. GPCR responses in vascular smooth muscle can occur predominantly through dual transactivation of kinase receptors and not classical Gαq protein signalling pathways.

    PubMed

    Little, Peter J

    2013-05-30

    GPCR signalling is well known to proceed through several linear pathways involving activation of G proteins and their downstream signalling pathways such as activation of phospholipase C. In addition, GPCRs signal via transactivation of Protein Tyrosine Kinase receptors such as that for Epidermal Growth Factor (EGF) and Platelet-Derived Growth Factor (PDGF) where GPCR agonists mediate increase levels of phosphorylated Erk (pErk) the immediate downstream product of the activation of EGF receptor. It has recently been shown that this paradigm can be extended to include the GPCR transactivation of a Protein Serine/Threonine Kinase receptor, specifically the Transforming Growth Factor β Type I receptor (also known as Alk V) (TβRI) in which case GPCR activation leads to the formation of carboxy terminal polyphosphorylated Smad2 (phosphoSmad2) being the immediate downstream product of the activation of TβRI. Growth factor and hormone regulation of proteoglycan synthesis in vascular smooth muscle cells represent one component of an in vitro model of atherosclerosis because modified proteoglycans show enhanced binding to lipoproteins as the initiating step in atherosclerosis. In the example of proteoglycan synthesis stimulated by GPCR agonists such as thrombin and endothelin-1, the transactivation pathways for the EGF receptor and TβRI are both active and together account for essentially all of the response to the GPCRs. In contrast, signalling downstream of GPCRs such as increased inositol 1,4,5 trisphosphate (IP3) and intracellular calcium do not have any effect on GPCR stimulated proteoglycan synthesis. These data lead to the conclusion that dual transactivation pathways for protein tyrosine and serine/threonine kinase receptors may play a far greater role in GPCR signalling than currently recognised.

  13. Identification of the transmitter and receptor mechanisms responsible for REM sleep paralysis.

    PubMed

    Brooks, Patricia L; Peever, John H

    2012-07-18

    During REM sleep the CNS is intensely active, but the skeletal motor system is paradoxically forced into a state of muscle paralysis. The mechanisms that trigger REM sleep paralysis are a matter of intense debate. Two competing theories argue that it is caused by either active inhibition or reduced excitation of somatic motoneuron activity. Here, we identify the transmitter and receptor mechanisms that function to silence skeletal muscles during REM sleep. We used behavioral, electrophysiological, receptor pharmacology and neuroanatomical approaches to determine how trigeminal motoneurons and masseter muscles are switched off during REM sleep in rats. We show that a powerful GABA and glycine drive triggers REM paralysis by switching off motoneuron activity. This drive inhibits motoneurons by targeting both metabotropic GABA(B) and ionotropic GABA(A)/glycine receptors. REM paralysis is only reversed when motoneurons are cut off from GABA(B), GABA(A) and glycine receptor-mediated inhibition. Neither metabotropic nor ionotropic receptor mechanisms alone are sufficient for generating REM paralysis. These results demonstrate that multiple receptor mechanisms trigger REM sleep paralysis. Breakdown in normal REM inhibition may underlie common sleep motor pathologies such as REM sleep behavior disorder.

  14. Mechanisms of disease: Toll-like receptors in cardiovascular disease.

    PubMed

    Frantz, Stefan; Ertl, Georg; Bauersachs, Johann

    2007-08-01

    The innate immune system detects highly conserved, relatively invariant structural motifs of pathogens. Toll-like receptors (TLRs) have been identified as the primary innate immune receptors. TLRs distinguish between different patterns of pathogens and activate a rapid innate immune response; however, TLRs can also be activated by host-derived molecules. In addition to being expressed in immune cells, TLRs are expressed in other tissues, such as those of the cardiovascular system. TLRs could, therefore, be a key link between cardiovascular disease development and the immune system. Indeed, evidence that TLR activation contributes to the development and progression of atherosclerosis, cardiac dysfunction in sepsis, and congestive heart failure, is convincing. Although much has been learned about TLR activation in cellular components of the cardiovascular system, the role individual TLR family members have in the pathophysiology of cardiovascular diseases and hence in clinical practice remains to be defined. Here we review the rapid progress that has been made in this field, which has improved our understanding of vascular as well as myocardial TLR function in basic and clinical science.

  15. A dual model of entertainment-based and community-based mechanisms to explore continued participation in online entertainment communities.

    PubMed

    Deng, Yun; Hou, Jinghui; Ma, Xiao; Cai, Shuqin

    2013-05-01

    Online entertainment communities have exploded in popularity and drawn attention from researchers. However, few studies have investigated what leads people to remain active in such communities at the postadoption stage. We proposed and tested a dual model of entertainment-based and community-based mechanisms to examine the factors that affect individuals' continued participation in online entertainment communities. Survival analysis was employed on a longitudinal dataset of 2,302 users collected over 2 years from an online game community. Our results were highly consistent with the theoretical model. Specifically, under the entertainment-based mechanism, our findings showed that the intensities of initial use and frequent use were positive predictors of players' activity lifespan. Under the community-based mechanism, the results demonstrated that the number of guilds a player was affiliated with and the average number of days of being a guild member positively predict players' lifespan in the game. Overall, our study suggests that the entertainment-based mechanism and community-based mechanism are two key drivers that determinate individuals' continued participation in online entertainment communities.

  16. Ligand Binding Mechanism in Steroid Receptors: From Conserved Plasticity to Differential Evolutionary Constraints.

    PubMed

    Edman, Karl; Hosseini, Ali; Bjursell, Magnus K; Aagaard, Anna; Wissler, Lisa; Gunnarsson, Anders; Kaminski, Tim; Köhler, Christian; Bäckström, Stefan; Jensen, Tina J; Cavallin, Anders; Karlsson, Ulla; Nilsson, Ewa; Lecina, Daniel; Takahashi, Ryoji; Grebner, Christoph; Geschwindner, Stefan; Lepistö, Matti; Hogner, Anders C; Guallar, Victor

    2015-12-01

    Steroid receptor drugs have been available for more than half a century, but details of the ligand binding mechanism have remained elusive. We solved X-ray structures of the glucocorticoid and mineralocorticoid receptors to identify a conserved plasticity at the helix 6-7 region that extends the ligand binding pocket toward the receptor surface. Since none of the endogenous ligands exploit this region, we hypothesized that it constitutes an integral part of the binding event. Extensive all-atom unbiased ligand exit and entrance simulations corroborate a ligand binding pathway that gives the observed structural plasticity a key functional role. Kinetic measurements reveal that the receptor residence time correlates with structural rearrangements observed in both structures and simulations. Ultimately, our findings reveal why nature has conserved the capacity to open up this region, and highlight how differences in the details of the ligand entry process result in differential evolutionary constraints across the steroid receptors.

  17. Diverse binding modes, same goal: the receptor recognition mechanism of botulinum neurotoxin

    PubMed Central

    Lam, Kwok-Ho; Yao, Guorui; Jin, Rongsheng

    2015-01-01

    Botulinum neurotoxins (BoNTs) are among the most deadly toxins known. They act rapidly in a highly specific manner to block neurotransmitter release by cleaving the soluble N-ethylmaleimide sensitive factor attachment protein receptor (SNARE) complex at neuromuscular junctions. The extreme toxicity of BoNTs relies predominantly on their neurotropism that is accomplished by recognition of two host receptors, a polysialo-ganglioside and in the majority of cases a synaptic vesicle protein, through their receptor-binding domains. Two proteins, synaptotagmin and synaptic vesicle glycoprotein 2, have been identified as the receptors for various serotypes of BoNTs. Here, we review recent breakthroughs in the structural studies of BoNT–protein receptor recognitions that highlight a range of diverse mechanisms by which BoNTs manipulate host neuronal proteins for highly specific uptake at neuromuscular junctions. PMID:25701633

  18. Understanding the deposition mechanism of pulsed laser deposited B-C films using dual-targets

    SciTech Connect

    Zhang, Song; He, Zhiqiang; Wang, Chuanbin; Shen, Qiang; Zhang, Lianmeng; Ji, Xiaoli; Lu, Wenzhong

    2014-04-21

    Boron carbide thin films with stoichiometry (boron-carbon atomic ratio) range of 0.1 ∼ 8.9 were fabricated via pulsed laser deposition by using boron-carbon dual-targets. However, this experimental data on stoichiometry were smaller than the computer simulation values. The discrepancy was investigated by studies on composition and microstructure of the thin films and targets by scanning electron microscopy, excitation laser Raman spectroscopy, and X-ray photoelectron spectroscopy. The results indicate that the boron liquid droplets were formed by phase explosion after laser irradiation on boron sector. Part of the boron droplets would be lost via ejection in the direction of laser beam, which is tilted 45° to the surface of substrate.

  19. Synthesis and Anticancer Mechanism Investigation of Dual Hsp27 and Tubulin Inhibitors

    PubMed Central

    Zhong, Bo; Chennamaneni, Snigdha; Lama, Rati; Yi, Xin; Geldenhuys, Werner J.; Pink, John J.; Dowlati, Afshin; Xu, Yan; Zhou, Aimin; Su, Bin

    2013-01-01

    Heat shock protein 27 (Hsp27) is a chaperone protein, and its expression is increased in response to various stress stimuli including anticancer chemotherapy, which allows the cells to survive and causes drug resistance. We previously identified lead compounds that bound to Hsp27 and tubulin via proteomic approaches. Systematic ligand based optimization in the current study significantly increased the cell growth inhibition and apoptosis inducing activities of the compounds. Compared to the lead compounds, one of the new derivatives exhibited much better potency to inhibit tubulin polymerization but a decreased activity to inhibit Hsp27 chaperone function, suggesting that the structural modification dissected the dual targeting effects of the compound. The most potent compounds 20 and 22 exhibited strong cell proliferation inhibitory activities at subnanomolar concentration against 60 human cancer cell lines conducted by Developmental Therapeutic Program at the National Cancer Institute and represented promising candidates for anticancer drug development. PMID:23767669

  20. Study on mechanism of amplitude fluctuation of dual-frequency beat in microchip Nd:YAG laser

    NASA Astrophysics Data System (ADS)

    Chen, Hao; Tan, Yidong; Zhang, Shulian; Sun, Liqun

    2017-01-01

    In the laser heterodyne interferometry based on the microchip Nd:YAG dual-frequency laser, the amplitude of the beat note periodically fluctuates in time domain, which leads to the instability of the measurement. On the frequency spectrums of the two mono-frequency components of the laser and their beat note, several weak sideband signals are observed on both sides of the beat note. It is proved that the sideband frequencies are associated with the relaxation oscillation frequencies of the laser. The mechanism for the relaxation oscillations inducing the occurrence of the sideband signals is theoretically analyzed, and the quantitative relationship between the intensity ratio of the beat note to the sideband signal and the level of the amplitude fluctuation is simulated with the derived mathematical model. The results demonstrate that the periodical amplitude fluctuation of the beat note is actually induced by the relaxation oscillation. And the level of the amplitude fluctuation is lower than 10% when the intensity ratio is greater than 32 dB. These conclusions are beneficial to reduce the amplitude fluctuation of the microchip Nd:YAG dual-frequency laser and improve the stability of the heterodyne interferometry.

  1. Mechanisms of the adenosine A2A receptor-induced sensitization of esophageal C fibers.

    PubMed

    Brozmanova, M; Mazurova, L; Ru, F; Tatar, M; Hu, Y; Yu, S; Kollarik, M

    2016-02-01

    Clinical studies indicate that adenosine contributes to esophageal mechanical hypersensitivity in some patients with pain originating in the esophagus. We have previously reported that the esophageal vagal nodose C fibers express the adenosine A2A receptor. Here we addressed the hypothesis that stimulation of the adenosine A2A receptor induces mechanical sensitization of esophageal C fibers by a mechanism involving transient receptor potential A1 (TRPA1). Extracellular single fiber recordings of activity originating in C-fiber terminals were made in the ex vivo vagally innervated guinea pig esophagus. The adenosine A2A receptor-selective agonist CGS21680 induced robust, reversible sensitization of the response to esophageal distention (10-60 mmHg) in a concentration-dependent fashion (1-100 nM). At the half-maximally effective concentration (EC50: ≈3 nM), CGS21680 induced an approximately twofold increase in the mechanical response without causing an overt activation. This sensitization was abolished by the selective A2A antagonist SCH58261. The adenylyl cyclase activator forskolin mimicked while the nonselective protein kinase inhibitor H89 inhibited mechanical sensitization by CGS21680. CGS21680 did not enhance the response to the purinergic P2X receptor agonist α,β-methylene-ATP, indicating that CGS21680 does not nonspecifically sensitize to all stimuli. Mechanical sensitization by CGS21680 was abolished by pretreatment with two structurally different TRPA1 antagonists AP18 and HC030031. Single cell RT-PCR and whole cell patch-clamp studies in isolated esophagus-specific nodose neurons revealed the expression of TRPA1 in A2A-positive C-fiber neurons and demonstrated that CGS21682 potentiated TRPA1 currents evoked by allylisothiocyanate. We conclude that stimulation of the adenosine A2A receptor induces mechanical sensitization of nodose C fibers by a mechanism sensitive to TRPA1 antagonists indicating the involvement of TRPA1.

  2. [Tapentadol is a new, strongly efficative analgeticum with dual effect mechanisms].

    PubMed

    Staahl, Camilla; Drewes, Asbjørn Mohr; Jensen, Niels-Henrik

    2011-06-20

    Tapentadol exerts its analgesic effects through opioid receptor agonism and noradrenaline reuptake inhibition in the central nervous system. Clinical studies show that tapentadol effectively relieves moderate to severe pain in both post-operative and chronic pain. In these trials with equianalgesic doses of tapentadol and oxycodone, treatment with tapentadol was associated with significantly fewer gastrointestinal-related adverse events. Furthermore, in a placebo-controlled study, tapentadol has shown good efficacy in painful diabetic polyneuropathy.

  3. Potentiation of morphine-induced mechanical antinociception by σ₁ receptor inhibition: role of peripheral σ₁ receptors.

    PubMed

    Sánchez-Fernández, Cristina; Nieto, Francisco Rafael; González-Cano, Rafael; Artacho-Cordón, Antonia; Romero, Lucía; Montilla-García, Ángeles; Zamanillo, Daniel; Baeyens, José Manuel; Entrena, José Manuel; Cobos, Enrique José

    2013-07-01

    We studied the modulation of morphine-induced mechanical antinociception and side effects by σ₁ receptor inhibition. Both wild-type (WT) and σ₁ receptor knockout (σ₁-KO) mice showed similar responses to paw pressure (100-600 g). The systemic (subcutaneous) or local (intraplantar) administration of σ₁ antagonists (BD-1063, BD-1047, NE-100 and S1RA) was devoid of antinociceptive effects in WT mice. However, σ₁-KO mice exhibited an enhanced mechanical antinociception in response to systemic morphine (1-16 mg/kg). Similarly, systemic treatment of WT mice with σ₁ antagonists markedly potentiated morphine-induced antinociception, and its effects were reversed by the selective σ₁ agonist PRE-084. Although the local administration of morphine (50-200 μg) was devoid of antinociceptive effects in WT mice, it induced dose-dependent antinociception in σ₁-KO mice. This effect was limited to the injected paw. Enhancement of peripheral morphine antinociception was replicated in WT mice locally co-administered with σ₁ antagonists and the opioid. None of the σ₁ antagonists tested enhanced morphine-antinociception in σ₁-KO mice, confirming a σ₁-mediated action. Morphine-induced side-effects (hyperlocomotion and inhibition of gastrointestinal transit) were unaltered in σ₁-KO mice. These results cannot be explained by a direct interaction of σ₁ ligands with μ-opioid receptors or adaptive changes of μ-receptors in σ₁-KO mice, given that [(3)H]DAMGO binding in forebrain, spinal cord, and hind-paw skin membranes was unaltered in mutant mice, and none of the σ₁ drugs tested bound to μ-opioid receptors. These results show that σ₁ receptor inhibition potentiates morphine-induced mechanical analgesia but not its acute side effects, and that this enhanced analgesia can be induced at peripheral level.

  4. Effects and mechanism of dual-frequency power ultrasound on the molecular weight distribution of corn gluten meal hydrolysates.

    PubMed

    Jin, Jian; Ma, Haile; Wang, Bei; Yagoub, Abu El-Gasim A; Wang, Kai; He, Ronghai; Zhou, Cunshan

    2016-05-01

    The impact of dual-frequency power ultrasound (DPU) on the molecular weight distribution (MWD) of corn gluten meal (CGM) hydrolysates and its mechanism were investigated in the present study. The mechanism was studied from aspects of structural and nano-mechanical characteristics of the major protein fractions of CGM, viz. zein and glutelin. The results of molecular weight distribution indicated that DPU pretreatment of CGM was beneficial to the preparation of peptides with molecular weights of 200-1000Da. Moreover, FTIR spectral analysis and atomic force microscopy characterization showed that the DPU pretreatment changed the contents of secondary structure of proteins, decreased the particle height and surface roughness of glutelin, reduced the Young's modulus and stiffness of zein while increased its adhesion force. In conclusion, DPU pretreatment of proteins before proteolysis is an efficient alternative method to produce short-chain peptides because of its positive effects originating from acoustic cavitation on the molecular conformation, nano-structures and nano-mechanical properties of proteins as well.

  5. Direct generation of 128-fs Gaussian pulses from a compensation-free fiber laser using dual mode-locking mechanisms

    NASA Astrophysics Data System (ADS)

    Peng, Junsong; Zhan, Li; Gu, Zhaochang; Qian, Kai; Luo, Shouyu; Shen, Qishun

    2012-03-01

    We have experimentally demonstrated the direct generation of 128-fs pulses in an all-anomalous-dispersion all-fiber mode-locked laser. The laser is free of dispersion compensation in the cavity based on standard single mode fiber (SMF). The time-bandwidth product is 0.536. The laser is achieved by using two mode-lockers, one is nonlinear polarization rotation (NPR), and the other is nonlinear amplifying loop mirror. The coexistence of dual mode-locking mechanisms can decrease the cavity length to 12-m, and also results in producing high-quality pulses with a Gaussian shape both on the pulse profile and spectrum, but without Kelly sidebands.

  6. Tumor necrosis factor inhibits ligand-stimulated EGF receptor activation through a TNF receptor 1-dependent mechanism

    PubMed Central

    McElroy, Steven J.; Frey, Mark R.; Yan, Fang; Edelblum, Karen L.; Goettel, Jeremy A.; John, Sutha; Polk, D. Brent

    2008-01-01

    Tumor necrosis factor (TNF) and epidermal growth factor (EGF) are key regulators in the intricate balance maintaining intestinal homeostasis. Previous work from our laboratory shows that TNF attenuates ligand-driven EGF receptor (EGFR) phosphorylation in intestinal epithelial cells. To identify the mechanisms underlying this effect, we examined EGFR phosphorylation in cells lacking individual TNF receptors. TNF attenuated EGF-stimulated EGFR phosphorylation in wild-type and TNFR2−/−, but not TNFR1−/−, mouse colon epithelial (MCE) cells. Reexpression of wild-type TNFR1 in TNFR1−/− MCE cells rescued TNF-induced EGFR inhibition, but expression of TNFR1 deletion mutant constructs lacking the death domain (DD) of TNFR1 did not, implicating this domain in EGFR downregulation. Blockade of p38 MAPK, but not MEK, activation of ERK rescued EGF-stimulated phosphorylation in the presence of TNF, consistent with the ability of TNFR1 to stimulate p38 phosphorylation. TNF promoted p38-dependent EGFR internalization in MCE cells, suggesting that desensitization is achieved by reducing receptor accessible to ligand. Taken together, these data indicate that TNF activates TNFR1 by DD- and p38-dependent mechanisms to promote EGFR internalization, with potential impact on EGF-induced proliferation and migration key processes that promote healing in inflammatory intestinal diseases. PMID:18467504

  7. An Fcγ receptor-dependent mechanism drives antibody-mediated target-receptor signaling in cancer cells.

    PubMed

    Wilson, Nicholas S; Yang, Becky; Yang, Annie; Loeser, Stefanie; Marsters, Scot; Lawrence, David; Li, Yun; Pitti, Robert; Totpal, Klara; Yee, Sharon; Ross, Sarajane; Vernes, Jean-Michel; Lu, Yanmei; Adams, Cam; Offringa, Rienk; Kelley, Bob; Hymowitz, Sarah; Daniel, Dylan; Meng, Gloria; Ashkenazi, Avi

    2011-01-18

    Antibodies to cell-surface antigens trigger activatory Fcγ receptor (FcγR)-mediated retrograde signals in leukocytes to control immune effector functions. Here, we uncover an FcγR mechanism that drives antibody-dependent forward signaling in target cells. Agonistic antibodies to death receptor 5 (DR5) induce cancer-cell apoptosis and are in clinical trials; however, their mechanism of action in vivo is not fully defined. Interaction of the DR5-agonistic antibody drozitumab with leukocyte FcγRs promoted DR5-mediated tumor-cell apoptosis. Whereas the anti-CD20 antibody rituximab required activatory FcγRs for tumoricidal function, drozitumab was effective in the context of either activatory or inhibitory FcγRs. A CD40-agonistic antibody required similar FcγR interactions to stimulate nuclear factor-κB activity in B cells. Thus, FcγRs can drive antibody-mediated receptor signaling in target cells.

  8. Functional receptor coupling to Gi is a mechanism of agonist-promoted desensitization of the beta2-adrenergic receptor.

    PubMed

    Tepe, N M; Liggett, S B

    2000-01-01

    The beta2-adrenergic receptor (beta2AR) couples to Gs activating adenylyl cyclase (AC) and increasing cAMP. Such signaling undergoes desensitization with continued agonist exposure. Beta2AR also couple to Gi after receptor phosphorylation by the cAMP dependent protein kinase A, but the efficiency of such coupling is not known. Given the PKA dependence of beta2AR-Gi coupling, we explored whether this may be a mechanism of agonist-promoted desensitization. HEK293 cells were transfected to express beta2AR or beta2AR and Gialpha2, and then treated with vehicle or the agonist isoproterenol to evoke agonist-promoted beta2AR desensitization. Membrane AC activities showed that Gialpha2 overexpression decreased basal levels, but the fold-stimulation of the AC over basal by agonist was not altered. However, with treatment of the cells with isoproterenol prior to membrane preparation, a marked decrease in agonist-stimulated AC was observed with the cells overexpressing Gialpha2. In the absence of such overexpression, beta2AR desensitization was 23+/-7%, while with 5-fold Gialpha2 overexpression desensitization was 58+/-5% (p<0.01, n=4). The effect of Gi on desensitization was receptor-specific, in that forskolin responses were not altered by G(i)alpha2 overexpression. Thus, acquired beta2AR coupling to Gi is an important mechanism of agonist-promoted desensitization, and pathologic conditions that increase Gi levels contribute to beta2AR dysfunction.

  9. Afr1p regulates the Saccharomyces cerevisiae alpha-factor receptor by a mechanism that is distinct from receptor phosphorylation and endocytosis.

    PubMed Central

    Davis, C; Dube, P; Konopka, J B

    1998-01-01

    The alpha-factor pheromone receptor activates a G protein signaling pathway that induces the conjugation of the yeast Saccharomyces cerevisiae. Our previous studies identified AFR1 as a gene that regulates this signaling pathway because overexpression of AFR1 promoted resistance to alpha-factor. AFR1 also showed an interesting genetic relationship with the alpha-factor receptor gene, STE2, suggesting that the receptor is regulated by Afr1p. To investigate the mechanism of this regulation, we tested AFR1 for a role in the two processes that are known to regulate receptor signaling: phosphorylation and down-regulation of ligand-bound receptors by endocytosis. AFR1 overexpression diminished signaling in a strain that lacks the C-terminal phosphorylation sites of the receptor, indicating that AFR1 acts independently of phosphorylation. The effects of AFR1 overexpression were weaker in strains that were defective in receptor endocytosis. However, AFR1 overexpression did not detectably influence receptor endocytosis or the stability of the receptor protein. Instead, gene dosage studies showed that the effects of AFR1 overexpression on signaling were inversely proportional to the number of receptors. These results indicate that AFR1 acts independently of endocytosis, and that the weaker effects of AFR1 in strains that are defective in receptor endocytosis were probably an indirect consequence of their increased receptor number caused by the failure of receptors to undergo ligand-stimulated endocytosis. Analysis of the ligand binding properties of the receptor showed that AFR1 overexpression did not alter the number of cell-surface receptors or the affinity for alpha-factor. Thus, Afr1p prevents alpha-factor receptors from activating G protein signaling by a mechanism that is distinct from other known pathways. PMID:9504911

  10. Mechanisms of inverse agonism of antipsychotic drugs at the D(2) dopamine receptor: use of a mutant D(2) dopamine receptor that adopts the activated conformation.

    PubMed

    Wilson, J; Lin, H; Fu, D; Javitch, J A; Strange, P G

    2001-04-01

    The antipsychotic drugs have been shown to be inverse agonists at the D(2) dopamine receptor. We have examined the mechanism of this inverse agonism by making mutations in residue T343 in the base of the sixth transmembrane spanning region of the receptor. T343R, T343S and T343K mutant D(2) dopamine receptors were made and the T343R mutant characterized in detail. The T343R mutant D(2) dopamine receptor exhibits properties of a receptor that resides more in the activated state, namely increased agonist binding affinity (independent of G-protein coupling and dependent on agonist efficacy), increased agonist potency in functional tests (adenylyl cyclase inhibition) and increased inverse agonist effects. The binding of agonists to the mutant receptor also shows sensitivity to sodium ions, unlike the native receptor, so that isomerization of the receptor to its inactive state may be driven by sodium ions. The binding of inverse agonists to the receptor is, however, unaffected by the mutation. We conclude that inverse agonism at this receptor is not achieved by the inverse agonist binding preferentially to the non-activated state of the receptor over the activated state. Rather the inverse agonist appears to bind to all forms of the receptor but then renders the receptor inactive.

  11. Mutation of Asp20 of human interleukin-2 reveals a dual role of the p55 alpha chain of the interleukin-2 receptor.

    PubMed

    Flemming, C L; Russell, S J; Collins, M K

    1993-04-01

    Mutation of Asp20 in human interleukin-2 (IL-2) to Lys is known to result in an IL-2 molecule with unchanged binding to the p55 subunit of the IL-2 receptor, but with greatly decreased affinity for the p75 subunit (Collins, L., Tsien, W.-H., Seals, C. et al. Proc. Natl. Acad. Sci USA 1988. 85: 7709). Here we demonstrate that Lys20 IL-2 competed with a reduced (10-fold) affinity for high-affinity IL-2 receptors on two murine cell lines HT2 and CTLL. In parallel with this difference in receptor interaction, Lys20 IL-2 stimulated half-maximal HT2 cell proliferation at a 10-fold higher concentration than wild-type IL-2. However, half-maximal stimulation of CTLL cells required a 100-fold higher concentration of Lys20 IL-2. A similar 100-fold reduction in bioactivity of Lys20 IL-2 was observed for primary, activated, human or murine lymphocytes. Anti-p55 antibodies increased the concentration of Lys20 IL-2 required to stimulate HT2 cells to that required for CTLL cells. These data suggest that CTLL cells, while able to bind Lys20 IL-2 with high affinity, are lacking a p55-dependent function necessary for optimal stimulation. Therefore, p55 has a dual role, being important both for high-affinity IL-2 binding and for optimal cell triggering.

  12. From empirical to mechanism-based discovery of clinically useful Selective Estrogen Receptor Modulators (SERMs)

    PubMed Central

    Wardell, Suzanne E.; Nelson, Erik R.; McDonnell, Donald P.

    2014-01-01

    Our understanding of the molecular mechanisms underlying the pharmacological actions of estrogen receptor (ER) ligands has evolved considerably in recent years. Much of this knowledge has come from a detailed dissection of the mechanism(s) of action of the Selective Estrogen Receptor Modulators (SERMs) tamoxifen and raloxifene, drugs whose estrogen receptor (ER) agonist/antagonist properties are influenced by the cell context in which they operate. These studies have revealed that notwithstanding differences in drug pharmokinetics, the activity of an ER ligand is determined primarily by (a) the impact that a given ligand has on the receptor conformation and (b) the ability of structurally distinct ER-ligand complexes to interact with functionally distinct coregulators. Exploitation of the established relationships between ER structure and activity has led to the development of improved SERMs with more favorable therapeutic properties and of tissue-selective estrogen complexes, drugs in which a SERM and an ER agonist are combined to yield a blended activity that results in distinct clinical profiles. Remarkably, endogenous ligands that exhibit SERM activity have also been identified. One of these ligands, 27-hydroxycholesterol (27HC), has been shown to manifest ER-dependent pathological activities in the cardiovascular system, bone and mammary gland. Whereas the physiological activity of 27HC remains to be determined, its discovery highlights how cells have adopted mechanisms to allow the same receptor ligand complex to manifest different activities in different cells, and also how these processes can be exploited for new drug development. PMID:25084324

  13. GR-127935-sensitive mechanism mediating hypotension in anesthetized rats: are 5-HT5B receptors involved?

    PubMed

    Sánchez-Maldonado, Carolina; López-Sánchez, Pedro; Anguiano-Robledo, Liliana; Leopoldo, Marcello; Lacivita, Enza; Terrón, José A

    2015-04-01

    The 5-HT1B/1D receptor antagonist, GR-127935, inhibits hypotensive responses produced by the 5-HT1A, 5-HT1B/1D and 5-HT7 receptor agonist, and 5-HT5A/5B receptor ligand, 5-carboxamidotryptamine (5-CT), in rats. This work further characterized the above mechanism using more selective 5-HT1B and 5-HT1D receptor antagonists. Also, expression of 5-HT5A and 5-HT5B receptor mRNAs in blood vessels was searched by reverse transcription polymerase chain reaction. Decreases in diastolic blood pressure induced by 5-CT (0.001-10 μg/kg, intravenously) were analyzed in anesthetized rats that had received intravenous vehicle (1 mL/kg), SB-224289 (5-HT1B antagonist; 0.3 and 1.0 mg/kg), BRL15572 (5-HT1D antagonist; 0.3 and 1.0 mg/kg), SB-224289 + BRL15572 (0.3 mg/kg, each), or SB-224289 + BRL15572 (0.3 mg/kg, each) + GR-127935 (1 mg/kg). Because only the latter treatment inhibited 5-CT-induced hypotension, suggestive of a mechanism unrelated to 5-HT1B/1D receptors, the effects of antagonists/ligands at 5-HT5A (SB-699551, 1 mg/kg), 5-HT6 (SB-399885, 1 mg/kg), and 5-HT1B/1D/5A/5B/7 receptors (ergotamine, 0.1 mg/kg) on 5-CT-induced hypotension were tested. Interestingly, only ergotamine blocked 5-CT-induced responses; this effect closely paralleled that of SB-224289 + BRL-15572 + GR-127935. Neither did ergotamine nor GR-127935 inhibit hypotensive responses induced by the 5-HT7 receptor agonist, LP-44. Faint but clear bands corresponding to 5-HT5A and 5-HT5B receptor mRNAs in aorta and mesenteric arteries were detected. Results suggest that the GR-127935-sensitive mechanism mediating hypotension in rats is unrelated to 5-HT1B, 5-HT1D, 5-HT5A, 5-HT6, and 5-HT7 receptors. This mechanism, however, resembles putative 5-HT5B receptors.

  14. Common mechanisms of spatial attention in memory and perception: a tactile dual-task study.

    PubMed

    Katus, Tobias; Andersen, Søren K; Müller, Matthias M

    2014-03-01

    Orienting attention to locations in mnemonic representations engages processes that functionally and anatomically overlap the neural circuitry guiding prospective shifts of spatial attention. The attention-based rehearsal account predicts that the requirement to withdraw attention from a memorized location impairs memory accuracy. In a dual-task study, we simultaneously presented retro-cues and pre-cues to guide spatial attention in short-term memory (STM) and perception, respectively. The spatial direction of each cue was independent of the other. The locations indicated by the combined cues could be compatible (same hand) or incompatible (opposite hands). Incompatible directional cues decreased lateralized activity in brain potentials evoked by visual cues, indicating interference in the generation of prospective attention shifts. The detection of external stimuli at the prospectively cued location was impaired when the memorized location was part of the perceptually ignored hand. The disruption of attention-based rehearsal by means of incompatible pre-cues reduced memory accuracy and affected encoding of tactile test stimuli at the retrospectively cued hand. These findings highlight the functional significance of spatial attention for spatial STM. The bidirectional interactions between both tasks demonstrate that spatial attention is a shared neural resource of a capacity-limited system that regulates information processing in internal and external stimulus representations.

  15. Molecular Mechanism Underlying the Plant NRT1.1 Dual-Affinity Nitrate Transporter

    PubMed Central

    Sun, Ji; Zheng, Ning

    2015-01-01

    Nitrate (NO3−) is one of the most important sources of mineral nitrogen, which also serves as a key signaling molecule for plant growth and development. To cope with nitrate fluctuation in soil that varies by up to four orders of magnitude, plants have evolved high- and low-affinity nitrate transporter systems, consisting of distinct families of transporters. Interestingly, the first cloned nitrate transporter in Arabidopsis, NRT1.1 functions as a dual-affinity transporter, which can change its affinity for nitrate in response to substrate availability. Phosphorylation of a threonine residue, Thr101, switches NRT1.1 from low- to high-affinity state. Recent structural studies have unveiled that the unmodified NRT1.1 transporter works as homodimers with Thr101 located in close proximity to the dimer interface. Modification on the Thr101 residue is shown to not only decouple the dimer configuration, but also increase structural flexibility, thereby, altering the substrate affinity of NRT1.1. The structure of NRT1.1 helps establish a novel paradigm in which protein oligomerzation and posttranslational modification can synergistically expand the functional capacity of the major facilitator superfamily (MFS) transporters. PMID:26733879

  16. Dual modulation of inward rectifier potassium currents in olfactory neuronal cells by promiscuous G protein coupling of the oxytocin receptor.

    PubMed

    Gravati, Marta; Busnelli, Marta; Bulgheroni, Elisabetta; Reversi, Alessandra; Spaiardi, Paolo; Parenti, Marco; Toselli, Mauro; Chini, Bice

    2010-09-01

    Oxytocin receptor is a seven transmembrane receptor widely expressed in the CNS that triggers G(i) or G(q) protein-mediated signaling cascades leading to the regulation of a variety of neuroendocrine and cognitive functions. We decided to investigate whether and how the promiscuous receptor/G protein coupling affects neuronal excitability. As an experimental model, we used the immortalized gonadotropin-releasing hormone-positive GN11 cell line displaying the features of immature, migrating olfactory neurons. Using RT-PCR analysis, we detected the presence of oxytocin receptors whose stimulation by oxytocin led to the accumulation of inositol phosphates and to the inhibition of cell proliferation, and the expression of several inward rectifier (IR) K+ channel subtypes. Moreover, electrophysiological and pharmacological inspections using whole-cell patch-clamp recordings evidenced that in GN11 cells, IR channel subtypes are responsive to oxytocin. In particular, we found that: (i) peptide activation of receptor either inhibited or stimulated IR conductances, and (ii) IR current inhibition was mediated by a pertussis toxin-resistant G protein presumably of the G(q/11) subtype, and by phospholipase C, whereas IR current activation was achieved via receptor coupling to a pertussis toxin-sensitive G(i/o) protein. The findings suggest that neuronal excitability might be tuned by a single peptide receptor that mediates opposing effects on distinct K+ channels through the promiscuous coupling to different G proteins.

  17. Desensitization and internalization of the m2 muscarinic acetylcholine receptor are directed by independent mechanisms.

    PubMed

    Pals-Rylaarsdam, R; Xu, Y; Witt-Enderby, P; Benovic, J L; Hosey, M M

    1995-12-01

    The phenomenon of acute desensitization of G-protein-coupled receptors has been associated with several events, including receptor phosphorylation, loss of high affinity agonist binding, receptor:G-protein uncoupling, and receptor internalization. However, the biochemical events underlying these processes are not fully understood, and their contributions to the loss of signaling remain correlative. In addition, the nature of the kinases and the receptor domains which are involved in modulation of activity have only begun to be investigated. In order to directly measure the role of G-protein-coupled receptor kinases (GRKs) in the desensitization of the m2 muscarinic acetylcholine receptor (m2 mAChR), a dominant-negative allele of GRK2 was used to inhibit receptor phosphorylation by endogenous GRK activity in a human embryonic kidney cell line. The dominant-negative GRK2K220R reduced agonist-dependent phosphorylation of the m2 mAChR by approximately 50% and prevented acute desensitization of the receptor as measured by the ability of the m2 mAChR to attenuate adenylyl cyclase activity. In contrast, the agonist-induced internalization of the m2 mAChR was unaffected by the GRK2K220R construct. Further evidence linking receptor phosphorylation to acute receptor desensitization was obtained when two deletions of the third intracellular loop were made which created m2 mAChRs that did not become phosphorylated in an agonist-dependent manner and did not desensitize. However, the mutant mAChRs retained the ability to internalize. These data provide the first direct evidence that GRK-mediated receptor phosphorylation is necessary for m2 mAChR desensitization; the likely sites of in vivo phosphorylation are in the central portion of the third intracellular loop (amino acids 282-323). These results also indicate that internalization of the m2 receptor is not a key event in desensitization and is mediated by mechanisms distinct from GRK phosphorylation of the receptor.

  18. Microcontroller Based Proportional Derivative Plus Conditional Integral Controller for Electro-Mechanical Dual Acting Pulley Continuously Variable Transmission Ratio Control

    NASA Astrophysics Data System (ADS)

    Budianto, A.; Tawi, K. B.; Hussein, M.; Supriyo, B.; Ariyono, S.; Che Kob, M. S.; Ezlamy Zulkifli, Mohd; K, Khairuldean A.; Daraoh, Aishah

    2012-09-01

    Electro-Mechanical Dual Acting Pulley (EMDAP) Continuously Variable Transmission (CVT) is a transmission utilized by electro-mechanical actuated system. It has a potential to reduce energy consumption because it only needs power during changing CVT ratio and no power is needed to maintain CVT ratio due to self lock mechanism design. This paper proposed simple proportional derivative plus conditional integral (PDCI) controller to control EMDAP CVT ratio which can be simply implemented on a microcontroller. This proposed controller used Astrom-Hagglund method and Ziegler-Nichols formula to tune PDCI gain. The Proportional Derivative controller is directly activated from the start but Integral controller is only activated when the error value reaches error value setting point. Simulation using Matlab/Simulink software was conducted to evaluate PDCI system performance. The simulation results showed PDCI controller has ability to perform maximum overshoot 0.1%, 0.001 steady state error and 0.5s settling time. For clamping condition, settling time is about 11.46s during changing ratio from 2.0 to 0.7, while for release condition, settling time is about 8.33s during changing ratio from 0.7 to 2.0.

  19. Estrogen receptor β in Alzheimer's disease: From mechanisms to therapeutics.

    PubMed

    Zhao, Liqin; Woody, Sarah K; Chhibber, Anindit

    2015-11-01

    Alzheimer's disease (AD) disproportionally affects women and men. The female susceptibility for AD has been largely associated with the loss of ovarian sex hormones during menopause. This review examines the current understanding of the role of estrogen receptor β (ERβ) in the regulation of neurological health and its implication in the development and intervention of AD. Since its discovery in 1996, research conducted over the last 15-20 years has documented a great deal of evidence indicating that ERβ plays a pivotal role in a broad spectrum of brain activities from development to aging. ERβ genetic polymorphisms have been associated with cognitive impairment and increased risk for AD predominantly in women. The role of ERβ in the intervention of AD has been demonstrated by the alteration of AD pathology in response to treatment with ERβ-selective modulators in transgenic models that display pronounced plaque and tangle histopathological presentations as well as learning and memory deficits. Future studies that explore the potential interactions between ERβ signaling and the genetic isoforms of human apolipoprotein E (APOE) in brain aging and development of AD-risk phenotype are critically needed. The current trend of lost-in-translation in AD drug development that has primarily been based on early-onset familial AD (FAD) models underscores the urgent need for novel models that recapitulate the etiology of late-onset sporadic AD (SAD), the most common form of AD representing more than 95% of the current human AD population. Combining the use of FAD-related models that generally have excellent face validity with SAD-related models that hold more reliable construct validity would together increase the predictive validity of preclinical findings for successful translation into humans.

  20. Oxytocin: its mechanism of action and receptor signalling in the myometrium.

    PubMed

    Arrowsmith, S; Wray, S

    2014-06-01

    Oxytocin is a nonapeptide hormone that has a central role in the regulation of parturition and lactation. In this review, we address oxytocin receptor (OTR) signalling and its role in the myometrium during pregnancy and in labour. The OTR belongs to the rhodopsin-type (Class 1) of the G-protein coupled receptor superfamily and is regulated by changes in receptor expression, receptor desensitisation and local changes in oxytocin concentration. Receptor activation triggers a number of signalling events to stimulate contraction, primarily by elevating intracellular calcium (Ca(2+) ). This includes inositol-tris-phosphate-mediated store calcium release, store-operated Ca(2+) entry and voltage-operated Ca(2+) entry. We discuss each mechanism in turn and also discuss Ca(2+) -independent mechanisms such as Ca(2+) sensitisation. Because oxytocin induces contraction in the myometrium, both the activation and the inhibition of its receptor have long been targets in the management of dysfunctional and preterm labours, respectively. We discuss current and novel OTR agonists and antagonists and their use and potential benefit in obstetric practice. In this regard, we highlight three clinical scenarios: dysfunctional labour, postpartum haemorrhage and preterm birth.

  1. Tachykinins and Their Receptors: Contributions to Physiological Control and the Mechanisms of Disease

    PubMed Central

    Steinhoff, Martin S.; von Mentzer, Bengt; Geppetti, Pierangelo; Pothoulakis, Charalabos; Bunnett, Nigel W.

    2014-01-01

    The tachykinins, exemplified by substance P, are one of the most intensively studied neuropeptide families. They comprise a series of structurally related peptides that derive from alternate processing of three Tac genes and are expressed throughout the nervous and immune systems. Tachykinins interact with three neurokinin G protein-coupled receptors. The signaling, trafficking, and regulation of neurokinin receptors have also been topics of intense study. Tachykinins participate in important physiological processes in the nervous, immune, gastrointestinal, respiratory, urogenital, and dermal systems, including inflammation, nociception, smooth muscle contractility, epithelial secretion, and proliferation. They contribute to multiple diseases processes, including acute and chronic inflammation and pain, fibrosis, affective and addictive disorders, functional disorders of the intestine and urinary bladder, infection, and cancer. Neurokinin receptor antagonists are selective, potent, and show efficacy in models of disease. In clinical trials there is a singular success: neurokinin 1 receptor antagonists to treat nausea and vomiting. New information about the involvement of tachykinins in infection, fibrosis, and pruritus justifies further trials. A deeper understanding of disease mechanisms is required for the development of more predictive experimental models, and for the design and interpretation of clinical trials. Knowledge of neurokinin receptor structure, and the development of targeting strategies to disrupt disease-relevant subcellular signaling of neurokinin receptors, may refine the next generation of neurokinin receptor antagonists. PMID:24382888

  2. Dual pH/redox responsive and CD44 receptor targeting hybrid nano-chrysalis based on new oligosaccharides of hyaluronan conjugates.

    PubMed

    Chen, Daquan; Dong, Xue; Qi, Mengjiao; Song, Xiaoyan; Sun, Jingfang

    2017-02-10

    A smart hybrid microenvironment-mediated dual pH/redox-responsive polymeric nanoparticles combined with inorganic calcium phosphate (CaP) was fabricated, which we term as armored nano-chrysalis inspired by butterfly pupa. The nano-chrysalis has an inner core composed of specially designed oligosaccharides of hyaluronan (oHA) targeting CD44 receptor. The inner core has two functions, i.e., the dual pH/redox responsive polymeric conjugate and the fluorescent curcumin-prodrug function. The prepared nano-chrysalis possessed a smaller size (102.5±4.6nm) than the unarmored nano-chrysalis (122.5±6.6nm). Interestingly, while the nano-chrysalis were stable under pH 7.4, when incubated under the tumor acidic conditions (pH 6.5) the outer CaP armor would dissolve in a pH-dependent, sustained manner. Moreover, nano-chrysalis was demonstrated to present the most effective antitumor efficacy than other formulations. This study provides a promising smart nano-carrier platform to enhance the stability, decrease the side effects, and improve the therapeutic efficacy of anticancer drugs.

  3. Examining the role of endogenous orexins in hypothalamus-pituitary-adrenal axis endocrine function using transient dual orexin receptor antagonism in the rat.

    PubMed

    Steiner, Michel A; Sciarretta, Carla; Brisbare-Roch, Catherine; Strasser, Daniel S; Studer, Rolf; Jenck, Francois

    2013-04-01

    The orexin neuropeptide system regulates wakefulness and contributes to physiological and behavioral stress responses. Moreover, a role for orexins in modulating hypothalamus-pituitary-adrenal (HPA) axis activity has been proposed. Brain penetrating dual orexin receptor (OXR) antagonists such as almorexant decrease vigilance and have emerged as a novel therapeutic class for the treatment of insomnia. Almorexant was used here as a pharmacological tool to examine the role of endogenous orexin signaling in HPA axis endocrine function under natural conditions. After confirming the expression of prepro-orexin and OXR-1 and OXR-2 mRNA in hypothalamus, pituitary and adrenal glands, the effects of systemic almorexant were investigated on peripheral HPA axis hormone release in the rat under baseline, stress and pharmacological challenge conditions. Almorexant did not alter basal or stress-induced corticosterone release despite affecting wake and sleep stages (detected by radiotelemetric electroencephalography/electromyography) during the stress exposure. Moreover, almorexant did not affect the release of adrenocorticotropin (ACTH) and corticosterone at different time points along the diurnal rhythm, nor corticotrophin-releasing hormone (CRH)- and ACTH-stimulated neuroendocrine responses, measured in vivo under stress-free conditions. These results illustrate that dual OXR antagonists, despite modulating stress-induced wakefulness, do not interfere with endocrine HPA axis function in the rat. They converge to suggest that endogenous orexin signaling plays a minor role in stress hormone release under basal conditions and under challenge.

  4. Cannabidiol inhibits human glioma cell migration through a cannabinoid receptor-independent mechanism

    PubMed Central

    Vaccani, Angelo; Massi, Paola; Colombo, Arianna; Rubino, Tiziana; Parolaro, Daniela

    2005-01-01

    We evaluated the ability of cannabidiol (CBD) to impair the migration of tumor cells stimulated by conditioned medium. CBD caused concentration-dependent inhibition of the migration of U87 glioma cells, quantified in a Boyden chamber. Since these cells express both cannabinoid CB1 and CB2 receptors in the membrane, we also evaluated their engagement in the antimigratory effect of CBD. The inhibition of cell was not antagonized either by the selective cannabinoid receptor antagonists SR141716 (CB1) and SR144528 (CB2) or by pretreatment with pertussis toxin, indicating no involvement of classical cannabinoid receptors and/or receptors coupled to Gi/o proteins. These results reinforce the evidence of antitumoral properties of CBD, demonstrating its ability to limit tumor invasion, although the mechanism of its pharmacological effects remains to be clarified. PMID:15700028

  5. Mechanism for activation of the EGF receptor catalytic domain by the juxtamembrane segment

    PubMed Central

    Jura, Natalia; Endres, Nicholas F.; Engel, Kate; Deindl, Sebastian; Das, Rahul; Lamers, Meindert H.; Wemmer, David E.; Zhang, Xuewu; Kuriyan, John

    2009-01-01

    Signaling by the epidermal growth factor receptor requires an allosteric interaction between the kinase domains of two receptors, whereby one activates the other. We show that the intracellular juxtamembrane segment of the receptor, known to potentiate kinase activity, is able to dimerize the kinase domains. The C-terminal half of the juxtamembrane segment latches the activated kinase domain to the activator, and the N-terminal half of this segment further potentiates dimerization, most likely by forming an antiparallel helical dimer that engages the transmembrane helices of the activated receptor. Our data are consistent with a mechanism in which the extracellular domains block the intrinsic ability of the transmembrane and cytoplasmic domains to dimerize and activate, with ligand binding releasing this block. The formation of the activating juxtamembrane latch is prevented by the C-terminal tails in a new structure of an inactive kinase domain dimer, suggesting how alternative dimers can prevent ligand-independent activation. PMID:19563760

  6. Dual Receptor-Targeted Theranostic Nanoparticles for Localized Delivery and Activation of Photodynamic Therapy Drug in Glioblastomas

    PubMed Central

    Dixit, Suraj; Miller, Kayla; Zhu, Yun; McKinnon, Emilie; Novak, Thomas; Kenney, Malcolm E.; Broome, Ann-Marie

    2015-01-01

    Targeting gold nanoparticles (AuNPs) with two or more receptor binding peptides has been proposed to address intratumoral heterogeneity of glioblastomas that overexpress multiple cell surface receptors to ultimately improve therapeutic efficacy. AuNPs conjugated with peptides against both the epidermal growth factor and transferrin receptors and loaded with the photosensitizer phthalocyanine 4 (Pc 4) have been designed and compared with monotargeted AuNPs for in vitro and in vivo studies. The (EGFpep+Tfpep)-AuNPs-Pc 4 with a particle size of ~41 nm improved both specificity and worked synergistically to decrease time of maximal accumulation in human glioma cells that overexpressed two cell surface receptors as compared to cells that overexpressed only one. Enhanced cellular association and increased cytotoxicity were achieved. In vivo studies show notable accumulation of these agents in the brain tumor regions. PMID:26198693

  7. Impaired wake-promoting mechanisms in ghrelin receptor-deficient mice.

    PubMed

    Esposito, Matthew; Pellinen, Jacob; Kapás, Levente; Szentirmai, Éva

    2012-01-01

    Ghrelin receptors are expressed by key components of the arousal system. Exogenous ghrelin induces behavioral activation, promotes wakefulness and stimulates eating. We hypothesized that ghrelin-sensitive mechanisms play a role in the arousal system. To test this, we investigated the responsiveness of ghrelin receptor knockout (KO) mice to two natural wake-promoting stimuli. Additionally, we assessed the integrity of their homeostatic sleep-promoting system using sleep deprivation. There was no significant difference in the spontaneous sleep-wake activity between ghrelin receptor KO and wild-type (WT) mice. WT mice mounted robust arousal responses to a novel environment and food deprivation. Wakefulness increased for 6 h after cage change accompanied by increases in body temperature and locomotor activity. Ghrelin receptor KO mice completely lacked the wake and body temperature responses to new environment. When subjected to 48 h food deprivation, WT mice showed marked increases in their waking time during the dark periods of both days. Ghrelin receptor KO mice failed to mount an arousal response on the first night and wake increases were attenuated on the second day. The responsiveness to sleep deprivation did not differ between the two genotypes. These results indicate that the ghrelin-receptive mechanisms play an essential role in the function of the arousal system but not in homeostatic sleep-promoting mechanisms.

  8. Proteinase activated receptor-2-mediated dual oxidase-2 up-regulation is involved in enhanced airway reactivity and inflammation in a mouse model of allergic asthma.

    PubMed

    Nadeem, Ahmed; Alharbi, Naif O; Vliagoftis, Harissios; Tyagi, Manoj; Ahmad, Sheikh F; Sayed-Ahmed, Mohamed M

    2015-07-01

    Airway epithelial cells (AECs) express a variety of receptors, which sense danger signals from various aeroallergens/pathogens being inhaled constantly. Proteinase-activated receptor 2 (PAR-2) is one such receptor and is activated by cockroach allergens, which have intrinsic serine proteinase activity. Recently, dual oxidases (DUOX), especially DUOX-2, have been shown to be involved in airway inflammation in response to Toll-like receptor activation. However, the association between PAR-2 and DUOX-2 has not been explored in airways of allergic mice. Therefore, this study investigated the contribution of DUOX-2/reactive oxygen species (ROS) signalling in airway reactivity and inflammation after PAR-2 activation. Mice were sensitized intraperitoneally with intact cockroach allergen extract (CE) in the presence of aluminium hydroxide followed by intranasal challenge with CE. Mice were then assessed for airway reactivity, inflammation, oxidative stress (DUOX-2, ROS, inducible nitric oxide synthase, nitrite, nitrotyrosine and protein carbonyls) and apoptosis (Bax, Bcl-2, caspase-3). Challenge with CE led to up-regulation of DUOX-2 and ROS in AECs with concomitant increases in airway reactivity/inflammation and parameters of oxidative stress, and apoptosis. All of these changes were significantly inhibited by intranasal administration of ENMD-1068, a small molecule antagonist of PAR-2 in allergic mice. Administration of diphenyliodonium to allergic mice also led to improvement of allergic airway responses via inhibition of the DUOX-2/ROS pathway; however, these effects were less pronounced than PAR-2 antagonism. The current study suggests that PAR-2 activation leads to up-regulation of the DUOX-2/ROS pathway in AECs, which is involved in regulation of airway reactivity and inflammation via oxidative stress and apoptosis.

  9. Incorporation of the purified epstein barr virus/C3d receptor (CR2) into liposomes and demonstration of its dual ligand binding functions

    SciTech Connect

    Mold, C.; Cooper, N.R.; Nemerow, G.R.

    1986-06-01

    The 145-kDA molecule that has been identified as the C3d receptor CR2 was isolated from lysates of Raji cells by affinity chromatography by using the monoclonal antibody (MoAb)HB-5. The purified protein was incorporated into /sup 14/C-phosphatidylcholine liposomes by deoxycholate dialysis followed by flotation on discontinuous sucrose gradients. Incorporation of the receptor was verified by testing the gradient fractions for CR2 by an enzyme-linked immunosorbent assay. Liposomes were shown to be unilamellar vesicles ranging in diameter from 25 to 100 nm by electron microscopy. The external orientation of CR2 in the membranes was demonstrated by immunoelectron microscopy. The functional activities of liposomes containing CR2 and liposomes without protein were compared. CR2 liposomes bound to EC3d, but not to E, and this binding was inhibited by the anti-CR2 MoAb OKB7 and by a MoAb specific for C3d. Control liposomes failed to bind to either E or EC3D. The ability of CR2 to function as a receptor for Epstein Barr virus (EBV) was tested in two ways. First, CR2 liposomes bound to B95-8, a cell line expressing EBV membrane antigens, but not to B95-8 cells treated with the viral DNA polymerase inhibitor phosphonoformic acid. Second, liposomes containing CR2 were shown by ultracentrifugal analyses to bind directly to purified EBV, and this binding was also inhibited by OKB7. Control liposomes did not bind to B95-8 cells or to EBV. These findings show that CR2 purified from detergent extracts of Raji cells can be reconstituted into lipid membranes with maintenance of its dual functions as a receptor for C3d and EBV.

  10. The dual effect of CA1 NMDA receptor modulation on ACPA-induced amnesia in step-down passive avoidance learning task.

    PubMed

    Nasehi, Mohammad; Amin-Yavari, Samaneh; Ebrahimi-Ghiri, Mohaddeseh; Torabi-Nami, Mohammad; Zarrindast, Mohammad-Reza

    2015-04-01

    It is well documented that cannabinoids play an important role in certain hippocampal memory processes in rodents. On the other hand, N-Methyl-d-aspartate receptors (NMDARs) mediate the synaptic plasticity related to learning and memory processes which take place in the hippocampus. Such insights prompted us to investigate the influence of dorsal hippocampal (CA1) NMDA receptor agents on amnesia induced by cannabinoid CB1 receptor agonist, arachidonylcyclopropylamide (ACPA) in male mice. One-trial step-down passive avoidance and hole-board apparatuses were used to examine the memory retrieval and exploratory behaviors, respectively. Based on our findings, pre-training intraperitoneal (i.p.) administration of ACPA (0.01mg/kg) decreased memory acquisition. Moreover, pre-training intra-CA1 infusion of NMDA (0.001, 0.0125, 0.025 and 0.2µg/mouse), d-AP7 (0.5 and 1µg/mouse) or AM251 (50ng/mouse) impaired the memory acquisition. Meanwhile, NMDA-treated animals at the doses of 0.0005, 0.05 and 0.1µg/mouse acquired memory formation. In addition, intra-CA1 microinjection of NMDA (0.0005) plus different doses of ACPA potentiated the ACPA response, while NMDA (0.1) plus the lower or the higher dose of ACPA potentiated or restored the ACPA response, respectively. Further investigation revealed that a subthreshold dose of d-AP7 could potentiate the memory acquisition impairment induced by ACPA. Moreover, the subthreshold dose of AM251 did not alter the ACPA response, while the effective dose of the drug restored the memory acquisition impairment induced by ACPA. According to these results, we concluded that activation of the NMDA receptors in the CA1 mediates a dual effect on ACPA-induced amnesia in step-down passive avoidance learning task.

  11. Mechanism of action of species-selective P2X7 receptor antagonists

    PubMed Central

    Michel, Anton D; Ng, Sin-Wei; Roman, Shilina; Clay, William C; Dean, David K; Walter, Daryl S

    2009-01-01

    Background and purpose: AZ11645373 and N-{2-methyl-5-[(1R, 5S)-9-oxa-3,7-diazabicyclo[3.3.1]non-3-ylcarbonyl]phenyl}-2-tricyclo[3.3.1.13,7]dec-1-ylacetamide hydrochloride (compound-22) are recently described P2X7 receptor antagonists. In this study we have further characterized these compounds to determine their mechanism of action and interaction with other species orthologues. Experimental approach: Antagonist effects at recombinant and chimeric P2X7 receptors were assessed by ethidium accumulation and radioligand-binding studies. Key results: AZ11645373 and compound-22 were confirmed as selective non-competitive antagonists of human or rat P2X7 receptors respectively. Both compounds were weak antagonists of the mouse and guinea-pig P2X7 receptors and, for each compound, their potency estimates at human and dog P2X7 receptors were similar. The potency of compound-22 was moderately temperature-dependent while that of AZ11645373 was not. The antagonist effects of both compounds were slowly reversible and were not prevented by decavanadate, suggesting that they were allosteric antagonists. Indeed, the compounds competed for binding sites labelled by an allosteric radio-labelled P2X7 receptor antagonist. The species selectivity of AZ11645373, but not compound-22, was influenced by the nature of the amino acid at position 95 of the P2X7 receptor. N2-(3,4-difluorophenyl)-N1-[2-methyl-5-(1-piperazinylmethyl)phenyl]glycinamide dihydrochloride, a positive allosteric modulator of the rat receptor, reduced the potency of compound-22 at the rat receptor but had little effect on the actions of AZ11645373. Conclusions: AZ11645373 and compound-22 are allosteric antagonists of human and rat P2X7 receptors respectively. The differential interaction of the two compounds with the receptor suggests there may be more than one allosteric regulatory site on the P2X7 receptor at which antagonists can bind and affect receptor function. PMID:19309360

  12. Naloxone potentiates the inotropic effects of isoproterenol in vitro by a nonopiate receptor mechanism.

    PubMed

    Lechner, R B

    1992-11-01

    Naloxone potentiates the effects of adrenergic agonists when administered to hypovolemic dogs, and it has been assumed that this effect is due to naloxone's action at opiate receptors. To help determine the site and mechanism of this interaction, we administered naloxone and its "d" stereo-isomer (which does not bind to opiate receptors) to guinea pig papillary muscles in the presence and absence of pharmacologic (isoproterenol) and physiologic (treppe) inotropic stimulation. In control muscles and in rapidly paced muscles, naloxone was without significant inotropic effect. In the presence of isoproterenol, d- and l-naloxone exerted significant positive inotropic effects that were dose dependent. We conclude that, since both d- and l-naloxone potentiated the inotropic effects of isoproterenol and this was seen in the absence of opioids, naloxone may increase contractility by a nonopiate receptor-mediated mechanism.

  13. Desensitization of the Y1 cell adrenocorticotropin receptor: evidence for a restricted heterologous mechanism implying a role for receptor-effector complexes.

    PubMed

    Baig, A H; Swords, F M; Noon, L A; King, P J; Hunyady, L; Clark, A J

    2001-11-30

    Receptor desensitization provides a potential mechanism for the regulation of adrenocortical adrenocorticotropin (ACTH) responsiveness. Using the mouse adrenocortical Y1 cell line we demonstrate that ACTH effectively desensitizes the cAMP response of its own receptor, the melanocortin 2 receptor (MC2R), in these cells with a maximal effect between 30 and 60 min. Neither forskolin nor isoproterenol (in Y1 cells stably transfected with the beta(2)-adrenergic receptor) desensitize this ACTH response. ACTH desensitizes its receptor at concentrations at which only a fraction of receptors are occupied, implying that this mechanism acts on agonist-unoccupied receptors. Y1 cells express G protein-coupled receptor kinase (GRK) 2 and 5, but stable expression of a dominant negative GRK2 (K220W) only marginally reduces the desensitization by ACTH. The protein kinase A (PKA) inhibitor, H89, extinguishes almost the entire desensitization response over the initial 30-min period at all concentrations of ACTH. A mutant MC2R in which the single consensus PKA phosphorylation site has been mutated (S208A) when expressed in MC2R-negative Y6 cells is also unable to desensitize. These data imply a heterologous, PKA-dependent, mode of desensitization, which is restricted to agonist-occupied and -unoccupied MC2R, possibly as a consequence of receptor/effector complexes that functionally compartmentalize this receptor.

  14. Active Joint Mechanism Driven by Multiple Actuators Made of Flexible Bags: A Proposal of Dual Structural Actuator

    PubMed Central

    Inou, Norio

    2013-01-01

    An actuator is required to change its speed and force depending on the situation. Using multiple actuators for one driving axis is one of the possible solutions; however, there is an associated problem of output power matching. This study proposes a new active joint mechanism using multiple actuators. Because the actuator is made of a flexible bag, it does not interfere with other actuators when it is depressurized. The proposed joint achieved coordinated motion of multiple actuators. This report also discusses a new actuator which has dual cylindrical structure. The cylinders are composed of flexible bags with different diameters. The joint torque is estimated based on the following factors: empirical formula for the flexible actuator torque, geometric relationship between the joint and the actuator, and the principle of virtual work. The prototype joint mechanism achieves coordinated motion of multiple actuators for one axis. With this motion, small inner actuator contributes high speed motion, whereas large outer actuator generates high torque. The performance of the prototype joint is examined by speed and torque measurements. The joint showed about 30% efficiency at 2.0 Nm load torque under 0.15 MPa air input. PMID:24385868

  15. Active joint mechanism driven by multiple actuators made of flexible bags: a proposal of dual structural actuator.

    PubMed

    Kimura, Hitoshi; Matsuzaki, Takuya; Kataoka, Mokutaro; Inou, Norio

    2013-01-01

    An actuator is required to change its speed and force depending on the situation. Using multiple actuators for one driving axis is one of the possible solutions; however, there is an associated problem of output power matching. This study proposes a new active joint mechanism using multiple actuators. Because the actuator is made of a flexible bag, it does not interfere with other actuators when it is depressurized. The proposed joint achieved coordinated motion of multiple actuators. This report also discusses a new actuator which has dual cylindrical structure. The cylinders are composed of flexible bags with different diameters. The joint torque is estimated based on the following factors: empirical formula for the flexible actuator torque, geometric relationship between the joint and the actuator, and the principle of virtual work. The prototype joint mechanism achieves coordinated motion of multiple actuators for one axis. With this motion, small inner actuator contributes high speed motion, whereas large outer actuator generates high torque. The performance of the prototype joint is examined by speed and torque measurements. The joint showed about 30% efficiency at 2.0 Nm load torque under 0.15 MPa air input.

  16. MECHANISMS OF ZN-INDUCED SIGNAL INITIATION THROUGH THE EPIDERMAL GROWTH FACTOR RECEPTOR (EGFR)

    EPA Science Inventory

    MECHANISMS OF Zn-INDUCED SIGNAL INITIATION THROUGH THE EPIDERMAL GROWTH FACTOR RECEPTOR (EGFR)
    James M. Samet*, Lee M. Graves? and Weidong Wu?. *Human Studies Division, NHEERL, ORD, Research Triangle Park, NC 27711, and ?Center for Environmental Medicine, University of North C...

  17. Endocrine disrupting chemicals targeting estrogen receptor signaling: Identification and mechanisms of action

    PubMed Central

    Shanle, Erin K.; Xu, Wei

    2011-01-01

    Many endocrine disrupting chemicals (EDCs) adversely impact estrogen signaling by interacting with two estrogen receptors (ERs): ERα and ERβ. Though the receptors have similar ligand binding and DNA binding domains, ERα and ERβ have some unique properties in terms of ligand selectivity and target gene regulation. EDCs that target ER signaling can modify genomic and non-genomic ER activity through direct interactions with ERs, indirectly through transcription factors like the aryl hydrocarbon receptor (AhR), or through modulation of metabolic enzymes that are critical for normal estrogen synthesis and metabolism. Many EDCs act through multiple mechanisms as exemplified by chemicals that bind both AhR and ER, such as 3-methylcholanthrene. Other EDCs that target ER signaling include phytoestrogens, bisphenolics, and organochlorine pesticides and many alter normal ER signaling through multiple mechanisms. EDCs can also display tissue-selective ER agonist and antagonist activities similar to selective estrogen receptor modulators (SERMs) designed for pharmaceutical use. Thus, biological effects of EDCs need to be carefully interpreted because EDCs can act through complex tissue-selective modulation of ERs and other signaling pathways in vivo. Current requirements by the U.S. Environmental Protection Agency require some in vitro and cell-based assays to identify EDCs that target ER signaling through direct and metabolic mechanisms. Additional assays may be useful screens for identifying EDCs that act through alternative mechanisms prior to further in vivo study. PMID:21053929

  18. Anaerobic reductive dechlorination of tetrachloroethene: how can dual Carbon-Chlorine isotopic measurements help elucidating the underlying reaction mechanism?

    NASA Astrophysics Data System (ADS)

    Badin, Alice; Buttet, Géraldine; Maillard, Julien; Holliger, Christof; Hunkeler, Daniel

    2014-05-01

    Chlorinated ethenes (CEs) such as tetrachloroethene (PCE) are common persistent groundwater contaminants. Among clean-up strategies applied to sites affected by such pollution, bioremediation has been considered with a growing interest as it represents a cost-effective, environmental friendly approach. This technique however sometimes leads to an incomplete and slow biodegradation of CEs resulting in an accumulation of toxic metabolites. Understanding the reaction mechanisms underlying anaerobic reductive dechlorination would thus help assessing PCE biodegradation in polluted sites. Stable isotope analysis can provide insight into reaction mechanisms. For chlorinated hydrocarbons, carbon (C) and chlorine (Cl) isotope data (δ13C and δ37Cl) tend to show a linear correlation with a slope (m ≡ ɛC/ɛCl) characteristic of the reaction mechanism [1]. This study hence aims at exploring the potential of a dual C-Cl isotope approach in the determination of the reaction mechanisms involved in PCE reductive dechlorination. C and Cl isotope fractionation were investigated during anaerobic PCE dechlorination by two bacterial consortia containing members of the Sulfurospirillum genus. The specificity in these consortia resides in the fact that they each conduct PCE reductive dechlorination catalysed by one different reductive dehalogenase, i.e. PceADCE which yields trichloroethene (TCE) and cis-dichloroethene (cDCE), and PceATCE which yields TCE only. The bulk C isotope enrichment factors were -3.6±0.3 o for PceATCE and -0.7±0.1o for PceADCE. The bulk Cl isotope enrichment factors were -1.3±0.2 o for PceATCE and -0.9±0.1 o for PceADCE. When applying the dual isotope approach, two m values of 2.7±0.1 and 0.7±0.2 were obtained for the reductive dehalogenases PceATCE and PceADCE, respectively. These results suggest that PCE can be degraded according to two different mechanisms. Furthermore, despite their highly similar protein sequences, each reductive dehalogenase seems

  19. Neurobiology of microglial action in CNS injuries: receptor-mediated signaling mechanisms and functional roles

    PubMed Central

    Hu, Xiaoming; Liou, Anthony K.F.; Leak, Rehana K.; Xu, Mingyue; An, Chengrui; Suenaga, Jun; Shi, Yejie; Gao, Yanqin; Zheng, Ping; Chen, Jun

    2014-01-01

    Microglia are the first line of immune defense against central nervous system (CNS) injuries and disorders. These highly plastic cells play dualistic roles in neuronal injury and recovery and are known for their ability to assume diverse phenotypes. A broad range of surface receptors are expressed on microglia and mediate microglial ‘On’ or ‘Off’ responses to signals from other host cells as well as invading microorganisms. The integrated actions of these receptors result in tightly regulated biological functions, including cell mobility, phagocytosis, the induction of acquired immunity, and trophic factor/inflammatory mediator release. Over the last few years, significant advances have been made towards deciphering the signaling mechanisms related to these receptors and their specific cellular functions. In this review, we describe the current state of knowledge of the surface receptors involved in microglial activation, with an emphasis on their engagement of distinct functional programs and their roles in CNS injuries. It will become evident from this review that microglial homeostasis is carefully maintained by multiple counterbalanced strategies, including, but not limited to, ‘On’ and ‘Off’ receptor signaling. Specific regulation of theses microglial receptors may be a promising therapeutic strategy against CNS injuries. PMID:24923657

  20. Dual mechanisms regulate ecosystem stability under decade-long warming and hay harvest.

    PubMed

    Shi, Zheng; Xu, Xia; Souza, Lara; Wilcox, Kevin; Jiang, Lifen; Liang, Junyi; Xia, Jianyang; García-Palacios, Pablo; Luo, Yiqi

    2016-06-15

    Past global change studies have identified changes in species diversity as a major mechanism regulating temporal stability of production, measured as the ratio of the mean to the standard deviation of community biomass. However, the dominant plant functional group can also strongly determine the temporal stability. Here, in a grassland ecosystem subject to 15 years of experimental warming and hay harvest, we reveal that warming increases while hay harvest decreases temporal stability. This corresponds with the biomass of the dominant C4 functional group being higher under warming and lower under hay harvest. As a secondary mechanism, biodiversity also explains part of the variation in temporal stability of production. Structural equation modelling further shows that warming and hay harvest regulate temporal stability through influencing both temporal mean and variation of production. Our findings demonstrate the joint roles that dominant plant functional group and biodiversity play in regulating the temporal stability of an ecosystem under global change.

  1. Dual mechanisms regulate ecosystem stability under decade-long warming and hay harvest

    PubMed Central

    Shi, Zheng; Xu, Xia; Souza, Lara; Wilcox, Kevin; Jiang, Lifen; Liang, Junyi; Xia, Jianyang; García-Palacios, Pablo; Luo, Yiqi

    2016-01-01

    Past global change studies have identified changes in species diversity as a major mechanism regulating temporal stability of production, measured as the ratio of the mean to the standard deviation of community biomass. However, the dominant plant functional group can also strongly determine the temporal stability. Here, in a grassland ecosystem subject to 15 years of experimental warming and hay harvest, we reveal that warming increases while hay harvest decreases temporal stability. This corresponds with the biomass of the dominant C4 functional group being higher under warming and lower under hay harvest. As a secondary mechanism, biodiversity also explains part of the variation in temporal stability of production. Structural equation modelling further shows that warming and hay harvest regulate temporal stability through influencing both temporal mean and variation of production. Our findings demonstrate the joint roles that dominant plant functional group and biodiversity play in regulating the temporal stability of an ecosystem under global change. PMID:27302085

  2. Mechanism of single- and double-sided inhibition of dual topology fluoride channels by synthetic monobodies.

    PubMed

    Turman, Daniel L; Stockbridge, Randy B

    2017-04-03

    The Fluc family of proteins comprises small, electrodiffusive fluoride channels, which prevent accumulation of toxic F(-) ions in microorganisms. Recent crystal structures have confirmed their unusual architecture, in which a pair of antiparallel subunits convenes to form a dimer with a twofold symmetry axis parallel to the plane of the membrane. These structures have also revealed the interactions between Fluc channels and several different fibronectin domain monobodies that inhibit Fluc-mediated F(-) currents; in all structures, each channel binds to two monobodies symmetrically, one on either side of the membrane. However, these structures do not reveal the mechanism of monobody inhibition. Moreover, the results appear to diverge from a recent electrophysiological study indicating that monobody binding is negatively cooperative; that is, a bound monobody on one side of a Fluc channel decreases the affinity of an oppositely bound monobody by ∼10-fold. In this study, we reconcile these observations by probing the mechanism of monobody binding and its negative cooperativity using electrophysiological experiments in planar lipid bilayers. Our results indicate that monobody inhibition occurs via a pore-blocking mechanism and that negative cooperativity arises from electrostatic repulsion between the oppositely bound monobodies. A single glutamate residue, on a loop of the monobody that extends into the channel interior, is responsible for negatively cooperative binding. This glutamate side chain also confers voltage dependence and sensitivity to the concentration of trans-F(-) ion to monobody binding. Neutralization by mutation to glutamine abolishes these electrostatic effects. Monobodies that are amenable to cocrystallization with Fluc channels lack an analogous negatively charged side chain and bind independently to opposite sides of the channel. Thus, this work reveals the source of voltage dependence and negative cooperativity of monobody binding to Fluc

  3. A dual flow bioreactor with controlled mechanical stimulation for cartilage tissue engineering.

    PubMed

    Spitters, Tim W G M; Leijten, Jeroen C H; Deus, Filipe D; Costa, Ines B F; van Apeldoorn, Aart A; van Blitterswijk, Clemens A; Karperien, Marcel

    2013-10-01

    In cartilage, tissue engineering bioreactors can create a controlled environment to study chondrocyte behavior under mechanical stimulation or produce chondrogenic grafts of clinically relevant size. Here we present a novel bioreactor that combines mechanical stimulation with a two compartment system through which nutrients can be supplied solely by diffusion from opposite sides of a tissue-engineered construct. This design is based on the hypothesis that creating gradients of nutrients, growth factors, and growth factor antagonists can aid in the generation of zonal tissue-engineered cartilage. Computational modeling predicted that the design facilitates the creation of a biologically relevant glucose gradient. This was confirmed by quantitative glucose measurements in cartilage explants. In this system, it is not only possible to create gradients of nutrients, but also of anabolic or catabolic factors. Therefore, the bioreactor design allows control over nutrient supply and mechanical stimulation useful for in vitro generation of cartilage constructs that can be used for the resurfacing of articulated joints or as a model for studying osteoarthritis disease progression.

  4. Thermodynamic evidence for a dual transport mechanism in a POT peptide transporter.

    PubMed

    Parker, Joanne L; Mindell, Joseph A; Newstead, Simon

    2014-12-02

    Peptide transport plays an important role in cellular homeostasis as a key route for nitrogen acquisition in mammalian cells. PepT1 and PepT2, the mammalian proton coupled peptide transporters (POTs), function to assimilate and retain diet-derived peptides and play important roles in drug pharmacokinetics. A key characteristic of the POT family is the mechanism of peptide selectivity, with members able to recognise and transport >8000 different peptides. In this study, we present thermodynamic evidence that in the bacterial POT family transporter PepTSt, from Streptococcus thermophilus, at least two alternative transport mechanisms operate to move peptides into the cell. Whilst tri-peptides are transported with a proton:peptide stoichiometry of 3:1, di-peptides are co-transported with either 4 or 5 protons. This is the first thermodynamic study of proton:peptide stoichiometry in the POT family and reveals that secondary active transporters can evolve different coupling mechanisms to accommodate and transport chemically and physically diverse ligands across the membrane.

  5. Evaluation of Thermo-Mechanical Stability of COTS Dual-Axis MEMS Accelerometers for Space Applications

    NASA Technical Reports Server (NTRS)

    Sharma, Ashok K.; Teverovksy, Alexander; Day, John H. (Technical Monitor)

    2000-01-01

    Microelectromechanical systems in MEMS is one of the fastest growing technologies in microelectronics, and is of great interest for military and aerospace applications. Accelerometers are the earliest and most developed representatives of MEMS. First demonstrated in 1979, micromachined accelerometers were used in automobile industry for air bag crash- sensing applications since 1990. In 1999, N4EMS accelerometers were used in NASA-JPL Mars Microprobe. The most developed accelerometers for airbag crash- sensing are rated for a full range of +/- 50 G. The range of sensitivity for accelerometers required for military or aerospace applications is much larger, varying from 20,000 G (to measure acceleration during gun and ballistic munition launches), and to 10(exp -6) G, when used as guidance sensors (to measure attitude and position of a spacecraft). The presence of moving parts on the surface of chip is specific to MEMS, and particularly, to accelerometers. This characteristic brings new reliability issues to micromachined accelerometers, including cyclic fatigue cracking of polysilicon cantilevers and springs, mechanical stresses that are caused by packaging and contamination in the internal cavity of the package. Studies of fatigue cracks initiation and growth in polysilicon showed that the fatigue damage may influence MEMS device performance, and the presence of water vapor significantly enhances crack initiation and growth. Environmentally induced failures, particularly, failures due to thermal cycling and mechanical shock are considered as one of major reliability concerns in MEMS. These environmental conditions are also critical for space applications of the parts. For example, the Mars pathfinder mission had experienced 80 mechanical shock events during the pyrotechnic separation processes.

  6. Dual mechanism of daunorubicin-induced cell death in both sensitive and MDR-resistant HL-60 cells

    PubMed Central

    Côme, M-G; Skladanowski, A; Larsen, A K; Laurent, G

    1999-01-01

    Exposure of some acute myeloid leukaemia (AML) cells to daunorubicin leads to rapid cell death, whereas other AML cells show natural drug resistance. This has been attributed to expression of functional P-glycoprotein resulting in reduced drug accumulation. However, it has also been proposed that P-glycoprotein-expressing multidrug-resistant (MDR) cells are inherently defective for apoptosis. To distinguish between these different possibilities, we have compared the cell death process in a human AML cell line (HL-60) with a MDR subline (HL-60/Vinc) at doses that yield either similar intracellular daunorubicin concentrations or comparable cytotoxicity. Adjustment of the dose to obtain the same intracellular drug accumulation in the two cell lines did not result in equal cytotoxicity, suggesting the presence of additional resistance mechanisms in the P-glycoprotein-expressing HL-60/Vinc cells. However, at equitoxic doses, similar cell death pathways were observed. In HL-60 cells, daunorubicin induced rapid apoptosis at 0.5–1 μM and delayed mitotic cell death at 0.1 μM. These concentrations are within the clinical dose range. Similarly, HL-60/Vinc cells underwent apoptosis at 50–100 μM daunorubicin and mitotic cell death at 10 μM. These results show, for the first time, that anthracyclines can induce cell death by a dual mechanism in both sensitive and MDR cells. Our results also show that not only the cytotoxicity, but also the kinetics and mechanism of cell death, are dose dependent. Interestingly, regrowth was observed only in association with delayed cell death and the formation of enlarged, often polyploid, cells with micronucleation, suggesting that morphological criteria may be useful to evaluate treatment efficacy in patients with myeloid leukaemias. © 1999 Cancer Research Campaign PMID:10098741

  7. Mechanism of HSV infection through soluble adapter-mediated virus bridging to the EGF receptor

    SciTech Connect

    Nakano, Kenji; Kobayashi, Masatoshi; Nakamura, Kei-ichiro; Nakanishi, Takeshi; Asano, Ryutaro; Kumagai, Izumi; Tahara, Hideaki; Kuwano, Michihiko; Cohen, Justus B.; Glorioso, Joseph C.

    2011-04-25

    Herpes simplex virus entry into cells requires the binding of envelope glycoprotein D (gD) to an entry receptor. Depending on the cell, entry occurs by different mechanisms, including fusion at the cell surface or endocytosis. Here we examined the entry mechanism through a non-HSV receptor mediated by a soluble bi-specific adapter protein composed of recognition elements for gD and the EGF receptor (EGFR). Virus entered into endosomes using either EGF or an EGFR-specific single chain antibody (scFv) for receptor recognition. Infection was less efficient with the EGF adapter which could be attributed to its weaker binding to a viral gD. Infection mediated by the scFv adapter was pH sensitive, indicating that gD-EGFR bridging alone was insufficient for capsid release from endosomes. We also show that the scFv adapter enhanced infection of EGFR-expressing tumor tissue in vivo. Our results indicate that adapters may retarget HSV infection without drastically changing the entry mechanism.

  8. Participation of central GABAA receptors in the trigeminal processing of mechanical allodynia in rats

    PubMed Central

    Kim, Min Ji; Park, Young Hong; Yang, Kui Ye; Ju, Jin Sook; Bae, Yong Chul

    2017-01-01

    Here we investigated the central processing mechanisms of mechanical allodynia and found a direct excitatory link with low-threshold input to nociceptive neurons. Experiments were performed on male Sprague-Dawley rats weighing 230-280 g. Subcutaneous injection of interleukin 1 beta (IL-1β) (1 ng/10 µL) was used to produce mechanical allodynia and thermal hyperalgesia. Intracisternal administration of bicuculline, a gamma aminobutyric acid A (GABAA) receptor antagonist, produced mechanical allodynia in the orofacial area under normal conditions. However, intracisternal administration of bicuculline (50 ng) produced a paradoxical anti-allodynic effect under inflammatory pain conditions. Pretreatment with resiniferatoxin (RTX), which depletes capsaicin receptor protein in primary afferent fibers, did not alter the paradoxical anti-allodynic effects produced by the intracisternal injection of bicuculline. Intracisternal injection of bumetanide, an Na-K-Cl cotransporter (NKCC 1) inhibitor, reversed the IL-1β-induced mechanical allodynia. In the control group, application of GABA (100 µM) or muscimol (3 µM) led to membrane hyperpolarization in gramicidin perforated current clamp mode. However, in some neurons, application of GABA or muscimol led to membrane depolarization in the IL-1β-treated rats. These results suggest that some large myelinated Aβ fibers gain access to the nociceptive system and elicit pain sensation via GABAA receptors under inflammatory pain conditions. PMID:28066142

  9. Dual wavelength laser damage mechanisms in the ultra-short pulse regime

    NASA Astrophysics Data System (ADS)

    Gyamfi, Mark; Costella, Marion; Willemsen, Thomas; Jürgens, Peter; Mende, Mathias; Jensen, Lars; Ristau, Detlev

    2016-12-01

    New ultrashort pulse laser systems exhibit an ever increasing performance which includes shorter pulses and higher pulse energies. Optical components used in these systems are facing increasing requirements regarding their durability, and therefore understanding of the damage mechanism is crucial. In the ultra-short pulse regime electron ionization processes control the damage mechanisms. For the single wavelength, single pulse regime the Keldysh [1] and the Drude model [2] allow a quantitative description of these ionization processes. However, in this model, the electrical field is restricted to a single wavelength, and therefore it cannot be applied in the case of irradiation with two pulses at different wavelengths. As frequency conversion is becoming more common in ultra-short pulse applications, further research is needed in this field to predict the damage resistance of optical components. We investigate the damage behavior of high reflective mirrors made of different metal oxide materials under simultaneous exposure to ultra-short pulses at the wavelengths 387.5 nm and 775 nm, respectively.

  10. Dual mechanism of Rag gene repression by c-Myb during pre-B cell proliferation.

    PubMed

    Timblin, Greg A; Xie, Liangqi; Tjian, Robert; Schlissel, Mark S

    2017-04-03

    Developing B lymphocytes undergo clonal expansion following successful immunoglobulin heavy chain gene rearrangement. During this proliferative burst, expression of the Rag genes is transiently repressed to prevent the generation of dsDNA breaks in cycling large pre-B cells. The Rag genes are then re-expressed in small resting pre-B cells for immunoglobulin light chain gene rearrangement. We previously identified c-Myb as a repressor of Rag transcription during clonal expansion using Abelson murine leukemia virus-transformed B cells. Nevertheless, the molecular mechanisms by which c-Myb achieved precise spatiotemporal repression of Rag expression remained obscure. Here we identify two mechanisms by which c-Myb represses Rag transcription. First, c-Myb negatively regulates the expression of the Rag activator Foxo1, an activity dependent on M303 in c-Myb's transactivation domain and likely the recruitment of corepressors to the Foxo1 locus by c-Myb. Second, c-Myb represses Rag transcription directly by occupying the Erag enhancer and antagonizing Foxo1 binding to a consensus forkhead site in this cis regulatory element that we show is crucial for Rag expression in Abelson pre-B cell lines. This work provides important mechanistic insight into how spatiotemporal expression of the Rag genes is tightly controlled during B lymphocyte development to prevent mistimed dsDNA breaks and their deleterious consequences.

  11. Dual mechanism of ion permeation through VDAC revealed with inorganic phosphate ions and phosphate metabolites.

    PubMed

    Krammer, Eva-Maria; Vu, Giang Thi; Homblé, Fabrice; Prévost, Martine

    2015-01-01

    In the exchange of metabolites and ions between the mitochondrion and the cytosol, the voltage-dependent anion channel (VDAC) is a key element, as it forms the major transport pathway for these compounds through the mitochondrial outer membrane. Numerous experimental studies have promoted the idea that VDAC acts as a regulator of essential mitochondrial functions. In this study, using a combination of molecular dynamics simulations, free-energy calculations, and electrophysiological measurements, we investigated the transport of ions through VDAC, with a focus on phosphate ions and metabolites. We showed that selectivity of VDAC towards small anions including monovalent phosphates arises from short-lived interactions with positively charged residues scattered throughout the pore. In dramatic contrast, permeation of divalent phosphate ions and phosphate metabolites (AMP and ATP) involves binding sites along a specific translocation pathway. This permeation mechanism offers an explanation for the decrease in VDAC conductance measured in the presence of ATP or AMP at physiological salt concentration. The binding sites occur at similar locations for the divalent phosphate ions, AMP and ATP, and contain identical basic residues. ATP features a marked affinity for a central region of the pore lined by two lysines and one arginine of the N-terminal helix. This cluster of residues together with a few other basic amino acids forms a "charged brush" which facilitates the passage of the anionic metabolites through the pore. All of this reveals that VDAC controls the transport of the inorganic phosphates and phosphate metabolites studied here through two different mechanisms.

  12. Fenfluramine releases serotonin from human brain nerve endings by a dual mechanism.

    PubMed

    Bonanno, G; Fassio, A; Severi, P; Ruelle, A; Raiteri, M

    1994-09-01

    Fenfluramine is the most widely used anorexigenic drug in humans. In animal experiments d-fenfluramine has been shown to act as a potent releaser of brain serotonin [5-hydroxytryptamine (5-HT)]. Here we have investigated the effects of d-fenfluramine on the release of [3H]5-HT from isolated nerve endings of human neocortex. The drug elicited release of unmetabolized [3H]5-HT, and this effect was concentration dependent. However, the mechanism of release seems to differ profoundly depending on the concentrations of d-fenfluramine used. At 5 microM, the release of [3H]5-HT was blocked by the 5-HT transporter inhibitor fluoxetine and was Ca2+ independent and insensitive to the human autoreceptor 5-HT1D agonist sumatriptan. The release of [3H]5-HT elicited by 0.5 microM d-fenfluramine was similarly blocked by fluoxetine, but it was strongly Ca2+ dependent and sensitive to sumatriptan. It is suggested that, at relatively high concentrations, d-fenfluramine largely diffuses into serotonergic terminals and causes release of 5-HT through the 5-HT carrier working in the inside-outside direction; at relatively low concentrations d-fenfluramine enters the terminals through the 5-HT transporter but elicits release of 5-HT by an exocytotic-like mechanism.

  13. mGlu2 metabotropic glutamate receptors restrain inflammatory pain and mediate the analgesic activity of dual mGlu2/mGlu3 receptor agonists.

    PubMed

    Zammataro, Magda; Chiechio, Santina; Montana, Michael C; Traficante, Anna; Copani, Agata; Nicoletti, Ferdinando; Gereau, Robert W

    2011-01-14

    Group II metabotropic glutamate receptors (mGluRs) couple to the inhibitory G-protein Gi. The group II mGluRs include two subtypes, mGlu2 and mGlu3, and their pharmacological activation produces analgesic effects in inflammatory and neuropathic pain states. However, the specific contribution of each one of the two subtypes has not been clarified due to the lack of selective orthosteric ligands that can discriminate between mGlu2 and mGlu3 subtypes.In this study we used mGlu2 or mGlu3 knock-out mice to dissect the specific role for these two receptors in the endogenous control of inflammatory pain and their specific contribution to the analgesic activity of mixed mGlu2/3 receptor agonists.Our results showed that mGlu2⁻(/)⁻ mice display a significantly greater pain response compared to their wild type littermates. Interestingly the increased pain sensitivity in mGlu2⁻(/)⁻ mice occurred only in the second phase of the formalin test. No differences were observed in the first phase. In contrast, mGlu3⁻(/)⁻ mice did not significantly differ from their wild type littermates in either phase of the formalin test.When systemically injected, a single administration of the mGlu2/3 agonist, LY379268 (3 mg/kg, ip), showed a significant reduction of both phases in wild-type mice and in mGlu3⁻(/)⁻ but not in mGlu2⁻(/)⁻ mice. However tolerance to the analgesic effect of LY379268 (3 mg/kg, ip) in mGlu3⁻(/)⁻ mice developed following 5 consecutive days of injection.Taken together, these results demonstrate that: (i) mGlu2 receptors play a predominant role over mGlu3 receptors in the control of inflammatory pain in mice; (ii) the analgesic activity of mixed mGlu2/3 agonists is entirely mediated by the activation of the mGlu2 subtype and (iii) the development of tolerance to the analgesic effect of mGlu2/3 agonists develops despite the lack of mGlu3 receptors.

  14. Beta adrenergic receptor blockade of feline myocardium. Cardiac mechanics, energetics, and beta adrenoceptor regulation.

    PubMed Central

    Cooper, G; Kent, R L; McGonigle, P; Watanabe, A M

    1986-01-01

    Myocardial oxygen consumption is regulated by interrelated mechanical and inotropic conditions; there is a parallel increase in the aerobic metabolism and inotropic state during beta-adrenergic stimulation under fixed mechanical conditions. In contrast, there is some evidence that beta-blockade may reduce oxygen consumption through effects independent of its influence on mechanical conditions and contractile state, and that prolonged beta-blockade may sensitize the myocardium to beta-adrenergic stimulation. To clarify these two points, the present study examined the relationship of myocardial energetics to mechanics and inotropism during acute beta-blockade and after the withdrawal of long-term beta-blockade, whereupon the basis for any effect observed was sought by characterizing the number, affinity, and affinity states of the beta-receptors as well as the coupling of activated beta-receptors to cyclic AMP generation. Studies of right ventricular papillary muscles from control and chronically beta-blocked cats demonstrated contractile and energetic properties as well as dose-response behavior and inotropic specificity suggestive of an increase in myocardial sensitivity to beta-adrenoceptor stimulation in the latter group. Assays of cardiac beta-adrenoceptors from further groups of control and pretreated cats, both in cardiac tissue and in isolated cardiac muscle cells, failed to define a difference between the two groups either in terms of receptor number and affinity or in terms of the proportion of receptors in the high-affinity state. However, coupling of the activated beta-adrenoceptors to cyclic AMP generation was enhanced in cardiac muscle cells from chronically beta-blocked cats. These data demonstrate that beta-adrenoceptor blockade (a) produces parallel effects on inotropic state and oxygen consumption without an independent effect on either and (b) increases myocardial sensitivity to beta-adrenergic stimulation after beta-blockade withdrawal, not by "up

  15. On the central mechanism underlying ghrelin's chronic pro-obesity effects in rats: new insights from studies exploiting a potent ghrelin receptor antagonist.

    PubMed

    Salomé, N; Hansson, C; Taube, M; Gustafsson-Ericson, L; Egecioglu, E; Karlsson-Lindahl, L; Fehrentz, J A; Martinez, J; Perrissoud, D; Dickson, S L

    2009-09-01

    In the present study, we explore the central nervous system mechanism underlying the chronic central effects of ghrelin with respect to increasing body weight and body fat. Specifically, using a recently developed ghrelin receptor antagonist, GHS-R1A (JMV2959), we investigate the role of GHS-R1A in mediating the effects of ghrelin on energy balance and on hypothalamic gene expression. As expected, in adult male rats, chronic central treatment with ghrelin for 14 days, when compared to vehicle-treated control rats, resulted in an increased body weight, lean mass and fat mass (assessed by dual X-ray absorptiometry), dissected white fat pad weight, cumulative food intake, food efficiency, respiratory exchange ratio and a decrease of energy expenditure. Co-administration of the ghrelin receptor antagonist JMV2959 suppressed/blocked the majority of these effects, with the notable exception of ghrelin-induced food intake and food efficiency. The hypothesis emerging from these data, namely that GHS-R1A mediates the chronic effects of ghrelin on fat accumulation, at least partly independent of food intake, is discussed in light of the accompanying data regarding the hypothalamic genes coding for peptides and receptors involved in energy balance regulation, which were found to have altered expression in these studies.

  16. Effects of the antitumor drug OSI-906, a dual inhibitor of IGF-1 receptor and insulin receptor, on the glycemic control, β-cell functions, and β-cell proliferation in male mice.

    PubMed

    Shirakawa, Jun; Okuyama, Tomoko; Yoshida, Eiko; Shimizu, Mari; Horigome, Yuka; Tuno, Takayuki; Hayasaka, Moe; Abe, Shiori; Fuse, Masahiro; Togashi, Yu; Terauchi, Yasuo

    2014-06-01

    The IGF-1 receptor has become a therapeutic target for the treatment of cancer. The efficacy of OSI-906 (linstinib), a dual inhibitor of IGF-1 receptor and insulin receptor, for solid cancers has been examined in clinical trials. The effects of OSI-906, however, on the blood glucose levels and pancreatic β-cell functions have not yet been reported. We investigated the impact of OSI-906 on glycemic control, insulin secretion, β-cell mass, and β-cell proliferation in male mice. Oral administration of OSI-906 worsened glucose tolerance in a dose-dependent manner in the wild-type mice. OSI-906 at a dose equivalent to the clinical daily dose (7.5 mg/kg) transiently evoked glucose intolerance and hyperinsulinemia. Insulin receptor substrate (IRS)-2-deficient mice and mice with diet-induced obesity, both models of peripheral insulin resistance, exhibited more severe glucose intolerance after OSI-906 administration than glucokinase-haploinsufficient mice, a model of impaired insulin secretion. Phloridzin improved the hyperglycemia induced by OSI-906 in mice. In vitro, OSI-906 showed no effect on insulin secretion from isolated islets. After daily administration of OSI-906 for a week to mice, the β-cell mass and β-cell proliferation rate were significantly increased. The insulin signals in the β-cells were apparently unaffected in those mice. Taken together, the results suggest that OSI-906 could exacerbate diabetes, especially in patients with insulin resistance. On the other hand, the results suggest that the β-cell mass may expand in response to chemotherapy with this drug.

  17. Research of the disbalance mechanism of dual collector lateral bipolar magnetotransistor

    NASA Astrophysics Data System (ADS)

    Tikhonov, R. D.; Polomoshnov, S. A.; Kozlov, A. V.; Krasukov, A. J.

    2008-03-01

    The experimental research initial disbalance potential collectors from the scheme of inclusion two-collector lateral bipolar magnetotransistor (BMT) NPN-type, generated in a well is lead. By means of device-technological modelling the mechanism of occurrence initial disbalance is investigated and the way of its reduction initial disbalance is certain at maintenance of preservation of high sensitivity. The choice of the operating mode bipolar magnetotransistor is based on the distributions of the emitter injected electron currents in two symmetrical base electrodes - two contacts to the base-well, in two contacts to a substrate, in two collectors. The mode of magnetotransistor based on the influence of a magnetic field. Reduction of initial disbalance allows to increase relative size of a output valid signal ΔU= UC1(B) - UC2(B) - UC1(0) + UC2(0).

  18. Dual Raman-Brillouin microscope for chemical and mechanical characterization and imaging (Conference Presentation)

    NASA Astrophysics Data System (ADS)

    Yakovlev, Vladislav V.

    2016-03-01

    We present a unique confocal microscope capable of measuring the Raman and Brillouin spectra simultaneously from a single spatial location. Raman and Brillouin scattering offer complementary information about a material's chemical and mechanical structure, respectively, and concurrent monitoring of both of these spectra would set a new standard for material characterization. We achieve this by applying recent innovations in Brillouin spectroscopy that reduce the necessary acquisition times to durations comparable to conventional Raman spectroscopy while attaining a high level of spectral accuracy. To demonstrate the potential of the system, we apply this system for imaging melanoma, arteriosclerotic plaque, embryonic development, bones and biomaterials. The developed instrument has the potential for very diverse analytical applications in basic biomedical science and biomedical diagnostics and imaging.

  19. Human formyl peptide receptor ligand binding domain(s). Studies using an improved mutagenesis/expression vector reveal a novel mechanism for the regulation of receptor occupancy.

    PubMed

    Perez, H D; Vilander, L; Andrews, W H; Holmes, R

    1994-09-09

    Recently, we reported the domain requirements for the binding of formyl peptide to its specific receptor. Based on experiments using receptor chimeras, we also postulated an importance for the amino-terminal domain of the receptor in ligand binding (Perez, H. D., Holmes, R., Vilander, L., Adams, R., Manzana, W., Jolley, D., and Andrews, W. H. (1993) J. Biol. Chem. 268, 2292-2295). We have begun to perform a detailed analysis of the regions within the formyl peptide receptor involved in ligand binding. To address the importance of the receptor amino-terminal domain, we substituted (or inserted) hydrophilic sequences within the amino-terminal domain, expressed the receptors, and determined their ability to bind ligand. A stretch of nine amino acids next to the initial methionine was identified as crucial for receptor occupancy. A peptide containing such a sequence specifically completed binding of the ligand to the receptor. Alanine screen mutagenesis of the second extracellular domain also identified amino acids involved in ligand binding as well as a disulfide bond (Cys98 to Cys176) crucial for maintaining the binding pocket. These studies provide evidence for a novel mechanism involved in regulation of receptor occupancy. Binding of the ligand induces conformational changes in the receptor that result in the apposition of the amino-terminal domain over the ligand, providing a lid to the binding pocket.

  20. Dual Mechanism of Ion Permeation through VDAC Revealed with Inorganic Phosphate Ions and Phosphate Metabolites

    PubMed Central

    Krammer, Eva-Maria; Vu, Giang Thi; Homblé, Fabrice; Prévost, Martine

    2015-01-01

    In the exchange of metabolites and ions between the mitochondrion and the cytosol, the voltage-dependent anion channel (VDAC) is a key element, as it forms the major transport pathway for these compounds through the mitochondrial outer membrane. Numerous experimental studies have promoted the idea that VDAC acts as a regulator of essential mitochondrial functions. In this study, using a combination of molecular dynamics simulations, free-energy calculations, and electrophysiological measurements, we investigated the transport of ions through VDAC, with a focus on phosphate ions and metabolites. We showed that selectivity of VDAC towards small anions including monovalent phosphates arises from short-lived interactions with positively charged residues scattered throughout the pore. In dramatic contrast, permeation of divalent phosphate ions and phosphate metabolites (AMP and ATP) involves binding sites along a specific translocation pathway. This permeation mechanism offers an explanation for the decrease in VDAC conductance measured in the presence of ATP or AMP at physiological salt concentration. The binding sites occur at similar locations for the divalent phosphate ions, AMP and ATP, and contain identical basic residues. ATP features a marked affinity for a central region of the pore lined by two lysines and one arginine of the N-terminal helix. This cluster of residues together with a few other basic amino acids forms a “charged brush” which facilitates the passage of the anionic metabolites through the pore. All of this reveals that VDAC controls the transport of the inorganic phosphates and phosphate metabolites studied here through two different mechanisms. PMID:25860993

  1. Concurrent Phosphorus Recovery and Energy Generation in Mediator-Less Dual Chamber Microbial Fuel Cells: Mechanisms and Influencing Factors.

    PubMed

    Almatouq, Abdullah; Babatunde, Akintunde O

    2016-03-29

    This study investigated the mechanism and key factors influencing concurrent phosphorus (P) recovery and energy generation in microbial fuel cells (MFC) during wastewater treatment. Using a mediator-less dual chamber microbial fuel cell operated for 120 days; P was shown to precipitate as struvite when ammonium and magnesium chloride solutions were added to the cathode chamber. Monitoring data for chemical oxygen demand (COD), pH, oxidation reduction potential (ORP) and aeration flow rate showed that a maximum 38% P recovery was achieved; and this corresponds to 1.5 g/L, pH > 8, -550 ± 10 mV and 50 mL/min respectively, for COD, pH(cathode), ORP and cathode aeration flow rate. More importantly, COD and aeration flow rate were shown to be the key influencing factors for the P recovery and energy generation. Results further show that the maximum P recovery corresponds to 72 mW/m² power density. However, the energy generated at maximum P recovery was not the optimum; this shows that whilst P recovery and energy generation can be concurrently achieved in a microbial fuel cell, neither can be at the optimal value.

  2. Mechanisms of direct hydrogen peroxide synthesis on silicon and phosphorus dual-doped graphene: a DFT-D study.

    PubMed

    Li, Shuo; Lu, Zhansheng; Zhang, Yi; Ma, Dongwei; Yang, Zongxian

    2017-03-17

    Hydrogen peroxide (H2O2) is an important chemical commodity, with demand growing significantly in chemical synthesis due to its green characteristics. The mechanisms of the direct synthesis of hydrogen peroxide (DSHP) on metal-free silicon and phosphorus dual-doped graphene (Si-P-G) catalyst, based on a dispersion-corrected density functional theory (DFT-D) method, are systematically investigated. The most stable Si-P-G catalyst is presented, with the local region of dopants shown to play an important role in the adsorption and reduction of oxygen. A two-electron pathway is probable for DSHP on Si-P-G according to kinetic and thermodynamic analyses. The hydrogenation of O2 to OOH is the rate-limiting step, with a small barrier energy of 0.66 eV, and the potential energy surface is downhill by Gibbs free energy calculations. All results indicate that Si-P-G is a novel catalyst with high activity and good selectivity for DSHP.

  3. Inhibition of CD73 AMP hydrolysis by a therapeutic antibody with a dual, non-competitive mechanism of action

    PubMed Central

    Geoghegan, James C.; Diedrich, Gundo; Lu, Xiaojun; Rosenthal, Kim; Sachsenmeier, Kris F.; Wu, Herren; Dall'Acqua, William F.; Damschroder, Melissa M.

    2016-01-01

    ABSTRACT CD73 (ecto-5′-nucleotidase) has recently been established as a promising immuno-oncology target. Given its role in activating purinergic signaling pathways to elicit immune suppression, antagonizing CD73 (i.e., releasing the brake) offers a complimentary pathway to inducing anti-tumor immune responses. Here, we describe the mechanistic activity of a new clinical therapeutic, MEDI9447, a human monoclonal antibody that non-competitively inhibits CD73 activity. Epitope mapping, structural, and mechanistic studies revealed that MEDI9447 antagonizes CD73 through dual mechanisms of inter-CD73 dimer crosslinking and/or steric blocking that prevent CD73 from adopting a catalytically active conformation. To our knowledge, this is the first report of an antibody that inhibits an enzyme's function through 2 distinct modes of action. These results provide a finely mapped epitope that can be targeted for selective, potent, and non-competitive inhibition of CD73, as well as establish a strategy for inhibiting enzymes that function in both membrane-bound and soluble states. PMID:26854859

  4. Concurrent Phosphorus Recovery and Energy Generation in Mediator-Less Dual Chamber Microbial Fuel Cells: Mechanisms and Influencing Factors

    PubMed Central

    Almatouq, Abdullah; Babatunde, Akintunde O.

    2016-01-01

    This study investigated the mechanism and key factors influencing concurrent phosphorus (P) recovery and energy generation in microbial fuel cells (MFC) during wastewater treatment. Using a mediator-less dual chamber microbial fuel cell operated for 120 days; P was shown to precipitate as struvite when ammonium and magnesium chloride solutions were added to the cathode chamber. Monitoring data for chemical oxygen demand (COD), pH, oxidation reduction potential (ORP) and aeration flow rate showed that a maximum 38% P recovery was achieved; and this corresponds to 1.5 g/L, pH > 8, −550 ± 10 mV and 50 mL/min respectively, for COD, pHcathode, ORP and cathode aeration flow rate. More importantly, COD and aeration flow rate were shown to be the key influencing factors for the P recovery and energy generation. Results further show that the maximum P recovery corresponds to 72 mW/m2 power density. However, the energy generated at maximum P recovery was not the optimum; this shows that whilst P recovery and energy generation can be concurrently achieved in a microbial fuel cell, neither can be at the optimal value. PMID:27043584

  5. Formulating a Sulfonated Anti-Viral Dendrimer in a Vaginal Microbicidal Gel having Dual Mechanisms of Action

    PubMed Central

    Mumper, Russell J.; Bell, Michael A.; Worthen, David R.; Cone, Richard A.; Lewis, Gareth R.; Moench, Thomas R.

    2009-01-01

    SPL7013 is the sodium salt of a sulfonated dendrimer that has potent antiviral properties. VivaGel®, a topical gel containing 3% w/w SPL7013, has been shown to be safe and well-tolerated in human clinical studies. BufferGel® is a Carbopol®-based acidic buffering gel that enhances the natural protective action of the vagina to produce a broad-spectrum microbicidal environment. The positive attributes of both gels were combined into a combination vaginal microbicidal gel having dual mechanisms of action. A 3% w/w SPL7013 combination gel, pH 3.7, was developed and fully characterized, and was shown to have more than 2-fold greater acidic buffering capacity than BufferGel. Ultracentrifugation experiments demonstrated that SPL7013 was not sequestered or entropically trapped in the viscous gel, thereby confirming, along with viral challenge studies, that SPL7013 has sufficient mobility in the viscous gel to exert antiviral properties. PMID:19040181

  6. Structure-affinity relationships and pharmacological characterization of new alkyl-resorcinol cannabinoid receptor ligands: Identification of a dual cannabinoid receptor/TRPA1 channel agonist.

    PubMed

    Brizzi, Antonella; Aiello, Francesca; Marini, Pietro; Cascio, Maria Grazia; Corelli, Federico; Brizzi, Vittorio; De Petrocellis, Luciano; Ligresti, Alessia; Luongo, Livio; Lamponi, Stefania; Maione, Sabatino; Pertwee, Roger G; Di Marzo, Vincenzo

    2014-09-01

    In our ongoing program aimed at deeply investigating the endocannabinoid system (ES), a set of new alkyl-resorcinol derivatives was prepared focusing on the nature and the importance of the carboxamide functionality. Binding studies on CB1 and CB2 receptors, monoacylglycerol lipase (MAGL) and fatty acid amide hydrolase (FAAH) showed that some of the newly developed compounds behaved as very potent cannabinoid receptor ligands (Ki in the nanomolar range) while, however, none of them was able to inhibit MAGL and/or FAAH. Derivative 11 was a potent CB1 and CB2 ligand, with Ki values similar to WIN 55,212, exhibiting a CB1 and CB2 agonist profile in vitro. In the formalin test of peripheral acute and inflammatory pain in mice, this compound showed a weak and delayed antinociceptive effect against the second phase of the nocifensive response, exhibiting, interestingly, a quite potent transient receptor potential ankyrin type-1 (TRPA1) channel agonist activity. Moreover, derivative 14, characterized by lower affinity but higher CB2 selectivity than 11, proved to behave as a weak CB2 competitive inverse agonist.

  7. Transmembrane chemokines act as receptors in a novel mechanism termed inverse signaling

    PubMed Central

    Hattermann, Kirsten; Gebhardt, Henrike; Krossa, Sebastian; Ludwig, Andreas; Lucius, Ralph

    2016-01-01

    The transmembrane chemokines CX3CL1/fractalkine and CXCL16 are widely expressed in different types of tumors, often without an appropriate expression of their classical receptors. We observed that receptor-negative cancer cells could be stimulated by the soluble chemokines. Searching for alternative receptors we detected that all cells expressing or transfected with transmembrane chemokine ligands bound the soluble chemokines with high affinity and responded by phosphorylation of intracellular kinases, enhanced proliferation and anti-apoptosis. This activity requires the intracellular domain and apparently the dimerization of the transmembrane chemokine ligand. Thus, shed soluble chemokines can generate auto- or paracrine signals by binding and activating their transmembrane forms. We term this novel mechanism “inverse signaling”. We suppose that inverse signaling is an autocrine feedback and fine-tuning system in the communication between cells that in tumors supports stabilization and proliferation. DOI: http://dx.doi.org/10.7554/eLife.10820.001 PMID:26796342

  8. Cannabinoid and heroin activation of mesolimbic dopamine transmission by a common mu1 opioid receptor mechanism.

    PubMed

    Tanda, G; Pontieri, F E; Di Chiara, G

    1997-06-27

    The effects of the active ingredient of Cannabis, Delta9-tetrahydrocannabinol (Delta9-THC), and of the highly addictive drug heroin on in vivo dopamine transmission in the nucleus accumbens were compared in Sprague-Dawley rats by brain microdialysis. Delta9-THC and heroin increased extracellular dopamine concentrations selectively in the shell of the nucleus accumbens; these effects were mimicked by the synthetic cannabinoid agonist WIN55212-2. SR141716A, an antagonist of central cannabinoid receptors, prevented the effects of Delta9-THC but not those of heroin. Naloxone, a generic opioid antagonist, administered systemically, or naloxonazine, an antagonist of micro1 opioid receptors, infused into the ventral tegmentum, prevented the action of cannabinoids and heroin on dopamine transmission. Thus, Delta9-THC and heroin exert similar effects on mesolimbic dopamine transmission through a common mu1 opioid receptor mechanism located in the ventral mesencephalic tegmentum.

  9. CD44 Antibody Inhibition of Macrophage Phagocytosis Targets Fcγ Receptor- and Complement Receptor 3-Dependent Mechanisms.

    PubMed

    Amash, Alaa; Wang, Lin; Wang, Yawen; Bhakta, Varsha; Fairn, Gregory D; Hou, Ming; Peng, Jun; Sheffield, William P; Lazarus, Alan H

    2016-04-15

    Targeting CD44, a major leukocyte adhesion molecule, using specific Abs has been shown beneficial in several models of autoimmune and inflammatory diseases. The mechanisms contributing to the anti-inflammatory effects of CD44 Abs, however, remain poorly understood. Phagocytosis is a key component of immune system function and can play a pivotal role in autoimmune states where CD44 Abs have shown to be effective. In this study, we show that the well-known anti-inflammatory CD44 Ab IM7 can inhibit murine macrophage phagocytosis of RBCs. We assessed three selected macrophage phagocytic receptor systems: Fcγ receptors (FcγRs), complement receptor 3 (CR3), and dectin-1. Treatment of macrophages with IM7 resulted in significant inhibition of FcγR-mediated phagocytosis of IgG-opsonized RBCs. The inhibition of FcγR-mediated phagocytosis was at an early stage in the phagocytic process involving both inhibition of the binding of the target RBC to the macrophages and postbinding events. This CD44 Ab also inhibited CR3-mediated phagocytosis of C3bi-opsonized RBCs, but it did not affect the phagocytosis of zymosan particles, known to be mediated by the C-type lectin dectin-1. Other CD44 Abs known to have less broad anti-inflammatory activity, including KM114, KM81, and KM201, did not inhibit FcγR-mediated phagocytosis of RBCs. Taken together, these findings demonstrate selective inhibition of FcγR and CR3-mediated phagocytosis by IM7 and suggest that this broadly anti-inflammatory CD44 Ab inhibits these selected macrophage phagocytic pathways. The understanding of the immune-regulatory effects of CD44 Abs is important in the development and optimization of therapeutic strategies for the potential treatment of autoimmune conditions.

  10. Intrathecal alpha2 adrenoceptor agonist clonidine inhibits mechanical transmission in mouse spinal cord via activation of muscarinic M1 receptors.

    PubMed

    Honda, Kenji; Koga, Kohei; Moriyama, Tomoko; Koguchi, Masako; Takano, Yukio; Kamiya, Hiro-o

    2002-04-12

    We examined the role of the spinal muscarinic receptor subtype in the anti-nociceptive effect of intrathecal (i.t.) alpha2 adrenoceptor agonist clonidine in mice. I.t. injection of the muscarinic receptor antagonist atropine completely inhibited i.t. clonidine-induced increase in the mechanical threshold, but did not affect the increase in tail-flick latency induced by i.t. clonidine. The clonidine-induced increase in mechanical threshold was inhibited by i.t. injection of the M1 receptor antagonist pirenzepine in a dose-dependent manner, and by the M3 receptor antagonist 4-DAMP, but not by the M2 receptor antagonist methoctramine. The potency of pirenzepine was greater than that of 4-DAMP. These results suggest that the clonidine-induced increase in mechanical threshold is mediated via the activation of M1 receptors in the spinal cord.

  11. Symposium overview: mechanism of action of nicotine on neuronal acetylcholine receptors, from molecule to behavior.

    PubMed

    Narahashi, T; Fenster, C P; Quick, M W; Lester, R A; Marszalec, W; Aistrup, G L; Sattelle, D B; Martin, B R; Levin, E D

    2000-10-01

    Nicotine has long been known to interact with nicotinic acetylcholine (ACh) receptors since Langley used it extensively to chart sympathetic ganglia a century ago. It has also been used as an effective insecticide. However, it was not until the 1990s that the significance of nicotine was increasingly recognized from the toxicological, pharmacological, and environmental points of view. This is partly because studies of neuronal nicotinic ACh receptors are rapidly emerging from orphan status, fueled by several lines of research. Since Alzheimer's disease is known to be associated with down-regulation of cholinergic activity in the brain, a variety of nicotine derivatives are being tested and developed for treatment of the disease. Public awareness of the adverse effects of nicotine has reached the highest level recently. Since insect resistance to insecticides is one of the most serious issues in the pest-control arena, it is an urgent requirement to develop new insecticides that act on target sites not shared by the existing insecticides. The neuronal nicotinic ACh receptor is one of them, and new nicotinoids are being developed. Thus, the time is ripe to discuss the mechanism of action of nicotine from a variety of angles, including the molecular, physiological, and behavioral points of view. This Symposium covered a wide area of nicotine studies: genetic, genomic, and functional aspects of nicotinic ACh receptors were studied, as related to anthelmintics and insecticides; interactions between ethanol and nicotine out the ACh receptor were analyzed, in an attempt to explain the well-known heavy drinker-heavy smoker correlation; the mechanisms that underlie the desensitization of ACh receptors were studied as related to nicotine action; selective pharmacological profiles of nicotine, and descriptions of some derivatives were described; and chronic nicotine infusion effects on memory were examined using animal models.

  12. Effect of grain size on the mechanical properties of dual phase Fe/Si/C steels

    SciTech Connect

    Ahn, J.H.

    1983-08-01

    For an Fe/2Si/0.1C steel with an intermediate quenching heat treatment, it was found that as the prior austenite grain size is refined, significant improvements in total elongation, reduction in area and impact toughness can be achieved, while uniform elongation, yield and tensile strengths are not affected. These improvements are analyzed in terms of microstructure and fracture characteristics. The cleavage cracks propagate nearly straight without deviation at the ferrite/martensite interfaces within the sub-units of the DFM structure, but change their path at high angle sub-unit boundaries. The crack is less likely to be deflected at the ferrite/martensite interface because the interface is coherent. Comparison of optical micrographs and SEM fractographs has shown that there is close agreement between the sub-unit size and cleavage facet size. The observations lead to the conclusion that the sub-unit size is the basic microstructure unit controlling the fracture behavior of DFM steels produced by the intermediate quenching heat treatment. A controlled rolling process was undertaken to obtain grain refined DFM steels. Results showed that this produces micro-duplex structures with attractive mechanical properties in an economicl way.

  13. Design of a dual nanostructured lipid carrier formulation based on physicochemical, rheological, and mechanical properties

    NASA Astrophysics Data System (ADS)

    Vitorino, Carla; Alves, Luís; Antunes, Filipe E.; Sousa, João J.; Pais, Alberto A. C. C.

    2013-10-01

    The synergy between nanostructured lipid carriers (NLC) and chemical penetration enhancers provides the basis for a promising strategy to effectively deliver drugs through the skin. In the present work, focus is given to the study of the interaction of limonene, ethanol, and Carbopol Ultrez® 10NF, as the gelling agent, with a co-encapsulating NLC dispersion, containing both olanzapine and simvastatin. The analysis is based on rheological, mechanical, and physicochemical properties. The nanoparticle size ranged from 130 to 400 nm, depending on the system considered. The inclusion of carbopol resulted in a considerable increase in the NLC particle size, which was attributed to carbomer bridging nanoparticles, resulting in some particle aggregation. Rheological measurements indicated that the viscosity of the neat carbopol hydrogel was reduced by the enhancers and to a higher extent by the presence of NLC. A more apolar medium can be pointed out as a general explanation, while a polymer/nanoparticle hydrophobic interaction coupled with surfactant/polymer H-bonding provides the rationale for the NLC effect. The inclusion of enhancers and a carbomer largely contributed to the physicochemical stability of the NLC formulation, as revealed by the low transmission profiles and more negative zeta potential values.

  14. Anticalculus effects of a novel, dual-phase polypyrophosphate dentifrice: chemical basis, mechanism, and clinical response.

    PubMed

    White, D J; Gerlach, R W

    2000-11-15

    A primary patient motivation for oral hygiene is effective cleaning. Dentifrice serves this function by including ingredients such as abrasives, surfactants, and specialized cleaning ingredients such as anticalculus agents. This introductory article aims to introduce professionals, educators, and researchers on the rationale behind the development of an improved cleaning dentifrice formulation, Crest Multicare Advanced Cleaning. This new dentifrice is based upon the application of an improved tartar control/cleaning ingredient that is a polymeric adjunct of a pyrophosphate anion commonly applied in tartar control and stain control whitening dentifrices. The polypyrophosphate anion, also referred to as sodium hexametaphosphate, produces superior activity and substantivity on oral surfaces as compared to both pyrophosphate and some other commonly used dental cleaning ingredients and cleaning/conditioning adjuncts. The increased activity and substantivity translate into significant improvements in the prevention of dental stains and supragingival calculus and in the non-abrasive removal of dental stains. This article describes the structure of polypyrophosphate as compared to the parent pyrophosphate molecule, the rationale for its improved chemistry, and, in particular, its tartar control chemistry. In addition, the fundamental mechanisms of calculus formation and inhibition are reviewed. Lastly, a preliminary clinical study evaluating the improved efficacy of a polypyrophosphate dentifrice is described where the tartar control activity of the polypyrophosphate dentifrice is shown to be superior to that of a clinically established and marketed industry standard pyrophosphate dentifrice.

  15. A dual inhibitory mechanism sufficient to maintain cell cycle restricted CENP-A assembly

    PubMed Central

    Stankovic, Ana; Guo, Lucie Y.; Mata, João F.; Bodor, Dani L.; Cao, Xing-Jun; Bailey, Aaron O.; Shabanowitz, Jeffrey; Hunt, Donald F.; Garcia, Benjamin A.; Black, Ben E.; Jansen, Lars E.T

    2017-01-01

    Summary Chromatin featuring the H3 variant CENP-A at the centromere is critical for its mitotic function and epigenetic maintenance. Assembly of centromeric chromatin is restricted to G1 phase through inhibitory action of Cdk1/2 kinases in other phases of the cell cycle. Here, we identify the two key targets sufficient to maintain cell cycle control of CENP-A assembly. We uncovered a single phosphorylation site in the licensing factor M18BP1 and a cyclin A binding site in the CENP-A chaperone, HJURP, mediating specific inhibitory phosphorylation. Simultaneous expression of mutant proteins lacking these residues, results in complete uncoupling from the cell cycle. Consequently, CENP-A assembly is fully recapitulated under high Cdk activities, indistinguishable from G1 assembly. We find that Cdk-mediated inhibition is exerted by sequestering active factors away from the centromere. Finally, we show that displacement of M18BP1 from the centromere is critical for the assembly mechanism of CENP-A. PMID:28017591

  16. Dual Mechanisms of Ion Absorption in Relation to Long Distance Transport in Plants

    PubMed Central

    Luttge, Ulrich; Laties, George G.

    1966-01-01

    The characteristics of ion transport to the shoots of young corn seedlings were studied with respect to the nature of the isotherm through a wide concentration range, the competitive influence of closely related ions upon the transport of a given ion, and the influence of the counter-ion. Both with respect to 36Cl and 86Rb transport, the characteristics of the process in every way resemble uptake by non-vacuolate root tips wherein the plasma membrane is the only membrane involved in absorption, and where system 1 — of the 2 systems which can be shown to participate in absorption by vacuolate tissue — is the only system operative. Net ion uptake by the roots per se was shown to display both the high affinity (system 1) and low affinity (system 2) mechanisms. It is concluded that the symplastic theory of ion movement to the xylem is valid, and that the contention that system 1 operates at the plasma membrane while system 2 functions at the tonoplast is strengthened. PMID:16656435

  17. Dual strain mechanisms in a lead-free morphotropic phase boundary ferroelectric

    PubMed Central

    Walker, Julian; Simons, Hugh; Alikin, Denis O.; Turygin, Anton P.; Shur, Vladimir Y.; Kholkin, Andrei L.; Ursic, Hana; Bencan, Andreja; Malic, Barbara; Nagarajan, Valanoor; Rojac, Tadej

    2016-01-01

    Electromechanical properties such as d33 and strain are significantly enhanced at morphotropic phase boundaries (MPBs) between two or more different crystal structures. Many actuators, sensors and MEMS devices are therefore systems with MPBs, usually between polar phases in lead (Pb)-based ferroelectric ceramics. In the search for Pb-free alternatives, systems with MPBs between polar and non-polar phases have recently been theorized as having great promise. While such an MPB was identified in rare-earth (RE) modified bismuth ferrite (BFO) thin films, synthesis challenges have prevented its realization in ceramics. Overcoming these, we demonstrate a comparable electromechanical response to Pb-based materials at the polar-to-non-polar MPB in Sm modified BFO. This arises from ‘dual’ strain mechanisms: ferroelectric/ferroelastic switching and a previously unreported electric-field induced transition of an anti-polar intermediate phase. We show that intermediate phases play an important role in the macroscopic strain response, and may have potential to enhance electromechanical properties at polar-to-non-polar MPBs. PMID:26791098

  18. A simple but effective dual redox and fluorescent ion pair receptor based on a ferrocene-imidazopyrene dyad.

    PubMed

    Alfonso, María; Espinosa, Arturo; Tárraga, Alberto; Molina, Pedro

    2011-04-15

    The ferrocene-imidazopyrene dyad, bearing the imidazole ring as the only receptor site, acts as a redox and optical molecular sensor for ion pairs, exhibiting an easily detectable signal change in the redox potential of the ferrocene/ferrocinium redox couple and in the emission spectrum. Perturbation of the emission spectrum follows the order Pb(2+) > Hg(2+) > Zn(2+) for cations and H(2)PO(4)(-) > AcO(-) for anions.

  19. A gate-latch-lock mechanism for hormone signalling by abscisic acid receptors

    SciTech Connect

    Melcher, Karsten; Ng, Ley-Moy; Zhou, X Edward; Soon, Fen-Fen; Xu, Yong; Suino-Powell, Kelly M; Park, Sang-Youl; Weiner, Joshua J; Fujii, Hiroaki; Chinnusamy, Viswanathan; Kovach, Amanda; Li, Jun; Wang, Yonghong; Li, Jiayang; Peterson, Francis C; Jensen, Davin R; Yong, Eu-Leong; Volkman, Brian F; Cutler, Sean R; Zhu, Jian-Kang; Xu, H Eric

    2010-01-12

    Abscisic acid (ABA) is a ubiquitous hormone that regulates plant growth, development and responses to environmental stresses. Its action is mediated by the PYR/PYL/RCAR family of START proteins, but it remains unclear how these receptors bind ABA and, in turn, how hormone binding leads to inhibition of the downstream type 2C protein phosphatase (PP2C) effectors. Here we report crystal structures of apo and ABA-bound receptors as well as a ternary PYL2-ABA-PP2C complex. The apo receptors contain an open ligand-binding pocket flanked by a gate that closes in response to ABA by way of conformational changes in two highly conserved β-loops that serve as a gate and latch. Moreover, ABA-induced closure of the gate creates a surface that enables the receptor to dock into and competitively inhibit the PP2C active site. A conserved tryptophan in the PP2C inserts directly between the gate and latch, which functions to further lock the receptor in a closed conformation. Together, our results identify a conserved gate-latch-lock mechanism underlying ABA signalling.

  20. Structural mechanism of ligand activation in human calcium-sensing receptor

    SciTech Connect

    Geng, Yong; Mosyak, Lidia; Kurinov, Igor; Zuo, Hao; Sturchler, Emmanuel; Cheng, Tat Cheung; Subramanyam, Prakash; Brown, Alice P.; Brennan, Sarah C.; Mun, Hee-chang; Bush, Martin; Chen, Yan; Nguyen, Trang X.; Cao, Baohua; Chang, Donald D.; Quick, Matthias; Conigrave, Arthur D.; Colecraft, Henry M.; McDonald, Patricia; Fan, Qing R.

    2016-07-19

    Human calcium-sensing receptor (CaSR) is a G-protein-coupled receptor (GPCR) that maintains extracellular Ca2+homeostasis through the regulation of parathyroid hormone secretion. It functions as a disulfide-tethered homodimer composed of three main domains, the Venus Flytrap module, cysteine-rich domain, and seven-helix transmembrane region. Here, we present the crystal structures of the entire extracellular domain of CaSR in the resting and active conformations. We provide direct evidence that L-amino acids are agonists of the receptor. In the active structure, L-Trp occupies the orthosteric agonist-binding site at the interdomain cleft and is primarily responsible for inducing extracellular domain closure to initiate receptor activation. Our structures reveal multiple binding sites for Ca2+and PO43-ions. Both ions are crucial for structural integrity of the receptor. While Ca2+ions stabilize the active state, PO43-ions reinforce the inactive conformation. The activation mechanism of CaSR involves the formation of a novel dimer interface between subunits.

  1. Histamine enhances inhibitory avoidance memory consolidation through a H2 receptor-dependent mechanism.

    PubMed

    da Silva, Weber C; Bonini, Juliana S; Bevilaqua, Lia R M; Izquierdo, Iván; Cammarota, Martín

    2006-07-01

    Several evidences suggest that brain histamine is involved in memory consolidation but the actual contribution of the hippocampal histaminergic system to this process remains controversial. Here, we show that when infused into the CA1 region of the dorsal hippocampus immediately after training in an inhibitory avoidance task, but not later, histamine induced a dose-dependent promnesic effect without altering locomotor activity, exploratory behavior, anxiety state or retrieval of the avoidance response. The facilitatory effect of intra-CA1 histamine was mimicked by the histamine N-methyltransferase inhibitor SKF-91844 as well as by the H2 receptor agonist dimaprit and it was blocked completely by the H2 receptor antagonist ranitidine. Conversely, the promnesic action of histamine was unaffected by the H1 receptor antagonist pyrilamine, the H3 receptor antagonist, thioperamide, and the NMDAr polyamine-binding site antagonist ifenprodil. By themselves, ranitidine, pyrilamine, thioperamide, and ifenprodil did not affect IA memory consolidation. Our data indicate that, when given into CA1, histamine enhances memory consolidation through a mechanism that involves activation of H2 receptors; however, endogenous CA1 histamine does not seem to participate in the consolidation of IA memory at least at the post-training times analyzed.

  2. Pathogenic Mechanism of an Autism-Associated Neuroligin Mutation Involves Altered AMPA-Receptor Trafficking

    PubMed Central

    Chanda, Soham; Aoto, Jason; Lee, Sung-Jin; Wernig, Marius; Südhof, Thomas C.

    2015-01-01

    Neuroligins are postsynaptic cell-adhesion molecules that bind to presynaptic neurexins. Although the general synaptic role of neuroligins is undisputed, their specific functions at a synapse remain unclear, even controversial. Moreover, many neuroligin gene mutations were associated with autism, but the pathophysiological relevance of these mutations is often unknown, and their mechanisms of action uninvestigated. Here, we examine the synaptic effects of an autism-associated neuroligin-4 substitution (called R704C) which mutates a cytoplasmic arginine residue that is conserved in all neuroligins. We show that the R704C mutation, when introduced into neuroligin-3, enhances the interaction between neuroligin-3 and AMPA-receptors, increases AMPA-receptor internalization, and decreases postsynaptic AMPA-receptor levels. When introduced into neuroligin-4, conversely, the R704C mutation unexpectedly elevated AMPA-receptor mediated synaptic responses. These results suggest a general functional link between neuroligins and AMPA-receptors, indicate that both neuroligin-3 and -4 act at excitatory synapses but perform surprisingly distinct functions, and demonstrate that the R704C mutation significantly impairs the normal function of neuroligin-4, thereby validating its pathogenicity. PMID:25778475

  3. Molecular mechanism of ATP binding and ion channel activation in P2X receptors

    SciTech Connect

    Hattori, Motoyuki; Gouaux, Eric

    2012-10-24

    P2X receptors are trimeric ATP-activated ion channels permeable to Na{sup +}, K{sup +} and Ca{sup 2+}. The seven P2X receptor subtypes are implicated in physiological processes that include modulation of synaptic transmission, contraction of smooth muscle, secretion of chemical transmitters and regulation of immune responses. Despite the importance of P2X receptors in cellular physiology, the three-dimensional composition of the ATP-binding site, the structural mechanism of ATP-dependent ion channel gating and the architecture of the open ion channel pore are unknown. Here we report the crystal structure of the zebrafish P2X4 receptor in complex with ATP and a new structure of the apo receptor. The agonist-bound structure reveals a previously unseen ATP-binding motif and an open ion channel pore. ATP binding induces cleft closure of the nucleotide-binding pocket, flexing of the lower body {beta}-sheet and a radial expansion of the extracellular vestibule. The structural widening of the extracellular vestibule is directly coupled to the opening of the ion channel pore by way of an iris-like expansion of the transmembrane helices. The structural delineation of the ATP-binding site and the ion channel pore, together with the conformational changes associated with ion channel gating, will stimulate development of new pharmacological agents.

  4. Structural mechanism of ligand activation in human calcium-sensing receptor

    PubMed Central

    Geng, Yong; Mosyak, Lidia; Kurinov, Igor; Zuo, Hao; Sturchler, Emmanuel; Cheng, Tat Cheung; Subramanyam, Prakash; Brown, Alice P; Brennan, Sarah C; Mun, Hee-chang; Bush, Martin; Chen, Yan; Nguyen, Trang X; Cao, Baohua; Chang, Donald D; Quick, Matthias; Conigrave, Arthur D; Colecraft, Henry M; McDonald, Patricia; Fan, Qing R

    2016-01-01

    Human calcium-sensing receptor (CaSR) is a G-protein-coupled receptor (GPCR) that maintains extracellular Ca2+ homeostasis through the regulation of parathyroid hormone secretion. It functions as a disulfide-tethered homodimer composed of three main domains, the Venus Flytrap module, cysteine-rich domain, and seven-helix transmembrane region. Here, we present the crystal structures of the entire extracellular domain of CaSR in the resting and active conformations. We provide direct evidence that L-amino acids are agonists of the receptor. In the active structure, L-Trp occupies the orthosteric agonist-binding site at the interdomain cleft and is primarily responsible for inducing extracellular domain closure to initiate receptor activation. Our structures reveal multiple binding sites for Ca2+ and PO43- ions. Both ions are crucial for structural integrity of the receptor. While Ca2+ ions stabilize the active state, PO43- ions reinforce the inactive conformation. The activation mechanism of CaSR involves the formation of a novel dimer interface between subunits. DOI: http://dx.doi.org/10.7554/eLife.13662.001 PMID:27434672

  5. Interaction mechanism of insulin-like peptide 5 with relaxin family peptide receptor 4.

    PubMed

    Hu, Meng-Jun; Wei, Dian; Shao, Xiao-Xia; Wang, Jia-Hui; Liu, Ya-Li; Xu, Zeng-Guang; Guo, Zhan-Yun

    2017-04-01

    Insulin-like peptide 5 (INSL5) is a gut peptide hormone belonging to the insulin/relaxin superfamily. It is implicated in the regulation of food intake and glucose homeostasis by activating relaxin family peptide receptor 4 (RXFP4). Previous studies have suggested that the B-chain is important for INSL5 activity against RXFP4. However, functionalities of the B-chain residues have not yet been systematically studied. In the present work, we conducted alanine-scanning mutagenesis of the B-chain residues of human INSL5 to obtain an overview of their contributions. Binding and activation assays of these INSL5 mutants with human RXFP4 identified two essential exposed B-chain C-terminal residues (B23Arg and B24Trp) and one important exposed central B-chain residue (B16Ile). These three determinant residues together with the C-terminal carboxylate moiety probably constitute a central receptor-binding patch that forms critical hydrophobic and electrostatic interactions with RXFP4 during INSL5 binding. Some other exposed residues, including B10Glu, B12Ile, B13Arg, B17Tyr, B21Ser, and B22Ser, made minor contributions to INSL5 function. These auxiliary residues are scattered around the edge of the central receptor-binding patch, and thus form a peripheral receptor-binding patch on the surface of INSL5. Our present work provides new insights into the interaction mechanism of INSL5 with its receptor RXFP4.

  6. X-ray structure, symmetry and mechanism of an AMPA-subtype glutamate receptor

    SciTech Connect

    Sobolevsky, Alexander I.; Rosconi, Michael P.; Gouaux, Eric

    2010-02-02

    Ionotropic glutamate receptors mediate most excitatory neurotransmission in the central nervous system and function by opening a transmembrane ion channel upon binding of glutamate. Despite their crucial role in neurobiology, the architecture and atomic structure of an intact ionotropic glutamate receptor are unknown. Here we report the crystal structure of the {alpha}-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA)-sensitive, homotetrameric, rat GluA2 receptor at 3.6 {angstrom} resolution in complex with a competitive antagonist. The receptor harbours an overall axis of two-fold symmetry with the extracellular domains organized as pairs of local dimers and with the ion channel domain exhibiting four-fold symmetry. A symmetry mismatch between the extracellular and ion channel domains is mediated by two pairs of conformationally distinct subunits, A/C and B/D. Therefore, the stereochemical manner in which the A/C subunits are coupled to the ion channel gate is different from the B/D subunits. Guided by the GluA2 structure and site-directed cysteine mutagenesis, we suggest that GluN1 and GluN2A NMDA (N-methyl-D-aspartate) receptors have a similar architecture, with subunits arranged in a 1-2-1-2 pattern. We exploit the GluA2 structure to develop mechanisms of ion channel activation, desensitization and inhibition by non-competitive antagonists and pore blockers.

  7. Mechanisms of Biased β-Arrestin-Mediated Signaling Downstream from the Cannabinoid 1 Receptor

    PubMed Central

    Delgado-Peraza, Francheska; Ahn, Kwang H.; Nogueras-Ortiz, Carlos; Mungrue, Imran N.; Mackie, Ken; Kendall, Debra A.

    2016-01-01

    Activation of G protein-coupled receptors results in multiple waves of signaling that are mediated by heterotrimeric G proteins and the scaffolding proteins β-arrestin 1/2. Ligands can elicit full or subsets of cellular responses, a concept defined as ligand bias or functional selectivity. However, our current understanding of β-arrestin-mediated signaling is still very limited. Here we provide a comprehensive view of β-arrestin-mediated signaling from the cannabinoid 1 receptor (CB1R). By using a signaling biased receptor, we define the cascades, specific receptor kinases, and molecular mechanism underlying β-arrestin-mediated signaling: We identify the interaction kinetics of CB1R and β-arrestin 1 during their endocytic trafficking as directly proportional to its efficacy. Finally, we demonstrate that signaling results in the control of genes clustered around prosurvival and proapoptotic functions among others. Together, these studies constitute a comprehensive description of β-arrestin-mediated signaling from CB1Rs and suggest modulation of receptor endocytic trafficking as a therapeutic approach to control β-arrestin-mediated signaling. PMID:27009233

  8. Dual mechanisms of NF-kappaB inhibition in carnosol-treated endothelial cells

    SciTech Connect

    Lian, K.-C.; Chuang, J.-J.; Hsieh, C.-W.; Wung, B.-S.; Huang, G.-D.; Jian, T.-Y.; Sun, Y.-W.

    2010-05-15

    The increased adhesion of monocytes to injured endothelial layers is a critical early event in atherogenesis. Under inflammatory conditions, there is increased expression of specific cell adhesion molecules on activated vascular endothelial cells, which increases monocyte adhesion. In our current study, we demonstrate a putative mechanism for the anti-inflammatory effects of carnosol, a diterpene derived from the herb rosemary. Our results show that both carnosol and rosemary essential oils inhibit the adhesion of TNFalpha-induced monocytes to endothelial cells and suppress the expression of ICAM-1 at the transcriptional level. Moreover, carnosol was found to exert its inhibitory effects by blocking the degradation of the inhibitory protein IkappaBalpha in short term pretreatments but not in 12 h pretreatments. Our data show that carnosol reduces IKK-beta phosphorylation in pretreatments of less than 3 h. In TNFalpha-treated ECs, NF-kappaB nuclear translocation and transcriptional activity was abolished by up to 12 h of carnosol pretreatment and this was blocked by Nrf-2 siRNA. The long-term inhibitory effects of carnosol thus appear to be mediated through its induction of Nrf-2-related genes. The inhibition of ICAM-1 expression and p65 translocation is reversed by HO-1 siRNA. Carnosol also upregulates the Nrf-2-related glutathione synthase gene and thereby increases the GSH levels after 9 h of exposure. Treating ECs with a GSH synthesis inhibitor, BSO, blocks the inhibitory effects of carnosol. In addition, carnosol increases p65 glutathionylation. Hence, our present findings indicate that carnosol suppresses TNFalpha-induced singling pathways through the inhibition of IKK-beta activity or the upregulation of HO-1 expression. The resulting GSH levels are dependent, however, on the length of the carnosol pretreatment period.

  9. Mechanism of allosteric regulation of β2-adrenergic receptor by cholesterol

    PubMed Central

    Manna, Moutusi; Niemelä, Miia; Tynkkynen, Joona; Javanainen, Matti; Kulig, Waldemar; Müller, Daniel J; Rog, Tomasz; Vattulainen, Ilpo

    2016-01-01

    There is evidence that lipids can be allosteric regulators of membrane protein structure and activation. However, there are no data showing how exactly the regulation emerges from specific lipid-protein interactions. Here we show in atomistic detail how the human β2-adrenergic receptor (β2AR) – a prototypical G protein-coupled receptor – is modulated by cholesterol in an allosteric fashion. Extensive atomistic simulations show that cholesterol regulates β2AR by limiting its conformational variability. The mechanism of action is based on the binding of cholesterol at specific high-affinity sites located near the transmembrane helices 5–7 of the receptor. The alternative mechanism, where the β2AR conformation would be modulated by membrane-mediated interactions, plays only a minor role. Cholesterol analogues also bind to cholesterol binding sites and impede the structural flexibility of β2AR, however cholesterol generates the strongest effect. The results highlight the capacity of lipids to regulate the conformation of membrane receptors through specific interactions. DOI: http://dx.doi.org/10.7554/eLife.18432.001 PMID:27897972

  10. Electrolocation and electrocommunication in pulse gymnotids: signal carriers, pre-receptor mechanisms and the electrosensory mosaic.

    PubMed

    Caputi, Angel A; Castelló, María E; Aguilera, Pedro; Trujillo-Cenóz, Omar

    2002-01-01

    Constraints introduced by signal carriers, pre-receptor mechanisms and receptor transduction are fundamental for shaping the signals used by the brain to build up perceptual images. This review analyses some of these constraints in the electrosensory system of pulse Gymnotids. First, it describes the characteristics and differences of electrolocation and electrocommunication carriers. Second, it analyses the role of electrogenic and non-electrogenic tissues of the fish body in the generation and conditioning of these carriers. Two pre-receptor mechanisms are discussed: (a) the funneling of currents to the perioral region and (b) a Mexican-hat profile involved in edge detection. Finally, some characteristics of the electroreceptor structure and the sensory mosaic are examined. We conclude that there is an electrosensory fovea at the perioral region where a large density and variety of receptors is stimulated by self- and conspecific-generated currents funneled there by non electrogenic tissues. Differences in carrier waveform may be used to distinguish between reafferent and communication signals.

  11. Fracture mechanisms in dual phase steels based on the acicular ferrite + martensite/austenite microstructure

    NASA Astrophysics Data System (ADS)

    Poruks, Peter

    The fracture mechanisms of low carbon microalloyed plate steels based on the acicular ferrite + marten site/austenite microstructure (AF + M/A) are investigated. The final microstructure consists of a dispersed phase of submicron equi-axed martensite particles with a bainitic ferrite matrix. A series of plates with M/A volume fractions of 0.076--0.179 are studied. Brittle fracture is investigated by Instrumented Charpy impact testing of samples at -196°C and subsequent metallography. The M/A particles are identified as the crack nucleation sites and the cleavage fracture stress calculated to be 2400 MPa in a complete AF microstrucuture. This value is significantly larger than in steels that contain significant proportions of conventional bainite. Standard Charpy and Instrumented Charpy impact testing is conducted through a temperature range from -80 to + 22°C to study ductile fracture behaviour. The total absorbed energy is separated into energies of crack nucleation and of crack propagation. It is found that the energy of crack nucleation is weakly dependent on the volume fraction of M/A and completely independent of temperature over the range studied. The crack propagation energy varies significantly with both variables, decreasing with increased volume fraction of M/A and with decreasing temperature. The peak load in the instrumented Charpy data is used to calculate the dynamic fracture toughness, KId, which is found to be 105--120 MPa-m1/2. The void nucleation and void growth stages of ductile fracture are studied by metallographic examination of tensile bars. The sites of void nucleation are identified as inclusions and M/A particles. Voids nucleate at the M/A particles by decohesion of the particle-matrix interface. A constant void nucleation strain of epsilon = 0.90 +/- 0.05 is measured for all of the samples independent of the volume fraction of M/A. A stress-based criterion is used to predict void nucleation and the interface strength is determined to be

  12. The role of activation of the 5-HT1A receptor and adenylate cyclase in the antidepressant-like effect of YL-0919, a dual 5-HT1A agonist and selective serotonin reuptake inhibitor.

    PubMed

    Qin, Juan-Juan; Chen, Hong-Xia; Zhao, Nan; Yuan, Li; Zhang, You-Zhi; Yang, Ri-Fang; Zhang, Li-Ming; Li, Yun-Feng

    2014-10-17

    This study aimed to explore the possible mechanisms underlying the antidepressant-like effect of YL-0919, a novel antidepressant candidate with dual activity as a 5-HT1A receptor agonist and a selective serotonin reuptake inhibitor. The animal models commonly used to evaluate potential antidepressants, i.e., tail suspension (TST) in mice and forced swimming test (FST) in mice were used to evaluate the antidepressant effect of YL-0919. The activity of adenylate cyclase (AC) on the synaptic membrane was determined by the homogeneous time-resolved fluorescence resonance energy transfer (TR-FRET) immunoassay. The results indicated that YL-0919 (1.25-2.5mg/kg, i.g.) significantly decreased the immobility time in both the tail suspension test and the forced swim test in a dose-dependent manner, demonstrating the antidepressant-like effect of YL-0919. Furthermore, this effect was completely antagonized by the co-administration of WAY-100635 (0.3mg/kg, s.c.), a 5-HT1A selective antagonist. YL-0919 (10(-9)-10(-5)mol/L) was also shown to activate AC in vitro in a dose-dependent manner in synaptic membranes extracted from the rat prefrontal cortex, and this effect (10(-7)-10(-5)mol/L) was antagonized by WAY-100635 (10(-7)mol/L). Finally, the antidepressant-like effect of YL-0919 (2.5mg/kg, i.g.) was also blocked by the co-administration of H-89 (3 μg/site, i.c.v.), a protein kinase A (PKA) selective inhibitor. These results indicate that the activation of 5-HT1A receptors and the subsequent activation of the AC-cAMP-PKA signaling pathway in the frontal cortex play a critical role in the antidepressant-like effect of YL-0919.

  13. Heteromerization of dopamine D2 receptors with dopamine D1 or D5 receptors generates intracellular calcium signaling by different mechanisms

    PubMed Central

    Hasbi, Ahmed; O’Dowd, Brian F.; George, Susan R.

    2009-01-01

    The repertoire of signal transduction pathways activated by dopamine in brain includes the increase of intracellular calcium. However the mechanism(s) by which dopamine activated this important second messenger system was unknown. Although we showed that activation of the D5 dopamine receptor increased calcium concentrations, the restricted anatomic distribution of this receptor made this unlikely to be the major mechanism in brain. We have identified novel heteromeric dopamine receptor complexes that are linked to calcium signaling. The calcium pathway activated through the D1–D2 receptor heteromer involved coupling to Gq, through phospholipase C and IP3 receptors to result in a rise in intracellular calcium. The calcium rise activated through the D2–D5 receptor heteromer involved a small rise in intracellular calcium through the Gq pathway that triggered a store operated channel mediated influx of extracellular calcium. These novel receptor heteromeric complexes, for the first time, establish the link between dopamine action and rapid calcium signaling. PMID:19897420

  14. Survival and ecological fitness of Pseudomonas fluorescens genetically engineered with dual biocontrol mechanisms.

    PubMed

    Bainton, N J; Lynch, J M; Naseby, D; Way, J A

    2004-10-01

    The antibiotic 2,4-diacetylphloroglucinol (Phl) is produced by a range of naturally occurring fluorescent pseudomonads. One isolate, Pseudomonas fluorescens F113, protects pea plants from the pathogenic fungus Pythium ultimum by reducing the number of pathogenic lesions on plant roots, but with a concurrent reduction in the emergence of plants such as pea. The genes responsible for Phl production have been shown to be functionally conserved between the wild-type (wt) P. fluorescens strains F113 and Q2-87. In this study the genes from F113 were isolated using an optimized long PCR method and a 6.7-kb gene cluster inserted into the chromosome of the non-Phl-producing P. fluorescens strain SBW25 EeZY6KX. This strain is a lacZY, km(R) marked derivative of the wt SBW25 which effects biological control against the plant pathogen Pythium ultimum by competitive exclusion as a result of its strong rhizosphere-colonizing ability. We describe here the integration of the Phl antifungal and competitive exclusion mechanisms into a single strain, and the impact this has on survival and plant emergence in microcosms. The insertion of the Phl biosynthetic genes from the F113 into the SBW25 chromosome gave a Phl-producing transformant (strain Pa21) able to suppress P. ultimum through antibiotic production. The growth of Pa21 was not reduced in flask culture at 20 degrees C compared with its parent strain. When inoculated on pea seedlings, the strain containing the Phl operon behaved similarly to the SBW25 EeZY6KX parent but did not show the tendency of the wt Phl producer F113 to cause lower pea seed emergence. Pea roots inoculated with SBW25 EeZY6KX have significantly lower indigenous populations than with F113 and the control. This is indicative of this strain's strong colonising presence. Pa21, the Phl-modified strain, is able to exclude the resident population from roots to the same degree as the SBW25 EeZY6KX from which it is derived. This suggests that it has maintained its

  15. New mechanisms of NOD-like receptor-associated inflammasome activation

    PubMed Central

    Wen, Haitao; Miao, Edward A.; Ting, Jenny P.-Y.

    2013-01-01

    A major function of a subfamily of NLR (nucleotide-binding domain, leucine rich repeat containing or NOD-like receptor) proteins is in inflammasome activation, which has been implicated in a multitude of disease models and human diseases. This work will highlight key progress in understanding the mechanisms which activates the best studied NLRs (NLRP3, NLRC4, NAIP and NLRP1) and in uncovering new inflammasome NLRs. PMID:24054327

  16. Anti-inflammatory effect of dual nociceptin and opioid receptor agonist, BU08070, in experimental colitis in mice

    PubMed Central

    Zielińska, Marta; Ben Haddou, Tanila; Cami-Kobeci, Gerta; Sałaga, Maciej; Jarmuż, Agata; Padysz, Milena; Kordek, Radzisław; Spetea, Mariana; Husbands, Stephen M.; Fichna, Jakub

    2016-01-01

    Endogenous opioid and nociceptin systems are widely distributed in the gastrointestinal tract where they seem to play a crucial role in maintaining homeostasis. The aim of our study was to assess whether activation of nociceptin (NOP) and μ-opioid (MOP) receptors by a mixed NOP/MOP receptor agonist, BU08070, induces anti-inflammatory response in experimental colitis. The anti-inflammatory effect of BU08070 (1 mg/kg i.p.) was characterized in the mouse model of 2,4,6-trinitrobenzenesulfonic acid (TNBS)-induced colitis, based on the assessment of the macroscopic and microscopic total damage scores and determination of MPO activity and TNF-α level in the colon. The effect of BU08070 on cell viability and NF-κB was characterized in THP-1 Blue cell line. The antinociceptive activity of BU08070 was examined in mustard oil-induced mouse model of abdominal pain. A potent anti-inflammatory effect of BU08070 (1 mg/kg i.p.) was observed as indicated by decrease in macroscopic damage score (1.88±0.39 vs. 5.19±0.43 units in TNBS alone treated mice), MPO activity (2.29±0.37 vs. 9.64±2.55 units) and TNF-α level in the colon (35.85±2.45 vs. 49.79±3.81 pg/ml). The anti-inflammatory effect of BU08070 was reversed by selective NOP and MOP receptor antagonists. BU08070 produced concentration-dependent inhibition of TNF-α and LPS-induced NF-κB activation. BU08070 exerted antinociceptive action in mice with experimental colitis. In conclusion, BU08070 significantly reduced the severity of colitis in TNBS-treated mice compared with controls. These results suggest that BU08070 is a potential therapeutic agent for IBD therapy. PMID:26404500

  17. Dual pathways for the intracellular processing of insulin. Relationship between retroendocytosis of intact hormone and the recycling of insulin receptors.

    PubMed

    Marshall, S

    1985-11-05

    Adipocytes process insulin through either of two pathways: a retroendocytotic pathway that culminates in the release of intact insulin, and a degradative pathway that terminates in the intracellular catabolism and release of degraded ligand. Mechanistically, these pathways were found to differ in several ways. First, temporal differences were found in the rate at which intact and degraded products were extruded. After 125I-insulin was preloaded into the cell interior, intact ligand was completely released during the first 10 min (t 1/2 = 2 min), whereas degraded insulin was released at a much slower rate over 1 h (t 1/2 greater than 8 min). Secondly, it was found that chloroquine profoundly inhibited the insulin degradative pathway, resulting in the intracellular accumulation of intact ligand and a reduction in the release of degraded products. In contrast, however, chloroquine was without effect on the retroendocytotic processing of insulin. Based on the known actions of chloroquine, it appears that retroendocytosis of insulin does not involve vesicular acidification or dissociation of the insulin-receptor complex and that insulin is most likely carried to the cell exterior in the same vesicles (either receptor-bound or free) as those mediating recycling receptors. Interestingly, accumulation of undergraded insulin within chloroquine-treated cells did not result in the release of additional intact ligand, suggesting that once insulin enters the degradative compartment it is committed to catabolism and cannot exit the cell through the retroendocytotic pathway. A third difference was revealed by the finding that extracellular unlabeled insulin (100 ng/ml) markedly accelerated the rate at which preloaded 125I-insulin was released from adipocytes (t 1/2 of 3 min versus 7 min in controls cells). Analysis of the composition of the released products revealed that extracellular insulin rapidly augmented (over 10 min) in a dose-dependent manner (5-200 ng/ml) the amount of

  18. Nanoscale imaging and mechanical analysis of Fc receptor-mediated macrophage phagocytosis against cancer cells.

    PubMed

    Li, Mi; Liu, Lianqing; Xi, Ning; Wang, Yuechao; Xiao, Xiubin; Zhang, Weijing

    2014-02-18

    Fc receptor-mediated macrophage phagocytosis against cancer cells is an important mechanism in the immune therapy of cancers. Traditional research about macrophage phagocytosis was based on optical microscopy, which cannot reveal detailed information because of the 200-nm-resolution limit. Quantitatively investigating the macrophage phagocytosis at micro- and nanoscale levels is still scarce. The advent of atomic force microscopy (AFM) offers an excellent analytical instrument for quantitatively investigating the biological processes at single-cell and single-molecule levels under native conditions. In this work, we combined AFM and fluorescence microscopy to visualize and quantify the detailed changes in cell morphology and mechanical properties during the process of Fc receptor-mediated macrophage phagocytosis against cancer cells. Lymphoma cells were discernible by fluorescence staining. Then, the dynamic process of phagocytosis was observed by time-lapse optical microscopy. Next, AFM was applied to investigate the detailed cellular behaviors during macrophage phagocytosis under the guidance of fluorescence recognition. AFM imaging revealed the distinct features in cellular ultramicrostructures for the different steps of macrophage phagocytosis. AFM cell mechanical property measurements indicated that the binding of cancer cells to macrophages could make macrophages become stiffer. The experimental results provide novel insights in understanding the Fc-receptor-mediated macrophage phagocytosis.

  19. GABA(B) receptors and opioid mechanisms involved in homotaurine-induced analgesia.

    PubMed

    Serrano, M I; Serrano, J S; Fernández, A; Asadi, I; Serrano-Martino, M C

    1998-03-01

    1. The involvement of GABA(B) receptors and opioid mechanisms in homotaurine-induced analgesia has been investigated in current models of nociception by using a GABA(B) receptor antagonist, morphine, and naloxone. CGP 35348 (50-200 mg/kg IP), a highly selective GABA(B) antagonist, was administered prior to carrying out a dose-response curve of homotaurine (22.6-445 mg/kg IP) antinociceptive effect in the abdominal constriction (mice) and tail flick (rats) tests. 2. The tail flick test was performed in animals pretreated with morphine (0.5 mg/kg SC) and naloxone (1 mg/kg), 15 min before amino acid. Animals treated with saline 10 ml/kg (mice) or 1.25 ml/kg (rats) were included as control for the vehicle used. 3. CGP 35348 antagonized the antinociceptive effect of homotaurine in both tests. The range of doses affected by the interaction depended on the test assayed, but it was coincident for the main part of the dose-response curve. 4. A subanalgesic dose of morphine potentiated the antinociceptive effect of lower doses of homotaurine in the tail flick test. Naloxone pretreatment inhibited the antinociceptive effect of homotaurine. 5. These data imply that GABA(B) receptor subpopulations and opiate mechanisms are involved in the antinociceptive effect of homotaurine. Because functional relationships have been found between GABAergic and opiate systems in analgesic effects, an interaction of the two mechanisms may be operating in the effects described for homotaurine.

  20. Blockade of metabotropic glutamate receptor 5 activation inhibits mechanical hypersensitivity following abdominal surgery.

    PubMed

    Dolan, Sharron; Nolan, Andrea Mary

    2007-08-01

    This study used the metabotropic glutamate 5 (mGlu5) receptor subtype-selective antagonist 2-methyl-6-(phenylethynyl)pyridine (MPEP) to characterise the contribution of mGlu5 receptor activity to pain and hypersensitivity in an animal model of post-surgical pain. Adult female Wistar rats (200-250g) were anaesthetised with isoflurane (2%) and underwent a midline laparotomy with gentle manipulation of the viscera, and the effects of pre- (30min) or post- (5h) operative treatment with MPEP (1, 3 or 10mgkg(-1); i.p.) or drug-vehicle on hindpaw withdrawal latency (in seconds) to thermal stimulation (Hargreave's Test) and response threshold (in grams) to mechanical stimulation (using a dynamic plantar aesthesiometer) were measured. Animals that underwent surgery displayed significant hypersensitivity to mechanical stimulation of the hindpaws. Hypersensitivity was maximum at 6h post-surgery (44.5+/-2.4% decrease; p<0.01 vs. anaesthesia only controls) and persisted for 48h. Surgery had no effect on thermal withdrawal latency. Both pre-operative and post-operative administration of 10mgkg(-1)MPEP blocked mechanical hypersensitivity induced by surgery (p<0.01 vs. vehicle treatment). MPEP had no effect on acute nociceptive thresholds in naïve animals. These data suggest that activity at mGlu5 receptors contributes to development of pain and hypersensitivity following surgery.

  1. Obesity and gastrointestinal hormones-dual effect of angiotensin II receptor blockade and a partial agonist of PPAR-γ.

    PubMed

    Nakagami, Hironori; Morishita, Ryuichi

    2011-03-01

    Obesity is strongly associated with type 2 diabetes, hypertension, and hyperlipidemia, which is one of the leading causes of mortality and morbidity worldwide. It is now clear that gut hormones play a role in the regulation of body weight and represent therapeutic targets for the future treatment of obesity. Recent evidence demonstrated that dysregulation of adipocytokine functions seen in abdominal obesity may be involved in the pathogenesis of the metabolic syndrome. Angiotensinogen, the precursor of angiotensin (Ang) II, is produced primarily in the liver, but also in adipose tissue, where it is up-regulated during the development of obesity and involved in blood pressure regulation and adipose tissue growth. Importantly, blockade of the RAS attenuates weight gain and adiposity by enhanced energy expenditure. The favorable metabolic effects of telmisartan have been related to its Ang II receptor blockade and action as a partial agonist of peroxisome proliferators activated receptor (PPAR)-γ. PPARγ plays an important role in regulating carbohydrate and lipid metabolism, and ligands for PPARγ can improve insulin sensitivity and reduce triglyceride levels. We designed a comparative study of telmisartan and losartan in ApoE-deficient mice. Treatment with telmisartan or losartan significantly reduced the development of lipid-rich plaque. However, treatment with telmisartan significantly improved endothelial dysfunction and inhibited lipid accumulation in the liver. These favorable characteristics of telmisartan might be due to its action as a partial agonist of PPAR-γ, beyond its blood pressure-lowering effect, through Ang II blockade, which may be called "metabosartan".

  2. Edg8/S1P5: an oligodendroglial receptor with dual function on process retraction and cell survival.

    PubMed

    Jaillard, C; Harrison, S; Stankoff, B; Aigrot, M S; Calver, A R; Duddy, G; Walsh, F S; Pangalos, M N; Arimura, N; Kaibuchi, K; Zalc, B; Lubetzki, C

    2005-02-09

    Endothelial differentiation gene (Edg) proteins are G-protein-coupled receptors activated by lysophospholipid mediators: sphingosine-1-phosphate (S1P) or lysophosphatidic acid. We show that in the CNS, expression of Edg8/S1P5, a high-affinity S1P receptor, is restricted to oligodendrocytes and expressed throughout development from the immature stages to the mature myelin-forming cell. S1P activation of Edg8/S1P5 on O4-positive pre-oligodendrocytes induced process retraction via a Rho kinase/collapsin response-mediated protein signaling pathway, whereas no retraction was elicited by S1P on these cells derived from Edg8/S1P5-deficient mice. Edg8/S1P5-mediated process retraction was restricted to immature cells and was no longer observed at later developmental stages. In contrast, S1P activation promoted the survival of mature oligodendrocytes but not of pre-oligodendrocytes. The S1P-induced survival of mature oligodendrocytes was mediated through a pertussis toxin-sensitive, Akt-dependent pathway. Our data demonstrate that Edg8/S1P5 activation on oligodendroglial cells modulates two distinct functional pathways mediating either process retraction or cell survival and that these effects depend on the developmental stage of the cell.

  3. ACTH Receptor (MC2R) Specificity: What Do We Know About Underlying Molecular Mechanisms?

    PubMed

    Fridmanis, Davids; Roga, Ance; Klovins, Janis

    2017-01-01

    Coincidentally, the release of this Research Topic in Frontiers in Endocrinology takes place 25 years after the discovery of the adrenocorticotropic hormone receptor (ACTHR) by Mountjoy and colleagues. In subsequent years, following the discovery of other types of mammalian melanocortin receptors (MCRs), ACTHR also became known as melanocortin type 2 receptor (MC2R). At present, five types of MCRs have been reported, all of which share significant sequence similarity at the amino acid level, and all of which specifically bind melanocortins (MCs)-a group of biologically active peptides generated by proteolysis of the proopiomelanocortin precursor. All MCs share an identical -H-F-R-W- pharmacophore sequence. α-Melanocyte-stimulating hormone (α-MSH) and adrenocorticotropic hormone (ACTH) are the most extensively studied MCs and are derived from the same region. Essentially, α-MSH is formed from the first 13 amino acid residues of ACTH. ACTHR is unique among MCRs because it binds one sole ligand-ACTH, which makes it a very attractive research object for molecular pharmacologists. However, much research has failed, and functional studies of this receptor are lagging behind other MCRs. The reason for these difficulties has already been outlined by Mountjoy and colleagues in their publication on ACTHR coding sequence discovery where the Cloudman S91 melanoma cell line was used for receptor expression because it was a "more sensitive assay system." Subsequent work showed that ACTHR could be successfully expressed only in endogenous MCR-expressing cell lines, since in other cell lines it is retained within the endoplasmic reticulum. The resolution of this methodological problem came in 2005 with the discovery of melanocortin receptor accessory protein, which is required for the formation of functionally active ACTHR. The decade that followed this discovery was filled with exciting research that provided insight into the molecular mechanisms underlying the action of

  4. Analysis of the activation mechanism of the guinea-pig Histamine H1-receptor

    NASA Astrophysics Data System (ADS)

    Straßer, Andrea; Wittmann, Hans-Joachim

    2007-09-01

    The Histamine H1-receptor (H1R), belonging to the amine receptor-class of family A of the G-protein coupled receptors (GPCRs) gets activated by agonists. The consequence is a conformational change of the receptor, which may involve the binding-pocket. So, for a good prediction of the binding-mode of an agonist, it is necessary to have knowledge about these conformational changes. Meanwhile some experimental data about the structural changes of GPCRs during activation exist. Based on homology modeling of the guinea-pig H1R (gpH1R), using the crystal structure of bovine rhodopsin as template, we performed several MD simulations with distance restraints in order to get an inactive and an active structure of the gpH1R. The calculations led to a Phe6.44/Trp6.48/Phe6.52-switch and linearization of the proline kinked transmembrane helix VI during receptor activation. Our calculations showed that the Trp6.48/Phe6.52-switch induces a conformational change in Phe6.44, which slides between transmembrane helices III and VI. Additionally we observed a hydrogen bond interaction of Ser3.39 with Asn7.45 in the inactive gpH1R, but because of a counterclockwise rotation of transmembrane helix III Ser3.39 establishes a water-mediated hydrogen bond to Asp2.50 in the active gpH1R. Additionally we simulated a possible mechanism for receptor activation with a modified LigPath-algorithm.

  5. Unraveling mechanisms underlying partial agonism in 5-HT3A receptors.

    PubMed

    Corradi, Jeremías; Bouzat, Cecilia

    2014-12-10

    Partial agonists have emerged as attractive therapeutic molecules. 2-Me-5HT and tryptamine have been defined as partial agonists of 5-HT3 receptors on the basis of macroscopic measurements. Because several mechanisms may limit maximal responses, we took advantage of the high-conductance form of the mouse serotonin type 3A (5-HT3A) receptor to understand their molecular actions. Individual 5-HT-bound receptors activate in long episodes of high open probability, consisting of groups of openings in quick succession. The activation pattern is similar for 2-Me-5HT only at very low concentrations since profound channel blockade takes place within the activating concentration range. In contrast, activation episodes are significantly briefer in the presence of tryptamine. Generation of a full activation scheme reveals that the fully occupied receptor overcomes transitions to closed preopen states (primed states) before opening. Reduced priming explains the partial agonism of tryptamine. In contrast, 2-Me-5HT is not a genuine partial agonist since priming is not dramatically affected and its low apparent efficacy is mainly due to channel blockade. The analysis also shows that the first priming step is the rate-limiting step and partial agonists require an increased number of priming steps for activation. Molecular docking suggests that interactions are similar for 5-HT and 2-Me-5HT but slightly different for tryptamine. Our study contributes to understanding 5-HT3A receptor activation, extends the novel concept of partial agonism within the Cys-loop family, reveals novel aspects of partial agonism, and unmasks molecular actions of classically defined partial agonists. Unraveling mechanisms underlying partial responses has implications in the design of therapeutic compounds.

  6. Mechanisms of Host Receptor Adaptation by Severe Acute Respiratory Syndrome Coronavirus

    SciTech Connect

    Wu, Kailang; Peng, Guiqing; Wilken, Matthew; Geraghty, Robert J.; Li, Fang

    2012-12-10

    The severe acute respiratory syndrome coronavirus (SARS-CoV) from palm civets has twice evolved the capacity to infect humans by gaining binding affinity for human receptor angiotensin-converting enzyme 2 (ACE2). Numerous mutations have been identified in the receptor-binding domain (RBD) of different SARS-CoV strains isolated from humans or civets. Why these mutations were naturally selected or how SARS-CoV evolved to adapt to different host receptors has been poorly understood, presenting evolutionary and epidemic conundrums. In this study, we investigated the impact of these mutations on receptor recognition, an important determinant of SARS-CoV infection and pathogenesis. Using a combination of biochemical, functional, and crystallographic approaches, we elucidated the molecular and structural mechanisms of each of these naturally selected RBD mutations. These mutations either strengthen favorable interactions or reduce unfavorable interactions with two virus-binding hot spots on ACE2, and by doing so, they enhance viral interactions with either human (hACE2) or civet (cACE2) ACE2. Therefore, these mutations were viral adaptations to either hACE2 or cACE2. To corroborate the above analysis, we designed and characterized two optimized RBDs. The human-optimized RBD contains all of the hACE2-adapted residues (Phe-442, Phe-472, Asn-479, Asp-480, and Thr-487) and possesses exceptionally high affinity for hACE2 but relative low affinity for cACE2. The civet-optimized RBD contains all of the cACE2-adapted residues (Tyr-442, Pro-472, Arg-479, Gly-480, and Thr-487) and possesses exceptionally high affinity for cACE2 and also substantial affinity for hACE2. These results not only illustrate the detailed mechanisms of host receptor adaptation by SARS-CoV but also provide a molecular and structural basis for tracking future SARS-CoV evolution in animals.

  7. Catalytic control in the EGF Receptor and its connection to general kinase regulatory mechanisms

    PubMed Central

    Jura, Natalia; Zhang, Xuewu; Endres, Nicholas F.; Seeliger, Markus A.; Schindler, Thomas; Kuriyan, John

    2011-01-01

    Summary In contrast to the active conformations of protein kinases, which are essentially the same for all kinases, inactive kinase conformations are structurally diverse. Some inactive conformations are, however, observed repeatedly in different kinases, perhaps reflecting an important role in catalysis. In this review, we analyze one of these recurring conformations, first identified in CDK and Src kinases, which turned out to be central to understanding of how kinase domain of the EGF receptor is activated. This mechanism, which involves the stabilization of the active conformation of an α helix, has features in common with mechanisms operative in several other kinases. PMID:21474065

  8. Mechanical stretch augments insulin-induced vascular smooth muscle cell proliferation by insulin-like growth factor-1 receptor

    SciTech Connect

    Liu, Gang; Hitomi, Hirofumi; Hosomi, Naohisa; Lei, Bai; Nakano, Daisuke; Deguchi, Kazushi; Mori, Hirohito; Masaki, Tsutomu; Ma, Hong; Griendling, Kathy K.; Nishiyama, Akira

    2011-10-15

    Insulin resistance and hypertension have been implicated in the pathogenesis of cardiovascular disease; however, little is known about the roles of insulin and mechanical force in vascular smooth muscle cell (VSMC) remodeling. We investigated the contribution of mechanical stretch to insulin-induced VSMC proliferation. Thymidine incorporation was stimulated by insulin in stretched VSMCs, but not in un-stretched VSMCs. Insulin increased 2-deoxy-glucose incorporation in both stretched and un-stretched VSMCs. Mechanical stretch augmented insulin-induced extracellular signal-regulated kinase (ERK) and Akt phosphorylation. Inhibitors of epidermal growth factor (EGF) receptor tyrosine kinase and Src attenuated insulin-induced ERK and Akt phosphorylation, as well as thymidine incorporation, whereas 2-deoxy-glucose incorporation was not affected by these inhibitors. Moreover, stretch augmented insulin-like growth factor (IGF)-1 receptor expression, although it did not alter the expression of insulin receptor and insulin receptor substrate-1. Insulin-induced ERK and Akt activation, and thymidine incorporation were inhibited by siRNA for the IGF-1 receptor. Mechanical stretch augments insulin-induced VSMC proliferation via upregulation of IGF-1 receptor, and downstream Src/EGF receptor-mediated ERK and Akt activation. Similar to in vitro experiment, IGF-1 receptor expression was also augmented in hypertensive rats. These results provide a basis for clarifying the molecular mechanisms of vascular remodeling in hypertensive patients with hyperinsulinemia. -- Highlights: {yields} Mechanical stretch augments insulin-induced VSMC proliferation via IGF-1 receptor. {yields} Src/EGFR-mediated ERK and Akt phosphorylation are augmented in stretched VSMCs. {yields} Similar to in vitro experiment, IGF-1 receptor is increased in hypertensive rats. {yields} Results provide possible mechanisms of vascular remodeling in hypertension with DM.

  9. Evidence toward a dual phosphatase mechanism that restricts Aurora A (Thr-295) phosphorylation during the early embryonic cell cycle.

    PubMed

    Kang, Qing; Srividhya, Jeyaraman; Ipe, Joseph; Pomerening, Joseph R

    2014-06-20

    The mitotic kinase Aurora A (AurA) is regulated by a complex network of factors that includes co-activator binding, autophosphorylation, and dephosphorylation. Dephosphorylation of AurA by PP2A (human, Ser-51; Xenopus, Ser-53) destabilizes the protein, whereas mitotic dephosphorylation of its T-loop (human, Thr-288; Xenopus, Thr-295) by PP6 represses AurA activity. However, AurA(Thr-295) phosphorylation is restricted throughout the early embryonic cell cycle, not just during M-phase, and how Thr-295 is kept dephosphorylated during interphase and whether or not this mechanism impacts the cell cycle oscillator were unknown. Titration of okadaic acid (OA) or fostriecin into Xenopus early embryonic extract revealed that phosphatase activity other than PP1 continuously suppresses AurA(Thr-295) phosphorylation during the early embryonic cell cycle. Unexpectedly, we observed that inhibiting a phosphatase activity highly sensitive to OA caused an abnormal increase in AurA(Thr-295) phosphorylation late during interphase that corresponded with delayed cyclin-dependent kinase 1 (CDK1) activation. AurA(Thr-295) phosphorylation indeed influenced this timing, because AurA isoforms retaining an intact Thr-295 residue further delayed M-phase entry. Using mathematical modeling, we determined that one phosphatase would be insufficient to restrict AurA phosphorylation and regulate CDK1 activation, whereas a dual phosphatase topology best recapitulated our experimental observations. We propose that two phosphatases target Thr-295 of AurA to prevent premature AurA activation during interphase and that phosphorylated AurA(Thr-295) acts as a competitor substrate with a CDK1-activating phosphatase in late interphase. These results suggest a novel relationship between AurA and protein phosphatases during progression throughout the early embryonic cell cycle and shed new light on potential defects caused by AurA overexpression.

  10. Monte-Carlo methods for chemical-mechanical planarization on multiple-layer and dual-material models

    NASA Astrophysics Data System (ADS)

    Chen, Yu; Kahng, Andrew B.; Robins, Gabriel; Zelikovsky, Alexander

    2002-07-01

    Chemical-mechanical planarization (CMP) and other manufacturing steps in very deep submicron VLSI have varying effects on device and interconnect features, depending on the local layout density. To improve manufacturability and performance predictability, we seek to make a layout uniform with respect to prescribed density criteria, by inserting area fill geometries in to the layout. We review previous research on single-layer fill for flat and hierarchical layout density control based on the Interlevel Dielectric CMP model. We also describe the recent combination of CMP physical modeling and linear programing for multiple-layer density control, as well as the Shallow Trench Isolation CMP model. Our work makes the following contributions for the Multiple-layer Interlevel Dielectric CMP model. First, we propose a new linear programming approach with a new objective for the multiple-layer fill problem. Second, we describe modified Monte-Carlo approaches for the multiple- layer fill problem. Comparisons with previous approaches show that the new linear programming method is more reasonable for manufacturability, and that the Monte-Carlo approach is efficient and yields more accurate results for large layouts. The CMP step in Shallow Trench Isolation (STI) is a dual-material polishing process, i.e., multiple materials are being polished simultaneously during the CMP process. Simple greedy methods were proposed for the non- linear problem with Min-Var and Min-Fill objectives, where the certain amount of dummy features are always added at a position with the smallest density. In this paper, we propose more efficient Monte-Carlo methods for the Min-Var objective, as well a improved Greedy and Monte-Carlo methods for the Min-Fill objective. Our experimental experience shows that they can get better solutions with respect to the objectives.

  11. Vandetanib (ZD6474), a dual inhibitor of vascular endothelial growth factor receptor (VEGFR) and epidermal growth factor receptor (EGFR) tyrosine kinases: current status and future directions.

    PubMed

    Morabito, Alessandro; Piccirillo, Maria Carmela; Falasconi, Fabiano; De Feo, Gianfranco; Del Giudice, Antonia; Bryce, Jane; Di Maio, Massimo; De Maio, Ermelinda; Normanno, Nicola; Perrone, Francesco

    2009-04-01

    Vandetanib is a novel, orally available inhibitor of different intracellular signaling pathways involved in tumor growth, progression, and angiogenesis: vascular endothelial growth factor receptor-2, epidermal growth factor receptor, and REarranged during Transfection tyrosine kinase activity. Phase I clinical trials have shown that vandetanib is well tolerated as a single agent at daily doses < or =300 mg. In the phase II setting, negative results were observed with vandetanib in small cell lung cancer, metastatic breast cancer, and multiple myeloma. In contrast, three randomized phase II studies showed that vandetanib prolonged the progression-free survival (PFS) time of patients with non-small cell lung cancer (NSCLC) as a single agent when compared with gefitinib or when added to chemotherapy. Rash, diarrhea, hypertension, fatigue, and asymptomatic QTc prolongation were the most common adverse events. Antitumor activity was also observed in medullary thyroid cancer. Four randomized phase III clinical trials in NSCLC are exploring the efficacy of vandetanib in combination with docetaxel, the Zactima in cOmbination with Docetaxel In non-small cell lung Cancer (ZODIAC) trial, or with pemetrexed, the Zactima Efficacy with Alimta in Lung cancer (ZEAL) trial, or as a single agent, the Zactima Efficacy when Studied versus Tarceva (ZEST) and the Zactima Efficacy trial for NSCLC Patients with History of EGFR-TKI chemo-Resistance (ZEPHYR) trials. Based on a press release by the sponsor of these trials, the PFS time was longer with vandetanib in the ZODIAC and ZEAL trials; the ZEST trial was negative for its primary superiority analysis, but was successful according to a preplanned noninferiority analysis of PFS. Ongoing phase II and III clinical trials will better define the appropriate schedule, the optimal setting of evaluation, and the safety of long-term use of vandetanib.

  12. A Genetic Approach to Identifying Signal Transduction Mechanisms Initiated by Receptors for TGF-B-Related Factors.

    DTIC Science & Technology

    1998-10-01

    resistant to TGF-ß-induced growth arrest suggest that both types of receptors are required for signaling (Boyd and Massague, 1989; Laiho et ah, 1990...II in TGF-ß- resistant cell mutants implicates both receptor types in signal transduction. J. Biol. Chem. 265, 18518-18524. Lechleider, R. J., de...I-1 « -J AD GRANT NUMBER DAMD17-94-J-4339 TITLE: A Genetic Approach to Identifying Signal Transduction Mechanisms Initiated by Receptors

  13. Role of Orexin-1 Receptor Mechanisms on Compulsive Food Consumption in a Model of Binge Eating in Female Rats

    PubMed Central

    Piccoli, Laura; Micioni Di Bonaventura, Maria Vittoria; Cifani, Carlo; Costantini, Vivian J A; Massagrande, Mario; Montanari, Dino; Martinelli, Prisca; Antolini, Marinella; Ciccocioppo, Roberto; Massi, Maurizio; Merlo-Pich, Emilio; Di Fabio, Romano; Corsi, Mauro

    2012-01-01

    Orexins (OX) and their receptors (OXR) modulate feeding, arousal, stress, and drug abuse. Neural systems that motivate and reinforce drug abuse may also underlie compulsive food seeking and intake. Therefore, the effects of GSK1059865 (5-bromo-N-[(2S,5S)-1-(3-fluoro-2-methoxybenzoyl)-5-methylpiperidin-2-yl]methyl-pyridin-2-amine), a selective OX1R antagonist, JNJ-10397049 (N-(2,4-dibromophenyl)-N′-[(4S,5S)-2,2-dimethyl-4-phenyl-1,3-dioxan-5-yl]urea), a selective OX2R antagonist, and SB-649868 (N-[((2S)-1-{[5-(4-fluorophenyl)-2-methyl-1,3-thiazol-4-yl]carbonyl}-2-piperidinyl)methyl]-1-benzofuran-4-carboxamide), a dual OX1/OX2R antagonist were evaluated in a binge eating (BE) model in female rats. BE of highly palatable food (HPF) was evoked by three cycles of food restriction followed by stress, elicited by exposing rats to HPF, but preventing them from having access to it for 15 min. Pharmacokinetic assessments of all compounds were obtained under the same experimental conditions used for the behavioral experiments. Topiramate was used as the reference compound as it selectively blocks BE in rats and humans. Dose-related thresholds for sleep-inducing effects of the OXR antagonists were measured using polysomnography in parallel experiments. SB-649868 and GSK1059865, but not JNJ-10397049, selectively reduced BE for HPF without affecting standard food pellet intake, at doses that did not induce sleep. These results indicate, for the first time, a major role of OX1R mechanisms in BE, suggesting that selective antagonism at OX1R could represent a novel pharmacological treatment for BE and possibly other eating disorders with a compulsive component. PMID:22569505

  14. Dual role for Fcγ receptors in host defense and disease in Borrelia burgdorferi-infected mice.

    PubMed

    Belperron, Alexia A; Liu, Nengyin; Booth, Carmen J; Bockenstedt, Linda K

    2014-01-01

    Arthritis in mice infected with the Lyme disease spirochete, Borrelia burgdorferi, results from the influx of innate immune cells responding to the pathogen in the joint and is influenced in part by mouse genetics. Production of inflammatory cytokines by innate immune cells in vitro is largely mediated by Toll-like receptor (TLR) interaction with Borrelia lipoproteins, yet surprisingly mice deficient in TLR2 or the TLR signaling molecule MyD88 still develop arthritis comparable to that seen in wild type mice after B. burgdorferi infection. These findings suggest that other, MyD88-independent inflammatory pathways can contribute to arthritis expression. Clearance of B. burgdorferi is dependent on the production of specific antibody and phagocytosis of the organism. As Fc receptors (FcγR) are important for IgG-mediated clearance of immune complexes and opsonized particles by phagocytes, we examined the role that FcγR play in host defense and disease in B. burgdorferi-infected mice. B. burgdorferi-infected mice deficient in the Fc receptor common gamma chain (FcεRγ(-/-) mice) harbored ~10 fold more spirochetes than similarly infected wild type mice, and this was associated with a transient increase in arthritis severity. While the elevated pathogen burdens seen in B. burgdorferi-infected MyD88(-/-) mice were not affected by concomitant deficiency in FcγR, arthritis was reduced in FcεRγ(-/-) MyD88(-/-) mice in comparison to wild type or single knockout mice. Gene expression analysis from infected joints demonstrated that absence of both MyD88 and FcγR lowers mRNA levels of proteins involved in inflammation, including Cxcl1 (KC), Xcr1 (Gpr5), IL-1beta, and C reactive protein. Taken together, our results demonstrate a role for FcγR-mediated immunity in limiting pathogen burden and arthritis in mice during the acute phase of B. burgdorferi infection, and further suggest that this pathway contributes to the arthritis that develops in B. burgdorferi-infected MyD88

  15. TRC120038, a Novel Dual AT1/ETA Receptor Blocker for Control of Hypertension, Diabetic Nephropathy, and Cardiomyopathy in ob-ZSF1 Rats

    PubMed Central

    Mohanan, Anookh; Gupta, Ram; Dubey, Amita; Jagtap, Vikrant; Mandhare, Appaji; Gupta, Ramesh C.; Chauthaiwale, Vijay; Dutt, Chaitanya

    2011-01-01

    In hypertensive subjects, angiotensin II and endothelin participate in a manner involving closely interwoven pathways in increasing blood pressure (BP) and inducing end organ damage. The primary objective of this study was to determine the effect of TRC120038, a novel dual AT1/ETA receptor blocker on BP, in obese Zucker spontaneously hypertensive fatty rats (ob-ZSF1), an animal model of moderate hypertension, diabetes with progressive renal and cardiac dysfunction. Ob-ZSF1 rats loaded with 0.5% salt were treated with TRC120038 (11.8 mg/kg bid.) or candesartan cilexetil (0.3 mg/kg od.) or vehicle control. Blood pressure (by radio-telemetry) and renal functional markers were monitored throughout the study. Cardiac function was assessed terminally by pressure volume catheter. Markers for renal dysfunction were measured and changes were evaluated histopathologically. TRC120038 showed greater fall in both systolic and diastolic BP in comparison to candesartan at its maximum antihypertensive dose. TRC120038 also reduced the severity of renal dysfunction and preserved cardiac function in ob-ZSF1 rat. PMID:22235363

  16. Enhanced Inhibition of Prostate Tumor Growth by Dual Targeting the Androgen Receptor and the Regulatory Subunit Type Iα of Protein Kinase A in Vivo

    PubMed Central

    Eder, Iris E.; Egger, Martina; Neuwirt, Hannes; Seifarth, Christof; Maddalo, Danilo; Desiniotis, Andreas; Schäfer, Georg; Puhr, Martin; Bektic, Jasmin; Cato, Andrew C. B.; Klocker, Helmut

    2013-01-01

    Progression to castration resistance is a major problem in the treatment of advanced prostate cancer and is likely to be driven by activation of several molecular pathways, including androgen receptor (AR) and cyclic AMP-dependent protein kinase A (PKA). In this study, we examined the therapeutic efficacy of a combined inhibition of the AR and the regulatory subunit type Iα (RIα) of protein kinase A with second generation antisense oligonucleotides (ODNs) in androgen-sensitive LNCaP and castration-resistant LNCaPabl tumors in vivo. We found that targeting the AR alone inhibited LNCaP, as well as LNCaPabl tumors. Combined inhibition resulted in an improved response over single targeting and even a complete tumor remission in LNCaPabl. Western blot analysis revealed that both ODNs were effective in reducing their target proteins when administered alone or in combination. In addition, treatment with the ODNs was associated with an induction of apoptosis. Our data suggest that dual targeting of the AR and PKARIα is more effective in inhibiting LNCaP and LNCaPabl tumor growth than single treatment and may give a treatment benefit, especially in castration-resistant prostate cancers. PMID:23736698

  17. D1-D2 Dopamine Receptor Synergy Promotes Calcium Signaling via Multiple Mechanisms

    PubMed Central

    Chun, Lani S.; Free, R. Benjamin; Doyle, Trevor B.; Huang, Xi-Ping; Rankin, Michele L.

    2013-01-01

    The D1 dopamine receptor (D1R) has been proposed to form a hetero-oligomer with the D2 dopamine receptor (D2R), which in turn results in a complex that couples to phospholipase C–mediated intracellular calcium release. We have sought to elucidate the pharmacology and mechanism of action of this putative signaling pathway. Dopamine dose-response curves assaying intracellular calcium mobilization in cells heterologously expressing the D1 and D2 subtypes, either alone or in combination, and using subtype selective ligands revealed that concurrent stimulation is required for coupling. Surprisingly, characterization of a putative D1-D2 heteromer-selective ligand, 6-chloro-2,3,4,5-tetrahydro-3-methyl-1-(3-methylphenyl)-1H-3-benzazepine-7,8-diol (SKF83959), found no stimulation of calcium release, but it did find a broad range of cross-reactivity with other G protein–coupled receptors. In contrast, SKF83959 appeared to be an antagonist of calcium mobilization. Overexpression of Gqα with the D1 and D2 dopamine receptors enhanced the dopamine-stimulated calcium response. However, this was also observed in cells expressing Gqα with only the D1R. Inactivation of Gi or Gs with pertussis or cholera toxin, respectively, largely, but not entirely, reduced the calcium response in D1R and D2R cotransfected cells. Moreover, sequestration of Gβγ subunits through overexpression of G protein receptor kinase 2 mutants either completely or largely eliminated dopamine-stimulated calcium mobilization. Our data suggest that the mechanism of D1R/D2R–mediated calcium signaling involves more than receptor-mediated Gq protein activation, may largely involve downstream signaling pathways, and may not be completely heteromer-specific. In addition, SKF83959 may not exhibit selective activation of D1-D2 heteromers, and its significant cross-reactivity to other receptors warrants careful interpretation of its use in vivo. PMID:23680635

  18. Ethanol Increases Mechanical Pain Sensitivity in Rats via Activation of GABAA Receptors in Medial Prefrontal Cortex.

    PubMed

    Geng, Kai-Wen; He, Ting; Wang, Rui-Rui; Li, Chun-Li; Luo, Wen-Jun; Wu, Fang-Fang; Wang, Yan; Li, Zhen; Lu, Yun-Fei; Guan, Su-Min; Chen, Jun

    2016-10-01

    Ethanol is widely known for its ability to cause dramatic changes in emotion, social cognition, and behavior following systemic administration in humans. Human neuroimaging studies suggest that alcohol dependence and chronic pain may share common mechanisms through amygdala-medial prefrontal cortex (mPFC) interactions. However, whether acute administration of ethanol in the mPFC can modulate pain perception is unknown. Here we showed that bilateral microinjections of ethanol into the prelimbic and infralimbic areas of the mPFC lowered the bilateral mechanical pain threshold for 48 h without influencing thermal pain sensitivity in adult rats. However, bilateral microinjections of artificial cerebrospinal fluid into the mPFC or bilateral microinjections of ethanol into the dorsolateral PFC (also termed as motor cortex area 1 in Paxinos and Watson's atlas of The Rat Brain. Elsevier Academic Press, Amsterdam, 2005) failed to do so, suggesting regional selectivity of the effects of ethanol. Moreover, bilateral microinjections of ethanol did not change the expression of either pro-apoptotic (caspase-3 and Bax) or anti-apoptotic (Bcl-2) proteins, suggesting that the dose was safe and validating the method used in the current study. To determine whether γ-aminobutyric acid A (GABAA) receptors are involved in mediating the ethanol effects, muscimol, a selective GABAA receptor agonist, or bicuculline, a selective GABAA receptor antagonist, was administered alone or co-administered with ethanol through the same route into the bilateral mPFC. The results showed that muscimol mimicked the effects of ethanol while bicuculline completely reversed the effects of ethanol and muscimol. In conclusion, ethanol increases mechanical pain sensitivity through activation of GABAA receptors in the mPFC of rats.

  19. Spinal alpha3beta2* nicotinic acetylcholine receptors tonically inhibit the transmission of nociceptive mechanical stimuli.

    PubMed

    Young, Tracey; Wittenauer, Shannon; McIntosh, J Michael; Vincler, Michelle

    2008-09-10

    The presence of non-alpha4beta2, non-alpha7 nicotinic acetylcholine receptors (nAChR) in the rat spinal cord has been suggested previously, but the identity of these nAChRs had not been shown. Intrathecal administration of the alpha3beta2*/alpha6beta2* selective alpha-conotoxin MII (alpha-CTX MII) dose- and time-dependently reduced paw withdrawal thresholds to mechanical pressure in normal rats. The pronociceptive effect of alpha-CTX MII was partially blocked by NMDA receptor antagonism and lost completely following ablation of C-fibers. The effect of spinal nerve ligation on alpha-CTX MII-induced mechanical hypersensitivity was also assessed. Sensitivity was lost in the hind paw ipsilateral to spinal nerve ligation, but maintained in the contralateral hind paw at control levels. Radioligand binding in spinal cord membranes revealed high and low affinity alpha-CTX MII binding sites. Spinal nerve ligation did not significantly alter alpha-CTX MII binding ipsilateral to ligation. Finally, no evidence for the presence of alpha6-containing nAChRs was identified. The results of these studies show the presence of 2 populations of alpha-CTX MII-sensitive nAChRs containing the alpha3 and beta2, but not the alpha6, subunits in the rat spinal cord that function to inhibit the transmission of nociceptive mechanical stimuli via inhibiting the release of glutamate from C-fibers. Spinal nerve ligation produces a unilateral loss of alpha-CTX MII-induced mechanical hypersensitivity without altering alpha-CTX MII binding sites. Our data support a peripheral injury-induced loss of a cholinergic inhibitory tone at spinal alpha3beta2* nAChRs, without the loss of the receptors themselves, which may contribute to mechanical hypersensitivity following spinal nerve ligation.

  20. Spinal α3β2* nicotinic acetylcholine receptors tonically inhibit the transmission of nociceptive mechanical stimuli

    PubMed Central

    Young, Tracey; Wittenauer, Shannon; McIntosh, J. Michael; Vincler, Michelle

    2008-01-01

    The presence of non-α4β2, non-α7 nicotinic acetylcholine receptors (nAChR) in the rat spinal cord has been suggested previously, but the identity of these nAChRs had not been shown. Intrathecal administration of the α3β2*/α6β2* selective α-conotoxin MII (α-CTX MII) dose- and time-dependently reduced paw withdrawal thresholds to mechanical pressure in normal rats. The pronociceptive effect of α-CTX MII was partially blocked by NMDA receptor antagonism and lost completely following ablation of C-fibers. The effect of spinal nerve ligation on α-CTX MII-induced mechanical hypersensitivity was also assessed. Sensitivity was lost in the hind paw ipsilateral to spinal nerve ligation, but maintained in the contralateral hind paw at control levels.. Radioligand binding in spinal cord membranes revealed high and low affinity α-CTX MII binding sites. Spinal nerve ligation did not significantly alter α-CTX MII binding ipsilateral to ligation. Finally, no evidence for the presence of α6-containing nAChRs was identified. The results of these studies show the presence of 2 populations of α-CTX MII-sensitive nAChRs containing the α3 and β2, but not the α6, subunits in the rat spinal cord that function to inhibit the transmission of nociceptive mechanical stimuli via inhibiting the release of glutamate from C-fibers. Spinal nerve ligation produces a unilateral loss of α-CTX MII-induced mechanical hypersensitivity without altering α-CTX MII binding sites. Our data support a peripheral injury-induced loss of a cholinergic inhibitory tone at spinal α3β2* nAChRs, without the loss of the receptors themselves, which may contribute to mechanical hypersensitivity following spinal nerve ligation. PMID:18634758

  1. Molecular Mechanisms of Cardiotoxicity Induced by ErbB Receptor Inhibitor Cancer Therapeutics

    PubMed Central

    Hervent, Anne-Sophie; De Keulenaer, Gilles W.

    2012-01-01

    The introduction of the so-called “targeted therapies”, particularly those drugs that inhibit the activity of tyrosine kinases, has represented a remarkable progress in the treatment of cancer. Although these drugs improve survival rates in cancer, significant cardiotoxicity, manifesting as left vertricular dysfunction and/or heart failure, has emerged. The ErbB receptor tyrosine kinases are being pursued as therapeutic targets because of their important roles in normal physiology and in cancer. Besides the fact that the ErbB receptors are indispensable during development and in normal adult physiology, epidermal growth factor (EGFR) and ErbB2 in particular have been implicated in the development of many human cancers. This review focuses on the rationale for targeting members of ErbB receptor family and numerous agents that are in use for inhibiting the pathway. We summarize the current knowledge on the physiological role of ErbB signaling in the ventricle and on structural aspects of ErbB receptor activation in cancer and cardiac cells. We examine the underlying mechanisms that result in on-target or off-target cardiotoxicities of ErbB inhibitors, which can influence the design of future anticancer therapies. PMID:23202898

  2. Dual role of SLP-76 in mediating T cell receptor-induced activation of phospholipase C-gamma1.

    PubMed

    Beach, Dvora; Gonen, Ronnie; Bogin, Yaron; Reischl, Ilona G; Yablonski, Deborah

    2007-02-02

    Phospholipase C-gamma1 (PLC-gamma1) activation depends on a heterotrimeric complex of adaptor proteins composed of LAT, Gads, and SLP-76. Upon T cell receptor stimulation, a portion of PLC-gamma1 is recruited to a detergent-resistant membrane fraction known as the glycosphingolipid-enriched membrane microdomains (GEMs), or lipid rafts, to which LAT is constitutively localized. In addition to LAT, PLC-gamma1 GEM recruitment depended on SLP-76, and, in particular, required the Gads-binding domain of SLP-76. The N-terminal tyrosine phosphorylation sites and P-I region of SLP-76 were not required for PLC-gamma1 GEM recruitment, but were required for PLC-gamma1 phosphorylation at Tyr(783). Thus, GEM recruitment can be insufficient for full activation of PLC-gamma1 in the absence of a second SLP-76-mediated event. Indeed, a GEM-targeted derivative of PLC-gamma1 depended on SLP-76 for T cell receptor-induced phosphorylation at Tyr783 and subsequent NFAT activation. On a biochemical level, SLP-76 inducibly associated with both Vav and catalytically active ITK, which efficiently phosphorylated a PLC-gamma1 fragment at Tyr783 in vitro. Both associations were disrupted upon mutation of the N-terminal tyrosine phosphorylation sites of SLP-76. The P-I region deletion disrupted Vav association and reduced SLP-76-associated kinase activity. A smaller deletion within the P-I region, which does not impair PLC-gamma1 activation, did not impair the association with Vav, but reduced SLP-76-associated kinase activity. These results provide new insight into the multiple roles of SLP-76 and the functional importance of its interactions with other signaling proteins.

  3. Loss of the membrane anchor of the target receptor is a mechanism of bioinsecticide resistance

    PubMed Central

    Darboux, Isabelle; Pauchet, Yannick; Castella, Claude; Silva-Filha, Maria Helena; Nielsen-LeRoux, Christina; Charles, Jean-François; Pauron, David

    2002-01-01

    The mosquitocidal activity of Bacillus sphaericus is because of a binary toxin (Bin), which binds to Culex pipiens maltase 1 (Cpm1), an α-glucosidase present in the midgut of Culex pipiens larvae. In this work, we studied the molecular basis of the resistance to Bin developed by a strain (GEO) of C. pipiens. Immunohistochemical and in situ hybridization experiments showed that Cpm1 was undetectable in the midgut of GEO larvae, although the gene was correctly transcribed. The sequence of the cpm1GEO cDNA differs from the sequence we previously reported for a susceptible strain (cpm1IP) by seven mutations: six missense mutations and a mutation leading to the premature termination of translation. When produced in insect cells, Cpm1IP was attached to the membrane by a glycosylphosphatidylinositol (GPI). In contrast, the premature termination of translation of Cpm1GEO resulted in the targeting of the protein to the extracellular compartment because of truncation of the GPI-anchoring site. The interaction between Bin and Cpm1GEO and the enzyme activity of the receptor were not affected. Thus, Bin is not toxic to GEO larvae because it cannot interact with the midgut cell membrane, even though its receptor site is unaffected. This mechanism contrasts with other known resistance mechanisms in which point mutations decrease the affinity of binding between the receptor and the toxin. PMID:11983886

  4. Sigma-1 Receptor Agonism Promotes Mechanical Allodynia After Priming the Nociceptive System with Capsaicin.

    PubMed

    Entrena, J M; Sánchez-Fernández, C; Nieto, F R; González-Cano, R; Yeste, S; Cobos, E J; Baeyens, J M

    2016-11-25

    Sigma-1 receptor antagonists promote antinociception in several models of pain, but the effects of sigma-1 agonists on nociception (particularly when the nociceptive system is primed) are not so well characterized; therefore we evaluated the effects of sigma-1 agonists on pain under different experimental conditions. The systemic administration of the selective sigma-1 agonists (+)-pentazocine and PRE-084, as well as the nonselective sigma-1 agonist carbetapentane (used clinically as an antitussive drug), did not alter sensitivity to mechanical stimulation under baseline conditions. However, they greatly promoted secondary mechanical allodynia after priming the nociceptive system with capsaicin. These effects of sigma-1 agonists were consistent in terms potency with the affinities of these drugs for sigma-1 receptors, were reversed by sigma-1 antagonists, and were not observed in sigma-1 knockout mice, indicating that they are sigma-1-mediated. Repeated systemic treatment with PRE-084 induced proallodynic effects even 24 h after treatment completion, but only after the nociceptive system was primed. However, neither the presence of this drug in the organism nor changes in sigma-1 receptor expression in areas involved in pain processing explains its long-term effects, suggesting that sustained sigma-1 agonism induces plastic changes in the nociceptive system that promote nociception.

  5. Gut bitter taste receptor signalling induces ABCB1 through a mechanism involving CCK.

    PubMed

    Jeon, Tae-Il; Seo, Young-Kyo; Osborne, Timothy F

    2011-08-15

    T2Rs (bitter taste-sensing type 2 receptors) are expressed in the oral cavity to prevent ingestion of dietary toxins through taste avoidance. They are also expressed in other cell types, including gut enteroendocrine cells, where their physiological role is enigmatic. Previously, we proposed that T2R-dependent CCK (cholecystokinin) secretion from enteroendocrine cells limits absorption of dietary toxins, but an active mechanism was lacking. In the present study we show that T2R signalling activates ABCB1 (ATP-binding cassette B1) in intestinal cells through a CCK signalling mechanism. PTC (phenylthiocarbamide), an agonist for the T2R38 bitter receptor, increased ABCB1 expression in both intestinal cells and mouse intestine. PTC induction of ABCB1 was decreased by either T2R38 siRNA (small interfering RNA) or treatment with YM022, a gastrin receptor antagonist. Thus gut ABCB1 is regulated through signalling by CCK/gastrin released in response to PTC stimulation of T2R38 on enteroendocrine cells. We also show that PTC increases the efflux activity of ABCB1, suggesting that T2R signalling limits the absorption of bitter tasting/toxic substances through modulation of gut efflux membrane transporters.

  6. Loss of the membrane anchor of the target receptor is a mechanism of bioinsecticide resistance.

    PubMed

    Darboux, Isabelle; Pauchet, Yannick; Castella, Claude; Silva-Filha, Maria Helena; Nielsen-LeRoux, Christina; Charles, Jean-François; Pauron, David

    2002-04-30

    The mosquitocidal activity of Bacillus sphaericus is because of a binary toxin (Bin), which binds to Culex pipiens maltase 1 (Cpm1), an alpha-glucosidase present in the midgut of Culex pipiens larvae. In this work, we studied the molecular basis of the resistance to Bin developed by a strain (GEO) of C. pipiens. Immunohistochemical and in situ hybridization experiments showed that Cpm1 was undetectable in the midgut of GEO larvae, although the gene was correctly transcribed. The sequence of the cpm1(GEO) cDNA differs from the sequence we previously reported for a susceptible strain (cpm1(IP)) by seven mutations: six missense mutations and a mutation leading to the premature termination of translation. When produced in insect cells, Cpm1(IP) was attached to the membrane by a glycosylphosphatidylinositol (GPI). In contrast, the premature termination of translation of Cpm1(GEO) resulted in the targeting of the protein to the extracellular compartment because of truncation of the GPI-anchoring site. The interaction between Bin and Cpm1(GEO) and the enzyme activity of the receptor were not affected. Thus, Bin is not toxic to GEO larvae because it cannot interact with the midgut cell membrane, even though its receptor site is unaffected. This mechanism contrasts with other known resistance mechanisms in which point mutations decrease the affinity of binding between the receptor and the toxin.

  7. Sigma-1 Receptor Agonism Promotes Mechanical Allodynia After Priming the Nociceptive System with Capsaicin

    PubMed Central

    Entrena, J. M.; Sánchez-Fernández, C.; Nieto, F. R.; González-Cano, R.; Yeste, S.; Cobos, E. J.; Baeyens, J. M.

    2016-01-01

    Sigma-1 receptor antagonists promote antinociception in several models of pain, but the effects of sigma-1 agonists on nociception (particularly when the nociceptive system is primed) are not so well characterized; therefore we evaluated the effects of sigma-1 agonists on pain under different experimental conditions. The systemic administration of the selective sigma-1 agonists (+)-pentazocine and PRE-084, as well as the nonselective sigma-1 agonist carbetapentane (used clinically as an antitussive drug), did not alter sensitivity to mechanical stimulation under baseline conditions. However, they greatly promoted secondary mechanical allodynia after priming the nociceptive system with capsaicin. These effects of sigma-1 agonists were consistent in terms potency with the affinities of these drugs for sigma-1 receptors, were reversed by sigma-1 antagonists, and were not observed in sigma-1 knockout mice, indicating that they are sigma-1-mediated. Repeated systemic treatment with PRE-084 induced proallodynic effects even 24 h after treatment completion, but only after the nociceptive system was primed. However, neither the presence of this drug in the organism nor changes in sigma-1 receptor expression in areas involved in pain processing explains its long-term effects, suggesting that sustained sigma-1 agonism induces plastic changes in the nociceptive system that promote nociception. PMID:27886264

  8. The morphology and mechanical sensitivity of lateral line receptors in zebrafish larvae (Danio rerio).

    PubMed

    Van Trump, William J; McHenry, Matthew J

    2008-07-01

    The lateral line system of fish and amphibians detects water flow with receptors on the surface of the body. Although differences in the shape of these receptors, called neuromasts, are known to influence their mechanics, it is unclear how neuromast morphology affects the sensitivity of the lateral line system. We examined the functional consequences of morphological variation by measuring the dimensions of superficial neuromasts in zebrafish larvae (Danio rerio) and mathematically modeling their mechanics. These measurements used a novel morphometric technique that recorded landmarks in three dimensions at a microscopic scale. The mathematical model predicted mechanical sensitivity as the ratio of neuromast deflection to flow velocity for a range of stimulus frequencies. These predictions suggest that variation in morphology within this species generates a greater than 30-fold range in the amplitude of sensitivity and more than a 200-fold range of variation in cut-off frequency. Most of this variation was generated by differences in neuromast height that do not correlate with body position. Our results suggest that natural variation in cupular height within a species is capable of generating large differences in their mechanical filtering and dynamic range.

  9. Kainate receptor pore‐forming and auxiliary subunits regulate channel block by a novel mechanism

    PubMed Central

    Brown, Patricia M. G. E.; Aurousseau, Mark R. P.; Musgaard, Maria; Biggin, Philip C.

    2016-01-01

    Key points Kainate receptor heteromerization and auxiliary subunits, Neto1 and Neto2, attenuate polyamine ion‐channel block by facilitating blocker permeation.Relief of polyamine block in GluK2/GluK5 heteromers results from a key proline residue that produces architectural changes in the channel pore α‐helical region.Auxiliary subunits exert an additive effect to heteromerization, and thus relief of polyamine block is due to a different mechanism.Our findings have broad implications for work on polyamine block of other cation‐selective ion channels. Abstract Channel block and permeation by cytoplasmic polyamines is a common feature of many cation‐selective ion channels. Although the channel block mechanism has been studied extensively, polyamine permeation has been considered less significant as it occurs at extreme positive membrane potentials. Here, we show that kainate receptor (KAR) heteromerization and association with auxiliary proteins, Neto1 and Neto2, attenuate polyamine block by enhancing blocker permeation. Consequently, polyamine permeation and unblock occur at more negative and physiologically relevant membrane potentials. In GluK2/GluK5 heteromers, enhanced permeation is due to a single proline residue in GluK5 that alters the dynamics of the α‐helical region of the selectivity filter. The effect of auxiliary proteins is additive, and therefore the structural basis of polyamine permeation and unblock is through a different mechanism. As native receptors are thought to assemble as heteromers in complex with auxiliary proteins, our data identify an unappreciated impact of polyamine permeation in shaping the signalling properties of neuronal KARs and point to a structural mechanism that may be shared amongst other cation‐selective ion channels. PMID:26682513

  10. Caffeine stimulates locomotor activity in the mammalian spinal cord via adenosine A1 receptor-dopamine D1 receptor interaction and PKA-dependent mechanisms.

    PubMed

    Acevedo, JeanMarie; Santana-Almansa, Alexandra; Matos-Vergara, Nikol; Marrero-Cordero, Luis René; Cabezas-Bou, Ernesto; Díaz-Ríos, Manuel

    2016-02-01

    Caffeine is a potent psychostimulant that can have significant and widely variable effects on the activity of multiple neuronal pathways. The most pronounced caffeine-induced behavioral effect seen in rodents is to increase locomotor activity which has been linked to a dose-dependent inhibition of A1 and A(2A) receptors. The effects of caffeine at the level of the lumbar spinal central pattern generator (CPG) network for hindlimb locomotion are lacking. We assessed the effects of caffeine to the locomotor function of the spinal CPG network via extracellular ventral root recordings using the isolated neonatal mouse spinal cord preparation. Addition of caffeine and of an A1 receptor antagonist significantly decreased the cycle period accelerating the ongoing locomotor rhythm, while decreasing burst duration reversibly in most preparations suggesting the role of A1 receptors as the primary target of caffeine. Caffeine and an A1 receptor antagonist failed to stimulate ongoing locomotor activity in the absence of dopamine or in the presence of a D1 receptor antagonist supporting A1/D1 receptor-dependent mechanism of action. The use of caffeine or an A1 receptor blocker failed to stimulate an ongoing locomotor rhythm in the presence of a blocker of the cAMP-dependent protein kinase (PKA) supporting the need of this intracellular pathway for the modulatory effects of caffeine to occur. These results support a stimulant effect of caffeine on the lumbar spinal network controlling hindlimb locomotion through the inhibition of A1 receptors and subsequent activation of D1 receptors via a PKA-dependent intracellular mechanism.

  11. Flunitrazepam rapidly reduces GABAA receptor subunit protein expression via a protein kinase C-dependent mechanism

    PubMed Central

    Johnston, Jonathan D; Price, Sally A; Bristow, David R

    1998-01-01

    Acute flunitrazepam (1 μM) exposure for 1 h reduced GABAA receptor α1 (22±4%, mean±s.e.mean) and β2/3 (21±4%) subunit protein levels in cultured rat cerebellar granule cells. This rapid decrease in subunit proteins was completely prevented by bisindolymaleimide 1 (1 μM), an inhibitor of protein kinase C, but not by N-[2-((p-bromocinnamyl)amino)ethyl]-5-isoquinolinesulfonamide (H-89, 4.8 μM), an inhibitor of protein kinases A and G. These results suggest the existence of a benzodiazepine-induced mechanism to rapidly alter GABAA receptor protein expression, that appears to be dependent on protein kinase C activity. PMID:9723942

  12. [Functional properties of taste bud cells. Mechanisms of afferent neurotransmission in Type II taste receptor cells].

    PubMed

    Romanov, R A

    2013-01-01

    Taste Bud cells are heterogeneous in their morphology and functionality. These cells are responsible for sensing a wide variety of substances and for associating detected compounds with a different taste: bitter, sweet, salty, sour and umami. Today we know that each of the five basic tastes corresponds to distinct cell populations organized into three basic morpho-functional cell types. In addition, some receptor cells of the taste bud demonstrate glia-related functions. In this article we expand on some properties of these three morphological receptor cell types. Main focus is devoted to the Type II cells and unusual mechanism for afferent neurotransmission in these cells. Taste cells of the Type II consist of three populations detecting bitter, sweet and umami tastes, and, thus, evoke a serious scientific interest.

  13. Progesterone modulates the LPS-induced nitric oxide production by a progesterone-receptor independent mechanism.

    PubMed

    Wolfson, Manuel Luis; Schander, Julieta Aylen; Bariani, María Victoria; Correa, Fernando; Franchi, Ana María

    2015-12-15

    Genital tract infections caused by Gram-negative bacteria induce miscarriage and are one of the most common complications of human pregnancy. LPS administration to 7-day pregnant mice induces embryo resorption after 24h, with nitric oxide playing a fundamental role in this process. We have previously shown that progesterone exerts protective effects on the embryo by modulating the inflammatory reaction triggered by LPS. Here we sought to investigate whether the in vivo administration of progesterone modulated the LPS-induced nitric oxide production from peripheral blood mononuclear cells from pregnant and non-pregnant mice. We found that progesterone downregulated LPS-induced nitric oxide production by a progesterone receptor-independent mechanism. Moreover, our results suggest a possible participation of glucocorticoid receptors in at least some of the anti-inflammatory effects of progesterone.

  14. The Protective Effect of Alpha 7 Nicotinic Acetylcholine Receptor Activation on Critical Illness and Its Mechanism

    PubMed Central

    REN, Chao; TONG, Ya-lin; LI, Jun-cong; LU, Zhong-qiu; YAO, Yong-ming

    2017-01-01

    Critical illnesses and injuries are recognized as major threats to human health, and they are usually accompanied by uncontrolled inflammation and dysfunction of immune response. The alpha 7 nicotinic acetylcholine receptor (α7nAchR), which is a primary receptor of cholinergic anti-inflammatory pathway (CAP), exhibits great benefits for critical ill conditions. It is composed of 5 identical α7 subunits that form a central pore with high permeability for calcium. This putative structure is closely associated with its functional states. Activated α7nAChR exhibits extensive anti-inflammatory and immune modulatory reactions, including lowered pro-inflammatory cytokines levels, decreased expressions of chemokines as well as adhesion molecules, and altered differentiation and activation of immune cells, which are important in maintaining immune homeostasis. Well understanding of the effects and mechanisms of α7nAChR will be of great value in exploring effective targets for treating critical diseases. PMID:28123345

  15. REVIEWMolecular mechanisms underlying physiological and receptor pleiotropic effects mediated by GLP-1R activation

    PubMed Central

    Pabreja, K; Mohd, M A; Koole, C; Wootten, D; Furness, S G B

    2014-01-01

    The incidence of type 2 diabetes in developed countries is increasing yearly with a significant negative impact on patient quality of life and an enormous burden on the healthcare system. Current biguanide and thiazolidinedione treatments for type 2 diabetes have a number of clinical limitations, the most serious long-term limitation being the eventual need for insulin replacement therapy (Table 1). Since 2007, drugs targeting the glucagon-like peptide-1 (GLP-1) receptor have been marketed for the treatment of type 2 diabetes. These drugs have enjoyed a great deal of success even though our underlying understanding of the mechanisms for their pleiotropic effects remain poorly characterized even while major pharmaceutical companies actively pursue small molecule alternatives. Coupling of the GLP-1 receptor to more than one signalling pathway (pleiotropic signalling) can result in ligand-dependent signalling bias and for a peptide receptor such as the GLP-1 receptor this can be exaggerated with the use of small molecule agonists. Better consideration of receptor signalling pleiotropy will be necessary for future drug development. This is particularly important given the recent failure of taspoglutide, the report of increased risk of pancreatitis associated with GLP-1 mimetics and the observed clinical differences between liraglutide, exenatide and the newly developed long-acting exenatide long acting release, albiglutide and dulaglutide. Linked ArticlesThis article is part of a themed section on Molecular Pharmacology of GPCRs. To view the other articles in this section visit http://dx.doi.org/10.1111/bph.2014.171.issue-5 PMID:23889512

  16. Mechanisms of dopamine D(1) and angiotensin type 2 receptor interaction in natriuresis.

    PubMed

    Padia, Shetal H; Kemp, Brandon A; Howell, Nancy L; Keller, Susanna R; Gildea, John J; Carey, Robert M

    2012-02-01

    Renal dopamine D(1)-like receptors (D(1)Rs) and angiotensin type 2 receptors (AT(2)Rs) are important natriuretic receptors counterbalancing angiotensin type 1 receptor-mediated tubular sodium reabsorption. Here we explore the mechanisms of D(1)R and AT(2)R interactions in natriuresis. In uninephrectomized, sodium-loaded Sprague-Dawley rats, direct renal interstitial infusion of the highly selective D(1)R agonist fenoldopam induced a natriuretic response that was abolished by the AT(2)R-specific antagonist PD-123319 or by microtubule polymerization inhibitor nocodazole but not by actin polymerization inhibitor cytochalasin D. By confocal microscopy and immunoelectron microscopy, fenoldopam translocated AT(2)Rs from intracellular sites to the apical plasma membranes of renal proximal tubule cells, and this translocation was abolished by nocodazole. Because D(1)R activation induces natriuresis via an adenylyl cyclase/cAMP signaling pathway, we explored whether this pathway is responsible for AT(2)R recruitment and AT(2)R-mediated natriuresis. Renal interstitial coinfusion of the adenylyl cyclase activator forskolin and 3-isobutly-1-methylxanthine induced natriuresis that was abolished either by PD-123319 or nocodazole but was unaffected by specific the D(1)R antagonist SCH-23390. Coadministration of forskolin and 3-isobutly-1-methylxanthine also translocated AT(2)Rs to the apical plasma membranes of renal proximal tubule cells; this translocation was abolished by nocodazole but was unaffected by SCH-23390. The results demonstrate that D(1)R-induced natriuresis requires AT(2)R recruitment to the apical plasma membranes of renal proximal tubule cells in a microtubule-dependent manner involving an adenylyl cyclase/cAMP signaling pathway. These studies provide novel insights regarding the mechanisms whereby renal D(1)Rs and AT(2)Rs act in concert to promote sodium excretion in vivo.

  17. New Hyperekplexia Mutations Provide Insight into Glycine Receptor Assembly, Trafficking, and Activation Mechanisms*

    PubMed Central

    Bode, Anna; Wood, Sian-Elin; Mullins, Jonathan G. L.; Keramidas, Angelo; Cushion, Thomas D.; Thomas, Rhys H.; Pickrell, William O.; Drew, Cheney J. G.; Masri, Amira; Jones, Elizabeth A.; Vassallo, Grace; Born, Alfred P.; Alehan, Fusun; Aharoni, Sharon; Bannasch, Gerald; Bartsch, Marius; Kara, Bulent; Krause, Amanda; Karam, Elie G.; Matta, Stephanie; Jain, Vivek; Mandel, Hanna; Freilinger, Michael; Graham, Gail E.; Hobson, Emma; Chatfield, Sue; Vincent-Delorme, Catherine; Rahme, Jubran E.; Afawi, Zaid; Berkovic, Samuel F.; Howell, Owain W.; Vanbellinghen, Jean-François; Rees, Mark I.; Chung, Seo-Kyung; Lynch, Joseph W.

    2013-01-01

    Hyperekplexia is a syndrome of readily provoked startle responses, alongside episodic and generalized hypertonia, that presents within the first month of life. Inhibitory glycine receptors are pentameric ligand-gated ion channels with a definitive and clinically well stratified linkage to hyperekplexia. Most hyperekplexia cases are caused by mutations in the α1 subunit of the human glycine receptor (hGlyR) gene (GLRA1). Here we analyzed 68 new unrelated hyperekplexia probands for GLRA1 mutations and identified 19 mutations, of which 9 were novel. Electrophysiological analysis demonstrated that the dominant mutations p.Q226E, p.V280M, and p.R414H induced spontaneous channel activity, indicating that this is a recurring mechanism in hGlyR pathophysiology. p.Q226E, at the top of TM1, most likely induced tonic activation via an enhanced electrostatic attraction to p.R271 at the top of TM2, suggesting a structural mechanism for channel activation. Receptors incorporating p.P230S (which is heterozygous with p.R65W) desensitized much faster than wild type receptors and represent a new TM1 site capable of modulating desensitization. The recessive mutations p.R72C, p.R218W, p.L291P, p.D388A, and p.E375X precluded cell surface expression unless co-expressed with α1 wild type subunits. The recessive p.E375X mutation resulted in subunit truncation upstream of the TM4 domain. Surprisingly, on the basis of three independent assays, we were able to infer that p.E375X truncated subunits are incorporated into functional hGlyRs together with unmutated α1 or α1 plus β subunits. These aberrant receptors exhibit significantly reduced glycine sensitivity. To our knowledge, this is the first suggestion that subunits lacking TM4 domains might be incorporated into functional pentameric ligand-gated ion channel receptors. PMID:24108130

  18. The aspartate receptor cytoplasmic domain: in situ chemical analysis of structure, mechanism and dynamics

    PubMed Central

    Bass, Randal B; Falke, Joseph J

    2010-01-01

    Background Site-directed sulfhydryl chemistry and spectroscopy can be used to probe protein structure, mechanism and dynamics in situ. The aspartate receptor of bacterial chemotaxis is representative of a large family of prokaryotic and eukaryotic receptors that regulate histidine kinases in two-component signaling pathways, and has become one of the best characterized transmembrane receptors. We report here the use of cysteine and disulfide scanning to probe the helix-packing architecture of the cytoplasmic domain of the aspartate receptor. Results A series of designed cysteine pairs have been used to detect proximities between cytoplasmic helices in the full-length, membrane-bound receptor by measurement of disulfide-bond formation rates. Upon mild oxidation, 25 disulfide bonds form rapidly between three specific pairs of helices, whereas other helix pairs yield no detectable disulfide-bond formation. Further constraints on helix packing are provided by 14 disulfide bonds that retain receptor function in an in vitro kinase regulation assay. Of these functional disulfides, seven lock the receptor in the conformation that constitutively stimulates kinase activity (‘lock on’), whereas the remaining seven retain normal kinase regulation. Finally, disulfide-trapping experiments in the absence of bound kinase reveal large-amplitude relative motions of adjacent helices, including helix translations and rotations of up to 19 Å and 180°, respectively. Conclusions The 25 rapidly formed and 14 functional disulfide bonds identify helix–helix contacts and theirregister in the full-length, membrane-bound receptor–kinase complex. The results reveal an extended, rather than compact, domain architecture in which the observed helix–helix interactions are best described by a four-helix bundle arrangement. A cluster of six lock-on disulfide bonds pinpoints a region of the subunit interface critical for kinase activation, whereas the signal-retaining disulfides indicate

  19. Structure-guided enzymology of the lipid A acyltransferase LpxM reveals a dual activity mechanism

    PubMed Central

    Dovala, Dustin; Rath, Christopher M.; Hu, Qijun; Sawyer, William S.; Shia, Steven; Elling, Robert A.; Knapp, Mark S.; Metzger, Louis E.

    2016-01-01

    Gram-negative bacteria possess a characteristic outer membrane, of which the lipid A constituent elicits a strong host immune response through the Toll-like receptor 4 complex, and acts as a component of the permeability barrier to prevent uptake of bactericidal compounds. Lipid A species comprise the bulk of the outer leaflet of the outer membrane and are produced through a multistep biosynthetic pathway conserved in most Gram-negative bacteria. The final steps in this pathway involve the secondary acylation of lipid A precursors. These are catalyzed by members of a superfamily of enzymes known as lysophospholipid acyltransferases (LPLATs), which are present in all domains of life and play important roles in diverse biological processes. To date, characterization of this clinically important class of enzymes has been limited by a lack of structural information and the availability of only low-throughput biochemical assays. In this work, we present the structure of the bacterial LPLAT protein LpxM, and we describe a high-throughput, label-free mass spectrometric assay to characterize acyltransferase enzymatic activity. Using our structure and assay, we identify an LPLAT thioesterase activity, and we provide experimental evidence to support an ordered-binding and “reset” mechanistic model for LpxM function. This work enables the interrogation of other bacterial acyltransferases’ structure–mechanism relationships, and the assay described herein provides a foundation for quantitatively characterizing the enzymology of any number of clinically relevant LPLAT proteins. PMID:27681620

  20. Structure-guided enzymology of the lipid A acyltransferase LpxM reveals a dual activity mechanism.

    PubMed

    Dovala, Dustin; Rath, Christopher M; Hu, Qijun; Sawyer, William S; Shia, Steven; Elling, Robert A; Knapp, Mark S; Metzger, Louis E

    2016-10-11

    Gram-negative bacteria possess a characteristic outer membrane, of which the lipid A constituent elicits a strong host immune response through the Toll-like receptor 4 complex, and acts as a component of the permeability barrier to prevent uptake of bactericidal compounds. Lipid A species comprise the bulk of the outer leaflet of the outer membrane and are produced through a multistep biosynthetic pathway conserved in most Gram-negative bacteria. The final steps in this pathway involve the secondary acylation of lipid A precursors. These are catalyzed by members of a superfamily of enzymes known as lysophospholipid acyltransferases (LPLATs), which are present in all domains of life and play important roles in diverse biological processes. To date, characterization of this clinically important class of enzymes has been limited by a lack of structural information and the availability of only low-throughput biochemical assays. In this work, we present the structure of the bacterial LPLAT protein LpxM, and we describe a high-throughput, label-free mass spectrometric assay to characterize acyltransferase enzymatic activity. Using our structure and assay, we identify an LPLAT thioesterase activity, and we provide experimental evidence to support an ordered-binding and "reset" mechanistic model for LpxM function. This work enables the interrogation of other bacterial acyltransferases' structure-mechanism relationships, and the assay described herein provides a foundation for quantitatively characterizing the enzymology of any number of clinically relevant LPLAT proteins.

  1. Simultaneous Activation of Induced Heterodimerization between CXCR4 Chemokine Receptor and Cannabinoid Receptor 2 (CB2) Reveals a Mechanism for Regulation of Tumor Progression.

    PubMed

    Coke, Christopher J; Scarlett, Kisha A; Chetram, Mahandranauth A; Jones, Kia J; Sandifer, Brittney J; Davis, Ahriea S; Marcus, Adam I; Hinton, Cimona V

    2016-05-06

    The G-protein-coupled chemokine receptor CXCR4 generates signals that lead to cell migration, cell proliferation, and other survival mechanisms that result in the metastatic spread of primary tumor cells to distal organs. Numerous studies have demonstrated that CXCR4 can form homodimers or can heterodimerize with other G-protein-coupled receptors to form receptor complexes that can amplify or decrease the signaling capacity of each individual receptor. Using biophysical and biochemical approaches, we found that CXCR4 can form an induced heterodimer with cannabinoid receptor 2 (CB2) in human breast and prostate cancer cells. Simultaneous, agonist-dependent activation of CXCR4 and CB2 resulted in reduced CXCR4-mediated expression of phosphorylated ERK1/2 and ultimately reduced cancer cell functions such as calcium mobilization and cellular chemotaxis. Given that treatment with cannabinoids has been shown to reduce invasiveness of cancer cells as well as CXCR4-mediated migration of immune cells, it is plausible that CXCR4 signaling can be silenced through a physical heterodimeric association with CB2, thereby inhibiting subsequent functions of CXCR4. Taken together, the data illustrate a mechanism by which the cannabinoid system can negatively modulate CXCR4 receptor function and perhaps tumor progression.

  2. Dual isotope plots reflect transformation pathways of pesticides: Potential to assess pesticide fate and elucidate transformation mechanisms

    NASA Astrophysics Data System (ADS)

    Meyer, Armin; Penning, Holger; Sorensen, Sebastian; Aamand, Jens; Elsner, Martin

    2010-05-01

    The degradation of pesticides in deeper soil layers and groundwater is of growing interest, because they have repeatedly been found in drinking water supply wells and may pose a risk to future water resources. Current assessment schemes face a common problem, however: natural degradation often cannot be reliably assessed by concentration measurements alone, since mass balances are difficult to establish and transformation cannot be distinguished from sorption or dilution. Even detection of metabolites may only give an incomplete picture. When several transformation pathways occur, some metabolites may be degraded or form bound residues so that the associated pathways may be missed. Our research shows that dual isotope plots derived from compound specific isotope analysis offer a novel approach to give additional, complementary insight into the natural degradation of pesticides. Detection of metabolites is not required, since the isotope fractionation can be fully observed in the pesticide itself. Specifically, different initial biotransformation reactions of the phenylurea herbicide isoproturon (3-(4-isopropylphenyl)-1,1-dimethylurea) in pure culture experiments with bacterial and fungal strains showed strongly pathway-dependent isotope fractionation. When analyzing isotopic changes in different parts of the isoproturon molecule, hydroxylation of the isopropyl group by fungi was found to be associated with C and H isotope fractionation. In contrast, hydrolysis by Arthrobacter globiformis D47 caused strong C and N isotope fractionation, albeit in a different manner than abiotic hydrolysis so that isotope measurements can distinguish between both modes of transformation. Likewise, we observed highly pathway-dependent C and N isotope fractionation of atrazine (1-chloro-3-ethylamino-5-isopropylamino-2,4,6-triazine). Desalkylation of atrazine by Rhodococcus sp. strain NI86/21 resulted in enrichment of both 13-C and 15-N in atrazine, whereas hydrolysis to hydroxyatrazine

  3. Metabotropic glutamate receptor-mediated long-term depression: molecular mechanisms.

    PubMed

    Gladding, Clare M; Fitzjohn, Stephen M; Molnár, Elek

    2009-12-01

    The ability to modify synaptic transmission between neurons is a fundamental process of the nervous system that is involved in development, learning, and disease. Thus, synaptic plasticity is the ability to bidirectionally modify transmission, where long-term potentiation and long-term depression (LTD) represent the best characterized forms of plasticity. In the hippocampus, two main forms of LTD coexist that are mediated by activation of either N-methyl-d-aspartic acid receptors (NMDARs) or metabotropic glutamate receptors (mGluRs). Compared with NMDAR-LTD, mGluR-LTD is less well understood, but recent advances have started to delineate the underlying mechanisms. mGluR-LTD at CA3:CA1 synapses in the hippocampus can be induced either by synaptic stimulation or by bath application of the group I selective agonist (R,S)-3,5-dihydroxyphenylglycine. Multiple signaling mechanisms have been implicated in mGluR-LTD, illustrating the complexity of this form of plasticity. This review provides an overview of recent studies investigating the molecular mechanisms underlying hippocampal mGluR-LTD. It highlights the role of key molecular components and signaling pathways that are involved in the induction and expression of mGluR-LTD and considers how the different signaling pathways may work together to elicit a persistent reduction in synaptic transmission.

  4. Mechanisms of protease-activated receptor 2-evoked hyperexcitability of nociceptive neurons innervating the mouse colon

    PubMed Central

    Kayssi, Ahmed; Amadesi, Silvia; Bautista, Francisco; Bunnett, Nigel W; Vanner, Stephen

    2007-01-01

    Agonists of protease-activated receptor 2 (PAR2) evoke hyperexcitability of dorsal root ganglia (DRG) neurons by unknown mechanisms. We examined the cellular mechanisms underlying PAR2-evoked hyperexcitability of mouse colonic DRG neurons to determine their potential role in pain syndromes such as visceral hyperalgesia. Colonic DRG neurons were identified by injecting Fast Blue and DiI retrograde tracers into the mouse colon. Using immunofluorescence, we found that DiI-labelled neurons contained PAR2 immunoreactivity, confirming the presence of receptors on colonic neurons. Whole-cell current-clamp recordings of acutely dissociated neurons demonstrated that PAR2 activation with a brief application (3 min) of PAR2 agonists, SLIGRL-NH2 and trypsin, evoked sustained depolarizations (up to 60 min) which were associated with increased input resistance and a marked reduction in rheobase (50% at 30 min). In voltage clamp, SLIGRL-NH2 markedly suppressed delayed rectifier IK currents (55% at 10 min), but had no effect on the transient IA current or TTX-resistant Na+ currents. In whole-cell current-clamp recordings, the sustained excitability evoked by PAR2 activation was blocked by the PKC inhibitor, calphostin, and the ERK1/2 inhibitor PD98059. Studies of ERK1/2 phosphorylation using confocal microscopy demonstrated that SLIGRL-NH2 increased levels of immunoreactive pERK1/2 in DRG neurons, particularly in proximity to the plasma membrane. Thus, activation of PAR2 receptors on colonic nociceptive neurons causes sustained hyperexcitability that is related, at least in part, to suppression of delayed rectifier IK currents. Both PKC and ERK1/2 mediate the PAR2-induced hyperexcitability. These studies describe a novel mechanism of sensitization of colonic nociceptive neurons that may be implicated in conditions of visceral hyperalgesia such as irritable bowel syndrome. PMID:17289784

  5. Novel Regulatory Mechanisms for Generation of the Soluble Leptin Receptor: Implications for Leptin Action

    PubMed Central

    Schaab, Michael; Kausch, Henriette; Klammt, Juergen; Nowicki, Marcin; Anderegg, Ulf; Gebhardt, Rolf; Rose-John, Stefan; Scheller, Juergen; Thiery, Joachim; Kratzsch, Juergen

    2012-01-01

    Background The adipokine leptin realizes signal transduction via four different membrane-anchored leptin receptor (Ob-R) isoforms in humans. However, the amount of functionally active Ob-R is affected by constitutive shedding of the extracellular domain via a so far unknown mechanism. The product of the cleavage process the so-called soluble leptin receptor (sOb-R) is the main binding protein for leptin in human blood and modulates its bioavailability. sOb-R levels are differentially regulated in metabolic disorders like type 1 diabetes mellitus or obesity and can, therefore, enhance or reduce leptin sensitivity. Methodology/Principal Findings To describe mechanisms of Ob-R cleavage and to investigate the functional significance of differential sOb-R levels we established a model of HEK293 cells transiently transfected with different human Ob-R isoforms. Using siRNA knockdown experiments we identified ADAM10 (A Disintegrin And Metalloproteinase 10) as a major protease for constitutive and activated Ob-R cleavage. Additionally, the induction of lipotoxicity and apoptosis led to enhanced shedding shown by increased levels of the soluble leptin receptor (sOb-R) in cell supernatants. Conversely, high leptin concentrations and ER stress reduced sOb-R levels. Decreased amounts of sOb-R due to ER stress were accompanied by impaired leptin signaling and reduced leptin binding. Conclusions Lipotoxicity and apoptosis increased Ob-R cleavage via ADAM10-dependent mechanisms. In contrast high leptin levels and ER stress led to reduced sOb-R levels. While increased sOb-R concentrations seem to directly block leptin action, reduced amounts of sOb-R may reflect decreased membrane expression of Ob-R. These findings could explain changes of leptin sensitivity which are associated with variations of serum sOb-R levels in metabolic diseases. PMID:22545089

  6. Allosteric mechanisms of G protein coupled receptor signaling: a structural perspective

    PubMed Central

    Thaker, Tarjani M.; Kaya, Ali I.; Preininger, Anita M.; Hamm, Heidi E.; Iverson, T.M.

    2012-01-01

    G protein-Coupled Receptors (GPCRs) use a complex series of intramolecular conformational changes to couple agonist binding to the binding and activation of cognate heterotrimeric G protein (Gαβγ). The mechanisms underlying this long-range activation have been identified using a variety of biochemical and structural approaches and have primarily used visual signal transduction via the GPCR rhodopsin and cognate heterotrimeric G protein transducin (Gt) as a model system. In this chapter, we will review the methods that have revealed allosteric signaling through rhodopsin and transducin. These methods can be applied to a variety of GPCR-mediated signaling pathways. PMID:22052489

  7. Role of GABAA and GABAB receptors and peripheral cholinergic mechanisms in the antinociceptive action of taurine.

    PubMed

    Serrano, M I; Serrano, J S; Guerrero, M R; Fernández, A

    1994-10-01

    1. Gabaergic and cholinergic mediation in the antinociceptive effect of taurine has been investigated in mice (acetic acid test) and rats (tail-flick test). 2. Scopolamine sulfate and methylnitrate exhibit intrinsic antinociceptive activity and increase the effect of taurine in mice. 3. Baclofen also increases the antinociceptive effect of taurine in mice. 4. Anticholinergic agents and bicuculline but not CGP 35348 antagonize the effect of taurine in rats. 5. These results suggest that the antinociceptive effect of taurine may be partly mediated by spinal GABAA receptors and peripheral cholinergic mechanisms.

  8. Receptor mutation is not a common mechanism of naturally occurring glucocorticoid resistance in leukaemia cell lines.

    PubMed

    Beesley, Alex H; Weller, Renae E; Senanayake, Saranga; Welch, Mathew; Kees, Ursula R

    2009-02-01

    Glucocorticoids (GCs) are among the most important drugs for the treatment of acute lymphoblastic leukaemia (ALL). Cell lines cultured in high GC concentrations typically contain mutated glucocorticoid receptor (GR), something that is rarely found in primary ALL specimens. We studied naturally occurring mechanisms of GC resistance and examined sensitivity to GC in 15 T-ALL cell lines grown without prior exposure to drugs. Resistance could not be attributed to mutations in GR or variations in levels of its expression. We conclude that this panel of cell lines provides a suitable in vitro model since it reflects GC resistance in primary ALL.

  9. Estradiol decreases cortical reactive astrogliosis after brain injury by a mechanism involving cannabinoid receptors.

    PubMed

    López Rodríguez, Ana Belén; Mateos Vicente, Beatriz; Romero-Zerbo, Silvana Y; Rodriguez-Rodriguez, Noé; Bellini, María José; Rodriguez de Fonseca, Fernando; Bermudez-Silva, Francisco Javier; Azcoitia, Iñigo; Garcia-Segura, Luis M; Viveros, María-Paz

    2011-09-01

    The neuroactive steroid estradiol reduces reactive astroglia after brain injury by mechanisms similar to those involved in the regulation of reactive gliosis by endocannabinoids. In this study, we have explored whether cannabinoid receptors are involved in the effects of estradiol on reactive astroglia. To test this hypothesis, the effects of estradiol, the cannabinoid CB1 antagonist/inverse agonist AM251, and the cannabinoid CB2 antagonist/inverse agonist AM630 were assessed in the cerebral cortex of male rats after a stab wound brain injury. Estradiol reduced the number of vimentin immunoreactive astrocytes and the number of glial fibrillary acidic protein immunoreactive astrocytes in the proximity of the wound. The effect of estradiol was significantly inhibited by the administration of either CB1 or CB2 receptor antagonists. The effect of estradiol may be in part mediated by alterations in endocannabinoid signaling because the hormone increased in the injured cerebral cortex the messenger RNA levels of CB2 receptors and of some of the enzymes involved in the synthesis and metabolism of endocannabinoids. These findings suggest that estradiol may decrease reactive astroglia in the injured brain by regulating the activity of the endocannabinoid system.

  10. Anti-infective mannose receptor immune mechanism in large yellow croaker (Larimichthys crocea).

    PubMed

    Dong, Xiangli; Li, Jiji; He, Jianyu; Liu, Wei; Jiang, Lihua; Ye, Yingying; Wu, Changwen

    2016-07-01

    Mannose receptor (MR) is a pattern recognition receptor (PRR) that plays a significant role in immunity responses. Its role has been described extensively in mammals, but very rarely in fish. Recently, with the rapid development of an aquaculture industry cultivating large yellow croaker (Larimichthys crocea), infectious diseases caused by viruses, bacteria and parasites are becoming more frequent and more severe, in particular bacterial infections caused by Vibrio anguillarum, resulting in great economical losses. Extensive use of antibiotics as conventional treatment has led to microenvironment imbalances, development of drug-resistant bacteria and deposition of drug residues, which cause environmental pollution and ultimately affect human health. The purpose of this pilot study was to detect the transcriptional levels of C-type mannose receptor genes MRC1 (4710-bp ORF; encoding 1437 aa; a signal peptide, a SMART RICIN domain, a SMART FN2 domain, eight SMART CLECT domain, and a transmembrane helix region) and MRC2 (3996-bp ORF; encoding 1484 aa; a SMART FN2 domain, eight SMART CLECT domains, and a transmembrane region) in the liver, kidney and spleen tissues of L. crocea challenged by V. anguillarum, to explore the effective domain and the molecular response mechanisms of MRC1 and MRC2, and, ultimately, to explore the possibility of developing a vaccine targeting V. anguillarum infections.

  11. Cardiac Alpha1-Adrenergic Receptors: Novel Aspects of Expression, Signaling Mechanisms, Physiologic Function, and Clinical Importance

    PubMed Central

    O’Connell, Timothy D.; Jensen, Brian C.; Baker, Anthony J.

    2014-01-01

    Adrenergic receptors (AR) are G-protein-coupled receptors (GPCRs) that have a crucial role in cardiac physiology in health and disease. Alpha1-ARs signal through Gαq, and signaling through Gq, for example, by endothelin and angiotensin receptors, is thought to be detrimental to the heart. In contrast, cardiac alpha1-ARs mediate important protective and adaptive functions in the heart, although alpha1-ARs are only a minor fraction of total cardiac ARs. Cardiac alpha1-ARs activate pleiotropic downstream signaling to prevent pathologic remodeling in heart failure. Mechanisms defined in animal and cell models include activation of adaptive hypertrophy, prevention of cardiac myocyte death, augmentation of contractility, and induction of ischemic preconditioning. Surprisingly, at the molecular level, alpha1-ARs localize to and signal at the nucleus in cardiac myocytes, and, unlike most GPCRs, activate “inside-out” signaling to cause cardioprotection. Contrary to past opinion, human cardiac alpha1-AR expression is similar to that in the mouse, where alpha1-AR effects are seen most convincingly in knockout models. Human clinical studies show that alpha1-blockade worsens heart failure in hypertension and does not improve outcomes in heart failure, implying a cardioprotective role for human alpha1-ARs. In summary, these findings identify novel functional and mechanistic aspects of cardiac alpha1-AR function and suggest that activation of cardiac alpha1-AR might be a viable therapeutic strategy in heart failure. PMID:24368739

  12. Pore conformations and gating mechanism of a Cys-loop receptor.

    PubMed

    Paas, Yoav; Gibor, Gilad; Grailhe, Regis; Savatier-Duclert, Nathalie; Dufresne, Virginie; Sunesen, Morten; de Carvalho, Lia Prado; Changeux, Jean-Pierre; Attali, Bernard

    2005-11-01

    Neurons regulate the propagation of chemoelectric signals throughout the nervous system by opening and closing ion channels, a process known as gating. Here, histidine-based metal-binding sites were engineered along the intrinsic pore of a chimeric Cys-loop receptor to probe state-dependent Zn(2+)-channel interactions. Patterns of Zn(2+) ion binding within the pore reveal that, in the closed state, the five pore-lining segments adopt an oblique orientation relative to the axis of ion conduction and constrict into a physical gate at their intracellular end. The interactions of Zn(2+) with the open state indicate that the five pore-lining segments should rigidly tilt to enable the movement of their intracellular ends away from the axis of ion conduction, so as to open the constriction (i.e., the gate). Alignment of the functional results with the 3D structure of an acetylcholine receptor allowed us to generate structural models accounting for the closed and open pore conformations and for a gating mechanism of a Cys-loop receptor.

  13. Mechanics of the trachea and behaviour of its slowly adapting stretch receptors.

    PubMed Central

    Mortola, J P; Sant'Ambrogio, G

    1979-01-01

    1. The trachea is constructed by a series of U-shaped cartilaginous rings supporting a membranous posterior wall. We have studied separately the pressure-volume relationships of the two components. 2. The motion of the membranous posterior wall contributes most to the tracheal volume change caused by any given transmural pressure change; the cartilaginous rings provide a semi-rigid support to the posterior wall and have a far greater compliance with negative than positive transmural pressure. 3. The response of tracheal stretch receptors to transmural pressure can be explained by the mechanical coupling between cartilages and posterior wall. They respond both to positive and negative transmural pressure, they are active at zero transmural pressure and have a point of least activity with small negative transmural pressures. 4. The stress-strain relationship of the posterior wall has been studied in static and dynamic conditions in control situations and after removal of either the tunica fibrosa or the trachealis muscle. Each of these two components contributes to the stiffness of the posterior wall, with the trachealis muscle providing most of its viscosity. 5. The response of tracheal stretch receptors to transverse traction of the posterior membranous wall has been studied in both static and dynamic conditions before and after removal of the tunica fibrosa. The behaviour of these receptors reflects the visco-elastic properties of the trachealis muscle in which they have been localized. PMID:439039

  14. Antiproliferative effects and mechanisms of liver X receptor ligands in pancreatic ductal adenocarcinoma cells.

    PubMed

    Candelaria, Nicholes R; Addanki, Sridevi; Zheng, Jine; Nguyen-Vu, Trang; Karaboga, Husna; Dey, Prasenjit; Gabbi, Chiara; Vedin, Lise-Lotte; Liu, Ka; Wu, Wanfu; Jonsson, Philip K; Lin, Jean Z; Su, Fei; Bollu, Lakshmi Reddy; Hodges, Sally E; McElhany, Amy L; Issazadeh, Mehdi A; Fisher, William E; Ittmann, Michael M; Steffensen, Knut R; Gustafsson, Jan-Åke; Lin, Chin-Yo

    2014-01-01

    Pancreatic ductal adenocarcinoma (PDAC) is difficult to detect early and is often resistant to standard chemotherapeutic options, contributing to extremely poor disease outcomes. Members of the nuclear receptor superfamily carry out essential biological functions such as hormone signaling and are successfully targeted in the treatment of endocrine-related malignancies. Liver X receptors (LXRs) are nuclear receptors that regulate cholesterol homeostasis, lipid metabolism, and inflammation, and LXR agonists have been developed to regulate LXR function in these processes. Intriguingly, these compounds also exhibit antiproliferative activity in diverse types of cancer cells. In this study, LXR agonist treatments disrupted proliferation, cell-cycle progression, and colony-formation of PDAC cells. At the molecular level, treatments downregulated expression of proteins involved in cell cycle progression and growth factor signaling. Microarray experiments further revealed changes in expression profiles of multiple gene networks involved in biological processes and pathways essential for cell growth and proliferation following LXR activation. These results establish the antiproliferative effects of LXR agonists and potential mechanisms of action in PDAC cells and provide evidence for their potential application in the prevention and treatment of PDAC.

  15. Distinct signaling mechanisms in multiple developmental pathways by the SCRAMBLED receptor of Arabidopsis.

    PubMed

    Kwak, Su-Hwan; Woo, Sooah; Lee, Myeong Min; Schiefelbein, John

    2014-10-01

    SCRAMBLED (SCM), a leucine-rich repeat receptor-like kinase in Arabidopsis (Arabidopsis thaliana), is required for positional signaling in the root epidermis and for tissue/organ development in the shoot. To further understand SCM action, we generated a series of kinase domain variants and analyzed their ability to complement scm mutant defects. We found that the SCM kinase domain, but not kinase activity, is required for its role in root epidermal patterning, supporting the view that SCM is an atypical receptor kinase. We also describe a previously uncharacterized role for SCM in fruit dehiscence, because mature siliques from scm mutants fail to open properly. Interestingly, the kinase domain of SCM appears to be dispensable for this developmental process. Furthermore, we found that most of the SCM kinase domain mutations dramatically inhibit inflorescence development. Because this process is not affected in scm null mutants, it is likely that SCM acts redundantly to regulate inflorescence size. The importance of distinct kinase residues for these three developmental processes provides an explanation for the maintenance of the conserved kinase domain in the SCM protein, and it may generally explain its conservation in other atypical kinases. Furthermore, these results indicate that individual leucine-rich repeat receptor-like kinases may participate in multiple pathways using distinct signaling mechanisms to mediate diverse cellular communication events.

  16. Internalization and recycling of 5-HT2A receptors activated by serotonin and protein kinase C-mediated mechanisms

    PubMed Central

    Bhattacharyya, Samarjit; Puri, Sapna; Miledi, Ricardo; Panicker, Mitradas M.

    2002-01-01

    Serotonin (5-HT), a major neurotransmitter, has a large number of G protein-coupled receptors in mammals. On activation by exposure to their ligand, 5-HT2 receptor subtypes increase IP3 levels and undergo desensitization and internalization. To visualize the receptor in cells during these processes, we have constructed a 5-HT2A-enhanced GFP (SR2-GFP) fusion receptor. We show that this fusion receptor undergoes internalization on exposure to its natural ligand, 5-HT. Because 5-HT2A receptors activate the phospholipase C pathway, we studied the effect of protein kinase C (PKC) on the internalization process and found that activation of PKC by its specific activator phorbol 12-myristate 13-acetate, in the absence of 5-HT, leads to internalization of the receptor. Moreover, inhibition of PKC by its inhibitor sphingosine in the presence of 5-HT prevents the internalization process, suggesting that activation of PKC is sufficient and necessary for the internalization of 5-HT2A receptors. We also show that SR2-GFP recycles back to the plasma membrane after 5-HT-dependent internalization, suggesting a mechanism for resensitization. In addition, receptors that have been internalized on addition of phorbol 12-myristate 13-acetate in the absence of 5-HT also recycle to the surface, with a time course similar to that seen after activation of the receptors by 5-HT. Our study suggests that 5-HT2A receptors internalize and return to the surface after both serotonin- and PKC-mediated processes. This study reveals a role for PKC in receptor internalization and also shows that 5-HT2A receptors are recycled. PMID:12388782

  17. Dual single-scission event analysis of constitutive transferrin receptor (TfR) endocytosis and ligand-triggered β2-adrenergic receptor (β2AR) or Mu-opioid receptor (MOR) endocytosis.

    PubMed

    Lampe, Marko; Pierre, Fabienne; Al-Sabah, Suleiman; Krasel, Cornelius; Merrifield, Christien J

    2014-10-01

    The dynamic relationship between constitutive and ligand-triggered clathrin-mediated endocytosis is only poorly characterized, and it remains controversial whether clathrin-coated pits specialize to internalize particular receptor cargo. Here we analyzed the ligand-triggered endocytosis of the model G-protein-coupled receptors (GPCRs) β2-adrenergic receptor (β2AR) and Mu-opioid receptor (MOR) at the level of individual endocytic events using a total internal reflection fluorescence microscopy (TIRFM)-based assay. Similar to the constitutive endocytosis of transferrin receptor (TfR), ligand- triggered endocytosis of β2AR occurs via quantized scission events hosted by clathrin spots and plaques of variable size and persistence. To address whether clathrin-coated structures (CCSs) specialize to internalize particular GPCRs, we adapted the TIRFM imaging assay to simultaneously quantify the internalization of TfR and the ligand- triggered endocytosis of the β2AR or MOR. Agonist-triggered β2AR or MOR endocytosis extended the maturation time of CCSs, as shown previously, but did not affect the rate of constitutive TfR endocytosis or loading of TfR into individual endocytic vesicles. Both the β2AR and the MOR receptors entered cells in the same vesicles as TfR, and the overall evidence for CCS specialization was weak. These data support a simple model in which different cargoes internalize through common CCSs.

  18. Dual single-scission event analysis of constitutive transferrin receptor (TfR) endocytosis and ligand-triggered β2-adrenergic receptor (β2AR) or Mu-opioid receptor (MOR) endocytosis

    PubMed Central

    Lampe, Marko; Pierre, Fabienne; Al-Sabah, Suleiman; Krasel, Cornelius; Merrifield, Christien J.

    2014-01-01

    The dynamic relationship between constitutive and ligand-triggered clathrin-mediated endocytosis is only poorly characterized, and it remains controversial whether clathrin-coated pits specialize to internalize particular receptor cargo. Here we analyzed the ligand-triggered endocytosis of the model G-protein–coupled receptors (GPCRs) β2-adrenergic receptor (β2AR) and Mu-opioid receptor (MOR) at the level of individual endocytic events using a total internal reflection fluorescence microscopy (TIRFM)–based assay. Similar to the constitutive endocytosis of transferrin receptor (TfR), ligand- triggered endocytosis of β2AR occurs via quantized scission events hosted by clathrin spots and plaques of variable size and persistence. To address whether clathrin-coated structures (CCSs) specialize to internalize particular GPCRs, we adapted the TIRFM imaging assay to simultaneously quantify the internalization of TfR and the ligand- triggered endocytosis of the β2AR or MOR. Agonist-triggered β2AR or MOR endocytosis extended the maturation time of CCSs, as shown previously, but did not affect the rate of constitutive TfR endocytosis or loading of TfR into individual endocytic vesicles. Both the β2AR and the MOR receptors entered cells in the same vesicles as TfR, and the overall evidence for CCS specialization was weak. These data support a simple model in which different cargoes internalize through common CCSs. PMID:25079691

  19. ACTH Receptor (MC2R) Specificity: What Do We Know About Underlying Molecular Mechanisms?

    PubMed Central

    Fridmanis, Davids; Roga, Ance; Klovins, Janis

    2017-01-01

    Coincidentally, the release of this Research Topic in Frontiers in Endocrinology takes place 25 years after the discovery of the adrenocorticotropic hormone receptor (ACTHR) by Mountjoy and colleagues. In subsequent years, following the discovery of other types of mammalian melanocortin receptors (MCRs), ACTHR also became known as melanocortin type 2 receptor (MC2R). At present, five types of MCRs have been reported, all of which share significant sequence similarity at the amino acid level, and all of which specifically bind melanocortins (MCs)—a group of biologically active peptides generated by proteolysis of the proopiomelanocortin precursor. All MCs share an identical –H–F–R–W– pharmacophore sequence. α-Melanocyte-stimulating hormone (α-MSH) and adrenocorticotropic hormone (ACTH) are the most extensively studied MCs and are derived from the same region. Essentially, α-MSH is formed from the first 13 amino acid residues of ACTH. ACTHR is unique among MCRs because it binds one sole ligand—ACTH, which makes it a very attractive research object for molecular pharmacologists. However, much research has failed, and functional studies of this receptor are lagging behind other MCRs. The reason for these difficulties has already been outlined by Mountjoy and colleagues in their publication on ACTHR coding sequence discovery where the Cloudman S91 melanoma cell line was used for receptor expression because it was a “more sensitive assay system.” Subsequent work showed that ACTHR could be successfully expressed only in endogenous MCR-expressing cell lines, since in other cell lines it is retained within the endoplasmic reticulum. The resolution of this methodological problem came in 2005 with the discovery of melanocortin receptor accessory protein, which is required for the formation of functionally active ACTHR. The decade that followed this discovery was filled with exciting research that provided insight into the molecular mechanisms underlying

  20. Dual modulation of both lipid oxidation and synthesis by peroxisome proliferator-activated receptor-gamma coactivator-1alpha and -1beta in cultured myotubes.

    PubMed

    Espinoza, Daniel O; Boros, Laszlo G; Crunkhorn, Sarah; Gami, Hiral; Patti, Mary-Elizabeth

    2010-04-01

    The peroxisome proliferator-activated receptor gamma coactivator-1 (PGC-1) family is a key regulator of mitochondrial function, and reduced mRNA expression may contribute to muscle lipid accumulation in obesity and type 2 diabetes. To characterize the effects of PGC-1 on lipid metabolism, we overexpressed PGC-1alpha and PGC-1beta in C2C12 myotubes using adenoviral vectors. Both PGC-1alpha and -1beta increased palmitate oxidation [31% (P<0.01) and 26% (P<0.05), respectively] despite reductions in cellular uptake [by 6% (P<0.05) and 21% (P<0.001)]. Moreover, PGC-1alpha and -1beta increased mRNA expression of genes regulating both lipid oxidation (e.g., CPT1b and ACADL/M) and synthesis (FAS, CS, ACC1/2, and DGAT1). To determine the net effect, we assessed lipid composition in PGC-1-expressing cells. Total lipid content decreased by 42% in palmitate-loaded serum-starved cells overexpressing PGC-1alpha (P<0.05). In contrast, in serum-replete cells, total lipid content was not significantly altered, but fatty acids C14:0, C16:0, C18:0, and C18:1 were increased 2- to 4-fold for PGC-1alpha/beta (P<0.05). Stable isotope-based dynamic metabolic profiling in serum-replete cells labeled with (13)C substrates revealed both increased de novo fatty acid synthesis from glucose and increased fatty acid synthesis by chain elongation with either PGC-1alpha or -1beta expression. These results indicate that PGC-1 can promote both lipid oxidation and synthesis, with net balance determined by the nutrient/hormonal environment.-Espinoza, D. O., Boros, L. G., Crunkhorn, S., Gami, H., Patti, M.-E. Dual Modulation of both lipid oxidation and synthesis by peroxisome proliferator-activated receptor-gamma coactivator-1alpha and -1beta in cultured myotubes.

  1. Molecular mechanism(s) of endocrine-disrupting chemicals and their potent oestrogenicity in diverse cells and tissues that express oestrogen receptors.

    PubMed

    Lee, Hye-Rim; Jeung, Eui-Bae; Cho, Myung-Haing; Kim, Tae-Hee; Leung, Peter C K; Choi, Kyung-Chul

    2013-01-01

    Endocrine-disrupting chemicals (EDCs) are natural or synthetic compounds present in the environment which can interfere with hormone synthesis and normal physiological functions of male and female reproductive organs. Most EDCs tend to bind to steroid hormone receptors including the oestrogen receptor (ER), progesterone receptor (PR) and androgen receptor (AR). As EDCs disrupt the actions of endogenous hormones, they may induce abnormal reproduction, stimulation of cancer growth, dysfunction of neuronal and immune system. Although EDCs represent a significant public health concern, there are no standard methods to determine effect of EDCs on human beings. The mechanisms underlying adverse actions of EDC exposure are not clearly understood. In this review, we highlighted the toxicology of EDCs and its effect on human health, including reproductive development in males and females as shown in in vitro and in vivo models. In addition, this review brings attention to the toxicity of EDCs via interaction of genomic and non-genomic signalling pathways through hormone receptors.

  2. Subunit-specific mechanisms and proton sensitivity of NMDA receptor channel block.

    PubMed

    Dravid, Shashank M; Erreger, Kevin; Yuan, Hongjie; Nicholson, Katherine; Le, Phuong; Lyuboslavsky, Polina; Almonte, Antoine; Murray, Ernest; Mosely, Cara; Barber, Jeremy; French, Adam; Balster, Robert; Murray, Thomas F; Traynelis, Stephen F

    2007-05-15

    We have compared the potencies of structurally distinct channel blockers at recombinant NR1/NR2A, NR1/NR2B, NR1/NR2C and NR1/NR2D receptors. The IC50 values varied with stereochemistry and subunit composition, suggesting that it may be possible to design subunit-selective channel blockers. For dizocilpine (MK-801), the differential potency of MK-801 stereoisomers determined at recombinant NMDA receptors was confirmed at native receptors in vitro and in vivo. Since the proton sensor is tightly linked both structurally and functionally to channel gating, we examined whether blocking molecules that interact in the channel pore with the gating machinery can differentially sense protonation of the receptor. Blockers capable of remaining trapped in the pore during agonist unbinding showed the strongest dependence on extracellular pH, appearing more potent at acidic pH values that promote channel closure. Determination of pK(a) values for channel blockers suggests that the ionization of ketamine but not of other blockers can influence its pH-dependent potency. Kinetic modelling and single channel studies suggest that the pH-dependent block of NR1/NR2A by (-)MK-801 but not (+)MK-801 reflects an increase in the MK-801 association rate even though protons reduce channel open probability and thus MK-801 access to its binding site. Allosteric modulators that alter pH sensitivity alter the potency of MK-801, supporting the interpretation that the pH sensitivity of MK-801 binding reflects the changes at the proton sensor rather than a secondary effect of pH. These data suggest a tight coupling between the proton sensor and the ion channel gate as well as unique subunit-specific mechanisms of channel block.

  3. A Common Molecular Motif Characterizes Extracellular Allosteric Enhancers of GPCR Aminergic Receptors and Suggests Enhancer Mechanism of Action

    PubMed Central

    Bernstein, Robert Root; Dillon, Patrick F

    2014-01-01

    Several classes of compounds that have no intrinsic activity on aminergic systems nonetheless enhance the potency of aminergic receptor ligands three-fold or more while significantly increasing their duration of activity, preventing tachyphylaxis and reversing fade. Enhancer compounds include ascorbic acid, ethylenediaminetetraacetic acid, cortico-steroids, opioid peptides, opiates and opiate antagonists. This paper provides the first review of aminergic enhancement, demonstrating that all enhancers have a common, inobvious molecular motif and work through a common mechanism that is manifested by three common characteristics. First, aminergic enhancers bind directly to the amines they enhance, suggesting that the common structural motif is reflected in common binding targets. Second, one common target is the first extracellular loop of aminergic receptors. Third, at least some enhancers are antiphosphodiesterases. These observations suggest that aminergic enhancers act on the extracellular surface of aminergic receptors to keep the receptor in its high affinity state, trapping the ligand inside the receptor. Enhancer binding produces allosteric modifications of the receptor structure that interfere with phosphorylation of the receptor, thereby inhibiting down-regulation of the receptor. The mechanism explains how enhancers potentiate aminergic activity and increase duration of activity and makes testable predictions about additional compounds that should act as aminergic enhancers. PMID:25174918

  4. Pregnane X receptor activation and silencing promote steatosis of human hepatic cells by distinct lipogenic mechanisms.

    PubMed

    Bitter, Andreas; Rümmele, Petra; Klein, Kathrin; Kandel, Benjamin A; Rieger, Jessica K; Nüssler, Andreas K; Zanger, Ulrich M; Trauner, Michael; Schwab, Matthias; Burk, Oliver

    2015-11-01

    In addition to its well-characterized role in the regulation of drug metabolism and transport by xenobiotics, pregnane X receptor (PXR) critically impacts on lipid homeostasis. In mice, both ligand-dependent activation and knockout of PXR were previously shown to promote hepatic steatosis. To elucidate the respective pathways in human liver, we generated clones of human hepatoma HepG2 cells exhibiting different PXR protein levels, and analyzed effects of PXR activation and knockdown on steatosis and expression of lipogenic genes. Ligand-dependent activation as well as knockdown of PXR resulted in increased steatosis in HepG2 cells. Activation of PXR induced the sterol regulatory element-binding protein (SREBP) 1-dependent lipogenic pathway via PXR-dependent induction of SREBP1a, which was confirmed in primary human hepatocytes. Inhibiting SREBP1 activity by blocking the cleavage-dependent maturation of SREBP1 protein impaired the induction of lipogenic SREBP1 target genes and triglyceride accumulation by PXR activation. On the other hand, PXR knockdown resulted in up-regulation of aldo-keto reductase (AKR) 1B10, which enhanced the acetyl-CoA carboxylase (ACC)-catalyzed reaction step of de novo lipogenesis. In a cohort of human liver samples histologically classified for non-alcoholic fatty liver disease, AKR1B10, SREBP1a and SREBP1 lipogenic target genes proved to be up-regulated in steatohepatitis, while PXR protein was reduced. In summary, our data suggest that activation and knockdown of PXR in human hepatic cells promote de novo lipogenesis and steatosis by induction of the SREBP1 pathway and AKR1B10-mediated increase of ACC activity, respectively, thus providing mechanistic explanations for a putative dual role of PXR in the pathogenesis of steatohepatitis.

  5. Emerging Mechanisms of Resistance to Androgen Receptor Inhibitors in Prostate Cancer

    PubMed Central

    Arora, Vivek K.

    2016-01-01

    Preface Over the past ten years, preclinical studies implicating sustained androgen receptor (AR) signaling as the primary driver of castration resistant prostate cancer (CRPC) led to the development of novel agents targeting the AR pathway that are now in widespread clinical use. These drugs prolong survival of patients with late stage prostate cancer but are not curative. In this review, we highlight emerging mechanisms of acquired resistance to these contemporary therapies, which fall into the three broad categories of restored AR signaling, AR bypass signaling and complete AR independence. This diverse spectrum of resistance mechanisms presents new challenges for long term disease control, which may be addressable through early use of combination therapies guided by recent insights from genomic landscape studies of CRPC. PMID:26563462

  6. Structure, signaling mechanism and regulation of the natriuretic peptide receptor guanylate cyclase.

    SciTech Connect

    Misono, K. S.; Philo, J. S.; Arakawa, T.; Ogata, C. M.; Qiu, Y.; Ogawa, H.; Young, H. S.

    2011-06-01

    Atrial natriuretic peptide (ANP) and the homologous B-type natriuretic peptide are cardiac hormones that dilate blood vessels and stimulate natriuresis and diuresis, thereby lowering blood pressure and blood volume. ANP and B-type natriuretic peptide counterbalance the actions of the renin-angiotensin-aldosterone and neurohormonal systems, and play a central role in cardiovascular regulation. These activities are mediated by natriuretic peptide receptor-A (NPRA), a single transmembrane segment, guanylyl cyclase (GC)-linked receptor that occurs as a homodimer. Here, we present an overview of the structure, possible chloride-mediated regulation and signaling mechanism of NPRA and other receptor GCs. Earlier, we determined the crystal structures of the NPRA extracellular domain with and without bound ANP. Their structural comparison has revealed a novel ANP-induced rotation mechanism occurring in the juxtamembrane region that apparently triggers transmembrane signal transduction. More recently, the crystal structures of the dimerized catalytic domain of green algae GC Cyg12 and that of cyanobacterium GC Cya2 have been reported. These structures closely resemble that of the adenylyl cyclase catalytic domain, consisting of a C1 and C2 subdomain heterodimer. Adenylyl cyclase is activated by binding of G{sub s}{alpha} to C2 and the ensuing 7{sup o} rotation of C1 around an axis parallel to the central cleft, thereby inducing the heterodimer to adopt a catalytically active conformation. We speculate that, in NPRA, the ANP-induced rotation of the juxtamembrane domains, transmitted across the transmembrane helices, may induce a similar rotation in each of the dimerized GC catalytic domains, leading to the stimulation of the GC catalytic activity.

  7. Cannabidiol potentiates pharmacological effects of Delta(9)-tetrahydrocannabinol via CB(1) receptor-dependent mechanism.

    PubMed

    Hayakawa, Kazuhide; Mishima, Kenichi; Hazekawa, Mai; Sano, Kazunori; Irie, Keiichi; Orito, Kensuke; Egawa, Takashi; Kitamura, Yoshihisa; Uchida, Naoki; Nishimura, Ryoji; Egashira, Nobuaki; Iwasaki, Katsunori; Fujiwara, Michihiro

    2008-01-10

    Cannabidiol, a non-psychoactive component of cannabis, has been reported to have interactions with Delta(9)-tetrahydrocannabinol (Delta(9)-THC). However, such interactions have not sufficiently been clear and may have important implications for understanding the pharmacological effects of marijuana. In the present study, we investigated whether cannabidiol modulates the pharmacological effects of Delta(9)-THC on locomotor activity, catalepsy-like immobilisation, rectal temperature and spatial memory in the eight-arm radial maze task in mice. In addition, we measured expression level of cannabinoid CB(1) receptor at striatum, cortex, hippocampus and hypothalamus. Delta(9)-THC (1, 3, 6 and 10 mg/kg) induced hypoactivity, catalepsy-like immobilisation and hypothermia in a dose-dependent manner. In addition, Delta(9)-THC (1, 3 and 6 mg/kg) dose-dependently impaired spatial memory in eight-arm radial maze. On the other hand, cannabidiol (1, 3, 10, 25 and 50 mg/kg) did not affect locomotor activity, catalepsy-like immobilisation, rectal temperature and spatial memory on its own. However, higher dose of cannabidiol (10 or 50 mg/kg) exacerbated pharmacological effects of lower dose of Delta(9)-THC, such as hypoactivity, hypothermia and impairment of spatial memory. Moreover, cannabidiol (50 mg/kg) with Delta(9)-THC (1 mg/kg) enhanced the expression level of CB(1) receptor expression in hippocampus and hypothalamus. Cannabidiol potentiated pharmacological effects of Delta(9)-THC via CB(1) receptor-dependent mechanism. These findings may contribute in setting the basis for interaction of cannabinoids and to find a cannabinoid mechanism in central nervous system.

  8. Studies on the mechanisms of action of picrotoxin, quercetin and pregnanolone at the GABAρ1 receptor

    PubMed Central

    Goutman, Juan D; Calvo, Daniel J

    2004-01-01

    The mechanisms of action of antagonists of the γ-aminobutyric acid C (GABAC) receptor picrotoxin, quercetin and pregnanolone were studied. Ionic currents (chloride), mediated through human homomeric GABAρ1 receptors expressed in Xenopus oocytes, were recorded by two-electrode voltage clamp. Dose–response (D–R) curves and kinetic measurements of GABAρ1 currents were carried out in the presence or absence of antagonists. Use-dependent actions were also evaluated. Picrotoxin, quercetin and pregnanolone exerted noncompetitive actions. IC50 values measured at the EC50 for GABA (1 μM) were as follows: picrotoxin 0.6±0.1 μM (Hill coefficient n=1.0±0.2); quercetin 4.4±0.4 μM (n=1.5±0.2); pregnanolone 2.1±0.5 μM (n=0.8±0.1). These antagonists produced changes only in the slope of the linear current–voltage relationships, which was indicative of voltage-independent effects. The effect of picrotoxin on GABAρ1 currents was use-dependent, strongly relied on agonist concentration and showed a slow onset and offset. The mechanism was compatible with an allosteric inhibition and receptor activation was a prerequisite for antagonism. The effect of quercetin was use-independent, showed relatively fast onset and offset, and resulted in a slowed time course of the GABA-evoked currents. The effect of pregnanolone was use-independent, presented fast onset and a very slow washout, and did not affect current activation. All the antagonists accelerated the time course of deactivation of the GABAρ1 currents. PMID:14732759

  9. The Nicotinic Receptor Alpha7 Impacts the Mouse Lung Response to LPS through Multiple Mechanisms

    PubMed Central

    Enioutina, Elena Y.; Myers, Elizabeth J.; Tvrdik, Petr; Hoidal, John R.; Rogers, Scott W.; Gahring, Lorise C.

    2015-01-01

    The nicotinic acetylcholine receptor alpha7 (α7) is expressed by neuronal and non-neuronal cells throughout the body. We examined the mechanisms of the lung inflammatory response to intranasal (i.n.) lipopolysaccharide (LPS) regulated by α7. This was done in mice using homologous recombination to introduce a point mutation in the α7 receptor that replaces the glutamate residue 260 that lines the pore with alanine (α7E260A), which has been implicated in controlling the exceptional calcium ion conductance of this receptor. The α7E260A mice exhibit normal inflammatory cell recruitment to the blood in response to i.n. LPS administration. This differs from the α7knock-out (α7KO) in which upstream signaling to initiate the recruitment to the blood following i.n. LPS is significantly impaired. While hematopoietic cells are recruited to the bloodstream in the α7E260A mouse, they fail to be recruited efficiently into both the interstitium and alveolar spaces of the lung. Bone marrow reconstitution experiments demonstrate that the responsiveness of both CD45+ and CD45- cells of the α7E260A mouse are impaired. The expression of several pro-inflammatory cytokine and chemokine RNAs including TNFα, IL-1α, Ccl2 and Cxcl10 are decreased in the α7E260A mouse. However, there is a substantial increase in IL-13 expression by CD45- lung interstitial cells in the α7E260A mouse. Our results support the conclusion that α7 functional pleiotropy contributes to modulating the tissue response to an inflammatory insult through impacting upon a variety of mechanisms reflecting the individual cell composition of the lung. PMID:25803612

  10. Nicotine Effects and Receptor Expression on Human Spermatozoa: Possible Neuroendocrine Mechanism

    PubMed Central

    Condorelli, Rosita A.; La Vignera, Sandro; Giacone, Filippo; Iacoviello, Linda; Mongioì, Laura M.; Li Volti, Giovanni; Barbagallo, Ignazio; Avola, Roberto; Calogero, Aldo E.

    2017-01-01

    The aim of this experimental study was to investigate the mechanism by which nicotine (NIC) alters spermatozoa and to evaluate the expression of nicotinic receptors (nAChR) subunits in human spermatozoa. We analyzed 30 healthy normozoospermic men. Spermatozoa were incubated with NIC 100 ng/ml and the nAChR antagonist, hexamethonium (HEX) (0, 100, 1,000, 10,000 ng/ml) for 3 and 24 h. The following sperm parameters evaluated: (a) progressive motility; (b) mitochondrial membrane potential (MMP); (c) chromatin compactness; (d) externalization of phosphatidylserine (PS); (e) late apoptosis; (f) viability; (g) DNA fragmentation; (h) degree of lipid peroxidation (LP) by flow cytometry; (i) nAChR subunits expression by quantitative Real Time PCR and (j) protein expression evaluation by Western blot analysis. HEX fully antagonized the effects of NIC both after 3 and 24 h of incubation with significant improvement (p < 0.05) of sperm progressive motility, MMP, abnormal chromatin compactness, PS externalization, late apoptosis and DNA fragmentation, already at the concentration of HEX 100 ng/ml. The degree of LP increased after incubation with NIC in raw semen but this effect was fully antagonized (p < 0.05) by HEX after 3 and 24 h of incubation. Finally, 8 nAChR subunits mRNA (α1, α3, α4, α6, α7, β2, β4, and δ) were found expressed in all samples examined, but only α7 subunit is translated, making an homomer receptor, in non-smokers subjects. The effects of NIC on sperm function are mediated by interaction with a specific nicotinic receptor. The presence of nAChR subunits suggests the presence of a neuroendocrine mechanism on human spermatozoa.

  11. An electrostatic selection mechanism controls sequential kinase signaling downstream of the T cell receptor

    PubMed Central

    Shah, Neel H; Wang, Qi; Yan, Qingrong; Karandur, Deepti; Kadlecek, Theresa A; Fallahee, Ian R; Russ, William P; Ranganathan, Rama; Weiss, Arthur; Kuriyan, John

    2016-01-01

    The sequence of events that initiates T cell signaling is dictated by the specificities and order of activation of the tyrosine kinases that signal downstream of the T cell receptor. Using a platform that combines exhaustive point-mutagenesis of peptide substrates, bacterial surface-display, cell sorting, and deep sequencing, we have defined the specificities of the first two kinases in this pathway, Lck and ZAP-70, for the T cell receptor ζ chain and the scaffold proteins LAT and SLP-76. We find that ZAP-70 selects its substrates by utilizing an electrostatic mechanism that excludes substrates with positively-charged residues and favors LAT and SLP-76 phosphosites that are surrounded by negatively-charged residues. This mechanism prevents ZAP-70 from phosphorylating its own activation loop, thereby enforcing its strict dependence on Lck for activation. The sequence features in ZAP-70, LAT, and SLP-76 that underlie electrostatic selectivity likely contribute to the specific response of T cells to foreign antigens. DOI: http://dx.doi.org/10.7554/eLife.20105.001 PMID:27700984

  12. Receptor Pre-Clustering and T cell Responses: Insights into Molecular Mechanisms.

    PubMed

    Castro, Mario; van Santen, Hisse M; Férez, María; Alarcón, Balbino; Lythe, Grant; Molina-París, Carmen

    2014-01-01

    T cell activation, initiated by T cell receptor (TCR) mediated recognition of pathogen-derived peptides presented by major histocompatibility complex class I or II molecules (pMHC), shows exquisite specificity and sensitivity, even though the TCR-pMHC binding interaction is of low affinity. Recent experimental work suggests that TCR pre-clustering may be a mechanism via which T cells can achieve such high sensitivity. The unresolved stoichiometry of the TCR makes TCR-pMHC binding and TCR triggering, an open question. We formulate a mathematical model to characterize the pre-clustering of T cell receptors (TCRs) on the surface of T cells, motivated by the experimentally observed distribution of TCR clusters on the surface of naive and memory T cells. We extend a recently introduced stochastic criterion to compute the timescales of T cell responses, assuming that ligand-induced cross-linked TCR is the minimum signaling unit. We derive an approximate formula for the mean time to signal initiation. Our results show that pre-clustering reduces the mean activation time. However, additional mechanisms favoring the existence of clusters are required to explain the difference between naive and memory T cell responses. We discuss the biological implications of our results, and both the compatibility and complementarity of our approach with other existing mathematical models.

  13. Receptor Pre-Clustering and T cell Responses: Insights into Molecular Mechanisms

    PubMed Central

    Castro, Mario; van Santen, Hisse M.; Férez, María; Alarcón, Balbino; Lythe, Grant; Molina-París, Carmen

    2014-01-01

    T cell activation, initiated by T cell receptor (TCR) mediated recognition of pathogen-derived peptides presented by major histocompatibility complex class I or II molecules (pMHC), shows exquisite specificity and sensitivity, even though the TCR–pMHC binding interaction is of low affinity. Recent experimental work suggests that TCR pre-clustering may be a mechanism via which T cells can achieve such high sensitivity. The unresolved stoichiometry of the TCR makes TCR–pMHC binding and TCR triggering, an open question. We formulate a mathematical model to characterize the pre-clustering of T cell receptors (TCRs) on the surface of T cells, motivated by the experimentally observed distribution of TCR clusters on the surface of naive and memory T cells. We extend a recently introduced stochastic criterion to compute the timescales of T cell responses, assuming that ligand-induced cross-linked TCR is the minimum signaling unit. We derive an approximate formula for the mean time to signal initiation. Our results show that pre-clustering reduces the mean activation time. However, additional mechanisms favoring the existence of clusters are required to explain the difference between naive and memory T cell responses. We discuss the biological implications of our results, and both the compatibility and complementarity of our approach with other existing mathematical models. PMID:24817867

  14. Molecular mechanisms deployed by virally encoded G protein-coupled receptors in human diseases.

    PubMed

    Montaner, Silvia; Kufareva, Irina; Abagyan, Ruben; Gutkind, J Silvio

    2013-01-01

    G protein-coupled receptors (GPCRs) represent the largest family of cell surface molecules involved in signal transduction. Surprisingly, open reading frames for multiple GPCRs were hijacked in the process of coevolution between Herpesviridae family viruses and their human and mammalian hosts. Virally encoded GPCRs (vGPCRs) evolved as parts of viral genomes, and this evolution allowed the power of host GPCR signaling circuitries to be harnessed in order to ensure viral replicative success. Phylogenetically, vGPCRs are distantly related to human chemokine receptors, although they feature several unique characteristics. Here, we describe the molecular mechanisms underlying vGPCR-mediated viral pathogenesis. These mechanisms include constitutive activity, aberrant coupling to human G proteins and β-arrestins, binding and activation by human chemokines, and dimerization with other GPCRs expressed in infected cells. The likely structural basis for these molecular events is described for the two closest viral homologs of human GPCRs. This information may aid in the development of novel targeted therapeutic strategies against viral diseases.

  15. Fc receptor-dependent mechanisms of monoclonal antibody therapy of cancer.

    PubMed

    Bakema, Jantine E; van Egmond, Marjolein

    2014-01-01

    Targeted therapies like treatment with monoclonal antibodies (mAbs) have entered the arsenal of modern anticancer drugs. mAbs combine specificity with multiple effector functions that can lead to reduction of tumour burden. Direct mechanisms of action, including induction of apoptosis or growth inhibition, depend on the biology of the target antigen. Fc tails of mAbs have furthermore the potential to initiate complement-dependent lysis as well as immune effector cell-mediated tumour cell killing via binding to Fc receptors. Natural killer cells can induce apoptosis via antibody-dependent cellular cytotoxicity (ADCC), whereas macrophages are able to phagocytose mAb-opsonized tumour cells (antibody-dependent cellular phagocytosis; ADCP). Finally, neutrophils can induce non-apoptotic tumour cell death, especially in the presence of immunoglobulin A (IgA) antitumour mAbs. In spite of promising clinical successes in some malignancies, improvement of mAb immunotherapy is required to achieve overall complete remission in cancer patients. New strategies to enhance Fc receptor-mediated mechanisms of action or to overcome the immunosuppressive microenvironment of the tumour in mAb therapy of cancer are therefore currently being explored and will be addressed in this chapter.

  16. Chronic methamphetamine treatment induces oxytocin receptor up-regulation in the amygdala and hypothalamus via an adenosine A2A receptor-independent mechanism.

    PubMed

    Zanos, Panos; Wright, Sherie R; Georgiou, Polymnia; Yoo, Ji Hoon; Ledent, Catherine; Hourani, Susanna M; Kitchen, Ian; Winsky-Sommerer, Raphaelle; Bailey, Alexis

    2014-04-01

    There is mounting evidence that the neuropeptide oxytocin is a possible candidate for the treatment of drug addiction. Oxytocin was shown to reduce methamphetamine self-administration, conditioned place-preference, hyperactivity and reinstatement in rodents, highlighting its potential for the management of methamphetamine addiction. Thus, we hypothesised that the central endogenous oxytocinergic system is dysregulated following chronic methamphetamine administration. We tested this hypothesis by examining the effect of chronic methamphetamine administration on oxytocin receptor density in mice brains with the use of quantitative receptor autoradiographic binding. Saline (4ml/kg/day, i.p.) or methamphetamine (1mg/kg/day, i.p.) was administered daily for 10 days to male, CD1 mice. Quantitative autoradiographic mapping of oxytocin receptors was carried out with the use of [(125)I]-vasotocin in brain sections of these animals. Chronic methamphetamine administration induced a region specific upregulation of oxytocin receptor density in the amygdala and hypothalamus, but not in the nucleus accumbens and caudate putamen. As there is evidence suggesting an involvement of central adenosine A2A receptors on central endogenous oxytocinergic function, we investigated whether these methamphetamine-induced oxytocinergic neuroadaptations are mediated via an A2A receptor-dependent mechanism. To test this hypothesis, autoradiographic oxytocin receptor binding was carried out in brain sections of male CD1 mice lacking A2A receptors which were chronically treated with methamphetamine (1mg/kg/day, i.p. for 10 days) or saline. Similar to wild-type animals, chronic methamphetamine administration induced a region-specific upregulation of oxytocin receptor binding in the amygdala and hypothalamus of A2A receptor knockout mice and no genotype effect was observed. These results indicate that chronic methamphetamine use can induce profound neuroadaptations of the oxytocinergic receptor

  17. Role of selective alpha and beta adrenergic receptor mechanisms in rat jejunal longitudinal muscle contractility.

    PubMed

    Seiler, Roland; Rickenbacher, Andreas; Shaw, Sidney; Haefliger, Simon; Balsiger, Bruno M

    2008-06-01

    Gut motility is modulated by adrenergic mechanisms. The aim of our study was to examine mechanisms of selective adrenergic receptors in rat jejunum. Spontaneous contractile activity of longitudinal muscle strips from rat jejunum was measured in 5-ml tissue chambers. Dose-responses (six doses, 10(-7) -3 x 10(-5)M) to norepinephrine (NE, nonspecific), phenylephrine (PH, alpha1), clonidine (C, alpha2), prenalterol (PR, beta1), ritodrine (RI, beta2), and ZD7714 (ZD, beta3) were evaluated with and without tetrodotoxin (TTX, nerve blocker). NE(3 x 10(-5)M) inhibited 74 +/- 5% (mean +/- SEM) of spontaneous activity. This was the maximum effect. The same dose of RI(beta2), PH(alpha1), or ZD(beta(3)) resulted in an inhibition of only 56 +/- 5, 43 +/- 4, 33 +/- 6, respectively. The calculated concentration to induce 50% inhibition (EC50) of ZD(beta3) was similar to NE, whereas higher concentrations of PH(alpha1) or RI(beta2) were required. C(alpha2) and PR(beta1) had no effect. TTX changed exclusively the EC50 of RI from 4.4 +/- 0.2 to 2.7 +/- 0.8% (p < 0.04). Contractility was inhibited by NE (nonspecific). PH(alpha1), RI(beta2), and ZD(beta3) mimic the effect of NE. TTX reduced the inhibition by RI. Our results suggest that muscular alpha1, beta2, and beta3 receptor mechanisms mediate adrenergic inhibition of contractility in rat jejunum. beta2 mechanisms seem to involve also neural pathways.

  18. Statistical and theoretical analysis of precipitates in dual-phase steels microalloyed with titanium and their effect on mechanical properties

    SciTech Connect

    Soto, R.; Saikaly, W.; Bano, X.; Issartel, C.; Rigaut, G.; Charai, A.

    1999-09-29

    Transmission electron microscopy was used to study the precipitation in four different dual-phase steels microalloyed with titanium. TiN, Ti{sub 4}C{sub 2}S{sub 2} and TiC were characterized. Detailed statistical analysis of hundreds of images of the various Ti compounds fit rather well with thermodynamical calculations and they show TiC particles (approximately 10 nm) to be the most dominant factor in strengthening.

  19. Polycythaemia-inducing mutations in the erythropoietin receptor (EPOR): mechanism and function as elucidated by epidermal growth factor receptor-EPOR chimeras.

    PubMed

    Gross, Mor; Ben-Califa, Nathalie; McMullin, Mary F; Percy, Melanie J; Bento, Celeste; Cario, Holger; Minkov, Milen; Neumann, Drorit

    2014-05-01

    Primary familial and congenital polycythaemia (PFCP) is a disease characterized by increased red blood cell mass, and can be associated with mutations in the intracellular region of the erythropoietin (EPO) receptor (EPOR). Here we explore the mechanisms by which EPOR mutations induce PFCP, using an experimental system based on chimeric receptors between epidermal growth factor receptor (EGFR) and EPOR. The design of the chimeras enabled EPOR signalling to be triggered by EGF binding. Using this system we analysed three novel EPOR mutations discovered in PFCP patients: a deletion mutation (Del1377-1411), a nonsense mutation (C1370A) and a missense mutation (G1445A). Three different chimeras, bearing these mutations in the cytosolic, EPOR region were generated; Hence, the differences in the chimera-related effects are specifically attributed to the mutations. The results show that the different mutations affect various aspects related to the signalling and metabolism of the chimeric receptors. These include slower degradation rate, higher levels of glycan-mature chimeric receptors, increased sensitivity to low levels of EGF (replacing EPO in this system) and extended signalling cascades. This study provides a novel experimental system to study polycythaemia-inducing mutations in the EPOR, and sheds new light on underlying mechanisms of EPOR over-activation in PFCP patients.

  20. The human fibroblast growth factor receptor genes: a common structural arrangement underlies the mechanisms for generating receptor forms that differ in their third immunoglobulin domain.

    PubMed Central

    Johnson, D E; Lu, J; Chen, H; Werner, S; Williams, L T

    1991-01-01

    To determine the mechanisms by which multiple forms of fibroblast growth factor (FGF) receptors are generated, we have mapped the arrangement of exons and introns in the human FGF receptor 1 (FGFR 1) gene (flg). We found three alternative exons encoding a portion of the third immunoglobulin (Ig)-like domain of the receptor. One of these alternatives encodes a sequence that is part of a secreted form of FGFR 1. The other two encode sequences that are likely part of transmembrane forms of FGFR 1. One of these forms has not been previously reported in published cDNAs. Also, we have determined the structural organization of a portion of the human FGFR 2 gene (bek) and found a similar arrangement of alternative exons for the third Ig-like domain. The arrangement of these genes suggests that there are conserved mechanisms governing the expression of secreted FGF receptors as well as the expression of at least two distinct membrane-spanning forms of the FGF receptors. The diverse forms appear to be generated by alternative splicing of mRNA and selective use of polyadenylation signals. Images PMID:1652059

  1. A model for electromigration-induced degradation mechanisms in dual-inlaid copper interconnects: Effect of interface bonding strength

    NASA Astrophysics Data System (ADS)

    Sukharev, Valeriy; Zschech, Ehrenfried

    2004-12-01

    A physical model and a simulation algorithm are used to predict an electromigration-(EM-) induced void nucleation and growth in dual-inlaid copper interconnect. Incorporation of all important atom migration driving forces into the mass balance equation and its solution together with solution of the coupled electromagnetics, heat transfer, and elasticity problems allows to simulate EM-induced degradation in a variety of dual-inlaid copper interconnect segments characterized by different dominant channels for mass transport. The interface bonding strengths, significantly influencing the interface diffusivity and consequently the mass transport along interfaces, result in completely different degradation and failure pictures for the weak and strengthened copper/capping layer interfaces. Strengthening of the top interface of inlaid copper interconnect metal line is a promising way to prolong the EM lifetime. The results of the numerical simulation have been proven experimentally by the EM degradation studies on the fully embedded dual-inlaid copper interconnect test structures. EM-induced void formation, movement, and growth in a copper interconnect were continuously monitored in an in situ scanning electron microscopy experiment. The correspondence between simulation results and experimental data indicates the applicability of the developed model for optimization of the physical and electrical design rules. Simulation-based optimization of the interconnect architecture, segment geometry, material properties, and some of the process parameters can generate on-chip interconnect systems with a high immunity to EM-induced failures.

  2. Activation of GABA(A) receptors in subthalamic neurons in vitro: properties of native receptors and inhibition mechanisms.

    PubMed

    Baufreton, J; Garret, M; Dovero, S; Dufy, B; Bioulac, B; Taupignon, A

    2001-07-01

    The subthalamic nucleus (STN) influences the output of the basal ganglia, thereby interfering with motor behavior. The main inputs to the STN are GABAergic. We characterized the GABA(A) receptors expressed in the STN and investigated the response of subthalamic neurons to the activation of GABA(A) receptors. Cell-attached and whole cell recordings were made from rat brain slices using the patch-clamp technique. The newly identified epsilon subunit confers atypical pharmacological properties on recombinant receptors, which are insensitive to barbiturates and benzodiazepines. We tested the hypothesis that native subthalamic GABA(A) receptors contain epsilon proteins. Applications of increasing concentrations of muscimol, a selective GABA(A) agonist, induced Cl(-) and HCO currents with an EC(50) of 5 microM. Currents induced by muscimol were fully blocked by the GABA(A) receptor antagonists, bicuculline and picrotoxin. They were strongly potentiated by the barbiturate, pentobarbital (+190%), and by the benzodiazepines, diazepam (+197%) and flunitrazepam (+199%). Spontaneous inhibitory postsynaptic currents were also significantly enhanced by flunitrazepam. Furthermore, immunohistological experiments with an epsilon subunit-specific antibody showed that the epsilon protein was not expressed within the STN. Native subthalamic GABA(A) receptors did not, therefore, display pharmacological or structural properties consistent with receptors comprising epsilon. Burst firing is a hallmark of Parkinson's disease. Half of the subthalamic neurons have the intrinsic capacity of switching from regular-firing to burst-firing mode when hyperpolarized by current injection. This raises the possibility that activation of GABA(A) receptors might trigger the switch. Statistical analysis of spiking activity established that 90% of intact neurons in vitro were in single-spike firing mode, whereas 10% were in burst-firing mode. Muscimol reversibly stopped recurrent electrical activity in

  3. Mechanical vibration inhibits osteoclast formation by reducing DC-STAMP receptor expression in osteoclast precursor cells.

    PubMed

    Kulkarni, Rishikesh N; Voglewede, Philip A; Liu, Dawei

    2013-12-01

    It is well known that physical inactivity leads to loss of muscle mass, but it also causes bone loss. Mechanistically, osteoclastogenesis and bone resorption have recently been shown to be regulated by vibration. However, the underlying mechanism behind the inhibition of osteoclast formation is yet unknown. Therefore, we investigated whether mechanical vibration of osteoclast precursor cells affects osteoclast formation by the involvement of fusion-related molecules such as dendritic cell-specific transmembrane protein (DC-STAMP) and P2X7 receptor (P2X7R). RAW264.7 (a murine osteoclastic-like cell line) cells were treated with 20ng/ml receptor activator of NF-κB ligand (RANKL). For 3 consecutive days, the cells were subjected to 1h of mechanical vibration with 20μm displacement at a frequency of 4Hz and compared to the control cells that were treated under the same condition but without the vibration. After 5days of culture, osteoclast formation was determined. Gene expression of DC-STAMP and P2X7R by RAW264.7 cells was determined after 1h of mechanical vibration, while protein production of the DC-STAMP was determined after 6h of postincubation after vibration. As a result, mechanical vibration of RAW264.7 cells inhibited the formation of osteoclasts. Vibration down-regulated DC-STAMP gene expression by 1.6-fold in the presence of RANKL and by 1.4-fold in the absence of RANKL. Additionally, DC-STAMP protein production was also down-regulated by 1.4-fold in the presence of RANKL and by 1.2-fold in the absence of RANKL in RAW264.7 cells in response to mechanical vibration. However, vibration did not affect P2X7R gene expression. Mouse anti-DC-STAMP antibody inhibited osteoclast formation in the absence of vibration. Our results suggest that mechanical vibration of osteoclast precursor cells reduces DC-STAMP expression in osteoclast precursor cells leading to the inhibition of osteoclast formation.

  4. Cellular and species resistance to murine amphotropic, gibbon ape, and feline subgroup C leukemia viruses is strongly influenced by receptor expression levels and by receptor masking mechanisms.

    PubMed

    Tailor, C S; Nouri, A; Kabat, D

    2000-10-01

    Chinese hamster ovary (CHO) cells are resistant to infections by gibbon ape leukemia virus (GALV) and amphotropic murine leukemia virus (A-MLV) unless they are pretreated with tunicamycin, an inhibitor of N-linked glycosylation. These viruses use the related sodium-phosphate symporters Pit1 and Pit2, respectively, as receptors in nonhamster cells, and evidence has suggested that the corresponding transporters of CHO cells may be masked by tunicamycin-sensitive secreted inhibitors. Although the E36 line of Chinese hamster cells was reported to secrete the putative Pit2 inhibitor and to be sensitive to the inhibitory CHO factors, E36 cells are highly susceptible to both GALV and A-MLV in the absence of tunicamycin. Moreover, expression of E36 Pit2 in CHO cells conferred tunicamycin-independent susceptibilities to both viruses. Based on the latter results, it was suggested that E36 Pit2 must functionally differ from the endogenous Pit2 of CHO cells. To test these ideas, we analyzed the receptor properties of CHO Pit1 and Pit2 in CHO cells. Surprisingly, and counterintuitively, transfection of a CHO Pit2 expression vector into CHO cells conferred strong susceptibility to both GALV and A-MLV, and similar overexpression of CHO Pit1 conferred susceptibility to GALV. Thus, CHO Pit2 is a promiscuous functional receptor for both viruses, and CHO Pit1 is a functional receptor for GALV. Similarly, we found that the natural resistance of Mus dunni tail fibroblasts to subgroup C feline leukemia viruses (FeLV-C) was eliminated simply by overexpression of the endogenous FeLV-C receptor homologue. These results demonstrate a novel and simple method to unmask latent retroviral receptor activities that occur in some cells. Specifically, resistances to retroviruses that are caused by subthreshold levels of receptor expression or by stoichiometrically limited masking or interference mechanisms can be efficiently overcome simply by overexpressing the endogenous receptors in the same

  5. Mechanisms of anabolic androgenic steroid inhibition of mammalian ɛ-subunit-containing GABAA receptors

    PubMed Central

    Jones, Brian L; Whiting, Paul J; Henderson, Leslie P

    2006-01-01

    GABAergic transmission regulates the activity of gonadotrophin-releasing hormone (GnRH) neurons in the preoptic area/hypothalamus that control the onset of puberty and the expression of reproductive behaviours. One of the hallmarks of illicit use of anabolic androgenic steroids (AAS) is disruption of behaviours under neuroendocrine control. GnRH neurons are among a limited population of cells that express high levels of the ɛ-subunit of the GABAA receptor. To better understand the actions of AAS on neuroendocrine mechanisms, we have characterized modulation of GABAA receptor-mediated currents in mouse native GnRH neurons and in heterologous cells expressing recombinant α2β3ɛ-receptors. GnRH neurons exhibited robust currents in response to millimolar concentrations of GABA and a picrotoxin (PTX)-sensitive, bicuculline-insensitive current that probably arises from spontaneous openings of GABAA receptors. The AAS 17α-methyltestosterone (17α-MeT) inhibited spontaneous and GABA-evoked currents in GnRH neurons. For recombinant α2β3ɛ-receptors, 17α-MeT inhibited phasic and tonic GABA-elicited responses, accelerated desensitization and slowed paired pulse response recovery. Single channel analysis indicated that GABA-evoked events could be described by three open dwell components and that 17α-MeT enhanced residence in the intermediate dwell state. This AAS also inhibited a PTX-sensitive, spontaneous current (open probability, ∼0.15–0.2) in a concentration-dependent fashion (IC50 ≈ 9 μm). Kinetic modelling indicated that the inhibition induced by 17α-MeT occurs by an allosteric block in which the AAS interacts preferentially with a closed state and promotes accumulation in that state. Finally, studies with a G302S mutant ɛ-subunit suggest that this residue within the transmembrane domain TM2 plays a role in mediating AAS binding and modulation. In sum, our results indicate that inclusion of the ɛ-subunit significantly alters the profile of AAS

  6. Low concentrations of bisphenol a suppress thyroid hormone receptor transcription through a nongenomic mechanism

    SciTech Connect

    Sheng, Zhi-Guo; Tang, Yuan; Liu, Yu-Xiang; Yuan, Ye; Zhao, Bao-Quan; Chao, Xi-Juan; Zhu, Ben-Zhan

    2012-02-15

    Bisphenol (BPA) is one of the highest-volume chemicals produced worldwide, and human exposure to BPA is thought to be ubiquitous. Various rodent and in vitro studies have shown that thyroid hormone (TH) function can be impaired by BPA. However, it is still unknown if low concentrations of BPA can suppress the thyroid hormone receptor (TR) transcription. The present study aims to investigate the possible suppressing effects of low concentrations of BPA on TR transcription and the involved mechanism(s) in CV-1 cells derived from cercopithecus aethiops monkey kidneys. Using gene reporter assays, BPA at concentrations as low as 10{sup −9} M suppresses TR or steroid receptor coactivator-1(SRC-1)-enhanced TR transcription, but not reducing TR/SRC-1 interaction in mammalian two-hybrid and glutathione S-transferase pull-down studies. It has been further shown that both nuclear receptor co-repressor (N-CoR) and silencing mediator for retinoid and thyroid hormone receptors (SMRT) are recruited to the TR-β1 by BPA in the presence of physiologic concentrations of T3 or T4. However, the overexpression of β3 integrin or c-Src significantly reduces BPA-induced recruitment of N-CoR/SMRT to TR or suppression of TR transcription. Furthermore, BPA inhibits the T3/T4-mediated interassociation of the β3 integrin/c-Src/MAPK/TR-β1 pathways by the co-immunoprecipitation. These results indicate that low concentrations of BPA suppress the TR transcription by disrupting physiologic concentrations of T3/T4-mediated β3 integrin/c-Src/MAPK/TR-β1 pathways, followed by recruiting N-CoR/SMRT to TR-β1, providing a novel insight regarding the TH disruption effects of low concentration BPA. -- Highlights: ► Environmentally relevant concentrations of BPA suppress TR transcription. ► BPA recruits the N-CoR/SMRT to TR under the physiologic concentrations of T3/T4. ► BPA disrupts T3/T4-mediated β3 integrin/c-Src/MAPK/TR-β1 pathways.

  7. Microstructure Evolution and Mechanical Behavior of a Hot-Rolled High-Manganese Dual-Phase Transformation-Induced Plasticity/Twinning-Induced Plasticity Steel

    NASA Astrophysics Data System (ADS)

    Fu, Liming; Shan, Mokun; Zhang, Daoda; Wang, Huanrong; Wang, Wei; Shan, Aidang

    2017-02-01

    The microstructures and deformation behavior were studied in a high-temperature annealed high-manganese dual-phase (28 vol pct δ-ferrite and 72 vol pct γ-austenite) transformation-induced plasticity/twinning-induced plasticity (TRIP/TWIP) steel. The results showed that the steel exhibits a special Lüders-like yielding phenomenon at room temperature (RT) and 348 K (75 °C), while it shows continuous yielding at 423 K, 573 K and 673 K (150 °C, 300 °C and 400 °C) deformation. A significant TRIP effect takes place during Lüders-like deformation at RT and 348 K (75 °C) temperatures. Semiquantitative analysis of the TRIP effect on the Lüders-like yield phenomenon proves that a softening effect of the strain energy consumption of strain-induced transformation is mainly responsible for this Lüders-like phenomenon. The TWIP mechanism dominates the 423 K (150 °C) deformation process, while the dislocation glide controls the plasticity at 573 K (300 °C) deformation. The delta-ferrite, as a hard phase in annealed dual-phase steel, greatly affects the mechanical stability of austenite due to the heterogeneous strain distribution between the two phases during deformation. A delta-ferrite-aided TRIP effect, i.e., martensite transformation induced by localized strain concentration of the hard delta-ferrite, is proposed to explain this kind of Lüders-like phenomenon. Moreover, the tensile curve at RT exhibits an upward curved behavior in the middle deformation stage, which is principally attributed to the deformation twinning of austenite retained after Lüders-like deformation. The combination of the TRIP effect during Lüders-like deformation and the subsequent TWIP effect greatly enhances the ductility in this annealed high-manganese dual-phase TRIP/TWIP steel.

  8. Dual blockade of the A1 and A2A adenosine receptor prevents amyloid beta toxicity in neuroblastoma cells exposed to aluminum chloride.

    PubMed

    Giunta, Salvatore; Andriolo, Violetta; Castorina, Alessandro

    2014-09-01

    In a previous work we have shown that exposure to aluminum (Al) chloride (AlCl3) enhanced the neurotoxicity of the amyloid beta(25-35) fragment (Abeta(25-35)) in neuroblastoma cells and affected the expression of Alzheimer's disease (AD)-related genes. Caffein, a compound endowed with beneficial effects against AD, exerts neuroprotection primarily through its antagonist activity on A2A adenosine receptors (A2AR), although it also inhibits A1Rs with similar potency. Still, studies on the specific involvement of these receptors in neuroprotection in a model of combined neurotoxicity (Abeta(25-35)+AlCl3) are missing. To address this issue, cultured SH-SY5Y cells exposed to Abeta(25-35)+AlCl3 were assessed for cell viability, morphology, intracellular ROS activity and expression of apoptosis-, stress- and AD-related proteins. To define the role of A1R and A2ARs, pretreatment with caffein, specific receptor antagonists (DPCPX or SCH58261) or siRNA-mediated gene knockdown were delivered. Results indicate that AlCl3 treatment exacerbated Abeta(25-35) toxicity, increased ROS production, lipid peroxidation, β-secretase-1 (BACE1) and amyloid precursor protein (APP). Interestingly, SCH58261 successfully prevented toxicity associated to Abeta(25-35) only, whereas pretreatment with both DPCPX and SCH58261 was required to fully avert Abeta(25-35)+AlCl3-induced damage, suggesting that A1Rs might also be critically involved in protection during combined toxicity. The effects of caffein were mimicked by both N-acetyl cysteine, an antioxidant, and desferrioxamine, likely acting through distinct mechanisms. Altogether, our data establish a novel protective function associated with A1R inhibition in the setting of combined Abeta(25-35)+AlCl3 neurotoxicity, and expand our current knowledge on the potential beneficial role of caffein to prevent AD progression in subjects environmentally exposed to aluminum.

  9. Uncovering the dual role of RHAMM as an HA receptor and a regulator of CD44 expression in RHAMM-expressing mesenchymal progenitor cells

    PubMed Central

    Veiseh, Mandana; Leith, Sean J.; Tolg, Cornelia; Elhayek, Sallie S.; Bahrami, S. Bahram; Collis, Lisa; Hamilton, Sara; McCarthy, James B.; Bissell, Mina J.; Turley, Eva

    2015-01-01

    The interaction of hyaluronan (HA) with mesenchymal progenitor cells impacts trafficking and fate after tissue colonization during wound repair and these events contribute to diseases such as cancer. How this interaction occurs is poorly understood. Using 10T½ cells as a mesenchymal progenitor model and fluorescent (F-HA) or gold-labeled HA (G-HA) polymers, we studied the role of two HA receptors, RHAMM and CD44, in HA binding and uptake in non-adherent and adherent mesenchymal progenitor (10T½) cells to mimic aspects of cell trafficking and tissue colonization. We show that fluorescent labeled HA (F-HA) binding/uptake was high in non-adherent cells but dropped over time as cells became increasingly adherent. Non-adherent cells displayed both CD44 and RHAMM but only function-blocking anti-RHAMM and not anti-CD44 antibodies significantly reduced F-HA binding/uptake. Adherent cells, which also expressed CD44 and RHAMM, primarily utilized CD44 to bind to F-HA since anti-CD44 but not anti-RHAMM antibodies blocked F-HA uptake. RHAMM overexpression in adherent 10T½ cells led to increased F-HA uptake but this increased binding remained CD44 dependent. Further studies showed that RHAMM-transfection increased CD44 mRNA and protein expression while blocking RHAMM function reduced expression. Collectively, these results suggest that cellular microenvironments in which these receptors function as HA binding proteins differ significantly, and that RHAMM plays at least two roles in F-HA binding by acting as an HA receptor in non-attached cells and by regulating CD44 expression and display in attached cells. Our findings demonstrate adhesion-dependent mechanisms governing HA binding/ uptake that may impact development of new mesenchymal cell-based therapies. PMID:26528478

  10. Uncovering the dual role of RHAMM as an HA receptor and a regulator of CD44 expression in RHAMM-expressing mesenchymal progenitor cells.

    PubMed

    Veiseh, Mandana; Leith, Sean J; Tolg, Cornelia; Elhayek, Sallie S; Bahrami, S Bahram; Collis, Lisa; Hamilton, Sara; McCarthy, James B; Bissell, Mina J; Turley, Eva

    2015-01-01

    The interaction of hyaluronan (HA) with mesenchymal progenitor cells impacts trafficking and fate after tissue colonization during wound repair and these events contribute to diseases such as cancer. How this interaction occurs is poorly understood. Using 10T½ cells as a mesenchymal progenitor model and fluorescent (F-HA) or gold-labeled HA (G-HA) polymers, we studied the role of two HA receptors, RHAMM and CD44, in HA binding and uptake in non-adherent and adherent mesenchymal progenitor (10T½) cells to mimic aspects of cell trafficking and tissue colonization. We show that fluorescent labeled HA (F-HA) binding/uptake was high in non-adherent cells but dropped over time as cells became increasingly adherent. Non-adherent cells displayed both CD44 and RHAMM but only function-blocking anti-RHAMM and not anti-CD44 antibodies significantly reduced F-HA binding/uptake. Adherent cells, which also expressed CD44 and RHAMM, primarily utilized CD44 to bind to F-HA since anti-CD44 but not anti-RHAMM antibodies blocked F-HA uptake. RHAMM overexpression in adherent 10T½ cells led to increased F-HA uptake but this increased binding remained CD44 dependent. Further studies showed that RHAMM-transfection increased CD44 mRNA and protein expression while blocking RHAMM function reduced expression. Collectively, these results suggest that cellular microenvironments in which these receptors function as HA binding proteins differ significantly, and that RHAMM plays at least two roles in F-HA binding by acting as an HA receptor in non-attached cells and by regulating CD44 expression and display in attached cells. Our findings demonstrate adhesion-dependent mechanisms governing HA binding/ uptake that may impact development of new mesenchymal cell-based therapies.

  11. Boldine enhances bile production in rats via osmotic and Farnesoid X receptor dependent mechanisms

    SciTech Connect

    Cermanova, Jolana; Kadova, Zuzana; Zagorova, Marie; Hroch, Milos; Tomsik, Pavel; Nachtigal, Petr; Kudlackova, Zdenka; Pavek, Petr; Dubecka, Michaela; Ceckova, Martina; Staud, Frantisek; Laho, Tomas; Micuda, Stanislav

    2015-05-15

    Boldine, the major alkaloid from the Chilean Boldo tree, is used in traditional medicine to support bile production, but evidence to support this function is controversial. We analyzed the choleretic potential of boldine, including its molecular background. The acute- and long-term effects of boldine were evaluated in rats either during intravenous infusion or after 28-day oral treatment. Infusion of boldine instantly increased the bile flow 1.4-fold in healthy rats as well as in animals with Mrp2 deficiency or ethinylestradiol induced cholestasis. This effect was not associated with a corresponding increase in bile acid or glutathione biliary excretion, indicating that the effect is not related to stimulation of either bile acid dependent or independent mechanisms of bile formation and points to the osmotic activity of boldine itself. We subsequently analyzed bile production under conditions of changing biliary excretion of boldine after bolus intravenous administration and found strong correlations between both parameters. HPLC analysis showed that bile concentrations of boldine above 10 μM were required for induction of choleresis. Importantly, long-term pretreatment, when the bile collection study was performed 24-h after the last administration of boldine, also accelerated bile formation despite undetectable levels of the compound in bile. The effect paralleled upregulation of the Bsep transporter and increased biliary clearance of its substrates, bile acids. We consequently confirmed the ability of boldine to stimulate the Bsep transcriptional regulator, FXR receptor. In conclusion, our study clarified the mechanisms and circumstances surrounding the choleretic activity of boldine. - Highlights: • Boldine may increase bile production by direct as well as indirect mechanisms. • Biliary concentrations of boldine above 10 μM directly stimulate bile production. • Long-term oral boldine administration increases bile acid (BA) biliary secretion. • Boldine

  12. Interaction of NMDA Receptor and Pacemaking Mechanisms in the Midbrain Dopaminergic Neuron

    PubMed Central

    Ha, Joon; Kuznetsov, Alexey

    2013-01-01

    Dopamine neurotransmission has been found to play a role in addictive behavior and is altered in psychiatric disorders. Dopaminergic (DA) neurons display two functionally distinct modes of electrophysiological activity: low- and high-frequency firing. A puzzling feature of the DA neuron is the following combination of its responses: N-methyl-D-aspartate receptor (NMDAR) activation evokes high-frequency firing, whereas other tonic excitatory stimuli (-amino-3-hydroxyl-5-methyl-4-isoxazolepropionate receptor (AMPAR) activation or applied depolarization) block firing instead. We suggest a new computational model that reproduces this combination of responses and explains recent experimental data. Namely, somatic NMDAR stimulation evokes high-frequency firing and is more effective than distal dendritic stimulation. We further reduce the model to a single compartment and analyze the mechanism of the distinct high-frequency response to NMDAR activation vs. other stimuli. Standard nullcline analysis shows that the mechanism is based on a decrease in the amplitude of calcium oscillations. The analysis confirms that the nonlinear voltage dependence provided by the magnesium block of the NMDAR determine its capacity to elevate the firing frequency. We further predict that the moderate slope of the voltage dependence plays the central role in the frequency elevation. Additionally, we suggest a repolarizing current that sustains calcium-independent firing or firing in the absence of calcium-dependent repolarizing currents. We predict that the ether–a-go-go current (ERG), which has been observed in the DA neuron, is the best fit for this critical role. We show that a calcium-dependent and a calcium-independent oscillatory mechanisms form a structure of interlocked negative feedback loops in the DA neuron. The structure connects research of DA neuron firing with circadian biology and determines common minimal models for investigation of robustness of oscillations, which is

  13. Influence of Mechanical Circulatory Support on Endothelin Receptor Expression in Human Left Ventricular Myocardium from Patients with Dilated Cardiomyopathy (DCM)

    PubMed Central

    Gärtner, Florian; Abraham, Getu; Kassner, Astrid; Baurichter, Daniela; Milting, Hendrik

    2017-01-01

    Background In terminal failing hearts ventricular assist devices (VAD) are implanted as a bridge to transplantation. Endothelin receptor (ETR) antagonists are used for treatment of secondary pulmonary hypertension in VAD patients. However, the cardiac ETR regulation in human heart failure and during VAD support is incompletely understood. Methods In paired left ventricular samples of 12 dilated cardiomyopathy patients we investigated the density of endothelin A (ETA) and B (ETB) receptors before VAD implantation and after device removal. Left ventricular samples of 12 non-failing donor hearts served as control. Receptor quantification was performed by binding of [125I]-ET-1 in the presence of nonselective and ETA selective ETR ligands as competitors. Additionally, the ETR mRNA expression was analyzed using quantitative real-time-PCR. Results The mRNA of ETA but not ETB receptors was significantly elevated in heart failure, whereas total ETR density analyzed by radioligand binding was significantly reduced due to ETB receptor down regulation. ETA and ETB receptor density showed poor correlation to mRNA data (spearman correlation factor: 0.43 and 0.31, respectively). VAD support had no significant impact on the density of both receptors and on mRNA expression of ETA whereas ETB mRNA increased during VAD. A meta-analysis reveals that the ETA receptor regulation in human heart failure appears to depend on non-failing hearts. Conclusions In deteriorating hearts of patients suffering from dilated cardiomyopathy the ETA receptor density is not changed whereas the ETB receptor is down regulated. The mRNA and the proteins of ETA and ETB show a weak correlation. Non-failing hearts might influence the interpretation of ETA receptor regulation. Mechanical unloading of the failing hearts has no impact on the myocardial ETR density. PMID:28095452

  14. Molecular mechanism of agonism and inverse agonism in the melanocortin receptors: Zn(2+) as a structural and functional probe.

    PubMed

    Holst, Birgitte; Schwartz, Thue W

    2003-06-01

    Among the rhodopsin-like 7TM receptors, the MC receptors are functionally unique because their high constitutive signaling activity is regulated not only by endogenous peptide agonists-MSH peptides-but also by endogenous inverse agonists, namely, the proteins agouti and AGRP. Moreover, the metal-ion Zn(2+) increases the signaling activity of at least the MC1 and MC4 receptors in three distinct ways: (1). by directly functioning as an agonist; (2). by potentiating the action of the endogenous agonist; and (3). by inhibiting the binding of the endogenous inverse agonist. Structurally the MC receptors are part of a small subset of 7TM receptors in which the main ligand-binding crevice, and especially extracellular loops 2 and 3, appear to be specially designed for easy ligand access and bias towards an active state of the receptor-i.e., constitutive activity. Thus, in the MC receptors extracellular loop 2 is ultrashort because TM-IV basically connects directly into TM-V, whereas extracellular loop 3 appears to be held in a particular, constrained conformation by a putative, internal disulfide bridge. The interaction mode for the small and well-defined zinc-ion between a third, free Cys residue in extracellular loop 3 and conceivably an Asp residue located at the inner face of TM-III gives important information concerning the activation mechanism for the MC receptors.

  15. The cannabinoid beta-caryophyllene (BCP) induces neuritogenesis in PC12 cells by a cannabinoid-receptor-independent mechanism.

    PubMed

    Santos, Neife Aparecida Guinaim; Martins, Nádia Maria; Sisti, Flávia Malvestio; Fernandes, Laís Silva; Ferreira, Rafaela Scalco; de Freitas, Osvaldo; Santos, Antônio Cardozo

    2017-01-05

    Beta-caryophyllene (BCP) is a phytocannabinoid whose neuroprotective activity has been mainly associated with selective activation of cannabinoid-type-2 (CB2) receptors, inhibition of microglial activation and decrease of inflammation. Here, we addressed the potential of BCP to induce neuritogenesis in PC12 cells, a model system for primary neuronal cells that express trkA receptors, respond to NGF and do not express CB2 receptors. We demonstrated that BCP increases the survival and activates the NGF-specific receptor trkA in NGF-deprived PC12 cells, without increasing the expression of NGF itself. The neuritogenic effect of BCP in PC12 cells was abolished by k252a, an inhibitor of the NGF-specific receptor trkA. Accordingly, BCP did not induce neuritogenesis in SH-SY5Y neuroblastoma cells, a neuronal model that does not express trkA receptors and do not respond to NGF. Additionally, we demonstrated that BCP increases the expression of axonal-plasticity-associated proteins (GAP-43, synapsin and synaptophysin) in PC12 cells. It is known that these proteins are up-regulated by NGF in neurons and neuron-like cells, such as PC12 cells. Altogether, these findings suggest that BCP activates trka receptors and induces neuritogenesis by a mechanism independent of NGF or cannabinoid receptors. This is the first study to show such effects of BCP and their beneficial role in neurodegenerative processes should be further investigated.

  16. Time-Dependence of Risperidone and Asenapine Sensitization and Associated D2 receptor Mechanism

    PubMed Central

    Gao, Jun; Li, Ming

    2013-01-01

    When an antipsychotic drug is given repeatedly and intermittently, there is often a long-term increase in its behavioral efficacy, termed antipsychotic sensitization. With the passage of time, the magnitude of antipsychotic sensitization may increase or decrease depending on the principle of Time-Dependent Sensitization (TDS) or memory decay, respectively. In the present study, we examined the time-dependent feature and possible dopamine D2 receptor mechanism of sensitization induced by the antipsychotics risperidone and asenapine in the conditioned avoidance response test. Well-trained male adult Sprague-Dawley rats were first repeatedly treated with risperidone (1.0 mg/kg) or asenapine (0.2 mg/kg) and tested for avoidance response daily for 5 consecutive days. Eight, 18 or 38 days after the 5th drug treatment, all rats were retested drug-free to assess the residual impact of prior risperidone or asenapine treatment. Drug-pretreated rats had significantly lower avoidance than vehicle-pretreated ones on this test, and the group differences increased with the passage of time. In the subsequent drug challenge test at 10, 20 or 40 days after the 5th drug treatment, all rats were injected with a low dose of risperidone (0.3 mg/kg) or asenapine (0.1 mg/kg). Drug-pretreated rats again made significantly less avoidances than controls, confirming the drug-induced sensitization effect. Finally, in the quinpirole (a D2/3 receptor agonist, 1.0 mg/kg, sc)-induced hyperlocomotion test, risperidone-pretreated rats exhibited a significantly higher level of motor activity than the vehicle-pretreated ones. These findings suggest that risperidone and asenapine sensitization is long-lasting, follows the TDS principle, and is likely mediated by D2 receptor supersensitivity. PMID:24103641

  17. ROLE OF ANGIOTENSIN AT2 RECEPTORS IN NATRIURESIS: INTRARENAL MECHANISMS AND THERAPEUTIC POTENTIAL

    PubMed Central

    Carey, Robert M.; Padia, Shetal H.

    2014-01-01

    SUMMARY The renin-angiotensin system is a coordinated hormonal cascade critical for the regulation of blood pressure (BP) and kidney function. Angiotensin II (Ang II), the major angiotensin effector peptide, binds to two major receptors, type-1 (AT1Rs) and type-2 (AT2Rs). AT1Rs engender antinatriuresis and raise BP, whereas AT2Rs oppose these effects, inducing natriuresis and reducing BPAT2Rs are highly expressed in the adult kidney, especially in the proximal tubule. In AT2R-null mice, long-term Ang II infusion results in pressor and antinatriuretic hypersensivivity compared to responses in wild-type animals.The major endogenous receptor ligand for AT2R-mediated natriuretic responses appears to be des-aspartyl1-Ang II (Ang III) instead of Ang II. Recent studies have demonstrated that Ang II requires metabolism to Ang III by aminopeptidase A in order to induce natriuresis and that inhibition of aminopeptidase N increases intrarenal Ang III and augments Ang III-induced natriuresis.The renal dopaminergic system is another important natriuretic pathway. Renal proximal tubule D1-like receptors (D1LIKERs) control approximately 50% of basal sodium (Na+) excretion. We have recently found that natriuresis induced by proximal tubule D1LIKERs requires AT2R activation and that D1LIKER stimulation induces recruitment of AT2Rs to the apical plasma membrane via a cyclic AMP-dependent mechanism.Initial studies employing potent AT2R non-peptide agonist Compound 21 demonstrate natriuresis in both the presence and absence of AT1R blockade indicating the therapeutic potential of this compound in fluid retaining states and hypertension. PMID:23336117

  18. The G2A receptor (GPR132) contributes to oxaliplatin-induced mechanical pain hypersensitivity.

    PubMed

    Hohmann, Stephan W; Angioni, Carlo; Tunaru, Sorin; Lee, Seungkyu; Woolf, Clifford J; Offermanns, Stefan; Geisslinger, Gerd; Scholich, Klaus; Sisignano, Marco

    2017-03-27

    Chemotherapy-induced peripheral neuropathic pain (CIPN) is a common and severe debilitating side effect of many widely used cytostatics. However, there is no approved pharmacological treatment for CIPN available. Among other substances, oxaliplatin causes CIPN in up to 80% of treated patients. Here, we report the involvement of the G-protein coupled receptor G2A (GPR132) in oxaliplatin-induced neuropathic pain in mice. We found that mice deficient in the G2A-receptor show decreased mechanical hypersensitivity after oxaliplatin treatment. Lipid ligands of G2A were found in increased concentrations in the sciatic nerve and dorsal root ganglia of oxaliplatin treated mice. Calcium imaging and patch-clamp experiments show that G2A activation sensitizes the ligand-gated ion channel TRPV1 in sensory neurons via activation of PKC. Based on these findings, we conclude that targeting G2A may be a promising approach to reduce oxaliplatin-induced TRPV1-sensitization and the hyperexcitability of sensory neurons and thereby to reduce pain in patients treated with this chemotherapeutic agent.

  19. [Current Topics on Vitamin D. Mechanism of molecular action of vitamin D via its nuclear receptor].

    PubMed

    Kato, Shigeaki; Morita, Tomohiro

    2015-03-01

    Most of vitamin D actions mediate expression of target genes regulated by nuclear vitamin D receptor (VDR). Regulation of chromatin environment has emerged to underlie gene regulation by liganded VDR. Active state of chromatin is defined by specific combination of post-translational modification of histone proteins, and chromatin remodelers as nuclear complexes conduct dynamic shift of chromatin sate. Among histone modifications, methylations of specific lysine residues located in the N-terminal tails of histone H3 are known to play pivotal roles in directing chromatin state, and the methylated lysine 4 and 9 in the histone H3 (H3K4me and H3K9me) are widely used as indicators of chromatin state. The histone modifying enzymes and chromatin remodelers are thus regulators for chromatin environment, and overtly co-regulate transcriptional regulations of a particular set of target genes by nuclear receptors including VDR. In this review, the molecular mechanism of regulated chromatin configuration is described by illustrating modifications of histone proteins and rearrangements of nucleosome array and their regulators.

  20. Mechanisms of anorexia-cachexia syndrome and rational for treatment with selective ghrelin receptor agonist.

    PubMed

    Esposito, Angela; Criscitiello, Carmen; Gelao, Lucia; Pravettoni, Gabriella; Locatelli, Marzia; Minchella, Ida; Di Leo, Maria; Liuzzi, Rita; Milani, Alessandra; Massaro, Mariangela; Curigliano, Giuseppe

    2015-11-01

    Cancer cachexia is a multi-organ, multifactorial and often irreversible syndrome affecting many patients with cancer. Cancer cachexia is invariably associated with weight loss, mainly from loss of skeletal muscle and body fat, conditioning a reduced quality of life due to asthenia, anorexia, anaemia and fatigue. Treatment options for treating cancer cachexia are limited. The approach is multimodal and may include: treatment of secondary gastrointestinal symptoms, nutritional treatments, drug, and non-drug treatments. Nutritional counselling and physical training may be beneficial in delaying or preventing the development of anorexia-cachexia. However, these interventions are limited in their effect, and no definitive pharmacological treatment is available to address the relevant components of the syndrome. Anamorelin is a first-in-class, orally active ghrelin receptor agonist that binds and stimulates the growth hormone secretagogue receptor centrally, thereby mimicking the appetite-enhancing and anabolic effects of ghrelin. It represents a new class of drug and an additional treatment option for this patient group, whose therapeutic options are currently limited. In this review we examine the mechanisms of anamorelin by which it contrasts catabolic states, its role in regulation of metabolism and energy homeostasis, the data of recent trials in the setting of cancer cachexia and its safety profile.