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Sample records for acetylcholine vesicular transporter

  1. Variants in SLC18A3, vesicular acetylcholine transporter, cause congenital myasthenic syndrome

    NARCIS (Netherlands)

    O'Grady, Gina L.; Verschuuren, Corien; Yuen, Michaela; Webster, Richard; Menezes, Manoj; Fock, Johanna M.; Pride, Natalie; Best, Heather A.; Damm, Tatiana Benavides; Turner, Christian; Lek, Monkol; Engel, Andrew G.; North, Kathryn N.; Clarke, Nigel F.; MacArthur, Daniel G.; Kamsteeg, Erik-Jan; Cooper, Sandra T.

    2016-01-01

    Objective: To describe the clinical and genetic characteristics of presynaptic congenital myasthenic syndrome secondary to biallelic variants in SLC18A3. Methods: Individuals from 2 families were identified with biallelic variants in SLC18A3, the gene encoding the vesicular acetylcholine transporter

  2. Spiroindolines identify the vesicular acetylcholine transporter as a novel target for insecticide action.

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    Ann Sluder

    Full Text Available The efficacy of all major insecticide classes continues to be eroded by the development of resistance mediated, in part, by selection of alleles encoding insecticide insensitive target proteins. The discovery of new insecticide classes acting at novel protein binding sites is therefore important for the continued protection of the food supply from insect predators, and of human and animal health from insect borne disease. Here we describe a novel class of insecticides (Spiroindolines encompassing molecules that combine excellent activity against major agricultural pest species with low mammalian toxicity. We confidently assign the vesicular acetylcholine transporter as the molecular target of Spiroindolines through the combination of molecular genetics in model organisms with a pharmacological approach in insect tissues. The vesicular acetylcholine transporter can now be added to the list of validated insecticide targets in the acetylcholine signalling pathway and we anticipate that this will lead to the discovery of novel molecules useful in sustaining agriculture. In addition to their potential as insecticides and nematocides, Spiroindolines represent the only other class of chemical ligands for the vesicular acetylcholine transporter since those based on the discovery of vesamicol over 40 years ago, and as such, have potential to provide more selective tools for PET imaging in the diagnosis of neurodegenerative disease. They also provide novel biochemical tools for studies of the function of this protein family.

  3. Elimination of the vesicular acetylcholine transporter in the striatum reveals regulation of behaviour by cholinergic-glutamatergic co-transmission.

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    Monica S Guzman

    2011-11-01

    Full Text Available Cholinergic neurons in the striatum are thought to play major regulatory functions in motor behaviour and reward. These neurons express two vesicular transporters that can load either acetylcholine or glutamate into synaptic vesicles. Consequently cholinergic neurons can release both neurotransmitters, making it difficult to discern their individual contributions for the regulation of striatal functions. Here we have dissected the specific roles of acetylcholine release for striatal-dependent behaviour in mice by selective elimination of the vesicular acetylcholine transporter (VAChT from striatal cholinergic neurons. Analysis of several behavioural parameters indicates that elimination of VAChT had only marginal consequences in striatum-related tasks and did not affect spontaneous locomotion, cocaine-induced hyperactivity, or its reward properties. However, dopaminergic sensitivity of medium spiny neurons (MSN and the behavioural outputs in response to direct dopaminergic agonists were enhanced, likely due to increased expression/function of dopamine receptors in the striatum. These observations indicate that previous functions attributed to striatal cholinergic neurons in spontaneous locomotor activity and in the rewarding responses to cocaine are mediated by glutamate and not by acetylcholine release. Our experiments demonstrate how one population of neurons can use two distinct neurotransmitters to differentially regulate a given circuitry. The data also raise the possibility of using VAChT as a target to boost dopaminergic function and decrease high striatal cholinergic activity, common neurochemical alterations in individuals affected with Parkinson's disease.

  4. (E)-[125I]-5-AOIBV: a SPECT radioligand for the vesicular acetylcholine transporter

    International Nuclear Information System (INIS)

    Emond, Patrick; Mavel, Sylvie; Zea-Ponce, Yolanda; Kassiou, Michael; Garreau, Lucette; Bodard, Sylvie; Drossard, Marie-Laure; Chalon, Sylvie; Guilloteau, Denis

    2007-01-01

    The premise that, over the course of Alzheimer's disease (AD), changes in the levels of the vesicular acetylcholine transporter (VAChT) occur in parallel with changes to other cholinergic marker proteins provides the basis for the applicability of benzovesamicol derivatives as radioligands for AD studies by single photon emission computed tomography or positron emission tomography. We report the synthesis of enantiopure benzovesamicol derivatives: (R,R) or (S,S)-(E)-2-hydroxy-5-(3-iodoprop-2-en-1-oxy)-3- (4-phenylpiperidino)tetralin [(R,R)-AOIBV: K d =0.45 nM or (S,S)-5-AOIBV: K d =4.3 nM] and their corresponding tributyltin precursors for radioiodination. (R,R or S,S)-5-AOIBV was labeled with iodine-125 from their corresponding n-tributyltin precursors. Both compounds were obtained with radiochemical and optical purity greater than 97% and in radiochemical yields ranging 34-36%. To determine if these compounds could provide an advantage when compared to [ 125 I]-iodo benzovesamicol (IBVM), IBVM was also labeled and used as the reference compound in all ex vivo experiments. Ex vivo biodistribution experiments in rats revealed that [ 125 I]-(R,R)-5-AOIBV displayed the most suitable pharmacological profile as the radioactivity distribution corresponded well with the known VAChT brain density. Moreover, pre-injection of vesamicol prevented the uptake of [ 125 I]-(R,R)-5-AOIBV in striatum, cortex and hippocampus, demonstrating selectivity for the VAChT. However, even if time activity curves of [ 125 I]-(R,R)-5-AOIBV confirmed that this compound could be used to visualize the VAChT in vivo, at each point of the kinetic study, [ 125 I]-(R,R)-5-AOIBV showed a lower specific binding compared to [ 125 I]-IBVM. These results made [ 125 I]-( R,R)-5-AOIBV inferior to [ 125 I]-IBVM for the VAChT exploration in vivo

  5. (E)-[{sup 125}I]-5-AOIBV: a SPECT radioligand for the vesicular acetylcholine transporter

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    Emond, Patrick [INSERM U619, 37000 Tours (France); Universite Francois-Rabelais de Tours, CHRU, Hopital Bretonneau, Service de Medecine nucleaire, 37000 Tours (France); Mavel, Sylvie [INSERM U619, 37000 Tours (France); Universite Francois-Rabelais de Tours, CHRU, Hopital Bretonneau, Service de Medecine nucleaire, 37000 Tours (France)], E-mail: sylvie.mavel@univ-tours.fr; Zea-Ponce, Yolanda [INSERM U619, 37000 Tours (France); Universite Francois-Rabelais de Tours, CHRU, Hopital Bretonneau, Service de Medecine nucleaire, 37000 Tours (France); Kassiou, Michael [Discipline of Medical Radiation Sciences, Brain and Mind Research Institute, University of Sydney, NSW 2050 (Australia); School of Chemistry, University of Sydney, NSW 2006 (Australia); Garreau, Lucette; Bodard, Sylvie; Drossard, Marie-Laure; Chalon, Sylvie; Guilloteau, Denis [INSERM U619, 37000 Tours (France); Universite Francois-Rabelais de Tours, CHRU, Hopital Bretonneau, Service de Medecine nucleaire, 37000 Tours (France)

    2007-11-15

    The premise that, over the course of Alzheimer's disease (AD), changes in the levels of the vesicular acetylcholine transporter (VAChT) occur in parallel with changes to other cholinergic marker proteins provides the basis for the applicability of benzovesamicol derivatives as radioligands for AD studies by single photon emission computed tomography or positron emission tomography. We report the synthesis of enantiopure benzovesamicol derivatives: (R,R) or (S,S)-(E)-2-hydroxy-5-(3-iodoprop-2-en-1-oxy)-3- (4-phenylpiperidino)tetralin [(R,R)-AOIBV: K{sub d}=0.45 nM or (S,S)-5-AOIBV: K{sub d}=4.3 nM] and their corresponding tributyltin precursors for radioiodination. (R,R or S,S)-5-AOIBV was labeled with iodine-125 from their corresponding n-tributyltin precursors. Both compounds were obtained with radiochemical and optical purity greater than 97% and in radiochemical yields ranging 34-36%. To determine if these compounds could provide an advantage when compared to [{sup 125}I]-iodo benzovesamicol (IBVM), IBVM was also labeled and used as the reference compound in all ex vivo experiments. Ex vivo biodistribution experiments in rats revealed that [{sup 125}I]-(R,R)-5-AOIBV displayed the most suitable pharmacological profile as the radioactivity distribution corresponded well with the known VAChT brain density. Moreover, pre-injection of vesamicol prevented the uptake of [{sup 125}I]-(R,R)-5-AOIBV in striatum, cortex and hippocampus, demonstrating selectivity for the VAChT. However, even if time activity curves of [{sup 125}I]-(R,R)-5-AOIBV confirmed that this compound could be used to visualize the VAChT in vivo, at each point of the kinetic study, [{sup 125}I]-(R,R)-5-AOIBV showed a lower specific binding compared to [{sup 125}I]-IBVM. These results made [{sup 125}I]-( R,R)-5-AOIBV inferior to [{sup 125}I]-IBVM for the VAChT exploration in vivo.

  6. Whole-Retina Reduced Electrophysiological Activity in Mice Bearing Retina-Specific Deletion of Vesicular Acetylcholine Transporter.

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    Jake Bedore

    Full Text Available Despite rigorous characterization of the role of acetylcholine in retinal development, long-term effects of its absence as a neurotransmitter are unknown. One of the unanswered questions is how acetylcholine contributes to the functional capacity of mature retinal circuits. The current study investigates the effects of disrupting cholinergic signalling in mice, through deletion of vesicular acetylcholine transporter (VAChT in the developing retina, pigmented epithelium, optic nerve and optic stalk, on electrophysiology and structure of the mature retina.A combination of electroretinography, optical coherence tomography imaging and histological evaluation assessed retinal integrity in mice bearing retina- targeted (embryonic day 12.5 deletion of VAChT (VAChTSix3-Cre-flox/flox and littermate controls at 5 and 12 months of age. VAChTSix3-Cre-flox/flox mice did not show any gross changes in nuclear layer cellularity or synaptic layer thickness. However, VAChTSix3-Cre-flox/flox mice showed reduced electrophysiological response of the retina to light stimulus under scotopic conditions at 5 and 12 months of age, including reduced a-wave, b-wave, and oscillatory potential (OP amplitudes and decreased OP peak power and total energy. Reduced a-wave amplitude was proportional to the reduction in b-wave amplitude and not associated with altered a-wave 10%-90% rise time or inner and outer segment thicknesses.This study used a novel genetic model in the first examination of function and structure of the mature mouse retina with disruption of cholinergic signalling. Reduced amplitude across the electroretinogram wave form does not suggest dysfunction in specific retinal cell types and could reflect underlying changes in the retinal and/or extraretinal microenvironment. Our findings suggest that release of acetylcholine by VAChT is essential for the normal electrophysiological response of the mature mouse retina.

  7. Quantitative autoradiography of brain binding sites for the vesicular acetylcholine transport blocker 2-(4-phenylpiperidino)cyclohexanol (AH5183)

    International Nuclear Information System (INIS)

    Marien, M.R.; Parsons, S.M.; Altar, C.A.

    1987-01-01

    2-(4-Phenylpiperidino)cyclohexanol (AH5183) is a noncompetitive and potent inhibitor of high-affinity acetylcholine transport into cholinergic vesicles. It is reported here that [ 3 H]AH5183 binds specifically and saturably to slide-mounted sections of the rat forebrain (Kd = 1.1 to 2.2 X 10(-8) M; Bmax = 286 to 399 fmol/mg of protein). The association and dissociation rate constants for [ 3 H]AH5183 binding are 8.6 X 10(6) M-1 X min-1 and 0.18 min-1, respectively. Bound [ 3 H]AH5183 can be displaced by nonradioactive AH5183 and by the structural analog (2 alpha,3 beta,4A beta,8A alpha)-decahydro-3-(4-phenyl-1-piperidinyl)-2- naphthalenol but not by 10 microM concentrations of the cholinergic drugs acetylcholine, choline, atropine, hexamethonium, eserine, or hemicholinium-3 or by the structurally related compounds 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine, 1-methyl-4-phenylpyridine, (+/-)-N-allylnormetazocine (SKF 10,047), levoxadrol, or dexoxadrol. Quantitative autoradiography reveals that [ 3 H]AH5183 binding sites are distributed heterogenously throughout the rat forebrain and are highly localized to cholinergic nerve terminal regions. At the level of the caudate nucleus-putamen, the highest concentrations of saturable [ 3 H]AH5183 binding (713-751 fmol/mg of protein) are found in the vertical limb of the diagonal band and the olfactory tubercle, with lesser amounts (334-516 fmol/mg of protein) in the caudate-putamen, nucleus accumbens, superficial layers of the cerebral cortex, and the primary olfactory cortex. At day 7 after transsection of the left fimbria, [ 3 H]AH5183 binding and choline acetyltransferase activity in the left hippocampus were reduced by 33 +/- 6% and 61 +/- 7%, respectively. These findings indicate that [ 3 H]AH5183 binds to a unique recognition site in rat brain that is topographically associated with cholinergic nerve terminals

  8. Mice deficient for striatal Vesicular Acetylcholine Transporter (VAChT) display impaired short-term but normal long-term object recognition memory.

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    Palmer, Daniel; Creighton, Samantha; Prado, Vania F; Prado, Marco A M; Choleris, Elena; Winters, Boyer D

    2016-09-15

    Substantial evidence implicates Acetylcholine (ACh) in the acquisition of object memories. While most research has focused on the role of the cholinergic basal forebrain and its cortical targets, there are additional cholinergic networks that may contribute to object recognition. The striatum contains an independent cholinergic network comprised of interneurons. In the current study, we investigated the role of this cholinergic signalling in object recognition using mice deficient for Vesicular Acetylcholine Transporter (VAChT) within interneurons of the striatum. We tested whether these striatal VAChT(D2-Cre-flox/flox) mice would display normal short-term (5 or 15min retention delay) and long-term (3h retention delay) object recognition memory. In a home cage object recognition task, male and female VAChT(D2-Cre-flox/flox) mice were impaired selectively with a 15min retention delay. When tested on an object location task, VAChT(D2-Cre-flox/flox) mice displayed intact spatial memory. Finally, when object recognition was tested in a Y-shaped apparatus, designed to minimize the influence of spatial and contextual cues, only females displayed impaired recognition with a 5min retention delay, but when males were challenged with a 15min retention delay, they were also impaired; neither males nor females were impaired with the 3h delay. The pattern of results suggests that striatal cholinergic transmission plays a role in the short-term memory for object features, but not spatial location. Copyright © 2016 Elsevier B.V. All rights reserved.

  9. Vesicular storage and release of acetylcholine in Torpedo electroplaque synapses

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    Suszkiw, J B; Zimmermann, H; Whittaker, V P [Max-Planck-Institut fuer Biophysikalische Chemie (Karl-Friedrich-Bonhoefer-Inst.), Goettingen (Germany, F.R.)

    1978-06-01

    The disposition of newly synthesized ACh subsequent to depletion of vesicular endogenous ACh by stimulation was studied in the electromotor nerve terminals of Torpedo marmorata using (/sup 3/H) acetate as a precursor of ACh. Little vesicular (/sup 3/H) ACh could be isolated from tissue immediately after stimulation at 1 Hz. After 3 h post-stimulation recovery the newly-synthesized (/sup 3/H) ACh is found predominantly in a subpopulation of vesicles distinct from the vesicles containing most of the endogenous poorly labelled ACh. Restimulation of the tissue causes release of highly labelled ACh with a specific radioactivity (SRA) comparable to that of the newly synthesized (/sup 3/H) ACh in the highly labelled subpopulation of vesicles and significantly greater than the SRA of ACh in the main vesicular pool of the total tissue.

  10. Are vesicular neurotransmitter transporters potential treatment targets for temporal lobe epilepsy?

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    Joeri eVan Liefferinge

    2013-08-01

    Full Text Available The vesicular neurotransmitter transporters (VNTs are small proteins responsible for packing synaptic vesicles with neurotransmitters thereby determining the amount of neurotransmitter released per vesicle through fusion in both neurons and glial cells. Each transporter subtype was classically seen as a specific neuronal marker of the respective nerve cells containing that particular neurotransmitter or structurally related neurotransmitters. More recently, however, it has become apparent that common neurotransmitters can also act as co-transmitters, adding complexity to neurotransmitter release and suggesting intriguing roles for VNTs therein. We will first describe the current knowledge on vesicular glutamate transporters (VGLUT1/2/3, the vesicular excitatory amino acid transporter (VEAT, the vesicular nucleotide transporter (VNUT, vesicular monoamine transporters (VMAT1/2, the vesicular acetylcholine transporter (VAChT and the vesicular γ-aminobutyric acid (GABA transporter (VGAT in the brain. We will focus on evidence regarding transgenic mice with disruptions in VNTs in different models of seizures and epilepsy. We will also describe the known alterations and reorganizations in the expression levels of these VNTs in rodent models for temporal lobe epilepsy (TLE and in human tissue resected for epilepsy surgery. Finally, we will discuss perspectives on opportunities and challenges for VNTs as targets for possible future epilepsy therapies.

  11. Model of reversible vesicular transport with exclusion

    International Nuclear Information System (INIS)

    Bressloff, Paul C; Karamched, Bhargav R

    2016-01-01

    A major question in neurobiology concerns the mechanics behind the motor-driven transport and delivery of vesicles to synaptic targets along the axon of a neuron. Experimental evidence suggests that the distribution of vesicles along the axon is relatively uniform and that vesicular delivery to synapses is reversible. A recent modeling study has made explicit the crucial role that reversibility in vesicular delivery to synapses plays in achieving uniformity in vesicle distribution, so called synaptic democracy (Bressloff et al 2015 Phys. Rev. Lett. 114 168101). In this paper we generalize the previous model by accounting for exclusion effects (hard-core repulsion) that may occur between molecular motor-cargo complexes (particles) moving along the same microtubule track. The resulting model takes the form of an exclusion process with four internal states, which distinguish between motile and stationary particles, and whether or not a particle is carrying vesicles. By applying a mean field approximation and an adiabatic approximation we reduce the system of ODEs describing the evolution of occupation numbers of the sites on a 1D lattice to a system of hydrodynamic equations in the continuum limit. We find that reversibility in vesicular delivery allows for synaptic democracy even in the presence of exclusion effects, although exclusion does exacerbate nonuniform distributions of vesicles in an axon when compared with a model without exclusion. We also uncover the relationship between our model and other models of exclusion processes with internal states. (paper)

  12. Armadillo motifs involved in vesicular transport.

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    Harald Striegl

    Full Text Available Armadillo (ARM repeat proteins function in various cellular processes including vesicular transport and membrane tethering. They contain an imperfect repeating sequence motif that forms a conserved three-dimensional structure. Recently, structural and functional insight into tethering mediated by the ARM-repeat protein p115 has been provided. Here we describe the p115 ARM-motifs for reasons of clarity and nomenclature and show that both sequence and structure are highly conserved among ARM-repeat proteins. We argue that there is no need to invoke repeat types other than ARM repeats for a proper description of the structure of the p115 globular head region. Additionally, we propose to define a new subfamily of ARM-like proteins and show lack of evidence that the ARM motifs found in p115 are present in other long coiled-coil tethering factors of the golgin family.

  13. Phenyl Ring-Substituted Lobelane Analogs: Inhibition of [3H]Dopamine Uptake at the Vesicular Monoamine Transporter-2

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    Nickell, Justin R.; Zheng, Guangrong; Deaciuc, Agripina G.; Crooks, Peter A.; Dwoskin, Linda P.

    2011-01-01

    Lobeline attenuates the behavioral effects of methamphetamine via inhibition of the vesicular monoamine transporter (VMAT2). To increase selectivity for VMAT2, chemically defunctionalized lobeline analogs, including lobelane, were designed to eliminate nicotinic acetylcholine receptor affinity. The current study evaluated the ability of lobelane analogs to inhibit [3H]dihydrotetrabenazine (DTBZ) binding to VMAT2 and [3H]dopamine (DA) uptake into isolated synaptic vesicles and determined the m...

  14. Vesicular and Plasma Membrane Transporters for Neurotransmitters

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    Blakely, Randy D.; Edwards, Robert H.

    2012-01-01

    The regulated exocytosis that mediates chemical signaling at synapses requires mechanisms to coordinate the immediate response to stimulation with the recycling needed to sustain release. Two general classes of transporter contribute to release, one located on synaptic vesicles that loads them with transmitter, and a second at the plasma membrane that both terminates signaling and serves to recycle transmitter for subsequent rounds of release. Originally identified as the target of psychoactive drugs, these transport systems have important roles in transmitter release, but we are only beginning to understand their contribution to synaptic transmission, plasticity, behavior, and disease. Recent work has started to provide a structural basis for their activity, to characterize their trafficking and potential for regulation. The results indicate that far from the passive target of psychoactive drugs, neurotransmitter transporters undergo regulation that contributes to synaptic plasticity. PMID:22199021

  15. Synaptic Democracy and Vesicular Transport in Axons

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    Bressloff, Paul C.; Levien, Ethan

    2015-04-01

    Synaptic democracy concerns the general problem of how regions of an axon or dendrite far from the cell body (soma) of a neuron can play an effective role in neuronal function. For example, stimulated synapses far from the soma are unlikely to influence the firing of a neuron unless some sort of active dendritic processing occurs. Analogously, the motor-driven transport of newly synthesized proteins from the soma to presynaptic targets along the axon tends to favor the delivery of resources to proximal synapses. Both of these phenomena reflect fundamental limitations of transport processes based on a localized source. In this Letter, we show that a more democratic distribution of proteins along an axon can be achieved by making the transport process less efficient. This involves two components: bidirectional or "stop-and-go" motor transport (which can be modeled in terms of advection-diffusion), and reversible interactions between motor-cargo complexes and synaptic targets. Both of these features have recently been observed experimentally. Our model suggests that, just as in human societies, there needs to be a balance between "efficiency" and "equality".

  16. Chromosomal localization of the human vesicular amine transporter genes

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    Peter, D.; Finn, P.; Liu, Y.; Roghani, A.; Edwards, R.H.; Klisak, I.; Kojis, T.; Heinzmann, C.; Sparkes, R.S. (UCLA School of Medicine, Los Angeles, CA (United States))

    1993-12-01

    The physiologic and behavioral effects of pharmacologic agents that interfere with the transport of monoamine neurotransmitters into vesicles suggest that vesicular amine transport may contribute to human neuropsychiatric disease. To determine whether an alteration in the genes that encode vesicular amine transport contributes to the inherited component of these disorders, the authors have isolated a human cDNA for the brain transporter and localized the human vesciular amine transporter genes. The human brain synaptic vesicle amine transporter (SVAT) shows unexpected conservation with rat SVAT in the regions that diverge extensively between rat SVAT and the rat adrenal chromaffin granule amine transporter (CGAT). Using the cloned sequences with a panel of mouse-human hybrids and in situ hybridization for regional localization, the adrenal CGAT gene (or VAT1) maps to human chromosome 8p21.3 and the brain SVAT gene (or VAT2) maps to chromosome 10q25. Both of these sites occur very close to if not within previously described deletions that produce severe but viable phenotypes. 26 refs., 3 figs., 1 tab.

  17. Distribution of vesicular glutamate transporters in the human brain

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    Erika eVigneault

    2015-03-01

    Full Text Available Glutamate is the major excitatory transmitter in the brain. Vesicular glutamate transporters (VGLUT1-3 are responsible for uploading glutamate into synaptic vesicles. VGLUT1 and VGLUT2 are considered as specific markers of canonical glutamatergic neurons, while VGLUT3 is found in neurons previously shown to use other neurotransmitters than glutamate. Although there exists a rich literature on the localization of these glutamatergic markers in the rodent brain, little is currently known about the distribution of VGLUT1-3 in the human brain. In the present study, using subtype specific probes and antisera, we examined the localization of the three vesicular glutamate transporters in the human brain by in situ hybridization, immunoautoradiography and immunohistochemistry. We found that the VGLUT1 transcript was highly expressed in the cerebral cortex, hippocampus and cerebellum, whereas VGLUT2 mRNA was mainly found in the thalamus and brainstem. VGLUT3 mRNA was localized in scarce neurons within the cerebral cortex, hippocampus, striatum and raphe nuclei. Following immunoautoradiographic labeling, intense VGLUT1- and VGLUT2-immunoreactivities were observed in all regions investigated (cerebral cortex, hippocampus, caudate-putamen, cerebellum, thalamus, amygdala, substantia nigra, raphe while VGLUT3 was absent from the thalamus and cerebellum. This extensive mapping of VGLUT1-3 in human brain reveals distributions that correspond for the most part to those previously described in rodent brains.

  18. Metabolic control of vesicular glutamate transport and release.

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    Juge, Narinobu; Gray, John A; Omote, Hiroshi; Miyaji, Takaaki; Inoue, Tsuyoshi; Hara, Chiaki; Uneyama, Hisayuki; Edwards, Robert H; Nicoll, Roger A; Moriyama, Yoshinori

    2010-10-06

    Fasting has been used to control epilepsy since antiquity, but the mechanism of coupling between metabolic state and excitatory neurotransmission remains unknown. Previous work has shown that the vesicular glutamate transporters (VGLUTs) required for exocytotic release of glutamate undergo an unusual form of regulation by Cl(-). Using functional reconstitution of the purified VGLUTs into proteoliposomes, we now show that Cl(-) acts as an allosteric activator, and the ketone bodies that increase with fasting inhibit glutamate release by competing with Cl(-) at the site of allosteric regulation. Consistent with these observations, acetoacetate reduced quantal size at hippocampal synapses and suppresses glutamate release and seizures evoked with 4-aminopyridine in the brain. The results indicate an unsuspected link between metabolic state and excitatory neurotransmission through anion-dependent regulation of VGLUT activity. Copyright © 2010 Elsevier Inc. All rights reserved.

  19. Expression of Vesicular Nucleotide Transporter in Rat Odontoblasts

    International Nuclear Information System (INIS)

    Ikeda, Erina; Goto, Tetsuya; Gunjigake, Kaori; Kuroishi, Kayoko; Ueda, Masae; Kataoka, Shinji; Toyono, Takashi; Nakatomi, Mitsushiro; Seta, Yuji; Kitamura, Chiaki; Nishihara, Tatsuji; Kawamoto, Tatsuo

    2016-01-01

    Several theories have been proposed regarding pain transmission mechanisms in tooth. However, the exact signaling mechanism from odontoblasts to pulp nerves remains to be clarified. Recently, ATP-associated pain transmission has been reported, but it is unclear whether ATP is involved in tooth pain transmission. In the present study, we focused on the vesicular nucleotide transporter (VNUT), a transporter of ATP into vesicles, and examined whether VNUT was involved in ATP release from odontoblasts. We examined the expression of VNUT in rat pulp by RT-PCR and immunostaining. ATP release from cultured odontoblast-like cells with heat stimulation was evaluated using ATP luciferase methods. VNUT was expressed in pulp tissue, and the distribution of VNUT-immunopositive vesicles was confirmed in odontoblasts. In odontoblasts, some VNUT-immunopositive vesicles were colocalized with membrane fusion proteins. Additionally P2X 3 , an ATP receptor, immunopositive axons were distributed between odontoblasts. The ATP release by thermal stimulation from odontoblast-like cells was inhibited by the addition of siRNA for VNUT. These findings suggest that cytosolic ATP is transported by VNUT and that the ATP in the vesicles is then released from odontoblasts to ATP receptors on axons. ATP vesicle transport in odontoblasts seems to be a key mechanism for signal transduction from odontoblasts to axons in the pulp

  20. Vesicular nucleotide transporter (VNUT): appearance of an actress on the stage of purinergic signaling.

    Science.gov (United States)

    Moriyama, Yoshinori; Hiasa, Miki; Sakamoto, Shohei; Omote, Hiroshi; Nomura, Masatoshi

    2017-09-01

    Vesicular storage of ATP is one of the processes initiating purinergic chemical transmission. Although an active transport mechanism was postulated to be involved in the processes, a transporter(s) responsible for the vesicular storage of ATP remained unidentified for some time. In 2008, SLC17A9, the last identified member of the solute carrier 17 type I inorganic phosphate transporter family, was found to encode the vesicular nucleotide transporter (VNUT) that is responsible for the vesicular storage of ATP. VNUT transports various nucleotides in a membrane potential-dependent fashion and is expressed in the various ATP-secreting cells. Mice with knockout of the VNUT gene lose vesicular storage and release of ATP from neurons and neuroendocrine cells, resulting in blockage of the initiation of purinergic chemical transmission. Thus, VNUT plays an essential role in the vesicular storage and release of ATP. The VNUT knockout mice exhibit resistance for neuropathic pain and a therapeutic effect against diabetes by way of increased insulin sensitivity. Thus, VNUT inhibitors and suppression of VNUT gene expression may be used for therapeutic purposes through suppression of purinergic chemical transmission. This review summarizes the studies to date on VNUT and discusses what we have learned about the relevance of vesicular ATP release as a potential drug target.

  1. Dual and Direction-Selective Mechanisms of Phosphate Transport by the Vesicular Glutamate Transporter

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    Julia Preobraschenski

    2018-04-01

    Full Text Available Summary: Vesicular glutamate transporters (VGLUTs fill synaptic vesicles with glutamate and are thus essential for glutamatergic neurotransmission. However, VGLUTs were originally discovered as members of a transporter subfamily specific for inorganic phosphate (Pi. It is still unclear how VGLUTs accommodate glutamate transport coupled to an electrochemical proton gradient ΔμH+ with inversely directed Pi transport coupled to the Na+ gradient and the membrane potential. Using both functional reconstitution and heterologous expression, we show that VGLUT transports glutamate and Pi using a single substrate binding site but different coupling to cation gradients. When facing the cytoplasm, both ions are transported into synaptic vesicles in a ΔμH+-dependent fashion, with glutamate preferred over Pi. When facing the extracellular space, Pi is transported in a Na+-coupled manner, with glutamate competing for binding but at lower affinity. We conclude that VGLUTs have dual functions in both vesicle transmitter loading and Pi homeostasis within glutamatergic neurons. : Preobraschenski et al. show that the vesicular glutamate transporter functions as a bi-directional phosphate transporter that is coupled with different cations in each direction and hence may play a key role in neuronal phosphate homeostasis. Keywords: VGLUT, SLC17 family, type I Na+-dependent inorganic phosphate transporter, ATPase, proteoliposomes, hybrid vesicles, anti-VGLUT1 nanobody

  2. Exclusion of close linkage between the synaptic vesicular monoamine transporter locus and schizophrenia spectrum disorders

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    Persico, A.M.; Uhl, G.R. [Johns Hopkins Univ. School of Medicine, Baltimore, MD (United States); Wang, Zhe Wu [Universitario Campus Bio-Medico, Rome (Italy)] [and others

    1995-12-18

    The principal brain synaptic vesicular monoamine transporter (VMAT2) is responsible for the reuptake of serotonin, dopamine, norepinephrine, epinephrine, and histamine from the cytoplasm into synaptic vesicles, thus contributing to determination of the size of releasable neurotransmitter vesicular pools. Potential involvement of VMAT2 gene variants in the etiology of schizophrenia and related disorders was tested using polymorphic VMAT2 gene markers in 156 subjects from 16 multiplex pedigrees with schizophrenia, schizophreniform, schizoaffective, and schizotypal disorders and mood incongruent psychotic depression. Assuming genetic homogeneity, complete ({theta} = 0.0) linkage to the schizophrenia spectrum was excluded under both dominant and recessive models. Allelic variants at the VMAT2 locus do not appear to provide major genetic contributions to the etiology of schizophrenia spectrum disorders in these pedigrees. 16 refs.

  3. Dopamine transporter and vesicular monoamine transporter knockout mice : implications for Parkinson's disease.

    Science.gov (United States)

    Miller, G W; Wang, Y M; Gainetdinov, R R; Caron, M G

    2001-01-01

    One of the most valuable methods for understanding the function of a particular protein is the generation of animals that have had the gene encoding for the protein of interest disrupted, commonly known as a "quo;knockout"quo; or null mutant. By incorporating a sequence of DNA (typically encoding antibiotic resistance to aid in the selection of the mutant gene) into embryonic stem cells by homologous recombination, the normal transcription of the gene is effectively blocked (Fig. 1). Since a particular protein is encoded by two copies of a gene, it is necessary to have the gene on both alleles "quo;knocked out."quo; This is performed by cross-breeding animals with one affected allele (heterozygote) to generate offspring that have inherited two mutant alleles (homozygote). This procedure has been used to generate animals lacking either the plasma membrane dopamine transporter (DAT; Fig. 2) or the vesicular monoamine transporter (VMAT2; Fig. 3). Both DAT and VMAT2 are essential for dopamine homeostasis and are thought to participate in the pathogenesis of Parkinson's disease (1-5). Fig. 1. Maps of the targeting vector and the mock construct. The mouse genomic fragment (clone 11) was isolated from a Stratagene 129 SvJ library by standard colony hybridization using a PCR probe from the 5' end of rat cDNA. The restriction site abbreviations are as follows: H, HindIII; N, NotI; Sc, SacI; Sn, SnaI; X, XbaI; and Xh, XhoI. The region between HindIII and SnaI on clone 11 containing the coding sequence from transmembrane domains 3 and 4 of VMAT2 was deleted and replaced with PGK-neo. The 3' fragment of clone 11 was reserved as an external probe for Southern analysis. To facilitate PCR screening of embryonic stem cell clones, a mock construct containing the SnaI/XbaI fragment and part of the Neo cassette was generated as a positive control. pPNT and pGEM4Z were used to construct knockout and mock vectors, respectively. (Reproduced with permission from ref. 1). Fig. 2. DAT and

  4. GLTP mediated non-vesicular GM1 transport between native membranes.

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    Ines Lauria

    Full Text Available Lipid transfer proteins (LTPs are emerging as key players in lipid homeostasis by mediating non-vesicular transport steps between two membrane surfaces. Little is known about the driving force that governs the direction of transport in cells. Using the soluble LTP glycolipid transfer protein (GLTP, we examined GM1 (monosialotetrahexosyl-ganglioside transfer to native membrane surfaces. With artificial GM1 donor liposomes, GLTP can be used to increase glycolipid levels over natural levels in either side of the membrane leaflet, i.e., external or cytosolic. In a system with native donor- and acceptor-membranes, we find that GLTP balances highly variable GM1 concentrations in a population of membranes from one cell type, and in addition, transfers lipids between membranes from different cell types. Glycolipid transport is highly efficient, independent of cofactors, solely driven by the chemical potential of GM1 and not discriminating between the extra- and intracellular membrane leaflet. We conclude that GLTP mediated non-vesicular lipid trafficking between native membranes is driven by simple thermodynamic principles and that for intracellular transport less than 1 µM GLTP would be required in the cytosol. Furthermore, the data demonstrates the suitability of GLTP as a tool for artificially increasing glycolipid levels in cellular membranes.

  5. Protective actions of the vesicular monoamine transporter 2 (VMAT2) in monoaminergic neurons.

    Science.gov (United States)

    Guillot, Thomas S; Miller, Gary W

    2009-04-01

    Vesicular monoamine transporters (VMATs) are responsible for the packaging of neurotransmitters such as dopamine, serotonin, norepinephrine, and epinephrine into synaptic vesicles. These proteins evolved from precursors in the major facilitator superfamily of transporters and are among the members of the toxin extruding antiporter family. While the primary function of VMATs is to sequester neurotransmitters within vesicles, they can also translocate toxicants away from cytosolic sites of action. In the case of dopamine, this dual role of VMAT2 is combined-dopamine is more readily oxidized in the cytosol where it can cause oxidative stress so packaging into vesicles serves two purposes: neurotransmission and neuroprotection. Furthermore, the deleterious effects of exogenous toxicants on dopamine neurons, such as MPTP, can be attenuated by VMAT2 activity. The active metabolite of MPTP can be kept within vesicles and prevented from disrupting mitochondrial function thereby sparing the dopamine neuron. The highly addictive drug methamphetamine is also neurotoxic to dopamine neurons by using dopamine itself to destroy the axon terminals. Methamphetamine interferes with vesicular sequestration and increases the production of dopamine, escalating the amount in the cytosol and leading to oxidative damage of terminal components. Vesicular transport seems to resist this process by sequestering much of the excess dopamine, which is illustrated by the enhanced methamphetamine neurotoxicity in VMAT2-deficient mice. It is increasingly evident that VMAT2 provides neuroprotection from both endogenous and exogenous toxicants and that while VMAT2 has been adapted by eukaryotes for synaptic transmission, it is derived from phylogenetically ancient proteins that originally evolved for the purpose of cellular protection.

  6. Brain dopamine-serotonin vesicular transport disease presenting as a severe infantile hypotonic parkinsonian disorder.

    Science.gov (United States)

    Jacobsen, Jessie C; Wilson, Callum; Cunningham, Vicki; Glamuzina, Emma; Prosser, Debra O; Love, Donald R; Burgess, Trent; Taylor, Juliet; Swan, Brendan; Hill, Rosamund; Robertson, Stephen P; Snell, Russell G; Lehnert, Klaus

    2016-03-01

    Two male siblings from a consanguineous union presented in early infancy with marked truncal hypotonia, a general paucity of movement, extrapyramidal signs and cognitive delay. By mid-childhood they had made little developmental progress and remained severely hypotonic and bradykinetic. They developed epilepsy and had problems with autonomic dysfunction and oculogyric crises. They had a number of orthopaedic problems secondary to their hypotonia. Cerebrospinal fluid (CSF) neurotransmitters were initially normal, apart from mildly elevated 5-hydroxyindolacetic acid, and the children did not respond favourably to a trial of levodopa-carbidopa. The youngest died from respiratory complications at 10 years of age. Repeat CSF neurotransmitters in the older sibling at eight years of age showed slightly low homovanillic acid and 5-hydroxyindoleacetic acid levels. Whole-exome sequencing revealed a novel mutation homozygous in both children in the monoamine transporter gene SLC18A2 (p.Pro237His), resulting in brain dopamine-serotonin vesicular transport disease. This is the second family to be described with a mutation in this gene. Treatment with the dopamine agonist pramipexole in the surviving child resulted in mild improvements in alertness, communication, and eye movements. This case supports the identification of the causal mutation in the original case, expands the clinical phenotype of brain dopamine-serotonin vesicular transport disease and confirms that pramipexole treatment may lead to symptomatic improvement in affected individuals.

  7. Vesicular Axonal Transport is Modified In Vivo by Tau Deletion or Overexpression in Drosophila

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    Yasmina Talmat-Amar

    2018-03-01

    Full Text Available Structural microtubule associated protein Tau is found in high amount in axons and is involved in several neurodegenerative diseases. Although many studies have highlighted the toxicity of an excess of Tau in neurons, the in vivo understanding of the endogenous role of Tau in axon morphology and physiology is poor. Indeed, knock-out mice display no strong cytoskeleton or axonal transport phenotype, probably because of some important functional redundancy with other microtubule-associated proteins (MAPs. Here, we took advantage of the model organism Drosophila, which genome contains only one homologue of the Tau/MAP2/MAP4 family to decipher (endogenous Tau functions. We found that Tau depletion leads to a decrease in microtubule number and microtubule density within axons, while Tau excess leads to the opposite phenotypes. Analysis of vesicular transport in tau mutants showed altered mobility of vesicles, but no change in the total amount of putatively mobile vesicles, whereas both aspects were affected when Tau was overexpressed. In conclusion, we show that loss of Tau in tau mutants not only leads to a decrease in axonal microtubule density, but also impairs axonal vesicular transport, albeit to a lesser extent compared to the effects of an excess of Tau.

  8. Functional genetic variants in the vesicular monoamine transporter 1 (VMAT1) modulate emotion processing

    Science.gov (United States)

    Lohoff, Falk W.; Hodge, Rachel; Narasimhan, Sneha; Nall, Aleksandra; Ferraro, Thomas N.; Mickey, Brian J.; Heitzeg, Mary M.; Langenecker, Scott A.; Zubieta, Jon-Kar; Bogdan, Ryan; Nikolova, Yuliya S.; Drabant, Emily; Hariri, Ahmad R.; Bevilacqua, Laura; Goldman, David; Doyle, Glenn A.

    2012-01-01

    SUMMARY Emotional behavior is in part heritable and often disrupted in psychopathology. Identification of specific genetic variants that drive this heritability may provide important new insight into molecular and neurobiological mechanisms involved in emotionality. Our results demonstrate that the presynaptic vesicular monoamine transporter 1 (VMAT1) Thr136Ile (rs1390938) polymorphism is functional in vitro, with the Ile allele leading to increased monoamine transport into presynaptic vesicles. Moreover, we show that the Thr136Ile variant predicts differential responses in emotional brain circuits consistent with its effects in vitro. Lastly, deep sequencing of bipolar disorder (BPD) patients and controls identified several rare novel VMAT1 variants. The variant Phe84Ser was only present in individuals with BPD and leads to marked increase monoamine transport in vitro. Taken together, our data show that VMAT1 polymorphisms influence monoamine signaling, the functional response of emotional brain circuits, and risk for psychopathology. PMID:23337945

  9. Regulation of the Dopamine and Vesicular Monoamine Transporters: Pharmacological Targets and Implications for Disease.

    Science.gov (United States)

    German, Christopher L; Baladi, Michelle G; McFadden, Lisa M; Hanson, Glen R; Fleckenstein, Annette E

    2015-10-01

    Dopamine (DA) plays a well recognized role in a variety of physiologic functions such as movement, cognition, mood, and reward. Consequently, many human disorders are due, in part, to dysfunctional dopaminergic systems, including Parkinson's disease, attention deficit hyperactivity disorder, and substance abuse. Drugs that modify the DA system are clinically effective in treating symptoms of these diseases or are involved in their manifestation, implicating DA in their etiology. DA signaling and distribution are primarily modulated by the DA transporter (DAT) and by vesicular monoamine transporter (VMAT)-2, which transport DA into presynaptic terminals and synaptic vesicles, respectively. These transporters are regulated by complex processes such as phosphorylation, protein-protein interactions, and changes in intracellular localization. This review provides an overview of 1) the current understanding of DAT and VMAT2 neurobiology, including discussion of studies ranging from those conducted in vitro to those involving human subjects; 2) the role of these transporters in disease and how these transporters are affected by disease; and 3) and how selected drugs alter the function and expression of these transporters. Understanding the regulatory processes and the pathologic consequences of DAT and VMAT2 dysfunction underlies the evolution of therapeutic development for the treatment of DA-related disorders. Copyright © 2015 by The American Society for Pharmacology and Experimental Therapeutics.

  10. Amyloid-Beta Induced Changes in Vesicular Transport of BDNF in Hippocampal Neurons

    Directory of Open Access Journals (Sweden)

    Bianca Seifert

    2016-01-01

    Full Text Available The neurotrophin brain derived neurotrophic factor (BDNF is an important growth factor in the CNS. Deficits in transport of this secretory protein could underlie neurodegenerative diseases. Investigation of disease-related changes in BDNF transport might provide insights into the cellular mechanism underlying, for example, Alzheimer’s disease (AD. To analyze the role of BDNF transport in AD, live cell imaging of fluorescently labeled BDNF was performed in hippocampal neurons of different AD model systems. BDNF and APP colocalized with low incidence in vesicular structures. Anterograde as well as retrograde transport of BDNF vesicles was reduced and these effects were mediated by factors released from hippocampal neurons into the extracellular medium. Transport of BDNF was altered at a very early time point after onset of human APP expression or after acute amyloid-beta(1-42 treatment, while the activity-dependent release of BDNF remained unaffected. Taken together, extracellular cleavage products of APP induced rapid changes in anterograde and retrograde transport of BDNF-containing vesicles while release of BDNF was unaffected by transgenic expression of mutated APP. These early transport deficits might lead to permanently impaired brain functions in the adult brain.

  11. Orf virus interferes with MHC class I surface expression by targeting vesicular transport and Golgi

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    Rohde Jörg

    2012-07-01

    Full Text Available Abstract Background The Orf virus (ORFV, a zoonotic Parapoxvirus, causes pustular skin lesions in small ruminants (goat and sheep. Intriguingly, ORFV can repeatedly infect its host, despite the induction of a specific immunity. These immune modulating and immune evading properties are still unexplained. Results Here, we describe that ORFV infection of permissive cells impairs the intracellular transport of MHC class I molecules (MHC I as a result of structural disruption and fragmentation of the Golgi apparatus. Depending on the duration of infection, we observed a pronounced co-localization of MHC I and COP-I vesicular structures as well as a reduction of MHC I surface expression of up to 50%. These subversion processes are associated with early ORFV gene expression and are accompanied by disturbed carbohydrate trimming of post-ER MHC I. The MHC I population remaining on the cell surface shows an extended half-life, an effect that might be partially controlled also by late ORFV genes. Conclusions The presented data demonstrate that ORFV down-regulates MHC I surface expression in infected cells by targeting the late vesicular export machinery and the structure and function of the Golgi apparatus, which might aid to escape cellular immune recognition.

  12. Unusual armadillo fold in the human general vesicular transport factor p115.

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    Harald Striegl

    Full Text Available The golgin family gives identity and structure to the Golgi apparatus and is part of a complex protein network at the Golgi membrane. The golgin p115 is targeted by the GTPase Rab1a, contains a large globular head region and a long region of coiled-coil which forms an extended rod-like structure. p115 serves as vesicle tethering factor and plays an important role at different steps of vesicular transport. Here we present the 2.2 A-resolution X-ray structure of the globular head region of p115. The structure exhibits an armadillo fold that is decorated by elongated loops and carries a C-terminal non-canonical repeat. This terminal repeat folds into the armadillo superhelical groove and allows homodimeric association with important implications for p115 mediated multiple protein interactions and tethering.

  13. Age-dependent methamphetamine-induced alterations in vesicular monoamine transporter-2 function: implications for neurotoxicity.

    Science.gov (United States)

    Truong, Jannine G; Wilkins, Diana G; Baudys, Jakub; Crouch, Dennis J; Johnson-Davis, Kamisha L; Gibb, James W; Hanson, Glen R; Fleckenstein, Annette E

    2005-09-01

    Tens of thousands of adolescents and young adults have used illicit methamphetamine. This is of concern since its high-dose administration causes persistent dopaminergic deficits in adult animal models. The effects in adolescents are less studied. In adult rodents, toxic effects of methamphetamine may result partly from aberrant cytosolic dopamine accumulation and subsequent reactive oxygen species formation. The vesicular monoamine transporter-2 (VMAT-2) sequesters cytoplasmic dopamine into synaptic vesicles for storage and perhaps protection against dopamine-associated oxidative consequences. Accordingly, aberrant VMAT-2 function may contribute to the methamphetamine-induced persistent dopaminergic deficits. Hence, this study examined effects of methamphetamine on VMAT-2 in adolescent (postnatal day 40) and young adult (postnatal day 90) rats. Results revealed that high-dose methamphetamine treatment caused greater acute (within 1 h) decreases in vesicular dopamine uptake in postnatal day 90 versus 40 rats, as determined in a nonmembrane-associated subcellular fraction. Greater basal levels of VMAT-2 at postnatal day 90 versus 40 in this purified fraction seemed to contribute to the larger effect. Basal tissue dopamine content was also greater in postnatal day 90 versus 40 rats. In addition, postnatal day 90 rats were more susceptible to methamphetamine-induced persistent dopaminergic deficits as assessed by measuring VMAT-2 activity and dopamine content 7 days after treatment, even if drug doses were adjusted for age-related pharmacokinetic differences. Together, these data demonstrate dynamic changes in VMAT-2 susceptibility to methamphetamine as a function of development. Implications with regard to methamphetamine-induced dopaminergic deficits, as well as dopamine-associated neurodegenerative disorders such as Parkinson's disease, are discussed.

  14. Vesicular transport of progeny parvovirus particles through ER and Golgi regulates maturation and cytolysis.

    Science.gov (United States)

    Bär, Séverine; Rommelaere, Jean; Nüesch, Jürg P F

    2013-09-01

    Progeny particles of non-enveloped lytic parvoviruses were previously shown to be actively transported to the cell periphery through vesicles in a gelsolin-dependent manner. This process involves rearrangement and destruction of actin filaments, while microtubules become protected throughout the infection. Here the focus is on the intracellular egress pathway, as well as its impact on the properties and release of progeny virions. By colocalization with cellular marker proteins and specific modulation of the pathways through over-expression of variant effector genes transduced by recombinant adeno-associated virus vectors, we show that progeny PV particles become engulfed into COPII-vesicles in the endoplasmic reticulum (ER) and are transported through the Golgi to the plasma membrane. Besides known factors like sar1, sec24, rab1, the ERM family proteins, radixin and moesin play (an) essential role(s) in the formation/loading and targeting of virus-containing COPII-vesicles. These proteins also contribute to the transport through ER and Golgi of the well described analogue of cellular proteins, the secreted Gaussia luciferase in absence of virus infection. It is therefore likely that radixin and moesin also serve for a more general function in cellular exocytosis. Finally, parvovirus egress via ER and Golgi appears to be necessary for virions to gain full infectivity through post-assembly modifications (e.g. phosphorylation). While not being absolutely required for cytolysis and progeny virus release, vesicular transport of parvoviruses through ER and Golgi significantly accelerates these processes pointing to a regulatory role of this transport pathway.

  15. Expression of vesicular glutamate transporters in peripheral vestibular structures and vestibular nuclear complex of rat.

    Science.gov (United States)

    Zhang, F X; Pang, Y W; Zhang, M M; Zhang, T; Dong, Y L; Lai, C H; Shum, D K Y; Chan, Y S; Li, J L; Li, Y Q

    2011-01-26

    Glutamate transmission from vestibular end organs to central vestibular nuclear complex (VNC) plays important role in transferring sensory information about head position and movements. Three isoforms of vesicular glutamate transporters (VGLUTs) have been considered so far the most specific markers for glutamatergic neurons/cells. In this study, VGLUT1 and VGLUT2 were immunohistochemically localized to axon terminals in VNC and somata of vestibular primary afferents in association with their central and peripheral axon endings, and VGLUT1 and VGLUT3 were co-localized to hair cells of otolith maculae and cristae ampullaris. VGLUT1 and VGLUT2 defined three subsets of Scarpa's neurons (vestibular ganglionic neurons): those co-expressing VGLUT1 and VGLUT2 or expressing only VGLUT2, and those expressing neither. In addition, many neurons located in all vestibular subnuclei were observed to contain hybridized signals for VGLUT2 mRNA and a few VNC neurons, mostly scattered in medial vestibular nucleus (MVe), displayed VGLUT1 mRNA labelling. Following unilateral ganglionectomy, asymmetries of VGLUT1-immunoreactivity (ir) and VGLUT2-ir occurred between two VNCs, indicating that the VNC terminals containing VGLUT1 and/or VGLUT2 are partly of peripheral origin. The present data indicate that the constituent cells/neurons along the vestibular pathway selectively apply VGLUT isoforms to transport glutamate into synaptic vesicles for glutamate transmission. © 2011 IBRO. Published by Elsevier Ltd. All rights reserved.

  16. Vesicular glutamate transporter-immunoreactivities in the vestibular nuclear complex of rat.

    Science.gov (United States)

    Deng, Jiao; Zhang, Fu-Xing; Pang, You-Wang; Li, Jin-Lian; Li, Yun-Qing

    2006-07-01

    Objective Aims to delineate the distribution profile of three isoforms of vesicular glutamate transporter (VGluT), viz. VGluT1-3, and their cellular localization within vestibular nuclear complex (VNC). Methods Brain sections from normal Sprague-Dawley rats were processed immunohistochemically for VGluT detection, employing avidin-biotinylated peroxidase complex method with 3-3'-diaminobenzidine (DAB) as chromogen. Results The whole VNC expressed all of the three transporters that were observed to be localized to the fiber endings. Compared with VGluT1 and VGluT3, VGluT2 demonstrated a relatively homogeneous distribution, with much higher density in VNC. VGluT3 displayed the highest density in lateral vestibular nucleus and group X, contrasting with the sparse immunostained puncta within vestibular medial and inferior nuclei. Conclusion Glutamtatergic pathways participate in the processing of vestibular signals within VNC mainly through the re-uptake of glutamate into synaptic vesicles by VGluT1 and 2, whereas VGluT3 may play a similar role mainly in areas other than medial and inferior nuclei of VNC.

  17. Vesicular glutamate transporter-immunoreactivities in the vestibular nuclear complex of rat

    Institute of Scientific and Technical Information of China (English)

    Jiao DENG; Fu-Xing ZHANG; You-Wang PANG; Jin-Lian LI; Yun-Qing LI

    2006-01-01

    Objective Aims to delineate the distribution profile of three isoforms of vesicular glutamate transporter (VGluT), viz. VGluT1~3, and their cellular localization within vestibular nuclear complex (VNC). Methods Brain sections from normal Sprague-Dawley rats were processed immunohistochemically for VGluT detection, employing avidinbiotinylated peroxidase complex method with 3-3'-diaminobenzidine (DAB) as chromogen. Results The whole VNC expressed all of the three transporters that were observed to be localized to the fiber endings. Compared with VGluT1 and VGluT3, VGluT2 demonstrated a relatively homogeneous distribution, with much higher density in VNC. VGluT3 displayed the highest density in lateral vestibular nucleus and group X, contrasting with the sparse immunostained puncta within vestibular medial and inferior nuclei. Conclusion Glutamtatergic pathways participate in the processing of vestibular signals within VNC mainly through the re-uptake of glutamate into synaptic vesicles by VGluT1 and 2, whereas VGluT3 may play a similar role mainly in areas other than medial and inferior nuclei of VNC.

  18. Effects of dopaminergic drug treatments on in vivo radioligand binding to brain vesicular monoamine transporters

    Energy Technology Data Exchange (ETDEWEB)

    Kilbourn, Michael R; Frey, Kirk A; Vander Borght, Thierry; Sherman, Phillip S

    1996-05-01

    The effects of various dopaminergic drug treatments on the in vivo regional brain distribution of high-affinity radioligands ([{sup 11}C]dihydrotetrabenazine and [{sup 11}C]methoxytetrabenazine) for the rat brain vesicular monoamine transporter (VMAT2) were determined. Acute treatments with reserpine (2 mg/kg i.p.), tetrabenazine (10 mg/kg i.v.) or related benzoisoquinolines significantly reduced radiotracer binding in vivo. In contrast, radiotracer distributions remained unchanged after treatments with other dopaminergic drugs, whether given by single injection (haloperidol, 1 mg/kg i.p., pargyline 80 mg/kg), repeatedly (pargyline, 80 mg/kg s.c., 14 days), or by continuous infusion (deprenyl, 10 mg/kg/day, 5 days; L-DOPA methyl ester 100 mg/kg/day, 5 days). Repeated injections of tetrabenazine (5 mg/kg i.p., twice daily, 3 days) did not alter in vivo radioligand binding measured after allowing drug washout from the brain. These studies support the proposal that in vivo PET imaging of VMAT2 radioligands in patients with extrapyramidal movement disorders will not be affected by concurrent use of L-DOPA or deprenyl.

  19. Preliminary evidence of apathetic-like behavior in aged vesicular monoamine transporter 2 deficient mice

    Directory of Open Access Journals (Sweden)

    Aron Baumann

    2016-11-01

    Full Text Available Apathy is considered to be a core feature of Parkinson’s disease (PD and has been associated with a variety of states and symptoms of the disease, such as increased severity of motor symptoms, impaired cognition, executive dysfunction, and dementia. Apart from the high prevalence of apathy in PD, which is estimated to be about 40%, the underlying pathophysiology remains poorly understood and current treatment approaches are unspecific and proved to be only partially effective. In animal models, apathy has been sub-optimally modeled, mostly by means of pharmacological and stress-induced methods, whereby concomitant depressive-like symptoms could not be ruled out. In the context of PD only a few studies on toxin-based models (i.e. 6-OHDA or MPTP claimed to have determined apathetic symptoms in animals. The assessment of apathetic symptoms in more elaborated and multifaceted genetic animal models of PD could help to understand the pathophysiological development of apathy in PD and eventually advance specific treatments for afflicted patients. Here we report the presence of behavioral signs of apathy in 12 months old mice that express only ~5% of the vesicular monoamine transporter 2 (VMAT2. Apathetic-like behavior in VMAT2 deficient (LO mice was evidenced by impaired burrowing and nest building skills, and a reduced preference for sweet solution in the saccharin preference test, while the performance in the forced swimming test was normal. Our preliminary results suggest that VMAT2 deficient mice show an apathetic-like phenotype that might be independent of depressive-like symptoms. Therefore VMAT2 LO mice could be a useful tool to study of the pathophysiological substrates of apathy and to test novel treatment strategies for apathy in the context of PD.

  20. Transportomics: screening for substrates of ABC transporters in body fluids using vesicular transport assays.

    NARCIS (Netherlands)

    Krumpochova, P; Sapthu, S.; Brouwers, J.F.H.M.; de Haas, M.; de Vos, R.; Borst, P.; van de Wetering, K.

    2013-01-01

    ABSTRACT The ATP-binding cassette (ABC) genes encode the largest family of transmembrane proteins. ABC transporters translocate a wide variety of substrates across membranes, but their physiological function is often incompletely understood. We describe a new method to study the substrate spectrum

  1. BDNF regulates the expression and distribution of vesicular glutamate transporters in cultured hippocampal neurons.

    Directory of Open Access Journals (Sweden)

    Carlos V Melo

    Full Text Available BDNF is a pro-survival protein involved in neuronal development and synaptic plasticity. BDNF strengthens excitatory synapses and contributes to LTP, presynaptically, through enhancement of glutamate release, and postsynaptically, via phosphorylation of neurotransmitter receptors, modulation of receptor traffic and activation of the translation machinery. We examined whether BDNF upregulated vesicular glutamate receptor (VGLUT 1 and 2 expression, which would partly account for the increased glutamate release in LTP. Cultured rat hippocampal neurons were incubated with 100 ng/ml BDNF, for different periods of time, and VGLUT gene and protein expression were assessed by real-time PCR and immunoblotting, respectively. At DIV7, exogenous application of BDNF rapidly increased VGLUT2 mRNA and protein levels, in a dose-dependent manner. VGLUT1 expression also increased but only transiently. However, at DIV14, BDNF stably increased VGLUT1 expression, whilst VGLUT2 levels remained low. Transcription inhibition with actinomycin-D or α-amanitine, and translation inhibition with emetine or anisomycin, fully blocked BDNF-induced VGLUT upregulation. Fluorescence microscopy imaging showed that BDNF stimulation upregulates the number, integrated density and intensity of VGLUT1 and VGLUT2 puncta in neurites of cultured hippocampal neurons (DIV7, indicating that the neurotrophin also affects the subcellular distribution of the transporter in developing neurons. Increased VGLUT1 somatic signals were also found 3 h after stimulation with BDNF, further suggesting an increased de novo transcription and translation. BDNF regulation of VGLUT expression was specifically mediated by BDNF, as no effect was found upon application of IGF-1 or bFGF, which activate other receptor tyrosine kinases. Moreover, inhibition of TrkB receptors with K252a and PLCγ signaling with U-73122 precluded BDNF-induced VGLUT upregulation. Hippocampal neurons express both isoforms during

  2. A glial variant of the vesicular monoamine transporter is required to store histamine in the Drosophila visual system.

    Directory of Open Access Journals (Sweden)

    Rafael Romero-Calderón

    2008-11-01

    Full Text Available Unlike other monoamine neurotransmitters, the mechanism by which the brain's histamine content is regulated remains unclear. In mammals, vesicular monoamine transporters (VMATs are expressed exclusively in neurons and mediate the storage of histamine and other monoamines. We have studied the visual system of Drosophila melanogaster in which histamine is the primary neurotransmitter released from photoreceptor cells. We report here that a novel mRNA splice variant of Drosophila VMAT (DVMAT-B is expressed not in neurons but rather in a small subset of glia in the lamina of the fly's optic lobe. Histamine contents are reduced by mutation of dVMAT, but can be partially restored by specifically expressing DVMAT-B in glia. Our results suggest a novel role for a monoamine transporter in glia that may be relevant to histamine homeostasis in other systems.

  3. Ion Transport in Human Pancreatic Duct Epithelium, Capan-1 Cells, Is Regulated by Secretin, VIP, Acetylcholine, and Purinergic Receptors

    DEFF Research Database (Denmark)

    Wang, Jing; Novak, Ivana

    2013-01-01

    OBJECTIVES: The objective of the study was to establish a solid model of polarized epithelium for human pancreatic ducts, where electrical parameters could be measured as indicators of ion transport. Further, we aimed to determine functional expression of several receptors, in particular, puriner...... transport in human pancreatic duct epithelium, Capan-1 cells, is regulated by secretin, VIP, acetylcholine, adenosine, and purinergic P2 receptors; and this human model has a good potential for studies of physiology and pathophysiology of pancreatic duct ion transport....

  4. Actin-based vesicular transport in the first 20 min after dusk is crucial for daily rhabdom synthesis in the compound eye of the grapsid crab Hemigrapsus sanguineus.

    Science.gov (United States)

    Matsushita, A; Arikawa, K

    1997-09-01

    In the crab Hemigrapsus sanguineus, maintained under a 12 h:12 h light:dark cycle, the amount of vesicular smooth endoplasmic reticulum (vesicular sER) in the photoreceptor cell body increases after the light is turned off. This paper demonstrates that actin filaments in the photoreceptor cell body are involved in the transport of vesicular sER towards the rhabdom. To specify the time of actin contribution to rhabdom synthesis, we disrupted the organization of actin filaments in the cell body with cytochalasin D at various time around dusk. We then measured the rhabdom size and also examined the ultrastructure of the photoreceptor cell body 3 h after extinguishing the light. When cytochalasin D was applied from either 1 h before or immediately after extinguishing the light, the rhabdom size did not increase, whereas vesicular sER accumulated in the cell body. In contrast, cytochalasin D applied to the eyes from 20 min after turning the light off did not inhibit rhabdom synthesis. These results indicate that the first 20 min after the light is turned off is particularly important for the transport of vesicular sER towards the rhabdom by the cell body actin filaments.

  5. Control of synthesis and release of radioactive acetylcholine in brain slices from the rat. Effects of neurotropic drugs

    Science.gov (United States)

    Grewaal, D. S.; Quastel, J. H.

    1973-01-01

    1. Studies of the synthesis and release of radioactive acetylcholine in rat brain-cortex slices incubated in Locke–bicarbonate–[U-14C]glucose media, containing paraoxon as cholinesterase inhibitor, revealed the following phenomena: (a) dependence of K+-or protoveratrine-stimulated acetylcholine synthesis and release on the presence of Na+ and Ca2+ in the incubation medium, (b) enhanced release of radioactive acetylcholine by substances that promote depolarization at the nerve cell membrane (e.g. high K+, ouabain, protoveratrine, sodium l-glutamate, high concentration of acetylcholine), (c) failure of acetylcholine synthesis to keep pace with acetylcholine release under certain conditions (e.g. the presence of ouabain or lack of Na+). 2. Stimulation by K+ of radioactive acetylcholine synthesis was directly proportional to the external concentration of Na+, but some synthesis and release of radioactive acetylcholine occurred in the absence of Na+ as well as in the absence of Ca2+. 3. The Na+ dependence of K+-stimulated acetylcholine synthesis was partly due to suppression of choline transport, as addition of small concentrations of choline partly neutralized the effect of Na+ lack, and partly due to the suppression of the activity of the Na+ pump. 4. Protoveratrine caused a greatly increased release of radioactive acetylcholine without stimulating total radioactive acetylcholine synthesis. Protoveratrine was ineffective in the absence of Ca2+ from the incubation medium. It completely blocked K+ stimulation of acetylcholine synthesis and release. 5. Tetrodotoxin abolished the effects of protoveratrine on acetylcholine release. It had blocking effects (partial or complete) on the action of high K+, sodium l-glutamate and lack of Ca2+ on acetylcholine synthesis and release. 6. Unlabelled exogenous acetylcholine did not diminish the content of labelled tissue acetylcholine, derived from labelled glucose, suggesting that no exchange with vesicular acetylcholine took

  6. The Effect of Albumin on MRP2 and BCRP in the Vesicular Transport Assay.

    Directory of Open Access Journals (Sweden)

    Feng Deng

    Full Text Available The ABC transporters multidrug resistance associated protein 2 (MRP2 and breast cancer resistance protein (BCRP are of interest in drug development, since they affect the pharmacokinetics of several drugs. Membrane vesicle transport assays are widely used to study interactions with these proteins. Since albumin has been found to affect the kinetics of metabolic enzymes in similar membrane preparations, we investigated whether albumin affects the kinetic parameters of efflux transport. We found that albumin increased the Vmax of 5(6-carboxy-2',7'-dichlorofluorescein (CDCF and estradiol-17-β-D-glucuronide uptake into MRP2 vesicles in the presence of 0.1% bovine serum albumin (BSA by 2 and 1.5-fold, respectively, while BSA increased Lucifer yellow uptake by 30% in BCRP vesicles. Km values increased slightly, but the change was not statistically significant. The effect of BSA on substrate uptake was dependent on the vesicle amount, while increasing BSA concentration did not significantly improve substrate uptake. These results indicate a minor effect of albumin on MRP2 and BCRP, but it should be considered if albumin is added to transporter assays for example as a solubilizer, since the effect may be substrate or transporter specific.

  7. Hyphal transport by a vesicular-arbuscular mycorrhizal fungus of N applied to the soil as ammonium or nitrate

    DEFF Research Database (Denmark)

    Johansen, A.; Jakobsen, I.; Jensen, E.S.

    1993-01-01

    Transport of N by hyphae of a vesicular-arbuscular mycorrhizal fungus was studied under controlled experimental conditions. The N source was applied to the soil as (NH4+)-N-15 or (NH3-)N-15. Cucumis sativus was grown for 25 days, either alone or in symbiosis with Glomus intraradices, in containers...... with a hyphal compartment separated from the root compartment by a fine nylon mesh. Mineral N was then applied to the hyphal compartment as (NH4+)-N-15 or (NO3-)-N-15 at 5 cm distance from the root compartment. Soil samples were taken from the hyphal compartment at 1, 3 and 5 cm distance from the root...... compartment at 7 and 12 days after labelling, and the concentration of mineral N in the samples was measured from 2 M KCl extracts. Mycorrhizal colonization did not affect plant dry weight. The recovery of N-15 in mycorrhizal plants was 38 or 40%, respectively, when (NH4+)-N-15 or (NO3-)-N-15 was applied...

  8. Expression profile of vesicular nucleotide transporter (VNUT, SLC17A9) in subpopulations of rat dorsal root ganglion neurons.

    Science.gov (United States)

    Nishida, Kentaro; Nomura, Yuka; Kawamori, Kanako; Moriyama, Yoshinori; Nagasawa, Kazuki

    2014-09-05

    ATP plays an important role in the signal transduction between sensory neurons and satellite cells in dorsal root ganglia (DRGs). In primary cultured DRG neurons, ATP is known to be stored in lysosomes via a vesicular nucleotide transporter (VNUT), and to be released into the intercellular space through exocytosis. DRGs consist of large-, medium- and small-sized neurons, which play different roles in sensory transmission, but there is no information on the expression profiles of VNUT in DRG subpopulations. Here, we obtained detailed expression profiles of VNUT in isolated rat DRG tissues. On immunohistochemical analysis, VNUT was found in DRG neurons, and was predominantly expressed by the small- and medium-sized DRG ones, as judged upon visual inspection, and this was compatible with the finding that the number of VNUT-positive DRG neurons in IB4-positive cells was greater than that in NF200-positive ones. These results suggest that VNUT play a role in ATP accumulation in DRG neurons, especially in small- and medium-sized ones, and might be involved in ATP-mediated nociceptive signaling in DRGs. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  9. How molecular motors are arranged on a cargo is important for vesicular transport.

    Directory of Open Access Journals (Sweden)

    Robert P Erickson

    2011-05-01

    Full Text Available The spatial organization of the cell depends upon intracellular trafficking of cargos hauled along microtubules and actin filaments by the molecular motor proteins kinesin, dynein, and myosin. Although much is known about how single motors function, there is significant evidence that cargos in vivo are carried by multiple motors. While some aspects of multiple motor function have received attention, how the cargo itself--and motor organization on the cargo--affects transport has not been considered. To address this, we have developed a three-dimensional Monte Carlo simulation of motors transporting a spherical cargo, subject to thermal fluctuations that produce both rotational and translational diffusion. We found that these fluctuations could exert a load on the motor(s, significantly decreasing the mean travel distance and velocity of large cargos, especially at large viscosities. In addition, the presence of the cargo could dramatically help the motor to bind productively to the microtubule: the relatively slow translational and rotational diffusion of moderately sized cargos gave the motors ample opportunity to bind to a microtubule before the motor/cargo ensemble diffuses out of range of that microtubule. For rapidly diffusing cargos, the probability of their binding to a microtubule was high if there were nearby microtubules that they could easily reach by translational diffusion. Our simulations found that one reason why motors may be approximately 100 nm long is to improve their 'on' rates when attached to comparably sized cargos. Finally, our results suggested that to efficiently regulate the number of active motors, motors should be clustered together rather than spread randomly over the surface of the cargo. While our simulation uses the specific parameters for kinesin, these effects result from generic properties of the motors, cargos, and filaments, so they should apply to other motors as well.

  10. Spreading of a prion domain from cell-to-cell by vesicular transport in Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Carmen I Nussbaum-Krammer

    2013-03-01

    Full Text Available Prion proteins can adopt self-propagating alternative conformations that account for the infectious nature of transmissible spongiform encephalopathies (TSEs and the epigenetic inheritance of certain traits in yeast. Recent evidence suggests a similar propagation of misfolded proteins in the spreading of pathology of neurodegenerative diseases including Alzheimer's or Parkinson's disease. Currently there is only a limited number of animal model systems available to study the mechanisms that underlie the cell-to-cell transmission of aggregation-prone proteins. Here, we have established a new metazoan model in Caenorhabditis elegans expressing the prion domain NM of the cytosolic yeast prion protein Sup35, in which aggregation and toxicity are dependent upon the length of oligopeptide repeats in the glutamine/asparagine (Q/N-rich N-terminus. NM forms multiple classes of highly toxic aggregate species and co-localizes to autophagy-related vesicles that transport the prion domain from the site of expression to adjacent tissues. This is associated with a profound cell autonomous and cell non-autonomous disruption of mitochondrial integrity, embryonic and larval arrest, developmental delay, widespread tissue defects, and loss of organismal proteostasis. Our results reveal that the Sup35 prion domain exhibits prion-like properties when expressed in the multicellular organism C. elegans and adapts to different requirements for propagation that involve the autophagy-lysosome pathway to transmit cytosolic aggregation-prone proteins between tissues.

  11. Activity-dependent formation of a vesicular inhibitory amino acid transporter gradient in the superior olivary complex of NMRI mice.

    Science.gov (United States)

    Ebbers, Lena; Weber, Maren; Nothwang, Hans Gerd

    2017-10-26

    In the mammalian superior olivary complex (SOC), synaptic inhibition contributes to the processing of binaural sound cues important for sound localization. Previous analyses demonstrated a tonotopic gradient for postsynaptic proteins mediating inhibitory neurotransmission in the lateral superior olive (LSO), a major nucleus of the SOC. To probe, whether a presynaptic molecular gradient exists as well, we investigated immunoreactivity against the vesicular inhibitory amino acid transporter (VIAAT) in the mouse auditory brainstem. Immunoreactivity against VIAAT revealed a gradient in the LSO and the superior paraolivary nucleus (SPN) of NMRI mice, with high expression in the lateral, low frequency processing limb and low expression in the medial, high frequency processing limb of both nuclei. This orientation is opposite to the previously reported gradient of glycine receptors in the LSO. Other nuclei of the SOC showed a uniform distribution of VIAAT-immunoreactivity. No gradient was observed for the glycine transporter GlyT2 and the neuronal protein NeuN. Formation of the VIAAT gradient was developmentally regulated and occurred around hearing-onset between postnatal days 8 and 16. Congenital deaf Claudin14 -/- mice bred on an NMRI background showed a uniform VIAAT-immunoreactivity in the LSO, whereas cochlear ablation in NMRI mice after hearing-onset did not affect the gradient. Additional analysis of C57Bl6/J, 129/SvJ and CBA/J mice revealed a strain-specific formation of the gradient. Our results identify an activity-regulated gradient of VIAAT in the SOC of NRMI mice. Its absence in other mouse strains adds a novel layer of strain-specific features in the auditory system, i.e. tonotopic organization of molecular gradients. This calls for caution when comparing data from different mouse strains frequently used in studies involving transgenic animals. The presence of strain-specific differences offers the possibility of genetic mapping to identify molecular

  12. Nicotinic acetylcholine receptor: subunit structure, functional binding sites, and ion transport properties

    International Nuclear Information System (INIS)

    Raftery, M.A.; Dunn, S.M.J.; Conti-Tronconi, B.M.; Middlemas, D.S.; Crawford, R.D.

    1983-01-01

    The structure of the nicotinic acetylcholine receptor has been highly conserved during animal evolution, and in all the species and tissues studied so far, including mammals, it is a pseudosymmetric, pentameric complex of related subunits with very similar physical properties. All subunits of these nicotinic receptors were derived from a common ancestral gene, probably by way of gene duplications occurring very early in animal evolution. 45 refs., 8 figs., 2 tabs

  13. Binding characteristics of 9-fluoropropyl-(+)-dihydrotetrabenzazine (AV-133) to the vesicular monoamine transporter type 2 in rats

    Energy Technology Data Exchange (ETDEWEB)

    Tsao, H.-H. [Department of Medical Imaging and Radiological Sciences, Chang Gung University, Taoyuan, Taiwan (China); Lin, K.-J. [Department of Medical Imaging and Radiological Sciences, Chang Gung University, Taoyuan, Taiwan (China); Department of Nuclear Medicine, Chang Gung University and Memorial Hospital, Taoyuan, Taiwan (China); Juang, J.-H. [Division of Endocrinology and Metabolism, Chung Gung University and Chung Gung Memorial Hospital, Taoyuan, Taiwan (China); Skovronsky, Daniel M. [Avid Radiopharmaceuticals, Philadelphia, PA (United States); Department of Radiology, University of Pennsylvania, Philadelphia, PA 19104 (United States); Yen, T.-C. [Department of Nuclear Medicine, Chang Gung University and Memorial Hospital, Taoyuan, Taiwan (China); Wey, S.-P. [Department of Medical Imaging and Radiological Sciences, Chang Gung University, Taoyuan, Taiwan (China); Kung, M.-P. [Department of Medical Imaging and Radiological Sciences, Chang Gung University, Taoyuan, Taiwan (China); Department of Radiology, University of Pennsylvania, Philadelphia, PA 19104 (United States)], E-mail: kungmp@sunmac.spect.upenn.edu

    2010-05-15

    The vesicular monoamine transporter type 2 (VMAT2) is highly expressed in pancreatic {beta}-cells and thus has been proposed to be a potential target for measuring {beta}-cell mass (BCM) by molecular imaging. C-11- and F-18-labeled tetrabenazine derivatives targeting VMAT2 have shown some promising results as potential biomarkers for BCM. In the present study, we examined the binding characteristics of 9-fluoropropyl-(+)-dihydrotetrabenzazine ([{sup 18}F]AV-133), a potential PET tracer for BCM imaging, in rat pancreas and rat brain. Methods: Pancreatic exocrine cells and pancreatic islet cells were isolated and purified from Sprague-Dawley rats. Membrane homogenates, prepared from both pancreatic exocrine and islet cells as well as from brain striatum and hypothalamus regions, were used for in vitro binding studies. In vitro and ex vivo autoradiography studies with [{sup 18}F]AV-133 were performed on rat brain and rat pancreas sections. Immunohistochemistry studies were performed to confirm the distribution of VMAT2 on islet {beta}-cells. Results: Excellent binding affinities of [{sup 18}F]AV-133 were observed in rat striatum and hypothalamus homogenates with K{sub d} values of 0.19 and 0.25 nM, respectively. In contrast to single-site binding observed in rat striatum homogenates, rat islet cell homogenates showed two saturable binding sites (site A: K{sub d}=6.76 nM, B{sub max}=60 fmol/mg protein; site B: K{sub d}=241 nM, B{sub max}=1500 fmol/mg protein). Rat exocrine pancreas homogenates showed only a single low-affinity binding site (K{sub d}=209 nM), which was similar to site B in islet cells. In vitro autoradiography of [{sup 18}F]AV-133 using frozen sections of rat pancreas showed specific labeling of islets, as evidenced by co-localization with anti-insulin antibody. Ex vivo VMAT2 pancreatic autoradiography in the rat, however, was not successful, in contrast to the excellent ex vivo autoradiography of VMAT2 binding sites in the brain. In vivo/ex vivo islet

  14. In vivo imaging of vesicular monoamine transporter 2 in pancreas using an {sup 18}F epoxide derivative of tetrabenazine

    Energy Technology Data Exchange (ETDEWEB)

    Kung, Hank F. [Department of Radiology, University of Pennsylvania, Philadelphia, PA 19104 (United States); Department of Pharmacology, University of Pennsylvania, Philadelphia, PA 19104 (United States)], E-mail: kunghf@sunmac.spect.upenn.edu; Lieberman, Brian P. [Department of Radiology, University of Pennsylvania, Philadelphia, PA 19104 (United States); Zhuang Zhiping [Avid Radiopharmaceuticals, Inc., Philadelphia, PA 19104 (United States); Oya, Shunichi; Kung Meiping [Department of Radiology, University of Pennsylvania, Philadelphia, PA 19104 (United States); Choi, Seok Rye [Avid Radiopharmaceuticals, Inc., Philadelphia, PA 19104 (United States); Poessl, Karl; Blankemeyer, Eric; Hou, Catherine [Department of Radiology, University of Pennsylvania, Philadelphia, PA 19104 (United States); Skovronsky, Daniel [Avid Radiopharmaceuticals, Inc., Philadelphia, PA 19104 (United States); Kilbourn, Michael [Department of Radiology, University of Michigan, Ann Arbor, MI 48109 (United States)

    2008-11-15

    Objectives: Development of imaging agents for pancreatic beta cell mass may provide tools for studying insulin-secreting beta cells and their relationship with diabetes mellitus. In this paper, a new imaging agent, [{sup 18}F](+)-2-oxiranyl-3-isobutyl-9-(3-fluoropropoxy)-10-methoxy-2,3,4,6,7, 11b-hexahydro-1H-pyrido[2,1-a]isoquinoline [{sup 18}F](+)4, which displays properties targeting vesicular monoamine transporter 2 (VMAT2) binding sites of beta cells in the pancreas, was evaluated as a positron emission tomography (PET) agent for estimating beta cell mass in vivo. The hydrolyzable epoxide group of (+)4 may provide a mechanism for shifting biodistribution from liver to kidney, thus reducing the background signal. Methods: Both {sup 18}F- and {sup 19}F-labeled (+) and (-) isomers of 4 were synthesized and evaluated. Organ distribution was carried out in normal rats. Uptake of [{sup 18}F](+)4 in pancreas of normal rats was measured and correlated with blocking studies using competing drugs, (+)dihydrotetrabenazine [(+)-DTBZ] or 9-fluoropropyl-(+)dihydro tetrabenazine [FP-(+)-DTBZ, (+)2]. Results: In vitro binding study of VMAT2 using rat brain striatum showed a K{sub i} value of 0.08 and 0.15 nM for the (+)4 and ({+-})4, respectively. The in vivo biodistribution of [{sup 18}F](+)4 in rats showed the highest uptake in the pancreas (2.68 %ID/g at 60 min postinjection). In vivo competition experiments with cold FP-(+)-DTBZ, (+)2, (3.5 mg/kg, 5 min iv pretreatment) led to a significant reduction of pancreas uptake (85% blockade at 60 min). The inactive isomer [{sup 18}F](-)4 showed significantly lower pancreas uptake (0.22 %ID/g at 30 min postinjection). Animal PET imaging studies of [{sup 18}F](+)4 in normal rats demonstrated an avid pancreatic uptake in rats. Conclusion: The preliminary results suggest that the epoxide, [{sup 18}F](+)4, is highly selective in binding to VMAT2 and it has an excellent uptake in the pancreas of rats. The liver uptake was significantly

  15. Vesicular transport route of horseradish C1a peroxidase is regulated by N- and C-terminal propeptides in tobacco cells.

    Science.gov (United States)

    Matsui, T; Nakayama, H; Yoshida, K; Shinmyo, A

    2003-10-01

    Peroxidases (PRX, EC 1.11.1.7) are widely distributed across microorganisms, plants, and animals; and, in plants, they have been implicated in a variety of secondary metabolic reactions. In particular, horseradish (Armoracia rusticana) root represents the main source of commercial PRX production. The prxC1a gene, which encodes horseradish PRX (HRP) C, is expressed mainly in the roots and stems of the horseradish plant. HRP C1a protein is shown to be synthesized as a preprotein with both a N-terminal (NTPP) and a C-terminal propeptide (CTPP). These propeptides, which might be responsible for intracellular localization or secretion, are removed before or concomitant with production of the mature protein. We investigated the functional role of HRP C1a NTPP and CTPP in the determination of the vesicular transport route, using an analytical system of transgenically cultured tobacco cells (Nicotiana tabacum, BY2). Here, we report that NTPP and CTPP are necessary and sufficient for accurate localization of mature HRP C1a protein to vacuoles of the vesicular transport system. We also demonstrate that HRP C1a derived from a preprotein lacking CTPP is shunted into the secretory pathway.

  16. Pathways of acetylcholine synthesis, transport and release as targets for treatment of adult-onset cognitive dysfunction.

    Science.gov (United States)

    Amenta, F; Tayebati, S K

    2008-01-01

    Acetylcholine (ACh) is a neurotransmitter widely diffused in central, peripheral, autonomic and enteric nervous system. This paper has reviewed the main mechanisms of ACh synthesis, storage, and release. Presynaptic choline transport supports ACh production and release, and cholinergic terminals express a unique transporter critical for neurotransmitter release. Neurons cannot synthesize choline, which is ultimately derived from the diet and is delivered through the blood stream. ACh released from cholinergic synapses is hydrolyzed by acetylcholinesterase into choline and acetyl coenzyme A and almost 50% of choline derived from ACh hydrolysis is recovered by a high-affinity choline transporter. Parallel with the development of cholinergic hypothesis of geriatric memory dysfunction, cholinergic precursor loading strategy was tried for treating cognitive impairment occurring in Alzheimer's disease. Controlled clinical studies denied clinical usefulness of choline and lecithin (phosphatidylcholine), whereas for other phospholipids involved in choline biosynthetic pathways such as cytidine 5'-diphosphocholine (CDP-choline) or alpha-glyceryl-phosphorylcholine (choline alphoscerate) a modest improvement of cognitive dysfunction in adult-onset dementia disorders is documented. These inconsistencies have probably a metabolic explanation. Free choline administration increases brain choline availability but it does not increase ACh synthesis/or release. Cholinergic precursors to serve for ACh biosynthesis should be incorporate and stored into phospholipids in brain. It is probable that appropriate ACh precursors and other correlated molecules (natural or synthesized) could represent a tool for developing therapeutic strategies by revisiting and updating treatments/supplementations coming out from this therapeutic stalemate.

  17. Fluoroethoxy-1,4-diphenethylpiperidine and piperazine derivatives: Potent and selective inhibitors of [3H]dopamine uptake at the vesicular monoamine transporter-2.

    Science.gov (United States)

    Hankosky, Emily R; Joolakanti, Shyam R; Nickell, Justin R; Janganati, Venumadhav; Dwoskin, Linda P; Crooks, Peter A

    2017-12-15

    A small library of fluoroethoxy-1,4-diphenethyl piperidine and fluoroethoxy-1,4-diphenethyl piperazine derivatives were designed, synthesized and evaluated for their ability to inhibit [ 3 H]dopamine (DA) uptake at the vesicular monoamine transporter-2 (VMAT2) and dopamine transporter (DAT), [ 3 H]serotonin (5-HT) uptake at the serotonin transporter (SERT), and [ 3 H]dofetilide binding at the human-ether-a-go-go-related gene (hERG) channel. The majority of the compounds exhibited potent inhibition of [ 3 H]DA uptake at VMAT2, Ki changes in the nanomolar range (K i  = 0.014-0.073 µM). Compound 15d exhibited the highest affinity (K i  = 0.014 µM) at VMAT2, and had 160-, 5-, and 60-fold greater selectivity for VMAT2 vs. DAT, SERT and hERG, respectively. Compound 15b exhibited the greatest selectivity (>60-fold) for VMAT2 relative to all the other targets evaluated, and 15b had high affinity for VMAT2 (K i  = 0.073 µM). Compound 15b was considered the lead compound from this analog series due to its high affinity and selectivity for VMAT2. Copyright © 2017 Elsevier Ltd. All rights reserved.

  18. Essential roles of aspartate aminotransferase 1 and vesicular glutamate transporters in β-cell glutamate signaling for incretin-induced insulin secretion.

    Directory of Open Access Journals (Sweden)

    Naoya Murao

    Full Text Available Incretins (GLP-1 and GIP potentiate insulin secretion through cAMP signaling in pancreatic β-cells in a glucose-dependent manner. We recently proposed a mechanistic model of incretin-induced insulin secretion (IIIS that requires two critical processes: 1 generation of cytosolic glutamate through the malate-aspartate (MA shuttle in glucose metabolism and 2 glutamate transport into insulin granules by cAMP signaling to promote insulin granule exocytosis. To directly prove the model, we have established and characterized CRISPR/Cas9-engineered clonal mouse β-cell lines deficient for the genes critical in these two processes: aspartate aminotransferase 1 (AST1, gene symbol Got1, a key enzyme in the MA shuttle, which generates cytosolic glutamate, and the vesicular glutamate transporters (VGLUT1, VGLUT2, and VGLUT3, gene symbol Slc17a7, Slc17a6, and Slc17a8, respectively, which participate in glutamate transport into secretory vesicles. Got1 knockout (KO β-cell lines were defective in cytosolic glutamate production from glucose and showed impaired IIIS. Unexpectedly, different from the previous finding that global Slc17a7 KO mice exhibited impaired IIIS from pancreatic islets, β-cell specific Slc17a7 KO mice showed no significant impairment in IIIS, as assessed by pancreas perfusion experiment. Single Slc17a7 KO β-cell lines also retained IIIS, probably due to compensatory upregulation of Slc17a6. Interestingly, triple KO of Slc17a7, Slc17a6, and Slc17a8 diminished IIIS, which was rescued by exogenously introduced wild-type Slc17a7 or Slc17a6 genes. The present study provides direct evidence for the essential roles of AST1 and VGLUTs in β-cell glutamate signaling for IIIS and also shows the usefulness of the CRISPR/Cas9 system for studying β-cells by simultaneous disruption of multiple genes.

  19. Decreased expression of vesicular glutamate transporter 1 and complexin II mRNAs in schizophrenia: further evidence for a synaptic pathology affecting glutamate neurons.

    Science.gov (United States)

    Eastwood, S L; Harrison, P J

    2005-03-01

    Synaptic protein gene expression is altered in schizophrenia. In the hippocampal formation there may be particular involvement of glutamatergic neurons and their synapses, but overall the profile remains unclear. In this in situ hybridization histochemistry (ISHH) study, we examined four informative synaptic protein transcripts: vesicular glutamate transporter (VGLUT) 1, VGLUT2, complexin I, and complexin II, in dorsolateral prefrontal cortex (DPFC), superior temporal cortex (STC), and hippocampal formation, in 13 subjects with schizophrenia and 18 controls. In these areas, VGLUT1 and complexin II are expressed primarily by excitatory neurons, whereas complexin I is mainly expressed by inhibitory neurons. In schizophrenia, VGLUT1 mRNA was decreased in hippocampal formation and DPFC, complexin II mRNA was reduced in DPFC and STC, and complexin I mRNA decreased in STC. Hippocampal VGLUT1 mRNA declined with age selectively in the schizophrenia group. VGLUT2 mRNA was not quantifiable due to its low level. The data provide additional evidence for a synaptic pathology in schizophrenia, in terms of a reduced expression of three synaptic protein genes. In the hippocampus, the loss of VGLUT1 mRNA supports data indicating that glutamatergic presynaptic deficits are prominent, whereas the pattern of results in temporal and frontal cortex suggests broadly similar changes may affect inhibitory and excitatory neurons. The impairment of synaptic transmission implied by the synaptic protein reductions may contribute to the dysfunction of cortical neural circuits that characterises the disorder.

  20. Vesicular monoamine transporter protein expression correlates with clinical features, tumor biology, and MIBG avidity in neuroblastoma: a report from the Children's Oncology Group

    International Nuclear Information System (INIS)

    Temple, William; Mendelsohn, Lori; Nekritz, Erin; Gustafson, W.C.; Matthay, Katherine K.; Kim, Grace E.; Lin, Lawrence; Giacomini, Kathy; Naranjo, Arlene; Van Ryn, Collin; Yanik, Gregory A.; Kreissman, Susan G.; Hogarty, Michael; DuBois, Steven G.

    2016-01-01

    Vesicular monoamine transporters 1 and 2 (VMAT1 and VMAT2) are thought to mediate MIBG uptake in adult neuroendocrine tumors. In neuroblastoma, the norepinephrine transporter (NET) has been investigated as the principal MIBG uptake protein, though some tumors without NET expression concentrate MIBG. We investigated VMAT expression in neuroblastoma and correlated expression with MIBG uptake and clinical features. We evaluated VMAT1 and VMAT2 expression by immunohistochemistry (IHC) in neuroblastoma tumors from 76 patients with high-risk metastatic disease treated in a uniform cooperative group trial (COG A3973). All patients had baseline MIBG diagnostic scans centrally reviewed. IHC results were scored as the product of intensity grading (0 - 3+) and percent of tumor cells expressing the protein of interest. The association between VMAT1 and VMAT2 scores and clinical and biological features was tested using Wilcoxon rank-sum tests. Patient characteristics were typical of high-risk neuroblastoma, though the cohort was intentionally enriched in patients with MIBG-nonavid tumors (n = 20). VMAT1 and VMAT2 were expressed in 62 % and 75 % of neuroblastoma tumors, respectively. VMAT1 and VMAT2 scores were both significantly lower in MYCN amplified tumors and in tumors with high mitotic karyorrhectic index. MIBG-avid tumors had significantly higher VMAT2 scores than MIBG-nonavid tumors (median 216 vs. 45; p = 0.04). VMAT1 expression did not correlate with MIBG avidity. VMAT1 and VMAT2 are expressed in the majority of neuroblastomas. Expression correlates with other biological features. The expression level of VMAT2 but not that of VMAT1 correlates with avidity for MIBG. (orig.)

  1. Patchy distributions of myelin and vesicular glutamate transporter 2 align with cytochrome oxidase blobs and interblobs in the superficial layers of the primary visual cortex

    Directory of Open Access Journals (Sweden)

    Rockoff EC

    2014-09-01

    Full Text Available Emily C Rockoff,1 Pooja Balaram,1 Jon H Kaas1,2 1Department of Psychology, 2Department of Cell and Molecular Biology, Vanderbilt University, Nashville, TN, USA Abstract: Blobs are a modular component of the primary visual cortex (area 17 of all primates, but not of other mammals closely related to primates. They are characterized as an even distribution of patches, puffs, or blobs of dense cytochrome oxidase (CO expression in layer III of area 17, and are now known to differ from surrounding, nonblob cortex in thalamic, intrinsic, and extrastriate connections. Previous studies have also recognized a blob-like pattern of myelin-dense patches in layer III of area 17 of primates, and more recently the vesicular glutamate transporter (VGLUT-2 isoform of the VGLUT family has been found to selectively distribute to layer III patches in a similar blob-like pattern. Here, we sought to determine if the blob-like patterns all identify the same modular structures in area 17 of primates by staining alternate brain sections cut parallel to the surface of area 17 of a prosimian primate (Otolemur garnettii for CO, myelin, and VGLUT2. By aligning the sections from the three preparations, we provide clear evidence that the three preparations all identify the same modular blob structures. The results provide a further understanding of the functional nature of the blobs by demonstrating that their higher level of CO activity is related to thalamic inputs from the lateral geniculate nucleus that use VGLUT2 as their main glutamate transporter, and via myelinated axons. Keywords: columns, modules, visual cortex, primates, prosimians

  2. Vesicular monoamine transporter protein expression correlates with clinical features, tumor biology, and MIBG avidity in neuroblastoma: a report from the Children's Oncology Group

    Energy Technology Data Exchange (ETDEWEB)

    Temple, William; Mendelsohn, Lori; Nekritz, Erin; Gustafson, W.C.; Matthay, Katherine K. [UCSF School of Medicine, Department of Pediatrics, San Francisco, CA (United States); UCSF Benioff Children' s Hospital, San Francisco, CA (United States); Kim, Grace E. [UCSF School of Medicine, Department of Pathology, San Francisco, CA (United States); Lin, Lawrence; Giacomini, Kathy [UCSF School of Pharmacy, Department of Bioengineering and Therapeutic Sciences, San Francisco, CA (United States); Naranjo, Arlene; Van Ryn, Collin [University of Florida, Children' s Oncology Group Statistics and Data Center, Gainesville, FL (United States); Yanik, Gregory A. [University of Michigan, CS Mott Children' s Hospital, Ann Arbor, MI (United States); Kreissman, Susan G. [Duke University Medical Center, Durham, NC (United States); Hogarty, Michael [University of Pennsylvania, Children' s Hospital of Philadelphia and Perelman School of Medicine, Philadelphia, PA (United States); DuBois, Steven G. [UCSF School of Medicine, Department of Pediatrics, San Francisco, CA (United States); UCSF Benioff Children' s Hospital, San Francisco, CA (United States); UCSF School of Medicine, San Francisco, CA (United States)

    2016-03-15

    Vesicular monoamine transporters 1 and 2 (VMAT1 and VMAT2) are thought to mediate MIBG uptake in adult neuroendocrine tumors. In neuroblastoma, the norepinephrine transporter (NET) has been investigated as the principal MIBG uptake protein, though some tumors without NET expression concentrate MIBG. We investigated VMAT expression in neuroblastoma and correlated expression with MIBG uptake and clinical features. We evaluated VMAT1 and VMAT2 expression by immunohistochemistry (IHC) in neuroblastoma tumors from 76 patients with high-risk metastatic disease treated in a uniform cooperative group trial (COG A3973). All patients had baseline MIBG diagnostic scans centrally reviewed. IHC results were scored as the product of intensity grading (0 - 3+) and percent of tumor cells expressing the protein of interest. The association between VMAT1 and VMAT2 scores and clinical and biological features was tested using Wilcoxon rank-sum tests. Patient characteristics were typical of high-risk neuroblastoma, though the cohort was intentionally enriched in patients with MIBG-nonavid tumors (n = 20). VMAT1 and VMAT2 were expressed in 62 % and 75 % of neuroblastoma tumors, respectively. VMAT1 and VMAT2 scores were both significantly lower in MYCN amplified tumors and in tumors with high mitotic karyorrhectic index. MIBG-avid tumors had significantly higher VMAT2 scores than MIBG-nonavid tumors (median 216 vs. 45; p = 0.04). VMAT1 expression did not correlate with MIBG avidity. VMAT1 and VMAT2 are expressed in the majority of neuroblastomas. Expression correlates with other biological features. The expression level of VMAT2 but not that of VMAT1 correlates with avidity for MIBG. (orig.)

  3. Effects of anesthetics on vesicular monoamine transporter type 2 binding to 18F-FP-(+)-DTBZ: a biodistribution study in rat brain

    International Nuclear Information System (INIS)

    Chen, Zhengping; Tang, Jie; Liu, Chunyi; Li, Xiaomin; Huang, Hongbo; Xu, Xijie; Yu, Huixin

    2016-01-01

    Objectives: The in vivo binding analysis of vesicular monoamine transporter type 2 (VMAT2) to radioligand has provided a means of investigating related disorders. Anesthesia is often inevitable when the investigations are performed in animals. In the present study, we tested effects of four commonly-used anesthetics: isoflurane, pentobarbital, chloral hydrate and ketamine, on in vivo VMAT2 binding to 18 F-FP-(+)-DTBZ, a specific VMAT2 radioligand, in rat brain. Methods: The transient equilibrium time window for in vivo binding of 18 F-FP-(+)-DTBZ after a bolus injection was firstly determined. The brain biodistribution studies under anesthetized and awake rats were then performed at the equilibrium time. Standard uptake values (SUVs) of the interest brain regions: the striatum (ST), hippocampus (HP), cortex (CX) and cerebellum (CB) were obtained; and ratios of tissue to cerebellum were calculated. Results: Isoflurane and pentobarbital did not alter distribution of 18 F-FP-(+)-DTBZ in the brain relative to the awake group; neither SUVs nor ratios of ST/CB and HP/CB were altered significantly. Chloral hydrate significantly increased SUVs of all the brain regions, but did not significantly alter ratios of ST/CB and HP/CB. Ketamine significantly increased SUVs of the striatum, hippocampus and cortex, and insignificantly increased the SUV of the cerebellum; consequently, ketamine significantly increased ratios of ST/CB and HP/CB. Conclusions: It is concluded that in vivo VMAT2 binding to 18 F-FP-(+)-DTBZ are not altered by isoflurane and pentobarbital, but altered by chloral hydrate and ketamine. Isoflurane and pentobarbital may be promising anesthetic compounds for investigating in vivo VMAT2 binding. Further studies are warranted to investigate the interactions of anesthetics with VMAT2 binding potential with in vivo PET studies.

  4. High-frequency stimulation of the subthalamic nucleus modifies the expression of vesicular glutamate transporters in basal ganglia in a rat model of Parkinson's disease.

    Science.gov (United States)

    Favier, Mathieu; Carcenac, Carole; Drui, Guillaume; Boulet, Sabrina; El Mestikawy, Salah; Savasta, Marc

    2013-12-05

    It has been suggested that glutamatergic system hyperactivity may be related to the pathogenesis of Parkinson's disease (PD). Vesicular glutamate transporters (VGLUT1-3) import glutamate into synaptic vesicles and are key anatomical and functional markers of glutamatergic excitatory transmission. Both VGLUT1 and VGLUT2 have been identified as definitive markers of glutamatergic neurons, but VGLUT 3 is also expressed by non glutamatergic neurons. VGLUT1 and VGLUT2 are thought to be expressed in a complementary manner in the cortex and the thalamus (VL/VM), in glutamatergic neurons involved in different physiological functions. Chronic high-frequency stimulation (HFS) of the subthalamic nucleus (STN) is the neurosurgical therapy of choice for the management of motor deficits in patients with advanced PD. STN-HFS is highly effective, but its mechanisms of action remain unclear. This study examines the effect of STN-HFS on VGLUT1-3 expression in different brain nuclei involved in motor circuits, namely the basal ganglia (BG) network, in normal and 6-hydroxydopamine (6-OHDA) lesioned rats. Here we report that: 1) Dopamine(DA)-depletion did not affect VGLUT1 and VGLUT3 expression but significantly decreased that of VGLUT2 in almost all BG structures studied; 2) STN-HFS did not change VGLUT1-3 expression in the different brain areas of normal rats while, on the contrary, it systematically induced a significant increase of their expression in DA-depleted rats and 3) STN-HFS reversed the decrease in VGLUT2 expression induced by the DA-depletion. These results show for the first time a comparative analysis of changes of expression for the three VGLUTs induced by STN-HFS in the BG network of normal and hemiparkinsonian rats. They provide evidence for the involvement of VGLUT2 in the modulation of BG cicuits and in particular that of thalamostriatal and thalamocortical pathways suggesting their key role in its therapeutic effects for alleviating PD motor symptoms.

  5. Melatonin mediated antidepressant-like effect in the hippocampus of chronic stress-induced depression rats: Regulating vesicular monoamine transporter 2 and monoamine oxidase A levels.

    Science.gov (United States)

    Stefanovic, Bojana; Spasojevic, Natasa; Jovanovic, Predrag; Jasnic, Nebojsa; Djordjevic, Jelena; Dronjak, Sladjana

    2016-10-01

    The hippocampus is sensitive to stress which activates norepinephrine terminals deriving from the locus coeruleus. Melatonin exerts positive effects on the hippocampal neurogenic process and on depressive-like behaviour. Thus, in the present study, an examination was made of the effect of chronic melatonin treatment on norepinephrine content, synthesis, uptake, vesicular transport and degradation in the hippocampus of rats exposed to CUMS. This entailed quantifying the norephinephrine, mRNA and protein levels of DBH, NET, VMAT 2, MAO-A and COMT. The results show that CUMS evoked prolonged immobility. Melatonin treatment decreased immobility in comparison with the placebo group, reflecting an antidepressant-like effect. Compared with the placebo group, a dramatic decrease in norepinephrine content, decreased VMAT2 mRNA and protein and increased MAO-A protein levels in the hippocampus of the CUMS rats were observed. However, no significant differences in the levels of DBH, NET, COMT mRNA and protein and MAO-A mRNA levels between the placebo and the stressed groups were found. The results showed the restorative effects of melatonin on the stress-induced decline in the norepinephrine content of the hippocampus. It was observed that melatonin treatment in the CUMS rats prevented the stress-induced decrease in VMAT2 mRNA and protein levels, whereas it reduced the increase of the mRNA of COMT and protein levels of MAO-A. Chronic treatment with melatonin failed to alter the gene expression of DBH or NET in the hippocampus of the CUMS rats. Additionally, the results show that melatonin enhances VMAT2 expression and norepinephrine storage, whilst it reduces norepinephrine degrading enzymes. Copyright © 2016 Elsevier B.V. and ECNP. All rights reserved.

  6. Sex-dependent role of vesicular glutamate transporter 3 in stress-regulation and related anxiety phenotype during the early postnatal period.

    Science.gov (United States)

    Balázsfi, Diána; Farkas, Lívia; Csikota, Péter; Fodor, Anna; Zsebők, Sándor; Haller, József; Zelena, Dóra

    2016-07-01

    Stress and related disorders are in the focus of interest and glutamate is one of the most important neurotransmitters that can affect these processes. Glutamatergic neurons are characterized by vesicular glutamate transporters (VGluT1-3) among which vGluT3 is unique contributing to the non-canonical, neuromodulatory effect of glutamate. We aimed to study the role of vGluT3 in stress axis regulation and related anxiety during the early postnatal period using knockout (KO) mice with special focus on sex differences. Anxiety was explored on postnatal day (PND) 7-8 by maternal separation-induced ultrasonic vocalization (USV). Stress-hormone levels were detected 60 min after intraperitoneal lipopolysaccharide (LPS) injection 7 days later. Both genotypes gained weight, but on PND 14-15 KO mice pups had smaller body weight compared to wild type (WT). vGluT3 KO mice reacted to an immune stressor with enhanced adrenocorticotropin (ACTH) and corticosterone secretion compared to WT. Although there was a tendency for enhanced anxiety measured by more emitted USV, this did not reach the level of significance. The only sex-related effect was the enhanced corticosterone reactivity in male pups. For the HPA axis regulation in neonates vGluT3 expression seems to be dispensable under basal conditions, but is required for optimal response to immune stressors, most probably through an interaction with other neurotransmitters. Disturbance of the fine balance between these systems may result in a borderline enhanced anxiety-like behavior in vGluT3 KO pups.

  7. Acetylcholine receptor antibody

    Science.gov (United States)

    ... medlineplus.gov/ency/article/003576.htm Acetylcholine receptor antibody To use the sharing features on this page, please enable JavaScript. Acetylcholine receptor antibody is a protein found in the blood of ...

  8. Effects of chronic inhalation of electronic cigarettes containing nicotine on glial glutamate transporters and α-7 nicotinic acetylcholine receptor in female CD-1 mice.

    Science.gov (United States)

    Alasmari, Fawaz; Crotty Alexander, Laura E; Nelson, Jessica A; Schiefer, Isaac T; Breen, Ellen; Drummond, Christopher A; Sari, Youssef

    2017-07-03

    Alteration in glutamate neurotransmission has been found to mediate the development of drug dependence, including nicotine. We and others, through using western blotting, have reported that exposure to drugs of abuse reduced the expression of glutamate transporter-1 (GLT-1) as well as cystine/glutamate antiporter (xCT), which consequently increased extracellular glutamate concentrations in the mesocorticolimbic area. However, our previous studies did not reveal any changes in glutamate/aspartate transporter (GLAST) following exposure to drugs of abuse. In the present study, for the first time, we investigated the effect of chronic exposure to electronic (e)-cigarette vapor containing nicotine, for one hour daily for six months, on GLT-1, xCT, and GLAST expression in frontal cortex (FC), striatum (STR), and hippocampus (HIP) in outbred female CD1 mice. In this study, we also investigated the expression of alpha-7 nicotinic acetylcholine receptor (α-7 nAChR), a major pre-synaptic nicotinic receptor in the glutamatergic neurons, which regulates glutamate release. We found that inhalation of e-cigarette vapor for six months increased α-7 nAChR expression in both FC and STR, but not in the HIP. In addition, chronic e-cigarette exposure reduced GLT-1 expression only in STR. Moreover, e-cigarette vapor inhalation induced downregulation of xCT in both the STR and HIP. We did not find any significant changes in GLAST expression in any brain region. Finally, using liquid chromatography-tandem mass spectrometry (LC-MS/MS) techniques, we detected high concentrations of nicotine and cotinine, a major metabolite of nicotine, in the FC tissues of e-cigarette exposed mice. These data provide novel evidence about the effects of chronic nicotine inhalation on the expression of key glial glutamate transporters as well as α-7 nAChR. Our work may suggest that nicotine exposure via chronic inhalation of e-cigarette vapor may be mediated in part by alterations in the glutamatergic

  9. The effect of acetylcholine on 14C-assimilates translocation of Isatis tinctoria L

    International Nuclear Information System (INIS)

    Yang Chongjun; Tang Feiyu; Zhang Ping; Guo Yuhai

    2004-01-01

    The effects of acetylcholine on 14 C-assimilates translocation are studied with source-channel-sink of Isatis tinctoria L. The experiments show that 0.01 mmol/L treatments of acetylcholine on the phloem, can improve the output of 14 C-assimilates in leaves indicating that acetylcholine enhances the activity of phloem transport. (authors)

  10. Transcriptomic effects of depleted uranium on acetylcholine and cholesterol metabolisms in Alzheimer's disease model; Effets transcriptomiques de l'uranium appauvri sur les metabolismes de l'acetylcholine et du cholesterol chez un modele de maladie d'Alzheimer

    Energy Technology Data Exchange (ETDEWEB)

    Lestaevel, Ph.; Bensoussan, H.; Racine, R.; Airault, F.; Gourmelon, P.; Souidi, M. [Direction de la radioprotection de l' Homme, service de radiobiologie et d' epidemiologie, laboratoire de radiotoxicologie experimentale, institut de radioprotection et de surete nucleaire, BP no 17, 92262 Fontenay-aux-Roses cedex (France)

    2011-02-15

    Some heavy metals, or aluminium, could participate in the development of Alzheimer disease (AD). Depleted uranium (DU), another heavy metal, modulates the cholinergic system and the cholesterol metabolism in the brain of rats, but without neurological disorders. The aim of this study was to determine what happens in organisms exposed to DU that will/are developing the AD. This study was thus performed on a transgenic mouse model for human amyloid precursor protein (APP), the Tg2576 strain. The possible effects of DU through drinking water (20 mg/L) over an 8-month period were analyzed on acetylcholine and cholesterol metabolisms at gene level in the cerebral cortex. The mRNA levels of choline acetyl transferase (ChAT) vesicular acetylcholine transporter (VAChT) and ATP-binding cassette transporter A1 (ABC A1) decreased in control Tg2576 mice in comparison with wild-type mice (respectively -89%, -86% and -44%, p < 0.05). Chronic exposure of Tg2576 mice to DU increased mRNA levels of ChAT (+189%, p < 0.05), VAChT (+120%, p < 0.05) and ABC A1 (+52%, p < 0.05) compared to control Tg2576 mice. Overall, these modifications of acetylcholine and cholesterol metabolisms did not lead to increased disturbances that are specific of AD, suggesting that chronic DU exposure did not worsen the pathology in this experimental model. (authors)

  11. Combined α7 nicotinic acetylcholine receptor agonism and partial serotonin transporter inhibition produce antidepressant-like effects in the mouse forced swim and tail suspension tests

    DEFF Research Database (Denmark)

    Andreasen, Jesper T; Redrobe, John P; Nielsen, Elsebet Ø

    2012-01-01

    Emerging evidence points to an involvement of nicotinic acetylcholine receptors (nAChRs) in major depression. Nicotine improves symptoms of depression in humans and shows antidepressant-like effects in rodents. Monoamine release is facilitated by nAChR stimulation, and nicotine-evoked serotonin (5...

  12. Pep3p/Pep5p complex: a putative docking factor at multiple steps of vesicular transport to the vacuole of Saccharomyces cerevisiae.

    OpenAIRE

    Srivastava, A; Woolford, C A; Jones, E W

    2000-01-01

    Pep3p and Pep5p are known to be necessary for trafficking of hydrolase precursors to the vacuole and for vacuolar biogenesis. These proteins are present in a hetero-oligomeric complex that mediates transport at the vacuolar membrane. PEP5 interacts genetically with VPS8, implicating Pep5p in the earlier Golgi to endosome step and/or in recycling from the endosome to the Golgi. To understand further the cellular roles of Pep3p and Pep5p, we isolated and characterized a set of pep3 conditional ...

  13. Current treatment of vesicular lithiasis

    International Nuclear Information System (INIS)

    Garcia Rodriguez, Oscar

    2010-01-01

    Surgical treatment of vesicular lithiasis has changed in past years. The addition of the new techniques in daily medical practice not always is immediate. Reasons relative to when to operate a patient presenting with gall bladder calculi are argued and documenting how this procedure is mainly reserved for symptomatic patients where pain is considered as a symptom par excellence. Also, it is exposed how this change has been faced. (author)

  14. Role of Intermediate Filaments in Vesicular Traffic

    Directory of Open Access Journals (Sweden)

    Azzurra Margiotta

    2016-04-01

    Full Text Available Intermediate filaments are an important component of the cellular cytoskeleton. The first established role attributed to intermediate filaments was the mechanical support to cells. However, it is now clear that intermediate filaments have many different roles affecting a variety of other biological functions, such as the organization of microtubules and microfilaments, the regulation of nuclear structure and activity, the control of cell cycle and the regulation of signal transduction pathways. Furthermore, a number of intermediate filament proteins have been involved in the acquisition of tumorigenic properties. Over the last years, a strong involvement of intermediate filament proteins in the regulation of several aspects of intracellular trafficking has strongly emerged. Here, we review the functions of intermediate filaments proteins focusing mainly on the recent knowledge gained from the discovery that intermediate filaments associate with key proteins of the vesicular membrane transport machinery. In particular, we analyze the current understanding of the contribution of intermediate filaments to the endocytic pathway.

  15. Renal epithelial cells can release ATP by vesicular fusion

    Directory of Open Access Journals (Sweden)

    Randi G Bjaelde

    2013-09-01

    Full Text Available Renal epithelial cells have the ability to release nucleotides as paracrine factors. In the intercalated cells of the collecting duct, ATP is released by connexin30 (cx30, which is selectively expressed in this cell type. However, ATP is released by virtually all renal epithelia and the aim of the present study was to identify possible alternative nucleotide release pathways in a renal epithelial cell model. We used MDCK (type1 cells to screen for various potential ATP release pathways. In these cells, inhibition of the vesicular H+-ATPases (bafilomycin reduced both the spontaneous and hypotonically (80%-induced nucleotide release. Interference with vesicular fusion using N-ethylamide markedly reduced the spontaneous nucleotide release, as did interference with trafficking from the endoplasmic reticulum to the Golgi apparatus (brefeldin A1 and vesicular transport (nocodazole. These findings were substantiated using a siRNA directed against SNAP-23, which significantly reduced spontaneous ATP release. Inhibition of pannexin and connexins did not affect the spontaneous ATP release in this cell type, which consists of ∼90% principal cells. TIRF-microscopy of either fluorescently-labeled ATP (MANT-ATP or quinacrine-loaded vesicles, revealed that spontaneous release of single vesicles could be promoted by either hypoosmolality (50% or ionomycin. This vesicular release decreased the overall cellular fluorescence by 5.8% and 7.6% respectively. In summary, this study supports the notion that spontaneous and induced ATP release can occur via exocytosis in renal epithelial cells.

  16. Transmission and pathogenesis of vesicular stomatitis viruses

    Science.gov (United States)

    Vesicular Stomatitis (VS) is caused by the Vesicular Stomatitis Virus (VSV), a negative single stranded RNA arthropod-borne virus member of the Family Rhabdoviridae. The virion is composed of the host derived plasma membrane, the envelope, and an internal ribonucleoprotein core. The envelope contain...

  17. Distribution of metallothionein I + II and vesicular zinc in the developing central nervous system: correlative study in the rat

    DEFF Research Database (Denmark)

    Penkowa, M; Nielsen, H; Hidalgo, J

    1999-01-01

    in hippocampal cortex, basal forebrain, neocortex, cerebellar cortex, and cranial nerve nuclei. MT I + II mRNAs were detected in regions of the brain that also displayed MT I + IIir, indicating transcriptional events. Vesicular Zn was recorded in neonatal brain solely in the dentate hi of the hippocampus...... candidates for chelating unbound Zn released from Zn-containing nerve terminals or transported into the brain. Whether vesicular Zn and MT I + II occur in identical regions of the developing brain is unknown. Accordingly, the developmental distribution of MT I + II and vesicular Zn was mapped. By using...

  18. Cholinergic neurotransmission in human corpus cavernosum. II. Acetylcholine synthesis

    International Nuclear Information System (INIS)

    Blanco, R.; De Tejada, S.; Goldstein, I.; Krane, R.J.; Wotiz, H.H.; Cohen, R.A.

    1988-01-01

    Physiological and histochemical evidence indicates that cholinergic nerves may participate in mediating penile erection. Acetylcholine synthesis and release was studied in isolated human corporal tissue. Human corpus cavernosum incubated with [ 3 H]choline accumulated [ 3 H]choline and synthesized [ 3 H]acethylcholine in an concentration-dependent manner. [ 3 H]Acetylcholine accumulation by the tissue was inhibited by hemicholinium-3, a specific antagonist of the high-affinity choline transport in cholinergic nerves. Transmural electrical field stimulation caused release of [ 3 H]acetylcholine which was significantly diminished by inhibiting neurotransmission with calcium-free physiological salt solution or tetrodotoxin. These observations provide biochemical and physiological evidence for the existence of cholinergic innervation in human corpus cavernosum

  19. Vesicular trafficking of immune mediators in human eosinophils revealed by immunoelectron microscopy

    Energy Technology Data Exchange (ETDEWEB)

    Melo, Rossana C.N., E-mail: rossana.melo@ufjf.edu.br [Laboratory of Cellular Biology, Department of Biology, ICB, Federal University of Juiz de Fora, UFJF, Rua José Lourenço Kelmer, Juiz de Fora, MG 36036-900 (Brazil); Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, 330 Brookline Avenue, CLS 943, Boston, MA 02215 (United States); Weller, Peter F. [Department of Medicine, Beth Israel Deaconess Medical Center, Harvard Medical School, 330 Brookline Avenue, CLS 943, Boston, MA 02215 (United States)

    2016-10-01

    Electron microscopy (EM)-based techniques are mostly responsible for our current view of cell morphology at the subcellular level and continue to play an essential role in biological research. In cells from the immune system, such as eosinophils, EM has helped to understand how cells package and release mediators involved in immune responses. Ultrastructural investigations of human eosinophils enabled visualization of secretory processes in detail and identification of a robust, vesicular trafficking essential for the secretion of immune mediators via a non-classical secretory pathway associated with secretory (specific) granules. This vesicular system is mainly organized as large tubular-vesicular carriers (Eosinophil Sombrero Vesicles – EoSVs) actively formed in response to cell activation and provides a sophisticated structural mechanism for delivery of granule-stored mediators. In this review, we highlight the application of EM techniques to recognize pools of immune mediators at vesicular compartments and to understand the complex secretory pathway within human eosinophils involved in inflammatory and allergic responses. - Highlights: • Application of EM to understand the complex secretory pathway in human eosinophils. • EM techniques reveal an active vesicular system associated with secretory granules. • Tubular vesicles are involved in the transport of granule-derived immune mediators.

  20. Vesicular trafficking of immune mediators in human eosinophils revealed by immunoelectron microscopy

    International Nuclear Information System (INIS)

    Melo, Rossana C.N.; Weller, Peter F.

    2016-01-01

    Electron microscopy (EM)-based techniques are mostly responsible for our current view of cell morphology at the subcellular level and continue to play an essential role in biological research. In cells from the immune system, such as eosinophils, EM has helped to understand how cells package and release mediators involved in immune responses. Ultrastructural investigations of human eosinophils enabled visualization of secretory processes in detail and identification of a robust, vesicular trafficking essential for the secretion of immune mediators via a non-classical secretory pathway associated with secretory (specific) granules. This vesicular system is mainly organized as large tubular-vesicular carriers (Eosinophil Sombrero Vesicles – EoSVs) actively formed in response to cell activation and provides a sophisticated structural mechanism for delivery of granule-stored mediators. In this review, we highlight the application of EM techniques to recognize pools of immune mediators at vesicular compartments and to understand the complex secretory pathway within human eosinophils involved in inflammatory and allergic responses. - Highlights: • Application of EM to understand the complex secretory pathway in human eosinophils. • EM techniques reveal an active vesicular system associated with secretory granules. • Tubular vesicles are involved in the transport of granule-derived immune mediators.

  1. Predicted overlapping microRNA regulators of acetylcholine packaging and degradation in neuroinflammation-related disorders

    Directory of Open Access Journals (Sweden)

    Bettina eNadorp

    2014-02-01

    Full Text Available MicroRNAs (miRNAs can notably control many targets each and regulate entire cellular pathways, but whether miRNAs can regulate complete neurotransmission processes is largely unknown. Here, we report that miRNAs with complementary sequence motifs to the key genes involved in acetylcholine (ACh synthesis and/or packaging show massive overlap with those regulating ACh degradation. To address this topic, we first searched for miRNAs that could target the 3’-untranslated regions of the choline acetyltransferase (ChAT gene that controls ACh synthesis; the vesicular ACh transporter (VAChT, encoded from an intron in the ChAT gene and the ACh hydrolyzing genes acetyl- and/or butyrylcholinesterase (AChE, BChE. Intriguingly, we found that many of the miRNAs targeting these genes are primate-specific, and that changes in their levels associate with inflammation, anxiety, brain damage, cardiac, neurodegenerative or pain-related syndromes. To validate the in vivo relevance of this dual interaction, we selected the evolutionarily conserved miR-186, which targets both the stress-inducible soluble readthrough variant AChE-R and the major peripheral cholinesterase BChE. We exposed mice to predator scent stress and searched for potential associations between consequent changes in their miR-186, AChE-R and BChE levels. Both intestinal miR-186 as well as BChE and AChE-R activities were conspicuously elevated one week post-exposure, highlighting the previously unknown involvement of miR-186 and BChE in psychological stress responses. Overlapping miRNA regulation emerges from our findings as a recently evolved surveillance mechanism over cholinergic neurotransmission in health and disease; and the corresponding miRNA details and disease relevance may serve as a useful resource for studying the molecular mechanisms underlying this surveillance.

  2. Regulation of vesicular traffic by a GTP-binding protein on the cytoplasmic surface of secretory vesicles in yeast

    International Nuclear Information System (INIS)

    Novick, P.J.; Goud, B.; Salminen, A.; Walworth, N.C.; Nair, J.; Potenza, M.

    1988-01-01

    Vesicular transport is an important mechanism for the intracellular traffic of proteins and lipids in eukaryotic cells. Vesicles mediate the passage of proteins between the various organelles of the secretory pathway and the exocytic release of these proteins into the extracellular environment. Vesicles also mediate the uptake of proteins and fluid from the external environment, delivering them to endosomes. Despite the generality of the vesicular transport mechanism, the process is not yet understood at a molecular level. The key questions that are addressed are (1) How are vesicles formed from the membrane of the donor organelle? (2) How are these vesicles transported? (3) How do the vesicles recognize the membrane of the target (acceptor) organelle? (4) How is membrane fusion accomplished? The genetic flexibility of yeast has been exploited to identify components of the cellular machinery required for vesicular transport

  3. Airstream Fractionation of Vesicular-Arbuscular Mycorrhizal Fungi: Concentration and Enumeration of Propagules

    OpenAIRE

    Tommerup, Inez C.

    1982-01-01

    Spores and fragments of vesicular-arbuscular mycorrhizal fungi in dry soils were concentrated up to 100-fold when the soils were partitioned by fluidization and elutriation with a series of upward airstreams at progressively increasing velocities. The propagules were transported with the finer soil particles according to their equivalent spherical diameters. The system was used to predict the transport of propagules by wind. Concentrated propagules were rapidly separated from the soil particl...

  4. Regulation of vesicular trafficking by Parkinson's disease-associated genes

    Directory of Open Access Journals (Sweden)

    Tsuyoshi Inoshita

    2015-10-01

    Full Text Available The regulatory mechanisms that control intracellular vesicular trafficking play important roles in cellular function and viability. Neurons have specific vesicular trafficking systems for synaptic vesicle formation, release and recycling. Synaptic vesicular trafficking impairments induce neuronal dysfunction and physiological and behavioral disorders. Parkinson's disease (PD is an age-dependent neurodegenerative disorder characterized by dopamine depletion and loss of dopamine neurons in the midbrain. The molecular mechanism responsible for the neurodegeneration that occurs during PD is still not understood; however, recent functional analyses of familial PD causative genes suggest that a number of PD causative genes regulate intracellular vesicular trafficking, including synaptic vesicular dynamics. This review focuses on recent insights regarding the functions of PD causative genes, their relationship with vesicular trafficking and how mutations associated with PD affect vesicular dynamics and neuronal survival.

  5. Vesicular stomatitis forecasting based on Google Trends.

    Science.gov (United States)

    Wang, JianYing; Zhang, Tong; Lu, Yi; Zhou, GuangYa; Chen, Qin; Niu, Bing

    2018-01-01

    Vesicular stomatitis (VS) is an important viral disease of livestock. The main feature of VS is irregular blisters that occur on the lips, tongue, oral mucosa, hoof crown and nipple. Humans can also be infected with vesicular stomatitis and develop meningitis. This study analyses 2014 American VS outbreaks in order to accurately predict vesicular stomatitis outbreak trends. American VS outbreaks data were collected from OIE. The data for VS keywords were obtained by inputting 24 disease-related keywords into Google Trends. After calculating the Pearson and Spearman correlation coefficients, it was found that there was a relationship between outbreaks and keywords derived from Google Trends. Finally, the predicted model was constructed based on qualitative classification and quantitative regression. For the regression model, the Pearson correlation coefficients between the predicted outbreaks and actual outbreaks are 0.953 and 0.948, respectively. For the qualitative classification model, we constructed five classification predictive models and chose the best classification predictive model as the result. The results showed, SN (sensitivity), SP (specificity) and ACC (prediction accuracy) values of the best classification predictive model are 78.52%,72.5% and 77.14%, respectively. This study applied Google search data to construct a qualitative classification model and a quantitative regression model. The results show that the method is effective and that these two models obtain more accurate forecast.

  6. Vesicular stomatitis forecasting based on Google Trends

    Science.gov (United States)

    Lu, Yi; Zhou, GuangYa; Chen, Qin

    2018-01-01

    Background Vesicular stomatitis (VS) is an important viral disease of livestock. The main feature of VS is irregular blisters that occur on the lips, tongue, oral mucosa, hoof crown and nipple. Humans can also be infected with vesicular stomatitis and develop meningitis. This study analyses 2014 American VS outbreaks in order to accurately predict vesicular stomatitis outbreak trends. Methods American VS outbreaks data were collected from OIE. The data for VS keywords were obtained by inputting 24 disease-related keywords into Google Trends. After calculating the Pearson and Spearman correlation coefficients, it was found that there was a relationship between outbreaks and keywords derived from Google Trends. Finally, the predicted model was constructed based on qualitative classification and quantitative regression. Results For the regression model, the Pearson correlation coefficients between the predicted outbreaks and actual outbreaks are 0.953 and 0.948, respectively. For the qualitative classification model, we constructed five classification predictive models and chose the best classification predictive model as the result. The results showed, SN (sensitivity), SP (specificity) and ACC (prediction accuracy) values of the best classification predictive model are 78.52%,72.5% and 77.14%, respectively. Conclusion This study applied Google search data to construct a qualitative classification model and a quantitative regression model. The results show that the method is effective and that these two models obtain more accurate forecast. PMID:29385198

  7. Vesicular stomatitis forecasting based on Google Trends.

    Directory of Open Access Journals (Sweden)

    JianYing Wang

    Full Text Available Vesicular stomatitis (VS is an important viral disease of livestock. The main feature of VS is irregular blisters that occur on the lips, tongue, oral mucosa, hoof crown and nipple. Humans can also be infected with vesicular stomatitis and develop meningitis. This study analyses 2014 American VS outbreaks in order to accurately predict vesicular stomatitis outbreak trends.American VS outbreaks data were collected from OIE. The data for VS keywords were obtained by inputting 24 disease-related keywords into Google Trends. After calculating the Pearson and Spearman correlation coefficients, it was found that there was a relationship between outbreaks and keywords derived from Google Trends. Finally, the predicted model was constructed based on qualitative classification and quantitative regression.For the regression model, the Pearson correlation coefficients between the predicted outbreaks and actual outbreaks are 0.953 and 0.948, respectively. For the qualitative classification model, we constructed five classification predictive models and chose the best classification predictive model as the result. The results showed, SN (sensitivity, SP (specificity and ACC (prediction accuracy values of the best classification predictive model are 78.52%,72.5% and 77.14%, respectively.This study applied Google search data to construct a qualitative classification model and a quantitative regression model. The results show that the method is effective and that these two models obtain more accurate forecast.

  8. Pancreatitis aguda grave asociada a gangrena vesicular

    OpenAIRE

    Arroyo-Sánchez, Abel S; Aguirre-Mejía, Rosa Y; Echenique-Martínez, Sergio E

    2014-01-01

    Se presenta el caso un paciente diabético que desarrolló un cuadro de pancreatitis aguda grave asociada a gangrena vesicular, en el que se evaluó la aplicabilidad de los criterios de clasificación y manejo de la hoja de ruta para pancreatitis aguda, así mismo se proponen algunos tópicos que pudieran ser investigados a futuro We present a diabetic patient who developed severe acute pancreatitis associated to gallbladder gangrene, in this case we assessed the applicability of classification ...

  9. Docking to flexible nicotinic acetylcholine receptors

    DEFF Research Database (Denmark)

    Sander, Tommy; Bruun, Anne T; Balle, Thomas

    2010-01-01

    Computational docking to nicotinic acetylcholine receptors (nAChRs) and other members of the Cys-loop receptor family is complicated by the flexibility of the so-called C-loop. As observed in the large number of published crystal structures of the acetylcholine binding protein (AChBP), a structural...

  10. Contribution of vesicular and cytosolic dopamine to the increased striatal dopamine efflux elicited by intrastriatal injection of SKF38393.

    NARCIS (Netherlands)

    Saigusa, T.; Aono, Y.; Sekino, R.; Uchida, T.; Takada, K.; Oi, Y.; Koshikawa, N.; Cools, A.R.

    2009-01-01

    Like dexamphetamine, SKF38393 induces an increase in striatal dopamine efflux which is insensitive for tetrodotoxin, Ca(2+) independent and prevented by a dopamine transporter inhibitor. The dexamphetamine-induced striatal dopamine efflux originates from both the reserpine-sensitive vesicular

  11. Neuromodulation: acetylcholine and memory consolidation.

    Science.gov (United States)

    Hasselmo

    1999-09-01

    Clinical and experimental evidence suggests that hippocampal damage causes more severe disruption of episodic memories if those memories were encoded in the recent rather than the more distant past. This decrease in sensitivity to damage over time might reflect the formation of multiple traces within the hippocampus itself, or the formation of additional associative links in entorhinal and association cortices. Physiological evidence also supports a two-stage model of the encoding process in which the initial encoding occurs during active waking and deeper consolidation occurs via the formation of additional memory traces during quiet waking or slow-wave sleep. In this article I will describe the changes in cholinergic tone within the hippocampus in different stages of the sleep-wake cycle and will propose that these changes modulate different stages of memory formation. In particular, I will suggest that the high levels of acetylcholine that are present during active waking might set the appropriate dynamics for encoding new information in the hippocampus, by partially suppressing excitatory feedback connections and so facilitating encoding without interference from previously stored information. By contrast, the lower levels of acetylcholine that are present during quiet waking and slow-wave sleep might release this suppression and thereby allow a stronger spread of activity within the hippocampus itself and from the hippocampus to the entorhinal cortex, thus facilitating the process of consolidation of separate memory traces.

  12. Antagonism of acetylcholine by adrenaline antagonists

    Science.gov (United States)

    Benfey, B. G.; Grillo, S. A.

    1963-01-01

    Phenoxybenzamine antagonized the inhibitory action of acetylcholine on the guinea-pig isolated atrium. The antagonism was slow in onset, very slowly reversible, and could be overcome by increased concentrations of acetylcholine. In contrast, atropine inhibited the action of acetylcholine quickly, and the effect disappeared soon after withdrawal. The pA10 of phenoxybenzamine (2 hr of contact) was 6.8, and that of atropine (30 min of contact) was 8.4. In the presence of atropine phenoxybenzamine did not exert a slowly reversible antagonism, and the dose-ratio of acetylcholine returned to normal soon after withdrawal of both drugs. Phenoxybenzamine also antagonized acetylcholine in the guinea-pig isolated ileum, but with higher concentrations acetylcholine did not overcome the antagonism. The pA10 (60 min of contact) was 6.6. The pA10 of chlorpromazine in the atrium (2 hr of contact) and ileum (60 min of contact) was 5.9. Phentolamine, 2-diethylaminomethylbenzo-1,4-dioxan hydrochloride (883 F), and yohimbine antagonized acetylcholine in the atrium and ileum but required higher concentrations than chlorpromazine. PMID:13967429

  13. Action potential-independent and pharmacologically unique vesicular serotonin release from dendrites

    Science.gov (United States)

    Colgan, Lesley A.; Cavolo, Samantha L.; Commons, Kathryn G.; Levitan, Edwin S.

    2012-01-01

    Serotonin released within the dorsal raphe nucleus (DR) induces feedback inhibition of serotonin neuron activity and consequently regulates mood-controlling serotonin release throughout the forebrain. Serotonin packaged in vesicles is released in response to action potentials by the serotonin neuron soma and terminals, but the potential for release by dendrites is unknown. Here three-photon (3P) microscopy imaging of endogenous serotonin in living rat brain slice, immunofluorescence and immuno-gold electron microscopy detection of VMAT2 (vesicular monoamine transporter 2) establish the presence of vesicular serotonin within DR dendrites. Furthermore, activation of glutamate receptors is shown to induce vesicular serotonin release from dendrites. However, unlike release from the soma and terminals, dendritic serotonin release is independent of action potentials, relies on L-type Ca2+ channels, is induced preferentially by NMDA, and displays distinct sensitivity to the selective serotonin reuptake inhibitor (SSRI) antidepressant fluoxetine. The unique control of dendritic serotonin release has important implications for DR physiology and the antidepressant action of SSRIs, dihydropyridines and NMDA receptor antagonists. PMID:23136413

  14. Vesicular glutamate release from central axons contributes to myelin damage.

    Science.gov (United States)

    Doyle, Sean; Hansen, Daniel Bloch; Vella, Jasmine; Bond, Peter; Harper, Glenn; Zammit, Christian; Valentino, Mario; Fern, Robert

    2018-03-12

    The axon myelin sheath is prone to injury associated with N-methyl-D-aspartate (NMDA)-type glutamate receptor activation but the source of glutamate in this context is unknown. Myelin damage results in permanent action potential loss and severe functional deficit in the white matter of the CNS, for example in ischemic stroke. Here, we show that in rats and mice, ischemic conditions trigger activation of myelinic NMDA receptors incorporating GluN2C/D subunits following release of axonal vesicular glutamate into the peri-axonal space under the myelin sheath. Glial sources of glutamate such as reverse transport did not contribute significantly to this phenomenon. We demonstrate selective myelin uptake and retention of a GluN2C/D NMDA receptor negative allosteric modulator that shields myelin from ischemic injury. The findings potentially support a rational approach toward a low-impact prophylactic therapy to protect patients at risk of stroke and other forms of excitotoxic injury.

  15. Transferosomes - A vesicular transdermal delivery system for enhanced drug permeation

    Directory of Open Access Journals (Sweden)

    Reshmy Rajan

    2011-01-01

    Full Text Available Transdermal administration of drugs is generally limited by the barrier function of the skin. Vesicular systems are one of the most controversial methods for transdermal delivery of active substances. The interest in designing transdermal delivery systems was relaunched after the discovery of elastic vesicles like transferosomes, ethosomes, cubosomes, phytosomes, etc. This paper presents the composition, mechanisms of penetration, manufacturing and characterization methods of transferosomes as transdermal delivery systems of active substances. For a drug to be absorbed and distributed into organs and tissues and eliminated from the body, it must pass through one or more biological membranes/barriers at various locations. Such a movement of drug across the membrane is called as drug transport. For the drugs to be delivered to the body, they should cross the membranous barrier. The concept of these delivery systems was designed in an attempt to concentrate the drug in the tissues of interest, while reducing the amount of drug in the remaining tissues. Hence, surrounding tissues are not affected by the drug. In addition, loss of drug does not happen due to localization of drug, leading to get maximum efficacy of the medication. Therefore, the phospholipid based carrier systems are of considerable interest in this era.

  16. Effect of vesicular arbuscular mycorrhizal fungus on the ...

    African Journals Online (AJOL)

    STORAGESEVER

    2008-10-06

    Oct 6, 2008 ... ... association between certain plants and microorganisms plays an important role in soil ..... an Agrostis capillaris population on a copper contaminated soil. Plant ... vesicular-arbuscular mycorrhizal fungi in Amazonian Peru.

  17. ATP is stored in lamellar bodies to activate vesicular P2X4 in an autocrine fashion upon exocytosis.

    Science.gov (United States)

    Fois, Giorgio; Winkelmann, Veronika Eva; Bareis, Lara; Staudenmaier, Laura; Hecht, Elena; Ziller, Charlotte; Ehinger, Konstantin; Schymeinsky, Jürgen; Kranz, Christine; Frick, Manfred

    2018-02-05

    Vesicular P2X 4 receptors are known to facilitate secretion and activation of pulmonary surfactant in the alveoli of the lungs. P2X 4 receptors are expressed in the membrane of lamellar bodies (LBs), large secretory lysosomes that store lung surfactant in alveolar type II epithelial cells, and become inserted into the plasma membrane after exocytosis. Subsequent activation of P2X 4 receptors by adenosine triphosphate (ATP) results in local fusion-activated cation entry (FACE), facilitating fusion pore dilation, surfactant secretion, and surfactant activation. Despite the importance of ATP in the alveoli, and hence lung function, the origin of ATP in the alveoli is still elusive. In this study, we demonstrate that ATP is stored within LBs themselves at a concentration of ∼1.9 mM. ATP is loaded into LBs by the vesicular nucleotide transporter but does not activate P2X 4 receptors because of the low intraluminal pH (5.5). However, the rise in intravesicular pH after opening of the exocytic fusion pore results in immediate activation of vesicular P2X 4 by vesicular ATP. Our data suggest a new model in which agonist (ATP) and receptor (P2X 4 ) are located in the same intracellular compartment (LB), protected from premature degradation (ATP) and activation (P2X 4 ), and ideally placed to ensure coordinated and timely receptor activation as soon as fusion occurs to facilitate surfactant secretion. © 2018 Fois et al.

  18. Organic cation transporter 2 (SLC22A2), a low-affinity and high-capacity choline transporter, is preferentially enriched on synaptic vesicles in cholinergic neurons.

    Science.gov (United States)

    Nakata, T; Matsui, T; Kobayashi, K; Kobayashi, Y; Anzai, N

    2013-11-12

    Organic cation transporters (OCTs) are expressed mainly in the kidney and liver. OCTs transport intrinsic organic cations, including monoamine, dopamine, serotonine and choline, across the plasma membrane. Here, we demonstrate that OCT2 (SLC22A2) is expressed in cholinergic neurons, motoneurons in the anterior horn of the spinal cord, and is implicated in acetylcholine (Ach) recycling in presynaptic terminals. Application of rabbit anti-peptide antibody revealed that OCT2 was expressed in the anterior horn of the spinal cord. Double immunostaining of muscle sections with anti-OCT2 and alpha-bungarotoxin (BTX) revealed that OCT2 was localized in the neuromuscular junctions (NMJs). Immunoelectron microscopy revealed that OCT2 was localized both in synaptic vesicles (SVs) in presynaptic terminals around the motoneurons (C-terminals) and in SVs in nerve terminals in NMJs. The similarity in the distribution of OCT2 in cholinergic neurons and that of vesicular acetyl choline transporter (VAchT), and the fact that OCT2 can transport choline suggest that OCT2 could work as a low-affinity and high-capacity choline transporter at presynaptic terminals in cholinergic neurons in a firing-dependent manner. Copyright © 2013 IBRO. Published by Elsevier Ltd. All rights reserved.

  19. Characterization of Vesicular Stomatitis Virus Recombinants That Express and Incorporate High Levels of Hepatitis C Virus Glycoproteins

    OpenAIRE

    Buonocore, Linda; Blight, Keril J.; Rice, Charles M.; Rose, John K.

    2002-01-01

    We generated recombinant vesicular stomatitis viruses (VSV) expressing genes encoding hybrid proteins consisting of the extracellular domains of hepatitis C virus (HCV) glycoproteins fused at different positions to the transmembrane and cytoplasmic domains of the VSV G glycoprotein (E1G and E2G). We show that these chimeric proteins are transported to the cell surface and incorporated into VSV virions efficiently. We also generated VSV recombinants in which the gene encoding the VSV G protein...

  20. REGULATED VESICULAR TRAFFICKING OF SPECIFIC PCDH15 AND VLGR1 VARIANTS IN AUDITORY HAIR CELLS

    Science.gov (United States)

    Zallocchi, Marisa; Delimont, Duane; Meehan, Daniel T.; Cosgrove, Dominic

    2012-01-01

    Usher syndrome is a genetically heterogeneous disorder characterized by hearing and balance dysfunction and progressive retinitis pigmentosa. Mouse models carrying mutations for the nine Usher-associated genes have splayed stereocilia and some show delayed maturation of ribbon synapses suggesting these proteins may play different roles in terminal differentiation of auditory hair cells. The presence of the Usher proteins at the basal and apical aspects of the neurosensory epithelia suggests the existence of regulated trafficking through specific transport proteins and routes. Immature mouse cochleae and UB/OC-1 cells were used in this work to address whether specific variants of PCDH15 and VLGR1 are being selectively transported to opposite poles of the hair cells. Confocal co-localization studies between apical and basal vesicular markers and the different PCDH15 and VLGR1 variants along with sucrose density gradients and the use of vesicle trafficking inhibitors show the existence of Usher protein complexes in at least two vesicular sub-pools. The apically trafficked pool co-localized with the early endosomal vesicle marker, rab5, while the basally trafficked pool associates with membrane microdomains and SNAP25. Moreover, co-immunoprecipitation experiments between SNAP25 and VLGR1 show a physical interaction of these two proteins in organ of Corti and brain. Collectively, these findings establish the existence of a differential vesicular trafficking mechanism for specific Usher protein variants in mouse cochlear hair cells, with the apical variants playing a potential role in endosomal recycling and stereocilia development/maintenance and the basolateral variants involved in vesicle docking and/or fusion through SNAP25-mediated interactions. PMID:23035094

  1. The α7 nicotinic acetylcholine receptor complex

    DEFF Research Database (Denmark)

    Thomsen, Morten Skøtt; Mikkelsen, Jens D

    2012-01-01

    The α7 nicotinic acetylcholine receptor (nAChR) is a promising drug target for a number of diseases ranging from schizophrenia and Alzheimer's disease to chronic pain and inflammatory diseases. Focusing on the central nervous system, we describe how endogenous and experimental compounds and prote......The α7 nicotinic acetylcholine receptor (nAChR) is a promising drug target for a number of diseases ranging from schizophrenia and Alzheimer's disease to chronic pain and inflammatory diseases. Focusing on the central nervous system, we describe how endogenous and experimental compounds...

  2. A Recombinant Vesicular Stomatitis Virus Ebola Vaccine.

    Science.gov (United States)

    Regules, Jason A; Beigel, John H; Paolino, Kristopher M; Voell, Jocelyn; Castellano, Amy R; Hu, Zonghui; Muñoz, Paula; Moon, James E; Ruck, Richard C; Bennett, Jason W; Twomey, Patrick S; Gutiérrez, Ramiro L; Remich, Shon A; Hack, Holly R; Wisniewski, Meagan L; Josleyn, Matthew D; Kwilas, Steven A; Van Deusen, Nicole; Mbaya, Olivier Tshiani; Zhou, Yan; Stanley, Daphne A; Jing, Wang; Smith, Kirsten S; Shi, Meng; Ledgerwood, Julie E; Graham, Barney S; Sullivan, Nancy J; Jagodzinski, Linda L; Peel, Sheila A; Alimonti, Judie B; Hooper, Jay W; Silvera, Peter M; Martin, Brian K; Monath, Thomas P; Ramsey, W Jay; Link, Charles J; Lane, H Clifford; Michael, Nelson L; Davey, Richard T; Thomas, Stephen J

    2017-01-26

    The worst Ebola virus disease (EVD) outbreak in history has resulted in more than 28,000 cases and 11,000 deaths. We present the final results of two phase 1 trials of an attenuated, replication-competent, recombinant vesicular stomatitis virus (rVSV)-based vaccine candidate designed to prevent EVD. We conducted two phase 1, placebo-controlled, double-blind, dose-escalation trials of an rVSV-based vaccine candidate expressing the glycoprotein of a Zaire strain of Ebola virus (ZEBOV). A total of 39 adults at each site (78 participants in all) were consecutively enrolled into groups of 13. At each site, volunteers received one of three doses of the rVSV-ZEBOV vaccine (3 million plaque-forming units [PFU], 20 million PFU, or 100 million PFU) or placebo. Volunteers at one of the sites received a second dose at day 28. Safety and immunogenicity were assessed. The most common adverse events were injection-site pain, fatigue, myalgia, and headache. Transient rVSV viremia was noted in all the vaccine recipients after dose 1. The rates of adverse events and viremia were lower after the second dose than after the first dose. By day 28, all the vaccine recipients had seroconversion as assessed by an enzyme-linked immunosorbent assay (ELISA) against the glycoprotein of the ZEBOV-Kikwit strain. At day 28, geometric mean titers of antibodies against ZEBOV glycoprotein were higher in the groups that received 20 million PFU or 100 million PFU than in the group that received 3 million PFU, as assessed by ELISA and by pseudovirion neutralization assay. A second dose at 28 days after dose 1 significantly increased antibody titers at day 56, but the effect was diminished at 6 months. This Ebola vaccine candidate elicited anti-Ebola antibody responses. After vaccination, rVSV viremia occurred frequently but was transient. These results support further evaluation of the vaccine dose of 20 million PFU for preexposure prophylaxis and suggest that a second dose may boost antibody responses

  3. Uptake of 3H-choline and synthesis of 3H-acetylcholine by human penile corpus cavernosum

    International Nuclear Information System (INIS)

    Blanco, R.; Saenz de Tejada, I.; Azadzoi, K.; Goldstein, I.; Krane, R.J.; Wotiz, H.H.; Cohen, R.A.

    1986-01-01

    The neuroeffectors which relax penile smooth muscle and lead to erection are unknown; physiological studies of human corpus cavernosum, in vitro, have suggested a significant role of cholinergic neurotransmission. To further characterize the importance of cholinergic nerves, biopsies of human corpus cavernosum were obtained at the time of penile prosthesis implantation. Tissues were incubated in 3 H-choline (10 -5 M, 80 Ci/mmol) in oxygenated physiological salt solution at 37 0 C, pH 7.4 for 1 hour. Radiolabelled compounds were extracted with perchloric acid (0.4 M) and acetylcholine and choline were separated by HPLC; 14 C-acetylcholine was used as internal standard. 3 H-choline was accumulated by the tissues (20 +/- 1.9 fmol/mg), and 3 H-acetylcholine was synthesized (4.0 +/- 1.1 fmol/mg). In control experiments, heating of the tissue blocked synthesis of 3 H-acetylcholine. Inhibition of high affinity choline transport by hemicholinium-3 (10 -5 M) diminished tissue accumulation of 3 H-choline and significantly reduced the synthesis of 3 H-acetylcholine (0.5 +/ 0.2 fmol/mg, p < 0.05). These results provide direct evidence of neuronal accumulation of choline and enzymatic conversion to acetylcholine in human corpus cavernosum. Taken together with the physiological studies, it can be concluded that cholinergic neurotransmission in human corpus cavernosum plays a role in penile erection

  4. Logistics in the cell: cargoes and transportation.

    Science.gov (United States)

    Nadezhdina, E S

    2014-09-01

    Eukaryotic cells are large and thus require a vesicular transport system. The system involves the formation of membrane transport containers, their short- and long-distance movements, recognition of destination points, and fusion with other membranes. Understanding the molecular mechanisms of these processes is of theoretical and practical significance. This special issue of Biochemistry (Moscow) collects surveys and experimental articles describing various aspects of vesicular transport.

  5. Structural modifications of vesicular aggregates following gamma-irradiation

    International Nuclear Information System (INIS)

    Mantaka-Marketou, A.E.; Domasou, A.S.

    1991-01-01

    The structural changes of the didodecyldimethylammonium bromide (DDAB) vesicular bilayers after γ-irradiation and under conditions where mainly OH radicals are present are reported. Alterations of the vesicular structure, such as polarity and fluidity, were detected after a dose of 0.65 kGy. A higher dose of ∼14kGy cause important damage to the well organized molecular structure and this is manifested by an important augmentation of the fluidity and polarity of the Stern region of the aggregates. Increased water penetration into the bilayer of the vesicle is probably the reason for these changes and electron micrographs support this hypothesis. (author)

  6. Structural Studies of Nicotinic Acetylcholine Receptors

    DEFF Research Database (Denmark)

    Shahsavar, Azadeh; Gajhede, Michael; Kastrup, Jette

    2016-01-01

    Nicotinic acetylcholine receptors (nAChRs) are members of the pentameric ligand-gated ion channel superfamily that play important roles in control of neurotransmitter release in the central and peripheral nervous system. These receptors are important therapeutic targets for development of drugs...

  7. Allosteric Modulation of Muscarinic Acetylcholine Receptors

    Czech Academy of Sciences Publication Activity Database

    Jakubík, Jan; El-Fakahany, E. E.

    2010-01-01

    Roč. 3, č. 9 (2010), s. 2838-2860 ISSN 1424-8247 R&D Projects: GA ČR GA305/09/0681 Institutional research plan: CEZ:AV0Z50110509 Keywords : muscarinic acetylcholine receptors * allosteric modulation * Alzheimer´s disease Subject RIV: CE - Biochemistry

  8. Porters and neurotransmitter transporters

    NARCIS (Netherlands)

    Nelson, Nathan; Lill, H

    1994-01-01

    Uptake of neurotransmitters involves multiple transporters acting in different brain locations under different physiological conditions. The vesicular transporters are driven by a proton-motive force generated by a V-ATPase and their substrates are taken up via proton/substrate exchange. The plasma

  9. Subaqueous non-vesicular to poorly-vesicular shards: hydroclastic fragmentation on seamounts and summit calderas

    Science.gov (United States)

    Mueller, W. U.; Dingwell, D. B.; Downey, W. S.; Mastin, L. G.

    2008-12-01

    Recognizing pyroclastic deposits that originate directly from magmatic and phreatomagmatic explosions in a subaqueous setting is based upon sedimentary structures, such as massive, stratified, and graded beds as well as (pyro)clast size. Ideally such deposits form ordered fining-and thinning-upward sequences. Pumice, scoria, glass shards, euhedral and broken crystals, and lithic fragments are constituents that support an explosive heritage. Recent deep-sea ROV and submersible dives have retrieved non-vesicular to vesicle- poor, mm-scale, mafic shards in 5-15 cm-thick massive and/or graded (stratified) deposits, for which a subaqueous explosive origin has been inferred. These sheet hyaloclastites with variable shard shapes were first documented on Seamount 6 as deep-sea Limu O Pele at water depths > 1000 m. We identified in Seamount 6 samples equant to blocky shards with angular to subrounded terminations, but also subordinate hair-like and contorted glassy filaments, warped shards and irregular shards. Shards display internal laminations (flow-banding?) and have local perlitic fractures. Bubble wall shards derived from scoria burst were rare. In combination with all the above and a poor shard vesicularity (tubes and ponded magma in depths > 1000 m. We envision that hydrostatic pressure commensurate with water depth played a significant role. The deposits can be readily explained by a hydroclastic process whereby fragmentation occurred at the milli-second (Limu) to second scale (hyaloclastite). Hence, hyperquenched glass shards or thread-like glass filaments need not require magmatic explosivity. Constant surface interaction between aphyric, low-viscosity, high temperature, magma-lava at depth with seawater causes fragmentation (granulation) that can generate such delicate shards. The transfer of heat to the ambient medium, seawater, favours turbulent convection causing strong water movement that strips glassy rinds and lofts the fine-grained shards and Limu O Pele

  10. Practical Microform Materials for Libraries: Silver, Diazo, Vesicular.

    Science.gov (United States)

    Veaner, Allen B.

    1982-01-01

    Remarks on the relative permanence and durability of three types of film in use in library microform reproduction (silver, diazo, and vesicular) and points out some technical and economic facts that govern the choice of microform materials for libraries. A 6-item reference list is included. (Author/JL)

  11. Reconstitution of the fusogenic activity of vesicular stomatitis virus

    NARCIS (Netherlands)

    Metsikkö, K.; van Meer, G.; Simons, K.

    1986-01-01

    Enveloped virus glycoproteins exhibit membrane fusion activity. We have analysed whether the G protein of vesicular stomatitis virus, reconstituted into liposomes, is able to fuse nucleated cells in a pH-dependent fashion. Proteoliposomes produced by octylglucoside dialysis did not exhibit cell

  12. The vesicular-arbuscular mycorrhizal symbiosis | Quilambo | African ...

    African Journals Online (AJOL)

    Vesicular-arbuscular mycorrhiza fungi are associated with the majority ot the terrestrial plants. Their function ranges from stress alleviation to bioremediation in soils polluted with heavy metals. However, our knowledge about this symbiosis is still limited. For the semi-arid tropics, where some african countries are located, ...

  13. The distribution of vesicular-arbuscular mycorrhizal fungi in India.

    Science.gov (United States)

    Rani, R; Mukerji, K G

    1990-01-01

    Vesicular-arbuscular mycorrhizal fungi are widely distributed throughout the area studied including different altitudes ranging from sea level to 2500 ft above sea level. VAM fungi were recorded from 88% of the sites examined with Glomus fasciculatum and Glomus macrocarpum being the most commonly recorded. Mean species diversity was found to be maximum in the areas thickly vegetated and undisturbed.

  14. Airstream fractionation of vesicular-arbuscular mycorrhizal fungi: concentration and enumeration of propagules.

    Science.gov (United States)

    Tommerup, I C

    1982-09-01

    Spores and fragments of vesicular-arbuscular mycorrhizal fungi in dry soils were concentrated up to 100-fold when the soils were partitioned by fluidization and elutriation with a series of upward airstreams at progressively increasing velocities. The propagules were transported with the finer soil particles according to their equivalent spherical diameters. The system was used to predict the transport of propagules by wind. Concentrated propagules were rapidly separated from the soil particles in each soil fraction by an aqueous flotation method. The technique is proposed as a quantitative method for estimating the numbers of spores and fragments of mycorrhizae. The scheme includes a viability test that was used to differentiate between potentially infective propagules and those that were either dormant or incapable of regrowth.

  15. Quantification of Water Flux in Vesicular Systems.

    Science.gov (United States)

    Hannesschläger, Christof; Barta, Thomas; Siligan, Christine; Horner, Andreas

    2018-06-04

    Water transport across lipid membranes is fundamental to all forms of life and plays a major role in health and disease. However, not only typical water facilitators like aquaporins facilitate water flux, but also transporters, ion channels or receptors represent potent water pathways. The efforts directed towards a mechanistic understanding of water conductivity determinants in transmembrane proteins, the development of water flow inhibitors, and the creation of biomimetic membranes with incorporated membrane proteins or artificial water channels depend on reliable and accurate ways of quantifying water permeabilities P f . A conventional method is to subject vesicles to an osmotic gradient in a stopped-flow device: Fast recordings of scattered light intensity are converted into the time course of vesicle volume change. Even though an analytical solution accurately acquiring P f from scattered light intensities exists, approximations potentially misjudging P f by orders of magnitude are used. By means of computational and experimental data we point out that erroneous results such as that the single channel water permeability p f depends on the osmotic gradient are direct results of such approximations. Finally, we propose an empirical solution of which calculated permeability values closely match those calculated with the analytical solution in the relevant range of parameters.

  16. Biochemical characterization of native Usher protein complexes from a vesicular subfraction of tracheal epithelial cells.

    Science.gov (United States)

    Zallocchi, Marisa; Sisson, Joseph H; Cosgrove, Dominic

    2010-02-16

    Usher syndrome is the major cause of deaf/blindness in the world. It is a genetic heterogeneous disorder, with nine genes already identified as causative for the disease. We noted expression of all known Usher proteins in bovine tracheal epithelial cells and exploited this system for large-scale biochemical analysis of Usher protein complexes. The dissected epithelia were homogenized in nondetergent buffer and sedimented on sucrose gradients. At least two complexes were evident after the first gradient: one formed by specific isoforms of CDH23, PCDH15, and VLGR-1 and a different one at the top of the gradient that included all of the Usher proteins and rab5, a transport vesicle marker. TEM analysis of these top fractions found them enriched in 100-200 nm vesicles, confirming a vesicular association of the Usher complex(es). Immunoisolation of these vesicles confirmed some of the associations already predicted and identified novel interactions. When the vesicles are lysed in the presence of phenylbutyrate, most of the Usher proteins cosediment into the gradient at a sedimentation coefficient of approximately 50 S, correlating with a predicted molecular mass of 2 x 10(6) Da. Although it is still unclear whether there is only one complex or several independent complexes that are trafficked within distinct vesicular pools, this work shows for the first time that native Usher protein complexes occur in vivo. This complex(es) is present primarily in transport vesicles at the apical pole of tracheal epithelial cells, predicting that Usher proteins may be directionally transported as complexes in hair cells and photoreceptors.

  17. BIOCHEMICAL CHARACTERIZATION OF NATIVE USHER PROTEIN COMPLEXES FROM A VESICULAR SUBFRACTION OF TRACHEAL EPITHELIAL CELLS†

    Science.gov (United States)

    Zallocchi, Marisa; Sisson, Joseph H.; Cosgrove, Dominic

    2010-01-01

    Usher syndrome is the major cause of deaf/blindness in the world. It is a genetic heterogeneous disorder, with nine genes already identified as causative for the disease. We noted expression of all known Usher proteins in bovine tracheal epithelial cells, and exploited this system for large-scale biochemical analysis of Usher protein complexes. The dissected epithelia were homogenized in non-detergent buffer, and sedimented on sucrose gradients. At least two complexes were evident after the first gradient: one formed by specific isoforms of CDH23, PCDH15 and VLGR-1, and a different one at the top of the gradient that included all the Usher proteins and rab5, a transport vesicle marker. TEM analysis of these top fractions found them enriched in 100–200 nm vesicles, confirming a vesicular association of the Usher complex(es). Immunoisolation of these vesicles confirmed some of the associations already predicted and identified novel interactions. When the vesicles are lysed in the presence of phenylbutyrate, most of the Usher proteins co-sediment into the gradient at a sedimentation coefficient of approximately 50S, correlating with a predicted molecular mass of 2 × 106 Daltons. Although it is still unclear whether there is only one complex or several independent complexes that are trafficked within distinct vesicular pools, this work shows for the first time that native Usher proteins complexes occur in vivo. This complex(es) is present primarily in transport vesicles at the apical pole of tracheal epithelial cells, predicting that Usher proteins may be directionally transported as complexes in hair cells and photoreceptors. PMID:20058854

  18. Vesicular thick-walled swollen hyphae in pulmonary zygomycosis.

    Science.gov (United States)

    Kimura, Masatomo; Ito, Hiroyuki

    2009-03-01

    An autopsy case of pulmonary zygomycosis in a patient with rheumatoid arthritis on immunosuppressive therapy is presented herein. There was a pulmonary cavitated infarct caused by mycotic thrombosis. Thin-walled narrow hyphae and vesicular thick-walled swollen hyphae were found on the pleural surface and in the necrotic tissue at the periphery of the cavity. Findings of such shaped fungal elements may cause erroneous histopathological diagnosis because pauciseptate broad thin-walled hyphae are usually the only detectable fungal elements in zygomycosis tissue. Although immunohistochemistry confirmed these unusual elements to be zygomycetous in the present case, it is important for the differential diagnosis to be aware that zygomycetes can form thin narrow hyphae and vesicular thick-walled swollen hyphae.

  19. An Atypical Local Vesicular Reaction to the Yellow Fever Vaccine

    OpenAIRE

    Wauters, Robert H.; Hernandez, Camellia L.; Petersen, Maureen M.

    2017-01-01

    Yellow fever vaccine is a live attenuated viral inoculation indicated for patients traveling to endemic areas. The vaccine is generally well tolerated with minimal adverse effects. Typical side effects include malaise, pain at the injection site, and, albeit rarely, immediate hypersensitivity reactions. We present a case of a rare adverse reaction to yellow fever vaccine in which a patient developed vesicular lesions resulting in bullae and circumferential hyperpigmentation.

  20. An Atypical Local Vesicular Reaction to the Yellow Fever Vaccine.

    Science.gov (United States)

    Wauters, Robert H; Hernandez, Camellia L; Petersen, Maureen M

    2017-09-19

    Yellow fever vaccine is a live attenuated viral inoculation indicated for patients traveling to endemic areas. The vaccine is generally well tolerated with minimal adverse effects. Typical side effects include malaise, pain at the injection site, and, albeit rarely, immediate hypersensitivity reactions. We present a case of a rare adverse reaction to yellow fever vaccine in which a patient developed vesicular lesions resulting in bullae and circumferential hyperpigmentation.

  1. Hyperforin inhibits vesicular uptake of monoamines by dissipating pH gradient across synaptic vesicle membrane.

    Science.gov (United States)

    Roz, Netta; Rehavi, Moshe

    2003-06-13

    Extracts of Hypericum perforatum (St. John's wort) have antidepressant properties in depressed patients and exert antidepressant-like action in laboratory animals. The phloroglucinol derivative hyperforin has become a topic of interest, as this Hypericum component is a potent inhibitor of monoamines reuptake. The molecular mechanism by which hyperforin inhibits monoamines uptake is yet unclear. In the present study we try to clarify the mechanism by which hyperforin inhibits the synaptic vesicle transport of monoamines. The pH gradient across the synaptic vesicle membrane, induced by vacuolar type H(+)-ATPase, is the major driving force for vesicular monoamines uptake and storage. We suggest that hyperforin, like the protonophore FCCP, dissipates an existing Delta pH generated by an efflux of inwardly pumped protons. Proton transport was measured by acridine orange fluorescence quenching. Adding Mg-ATP to a medium containing 130 mM KCl and synaptic vesicles caused an immediate decrease in fluorescence of acridine orange and the addition of 1 microM FCCP abolished this effect. H(+)-ATPase dependent proton pumping was inhibited by hyperforin in a dose dependent manner (IC(50) = 1.9 x 10(-7) M). Hyperforin acted similarly to the protonophore FCCP, abolishing the ATP induced fluorescence quenching (IC(50) = 4.3 x 10(-7) M). Hyperforin and FCCP had similar potencies for inhibiting rat brain synaptosomal uptake of [3H]monoamines as well as vesicular monoamine uptake. The efflux of [3H]5HT from synaptic vesicles was sensitive to both drugs, thus 50% of preloaded [3H]5HT was released in the presence of 2.1 x 10(-7) M FCCP and 4 x 10(-7) M hyperforin. The effect of hyperforin on the pH gradient in synaptic vesicle membrane may explain its inhibitory effect on monoamines uptake, but could only partially explain its antidepressant properties.

  2. Plasma concentration of acetylcholine in young women

    International Nuclear Information System (INIS)

    Kawashima, K.; Oohata, H.; Fujimoto, K.; Suzuki, T.

    1987-01-01

    A sensitive and specific radioimmunoassay was applied to the determination of acetylcholine (ACh) in plasma. The concentration of ACh in plasma sampled from 32 young women was 456.1+-53.1 (mean +-S.E.M.) pg/ml. No significant correlations were observed between plasma concentrations of ACh and acetylcholinesterase (AChE) activity, or gonadal hormones. These data demonstrate that an amount of ACh measurable by radioimmunoassay is present in plasma and plasma ACh is not regulated by AChE activity and the menstrual cycle in young women. The origin and physiological as well as pathophysiological significance of ACh in plasma remain to be clarified. 13 refs. (Author)

  3. α-4 subunit of nicotinic acetylcholine receptor polymorphisms exhibit ...

    African Journals Online (AJOL)

    Background: Smoking behavior is influenced by both genetic and environmental factors. Nicotine is the major addictive substance in cigarettes. Nicotinic acetylcholine receptors (nAChRs) are thought to play an important role in nicotine addiction of smokers. One of the genes, α-4 subunit of nicotinic acetylcholine receptor ...

  4. Studies of two naturally occurring compounds which effect release of acetylcholine from synaptosomes

    International Nuclear Information System (INIS)

    Koenig, M.L.

    1985-01-01

    Two naturally occurring compounds which effect the release of neurotransmitter from synaptosomes have been purified to apparent homogeneity. Iotrochotin (IOT) isolated from wound exudate of the Caribbean purple bleeder sponge promotes release in a manner that is independent of the extracellular Ca 2+ ion concentration. Leptinotarsin (LPT-d), a protein taken from hemolymph of the Colorado potato beetle, Leptinotarsa decemlineata, stimulates Ca 2+ -dependent release. IOT is slightly acidic and has a molecular weight of approximately 18 kD. [ 3 H]acetylcholine which has been introduced into synaptosomes as [ 3 H]choline can be released by IOT. The toxin releasable pool of labelled neurotransmitter is not depleted by depolarization of the synaptosomes with high potassium, and therefore seems to be primarily extravesicular. LPT-d is a larger protein (molecular weight = 45 kD) than IOT, and seems to effect primarily vesicular release by opening at least one type of presynaptic Ca 2+ channel. The facilitatory effects of the toxin on synaptosomal release can be inhibited by inorganic Ca 2+ channel antagonists, but are not generally affected by organic antagonists

  5. Dysregulated Homeostasis of Acetylcholine Levels in Immune Cells of RR-Multiple Sclerosis Patients

    Directory of Open Access Journals (Sweden)

    Maria Di Bari

    2016-11-01

    Full Text Available Multiple sclerosis (MS is characterized by pro-inflammatory cytokine production. Acetylcholine (ACh contributes to the modulation of central and peripheral inflammation. We studied the homeostasis of the cholinergic system in relation to cytokine levels in immune cells and sera of relapsing remitting-MS (RR-MS patients. We demonstrated that lower ACh levels in serum of RR-MS patients were inversely correlated with the increased activity of the hydrolyzing enzymes acetylcholinesterase (AChE and butyrylcholinesterase (BuChE. Interestingly, the expression of the ACh biosynthetic enzyme and the protein carriers involved in non-vesicular ACh release were found overexpressed in peripheral blood mononuclear cells of MS patients. The inflammatory state of the MS patients was confirmed by increased levels of TNFα, IL-12/IL-23p40, IL-18. The lower circulating ACh levels in sera of MS patients are dependent on the higher activity of cholinergic hydrolyzing enzymes. The smaller ratio of ACh to TNFα, IL-12/IL-23p40 and IL-18 in MS patients, with respect to healthy donors (HD, is indicative of an inflammatory environment probably related to the alteration of cholinergic system homeostasis.

  6. [3H]acetylcholine synthesis in cultured ciliary ganglion neurons: effects of myotube membranes

    International Nuclear Information System (INIS)

    Gray, D.B.; Tuttle, J.B.

    1987-01-01

    Avian ciliary ganglion neurons in cell culture were examined for the capacity to synthesize acetylcholine (ACh) from the exogenously supplied precursor, choline. Relevant kinetic parameters of the ACh synthetic system in cultured neurons were found to be virtually the same as those of the ganglionic terminals in the intact iris. Neurons were cultured in the presence of and allowed to innervate pectoral muscle; this results in an capacity for ACh synthesis. In particular, the ability to increase ACh synthesis upon demand after stimulation is affected by interaction with the target. This effect is shown to be an acceleration of the maturation of the cultured neurons. Lysed and washed membrane remnants of the muscle target were able to duplicate, in part, this effect of live target tissue on neuronal transmitter metabolism. Culture medium conditioned by muscle, and by the membrane remnants of muscle, was without significant effect. Thus, substances secreted into the medium do not play a major role in this interaction. Neurons cultured with either muscle or muscle membrane remnants formed large, elongate structures on the target membrane surface. These were not seen in the absence of the target at the times examined. This morphological difference in terminal-like structures may parallel the developmental increases in size and vesicular content of ciliary ganglion nerve terminals in the chick iris, and may relate to the increased ACh synthetic activity. The results suggest that direct contact with an appropriate target membrane has a profound, retrograde influence upon neuronal metabolic and morphological maturation

  7. The effect of ketamine on intraspinal acetylcholine release

    DEFF Research Database (Denmark)

    Abelson, Klas S P; Goldkuhl, Renée Röstlinger; Nylund, Anders

    2006-01-01

    The general anaesthetic ketamine affects the central cholinergic system in several manners, but its effect on spinal acetylcholine release, which may be an important transmitter in spinal antinociception, is unknown. This study aimed to investigate the effect of ketamine on spinal acetylcholine...... release. Microdialysis probes were placed intraspinally in male rats, and acetylcholine was quantified with HPLC. Anaesthesia was switched from isoflurane (1.3%) to ketamine (150 mg/kg h), which resulted in a 500% increased acetylcholine release. The increase was attenuated during nicotinic receptor...... blockade (50 microM mecamylamine). The nicotinic receptor agonist epibatidine (175 microM) produced a ten-fold higher relative increase of acetylcholine release during isoflurane anaesthesia compared to ketamine anaesthesia (270% to 27%). Intraspinal administration of ketamine and norketamine both...

  8. Expression and function of nicotinic acetylcholine receptors in stem cells

    Directory of Open Access Journals (Sweden)

    Herman S. Cheung

    2016-07-01

    Full Text Available Nicotinic acetylcholine receptors are prototypical ligand gated ion channels typically found in muscular and neuronal tissues. Functional nicotinic acetylcholine receptors, however, have also recently been identified on other cell types, including stem cells. Activation of these receptors by the binding of agonists like choline, acetylcholine, or nicotine has been implicated in many cellular changes. In regards to stem cell function, nicotinic acetylcholine receptor activation leads to changes in stem cell proliferation, migration and differentiation potential. In this review we summarize the expression and function of known nicotinic acetylcholine receptors in different classes of stem cells including: pluripotent stem cells, mesenchymal stem cells, periodontal ligament derived stem cells, and neural progenitor cells and discuss the potential downstream effects of receptor activation on stem cell function.

  9. The α7 nicotinic acetylcholine receptor complex

    DEFF Research Database (Denmark)

    Thomsen, Morten S; Mikkelsen, Jens D

    2012-01-01

    The α7 nicotinic acetylcholine receptor (nAChR) is a promising drug target for a number of diseases ranging from schizophrenia and Alzheimer's disease to chronic pain and inflammatory diseases. Focusing on the central nervous system, we describe how endogenous and experimental compounds and prote......The α7 nicotinic acetylcholine receptor (nAChR) is a promising drug target for a number of diseases ranging from schizophrenia and Alzheimer's disease to chronic pain and inflammatory diseases. Focusing on the central nervous system, we describe how endogenous and experimental compounds...... in diseases such as schizophrenia and Alzheimer's disease. Furthermore, α7 nAChR agonists and allosteric modulators differentially alter expression and functionality of the α7 nAChR with repeated administration, which suggests that there may be fundamentally different outcomes of long-term administration...... with these different types of compounds. Finally, we describe the special case of Aβ1-42 binding to the α7 nAChR, which may pose a unique challenge to drug development of α7 nAChR-specific ligands for Alzheimer's disease. Hopefully, a greater knowledge of the many factors influencing α7 nAChR function as well...

  10. Vesicular-Arbuscular Mycorrhiza in Field-Grown Crops

    DEFF Research Database (Denmark)

    Jakobsen, Iver

    1986-01-01

    The importance of vesicular-arbuscular mycorrhiza (VAM) and P fertilizer for P nutrition and dry matter production in field peas (Pisum sativum L.) was studied in moderately P-deficient soil. Half of the experimental plots were fumigated to reduce the level of VAM infection. Shoots and 0 to 30 cm...... in fumigated plots, although both it and P uptake were increased by adding P fertilizer. The possible reasons for this discrepancy are discussed. A supplementary survey on infection development at five other field sites showed that peas are extensively colonized by VAM fungi, even in soils where a standard...

  11. Acetylcholine Receptor: Complex of Homologous Subunits

    Science.gov (United States)

    Raftery, Michael A.; Hunkapiller, Michael W.; Strader, Catherine D.; Hood, Leroy E.

    1980-06-01

    The acetylcholine receptor from the electric ray Torpedo californica is composed of five subunits; two are identical and the other three are structurally related to them. Microsequence analysis of the four polypeptides demonstrates amino acid homology among the subunits. Further sequence analysis of both membrane-bound and Triton-solubilized, chromatographically purified receptor gave the stoichiometry of the four subunits (40,000:50,000:60,000:65,000 daltons) as 2:1:1:1, indicating that this protein is a pentameric complex with a molecular weight of 255,000 daltons. Genealogical analysis suggests that divergence from a common ancestral gene occurred early in the evolution of the receptor. This shared ancestry argues that each of the four subunits plays a functional role in the receptor's physiological action.

  12. Alcohol's actions on neuronal nicotinic acetylcholine receptors.

    Science.gov (United States)

    Davis, Tiffany J; de Fiebre, Christopher M

    2006-01-01

    Although it has been known for many years that alcoholism and tobacco addiction often co-occur, relatively little information is available on the biological factors that regulate the co-use and abuse of nicotine and alcohol. In the brain, nicotine acts at several different types of receptors collectively known as nicotinic acetylcholine receptors (nAChRs). Alcohol also acts on at least some of these receptors, enhancing the function of some nAChR subtypes and inhibiting the activity of others. Chronic alcohol and nicotine administration also lead to changes in the numbers of nAChRs. Natural variations (i.e., polymorphisms) in the genes encoding different nAChR subunits may be associated with individual differences in the sensitivity to some of alcohol's and nicotine's effects. Finally, at least one subtype of nAChR may help protect cells against alcohol-induced neurotoxicity.

  13. Impulsive behavior and nicotinic acetylcholine receptors.

    Science.gov (United States)

    Ohmura, Yu; Tsutsui-Kimura, Iku; Yoshioka, Mitsuhiro

    2012-01-01

    Higher impulsivity is thought to be a risk factor for drug addiction, criminal involvement, and suicide. Excessive levels of impulsivity are often observed in several psychiatric disorders including attention-deficit/hyperactivity disorder and schizophrenia. Previous studies have demonstrated that nicotinic acetylcholine receptors (nAChRs) are involved in impulsive behavior. Here, we introduce recent advances in this field and describe the role of the following nAChR-related brain mechanisms in modulating impulsive behavior: dopamine release in the ventral striatum; α4β2 nAChRs in the infralimbic cortex, which is a ventral part of the medial prefrontal cortex (mPFC); and dopamine release in the mPFC. We also suggest several potential therapeutic drugs to address these mechanisms in impulsivity-related disorders and explore future directions to further elucidate the roles of central nAChRs in impulsive behavior.

  14. Inhibition of acetylcholine synthesis in vitro

    International Nuclear Information System (INIS)

    O-Neill, J.J.; Capacio, B.; Doukas, P.H.; Leech, R.; Ricciardi, F.; Sterling, G.H.

    1986-01-01

    In order to better understand diseases that stem from deficiencies in cholinergic activity, reproducible in vitro and in vivo models displaying cholinergic hypofunction are desirable. This necessitates the availability of specific inhibitors. This paper examines the design, synthesis and evaluation of quinuclidinyl compounds with structural features previously reported, but with certain key differences. Structure activity studies with in vitro assay systems are presented. In a few studies, choline was held constant and acetyl-CoA concentration was varied, but with a constant amount of ( 14 C) - acetyl CoA. Acetylcholine synthesis and CO 2 production from labelled glucose were measured in cerebral cortex slices from male rats after decapitation. The nanomoles of ACh and CO 2 produced from ( 14 C) -glucose were calculated from glucose specific activity. Results are presented

  15. Acetylcholine receptors in the human retina

    International Nuclear Information System (INIS)

    Hutchins, J.B.; Hollyfield, J.G.

    1985-01-01

    Evidence for a population of acetylcholine (ACh) receptors in the human retina is presented. The authors have used the irreversible ligand 3 H-propylbenzilylcholine mustard ( 3 H-PrBCM) to label muscarinic receptors. 3 H- or 125 I-alpha-bungarotoxin (alpha-BTx) was used to label putative nicotinic receptors. Muscarinic receptors are apparently present in the inner plexiform layer of the retina. Autoradiographic grain densities are reduced in the presence of saturating concentrations of atropine, quinuclidinyl benzilate or scopolamine; this indicates that 3 H-PrBCM binding is specific for a population of muscarinic receptors in the human retina. Binding sites for radiolabeled alpha-BTx are found predominantly in the inner plexiform layer of the retina. Grain densities are reduced in the presence of d-tubocurarine, indicating that alpha-BTx may bind to a pharmacologically relevant nicotinic ACh receptor. This study provides evidence for cholinergic neurotransmission in the human retina

  16. Methodologic aspects of acetylcholine-evoked relaxation of rabbit aorta

    DEFF Research Database (Denmark)

    Larsen, Kirsten Vendelbo; Nedergaard, Ove A.

    1999-01-01

    The acetylcholine-evoked relaxation of rabbit isolated thoracic aorta precontracted by phenylephrine was studied. Phenylephrine caused a steady contraction that was maintained for 6 h. In the presence of calcium disodium ethylenediaminetetraacetate (EDTA) and ascorbic acid the contraction decreased...

  17. The Ghosts of Acetylcholine : structure-activity relationships of ...

    African Journals Online (AJOL)

    The Ghosts of Acetylcholine : structure-activity relationships of muscle relaxants : registrar communication. ... AJOL African Journals Online. HOW TO USE AJOL... for Researchers · for Librarians · for Authors · FAQ's · More about AJOL · AJOL's ...

  18. Effect of aerosolized acetylcholine on bronchial blood flow.

    Science.gov (United States)

    Charan, N B; Carvalho, P; Johnson, S R; Thompson, W H; Lakshminarayan, S

    1998-08-01

    We studied the effects of aerosolized as well as intravenous infusion of acetylcholine on bronchial blood flow in six anesthetized sheep. Intravenous infusion of acetylcholine, at a dose of 2 microg/kg, increased bronchial blood flow from 45 +/- 15 (SE) to 74 +/- 30 ml/min, and vascular conductance increased by 76 +/- 22%. In contrast, aerosolized acetylcholine at doses of 2 and 20 microg/kg decreased bronchial vascular conductance by approximately 10%. At an aerosolized dose of 200 microg/kg, the bronchial vascular conductance increased by approximately 15%, and there was no further increase in conductance when the aerosolized dose was increased to 2,000 microg/kg. Pretreatment of animals with a nitric oxide synthase inhibitor, Nomega-nitro-L-arginine methyl ester hydrochloride, partially blocked the vasodilatory effects of intravenous acetylcholine and completely blocked the vasodilatory effects of high-dose aerosolized acetylcholine. These data suggest that aerosolized acetylcholine does not readily penetrate the vascular wall of bronchial circulatory system and, therefore, has minimal vasodilatory effects on the bronchial vasculature.

  19. Vesicular-arbuscular mycorrhizal populations in stored topsoil

    Energy Technology Data Exchange (ETDEWEB)

    Harris, J.A.; Hunter, D.; Birch, P.; Short, K.C. (North East London Polytechnic, London (UK). Environment and Industry Research Unit, Dept. of Biology and Biochemistry)

    1987-01-01

    Two soil stores of different ages were sampled to investigate their vesicular-arbuscular mycorrhizal (VAM) populations. The soils collected were assessed for pH, moisture content, loss on ignition, spore numbers, number and size of root fragments present and percentage of these roots infected with VAM. A corn-root bioassay was used to determine soil infectivity. Root fragment number, size, % root infection and soil infectivity were negatively correlated with soil depth. VAM spore number was not significantly correlated with depth in either store. It appears that infected root fragments and fresh roots were the source of inoculum although there may have been a contribution from spores in the younger store. The infectivity of the older store soil was less than that of the younger store. 12 refs., 5 tabs.

  20. Transportation

    National Research Council Canada - National Science Library

    Adams, James; Carr, Ron; Chebl, Maroun; Coleman, Robert; Costantini, William; Cox, Robert; Dial, William; Jenkins, Robert; McGovern, James; Mueller, Peter

    2006-01-01

    ...., trains, ships, etc.) and maximizing intermodal efficiency. A healthy balance must be achieved between the flow of international commerce and security requirements regardless of transportation mode...

  1. Synaptically evoked Ca2+ release from intracellular stores is not influenced by vesicular zinc in CA3 hippocampal pyramidal neurones.

    Science.gov (United States)

    Evstratova, Alesya; Tóth, Katalin

    2011-12-01

    The co-release of neuromodulatory substances in combination with classic neurotransmitters such as glutamate and GABA from individual presynaptic nerve terminals has the capacity to dramatically influence synaptic efficacy and plasticity. At hippocampal mossy fibre synapses vesicular zinc is suggested to serve as a cotransmitter capable of regulating calcium release from internal stores in postsynaptic CA3 pyramidal cells. Here we investigated this possibility using combined intracellular ratiometric calcium imaging and patch-clamp recording techniques. In acute hippocampal slices a brief train of mossy fibre stimulation produced a large, delayed postsynaptic Ca(2+) wave that was spatially restricted to the proximal apical dendrites of CA3 pyramidal cells within stratum lucidum. This calcium increase was sensitive to intracellularly applied heparin indicating reliance upon release from internal stores and was triggered by activation of both group I metabotropic glutamate and NMDA receptors. Importantly, treatment of slices with the membrane-impermeant zinc chelator CaEDTA did not influence the synaptically evoked postsynaptic Ca(2+) waves. Moreover, mossy fibre stimulus evoked postsynaptic Ca(2+) signals were not significantly different between wild-type and zinc transporter 3 (ZnT3) knock-out animals. Considered together our data do not support a role for vesicular zinc in regulating mossy fibre evoked Ca(2+) release from CA3 pyramidal cell internal stores.

  2. Transportation

    International Nuclear Information System (INIS)

    Anon.

    1998-01-01

    Here is the decree of the thirtieth of July 1998 relative to road transportation, to trade and brokerage of wastes. It requires to firms which carry out a road transportation as well as to traders and to brokers of wastes to declare their operations to the prefect. The declaration has to be renewed every five years. (O.M.)

  3. 1995 epizootic of vesicular stomatitis (New Jersey serotype) in the western United States: an entomologic perspective.

    Science.gov (United States)

    Schmidtmann, E T; Tabachnick, W J; Hunt, G J; Thompson, L H; Hurd, H S

    1999-01-01

    Entomologic and epizootic data are reviewed concerning the potential for transmission of vesicular stomatitis (VS) virus by insects, including field data from case-positive premises in New Mexico and Colorado during the 1995 outbreak of the New Jersey serotype (VSNJ). As with previous outbreaks of VSNJ in the western United States, the 1995 epizootic illustrated that risk of exposure is seasonal, increasing during warm weather and decreasing with onset of cool weather; virus activity spread from south to north along river valleys of the southwestern and Rocky Mountain states; clinical disease was detected most commonly in horses, but also occurred in cattle and 1 llama; and most infections were subclinical. Overall, 367 case-positive premises were identified during the 1995 outbreak, with foci of virus activity along the Rio Grande River south of Albuquerque, NM, in southwestern Colorado, and along the Colorado River near Grand Junction, CO. The establishment of a 16-km (10-mile) radius zone of restricted animal movement around confirmed positive premises, along with imposition of state and international embargoes, created economic hardship for livestock owners and producers. The importance of defining the role of blood-feeding insects as biological vectors of VSNJ virus relative to risk factors that promote high levels of insect transmission, such as the presence of livestock along western river valleys, blood feeding activity, and frequent transport of animals for recreational purposes, is emphasized as a basis for developing effective disease management.

  4. The role of acetylcholine in cocaine addiction.

    Science.gov (United States)

    Williams, Mark J; Adinoff, Bryon

    2008-07-01

    Central nervous system cholinergic neurons arise from several discrete sources, project to multiple brain regions, and exert specific effects on reward, learning, and memory. These processes are critical for the development and persistence of addictive disorders. Although other neurotransmitters, including dopamine, glutamate, and serotonin, have been the primary focus of drug research to date, a growing preclinical literature reveals a critical role of acetylcholine (ACh) in the experience and progression of drug use. This review will present and integrate the findings regarding the role of ACh in drug dependence, with a primary focus on cocaine and the muscarinic ACh system. Mesostriatal ACh appears to mediate reinforcement through its effect on reward, satiation, and aversion, and chronic cocaine administration produces neuroadaptive changes in the striatum. ACh is further involved in the acquisition of conditional associations that underlie cocaine self-administration and context-dependent sensitization, the acquisition of associations in conditioned learning, and drug procurement through its effects on arousal and attention. Long-term cocaine use may induce neuronal alterations in the brain that affect the ACh system and impair executive function, possibly contributing to the disruptions in decision making that characterize this population. These primarily preclinical studies suggest that ACh exerts a myriad of effects on the addictive process and that persistent changes to the ACh system following chronic drug use may exacerbate the risk of relapse during recovery. Ultimately, ACh modulation may be a potential target for pharmacological treatment interventions in cocaine-addicted subjects. However, the complicated neurocircuitry of the cholinergic system, the multiple ACh receptor subtypes, the confluence of excitatory and inhibitory ACh inputs, and the unique properties of the striatal cholinergic interneurons suggest that a precise target of cholinergic

  5. Aspects of dopamine and acetylcholine release induced by glutamate receptors

    International Nuclear Information System (INIS)

    Paes, Paulo Cesar de Arruda

    2002-01-01

    The basal ganglia play an important role in the motor control of rats and humans. This control involves different neurotransmitters and the mutual control of these key elements has been subject to several studies. In this work we determined the role of glutamate on the release of radioactively labelled dopamine and acetylcholine from chopped striatal tissue in vitro. The values of Effective Concentration 50% for glutamate, NMDA, kainic, quisqualic acids and AMPA on the release of dopamine and acetylcholine were obtained. The inhibitory effects of magnesium, tetrodotoxin, MK-801, AP5 and MCPG, as well as the effects of glycin were evaluated. The results suggested that dopamine is influenced by the NMDA type glutamate receptor while acetylcholine seems to be influenced by NMDA, kainate and AMPA receptors. Tetrodotoxin experiments suggested that kainate receptors are both present in cholinergic terminals and cell bodies while AMPA and NMDA receptors are preferentially distributed in cell bodies. Magnesium effectively blocked the NMDA stimulation and unexpectedly also AMPA- and quisqualate-induced acetylcholine release. The latter could not be blocked by MCPG ruling out the participation of methabotropic receptors. MK-801 also blocked NMDA-receptors. Results point out the importance of the glutamic acid control of dopamine and acetylcholine release in striatal tissue. (author)

  6. Acetylcholine release and inhibitory interneuron activity in hippocampal CA1

    Directory of Open Access Journals (Sweden)

    A. Rory McQuiston

    2014-09-01

    Full Text Available Acetylcholine release in the central nervous system (CNS has an important role in attention, recall and memory formation. One region influenced by acetylcholine is the hippocampus, which receives inputs from the medial septum and diagonal band of Broca complex (MS/DBB. Release of acetylcholine from the MS/DBB can directly affect several elements of the hippocampus including glutamatergic and GABAergic neurons, presynaptic terminals, postsynaptic receptors and astrocytes. A significant portion of acetylcholine’s effect likely results from the modulation of GABAergic inhibitory interneurons, which have crucial roles in controlling excitatory inputs, synaptic integration, rhythmic coordination of principal neurons and outputs in the hippocampus. Acetylcholine affects interneuron function in large part by altering their membrane potential via muscarinic and nicotinic receptor activation. This minireview describes recent data from mouse hippocampus that investigated changes in CA1 interneuron membrane potentials following acetylcholine release. The interneuron subtypes affected, the receptor subtypes activated, and the potential outcome on hippocampal CA1 network function is discussed.

  7. Infection of guinea pigs with vesicular stomatitis New Jersey virus Transmitted by Culicoides sonorensis (Diptera: Ceratopogonidae).

    Science.gov (United States)

    Pérez De León, Adalberto A; O'Toole, Donal; Tabachnick, Walter J

    2006-05-01

    Intrathoracically inoculated Culicoides sonorensis Wirth & Jones were capable of transmitting vesicular stomatitis New Jersey virus (family Rhabdoviridae, genus Vesiculovirus, VSNJV) during blood feeding on the abdomen of six guinea pigs. None of the guinea pigs infected in this manner developed clinical signs of vesicular stomatitis despite seroconversion for VSNJV. Guinea pigs infected by intradermal inoculations of VSNJV in the abdomen also failed to develop clinical signs of vesicular stomatitis. Three guinea pigs given intradermal inoculations of VSNJV in the foot pad developed lesions typical of vesicular stomatitis. Transmission by the bite of C. sonorensis may have facilitated guinea pig infection with VSNJV because a single infected C. sonorensis caused seroconversion and all guinea pigs infected by insect bite seroconverted compared with 50% of the guinea pigs infected by intradermal inoculation with a higher titer VSNJV inoculum. The role of C. sonorensis in the transmission of VSNJV is discussed.

  8. Transportation

    National Research Council Canada - National Science Library

    Allshouse, Michael; Armstrong, Frederick Henry; Burns, Stephen; Courts, Michael; Denn, Douglas; Fortunato, Paul; Gettings, Daniel; Hansen, David; Hoffman, D. W; Jones, Robert

    2007-01-01

    .... The ability of the global transportation industry to rapidly move passengers and products from one corner of the globe to another continues to amaze even those wise to the dynamics of such operations...

  9. Epizootic vesicular stomatitis in Colorado, 1982: epidemiologic and entomologic studies.

    Science.gov (United States)

    Walton, T E; Webb, P A; Kramer, W L; Smith, G C; Davis, T; Holbrook, F R; Moore, C G; Schiefer, T J; Jones, R H; Janney, G C

    1987-01-01

    An epizootic of vesicular stomatitis (VS) caused by the New Jersey serotype of VS virus affected livestock and humans in 14 western states in 1982-1983. Epidemiological observations were made on at least 10% of the cattle in 4 dairy herds that were located in the vicinity of Grand Junction, Colorado. High rates of neutralizing antibody to the New Jersey serotype were seen in all cattle regardless of whether livestock in the dairy had clinical VS or a decrease in mild production. Antibody titers remained high in these cattle for as long as 2 years after the epizootic. No virus isolations were made from 32 humans with clinical signs compatible with viral disease. Entomological information was obtained during the epizootic from 23 premises in northwestern Colorado. Insect collections yielded 4 isolates from Culicoides spp. midges, 2 from C. variipennis, and 1 each from C. stellifer and C. (Selfia) spp. This is the first report of VS virus isolations from field-collected Culicoides.

  10. Oncotargeting by Vesicular Stomatitis Virus (VSV: Advances in Cancer Therapy

    Directory of Open Access Journals (Sweden)

    Suman Bishnoi

    2018-02-01

    Full Text Available Modern oncotherapy approaches are based on inducing controlled apoptosis in tumor cells. Although a number of apoptosis-induction approaches are available, site-specific delivery of therapeutic agents still remain the biggest hurdle in achieving the desired cancer treatment benefit. Additionally, systemic treatment-induced toxicity remains a major limiting factor in chemotherapy. To specifically address drug-accessibility and chemotherapy side effects, oncolytic virotherapy (OV has emerged as a novel cancer treatment alternative. In OV, recombinant viruses with higher replication capacity and stronger lytic properties are being considered for tumor cell-targeting and subsequent cell lysing. Successful application of OVs lies in achieving strict tumor-specific tropism called oncotropism, which is contingent upon the biophysical interactions of tumor cell surface receptors with viral receptors and subsequent replication of oncolytic viruses in cancer cells. In this direction, few viral vector platforms have been developed and some of these have entered pre-clinical/clinical trials. Among these, the Vesicular stomatitis virus (VSV-based platform shows high promise, as it is not pathogenic to humans. Further, modern molecular biology techniques such as reverse genetics tools have favorably advanced this field by creating efficient recombinant VSVs for OV; some have entered into clinical trials. In this review, we discuss the current status of VSV based oncotherapy, challenges, and future perspectives regarding its therapeutic applications in the cancer treatment.

  11. Oncotargeting by Vesicular Stomatitis Virus (VSV): Advances in Cancer Therapy.

    Science.gov (United States)

    Bishnoi, Suman; Tiwari, Ritudhwaj; Gupta, Sharad; Byrareddy, Siddappa N; Nayak, Debasis

    2018-02-23

    Modern oncotherapy approaches are based on inducing controlled apoptosis in tumor cells. Although a number of apoptosis-induction approaches are available, site-specific delivery of therapeutic agents still remain the biggest hurdle in achieving the desired cancer treatment benefit. Additionally, systemic treatment-induced toxicity remains a major limiting factor in chemotherapy. To specifically address drug-accessibility and chemotherapy side effects, oncolytic virotherapy (OV) has emerged as a novel cancer treatment alternative. In OV, recombinant viruses with higher replication capacity and stronger lytic properties are being considered for tumor cell-targeting and subsequent cell lysing. Successful application of OVs lies in achieving strict tumor-specific tropism called oncotropism, which is contingent upon the biophysical interactions of tumor cell surface receptors with viral receptors and subsequent replication of oncolytic viruses in cancer cells. In this direction, few viral vector platforms have been developed and some of these have entered pre-clinical/clinical trials. Among these, the Vesicular stomatitis virus (VSV)-based platform shows high promise, as it is not pathogenic to humans. Further, modern molecular biology techniques such as reverse genetics tools have favorably advanced this field by creating efficient recombinant VSVs for OV; some have entered into clinical trials. In this review, we discuss the current status of VSV based oncotherapy, challenges, and future perspectives regarding its therapeutic applications in the cancer treatment.

  12. Dynamics of vesicular-arbuscular mycorrhizae during old field succession.

    Science.gov (United States)

    Johnson, Nancy Collins; Zak, Donald R; Tilman, David; Pfleger, F L

    1991-05-01

    The species composition of vesicular-arbuscular mycorrhizal (VAM) fungal communities changed during secondary succession of abandoned fields based on a field to forest chronosequence. Twenty-five VAM fungal species were identified. Seven species were clearly early successional and five species were clearly late successional. The total number of VAM fungal species did not increase with successional time, but diversity as measured by the Shannon-Wiener index tended to increase, primarily because the community became more even as a single species, Glomus aggregatum, became less dominant in the older sites. Diversity of the VAM fungal community was positively correlated with soil C and N. The density of VAM fungi, as measured by infectivity and total spore count, first increased with time since abandonment and then decreased in the late successional forest sites. Within 12 abandoned fields, VAM fungal density increased with increasing soil pH, H 2 O soluble soil C, and root biomass, but was inversely related to extractable soil P and percent cover of non-host plant species. The lower abundance of VAM fungi in the forest sites compared with the field sites agrees with the findings of other workers and corresponds with a shift in the dominant vegetation from herbaceous VAM hosts to woody ectomycorrhizal hosts.

  13. First case report of vesicular stomatitis in the State of Paraíba, Brazil

    Directory of Open Access Journals (Sweden)

    Inácio José Clementino

    2014-10-01

    Full Text Available The present report describes the first case of vesicular stomatitis in the State of Paraíba, Brazil. Records from the Official Veterinary Services of the State of Paraíba were analyzed while responding to a suspected case of vesicular disease at a property (property I in the municipality of Pombal in which the cattle showed clinical signs and lesions of vesicular disease. Surveillance in the surrounding area revealed similar lesions in cattle at two other properties (II and III. Based on these events, the suspicion was considered well founded, and samples were collected for evaluation at the National Agricultural Laboratory of the State of Pará. The property was interdicted, and those located within a 3 km radius (perifocal from the focus were inspected. At property I, 42.86% (6/14 of the cattle showed vesicular disease lesions characterized by intense sialorrhea, ruptured oral vesicles, epithelial detachment of the tongue and muzzle, and vesicular lesions in the udder and interdigital space. Similar lesions were detected in cattle at properties II and III, affecting 80.95% (34/42 and 11.54% (3/26 of the animals, respectively. Tissue samples collected from the three properties were positive for the vesicular stomatitis virus (Indiana 3 subtype. The properties were monitored for 21 days after the last infected animal was cured, and afterwards, the three properties were released.

  14. Melatonin modulates rat myotube-acetylcholine receptors by inhibiting calmodulin.

    Science.gov (United States)

    de Almeida-Paula, Lidiana Duarte; Costa-Lotufo, Leticia V; Silva Ferreira, Zulma; Monteiro, Amanda Elisa G; Isoldi, Mauro Cesar; Godinho, Rosely O; Markus, Regina P

    2005-11-21

    Melatonin, the pineal gland hormone, modulates alpha-bungarotoxin sensitive nicotinic acetylcholine receptors in sympathetic nerve terminals, cerebellum and chick retina imposing a diurnal variation in functional responses [Markus, R.P., Zago, W.M., Carneiro, R.C., 1996. Melatonin modulation of presynaptic nicotinic acetylcholine receptors in the rat vas deferens. J. Pharmacol. Exp. Ther. 279, 18-22; Markus, R.P., Santos, J.M., Zago, W., Reno, L.A., 2003. Melatonin nocturnal surge modulates nicotinic receptors and nicotine-induced [3HI] glutamate release in rat cerebellum slices. J. Pharmacol. Exp. Ther. 305, 525-530; Sampaio, L.F.S., Hamassaki-Britto, D.E., Markus, R.P., 2005. Influence of melatonin on the development of functional nicotinic acetylcholine receptors in cultured chick retinal cells. Braz. J. Med. Biol. Res. 38, 603-613]. Here we show that in rat myotubes forskolin and melatonin reduced the number of nicotinic acetylcholine receptors expressed in plasma membrane. In addition, these cells expressed melatonin MT1 receptors, which are known to be coupled to G(i)-protein. However, the pharmacological profile of melatonin analogs regarding the reduction in cyclic AMP accumulation and number of nicotinic acetylcholine receptors did not point to a mechanism mediated by activation of G(i)-protein coupled receptors. On the other hand, calmidazolium, a classical inhibitor of calmodulin, reduced in a similar manner both effects. Considering that one isoform of adenylyl cyclase present in rat myotubes is regulated by Ca2+/calmodulin, we propose that melatonin modulates the number of nicotinic acetylcholine receptors via reduction in cyclic AMP accumulation.

  15. Flavonoids with M1 Muscarinic Acetylcholine Receptor Binding Activity

    Directory of Open Access Journals (Sweden)

    Meyyammai Swaminathan

    2014-06-01

    Full Text Available Muscarinic acetylcholine receptor-active compounds have potential for the treatment of Alzheimer’s disease. In this study, a series of natural and synthetic flavones and flavonols was assayed in vitro for their ability to inhibit radioligand binding at human cloned M1 muscarinic receptors. Several compounds were found to possess competitive binding affinity (Ki = 40–110 µM, comparable to that of acetylcholine (Ki = 59 µM. Despite the fact that these compounds lack a positively-charged ammonium group under physiological conditions, molecular modelling studies suggested that they bind to the orthosteric site of the receptor, mainly through non-polar interactions.

  16. Muscarinic acetylcholine receptor expression in aganglionic bowel.

    Science.gov (United States)

    Oue, T; Yoneda, A; Shima, H; Puri, P

    2000-01-01

    In Hirschsprung's disease (HD) there exists an overabundance of acetylcholine (ACh), which in turn stimulates excessive production of the enzyme acetylcholinesterase. Muscarinic ACh receptors (mAChRs) play an important role in smooth-muscle contraction. Recent studies have indicated five different subtypes of mAChRs encoded by five different genes, ml to m5. The purpose of this study was to investigate the expression of each mAChR subtype in aganglionic (AG) colon to further understand the pathophysiology of HD. Entire colon resected at the time of pull-through operation for HD was obtained from 14 patients. Specimens obtained at autopsy from 8 age-matched patients without gastrointestinal disease acted as controls. Frozen sections were used for indirect immunohistochemistry as well as in-situ hybridization. Immunohistochemistry was performed using specific antiserum against each mAChR subtype and in-situ hybridization was performed using specific oligonucleotide probes against ml to m5 subtypes. Messenger RNA (mRNA) was extracted from normoganglionic (NG) and AG bowel of HD patients and normal control bowel. Reverse transcription-polymerase chain reaction was performed to evaluate mRNA levels of each mAChR subtype. To adjust the levels of mRNA expression, a housekeeping gene G3PDH, known to be expressed normally, was used as an internal control. Strong m2 and m3 immunoreactivity was observed in the mucosal layer, smooth-muscle layers, and myenteric plexus of NG bowel, whereas ml immunoreactivity was only detected in the mucosal layer. The most striking finding was the abundance of m3-immunoreactive fibers in muscle layers of NG bowel while there was a total lack of m3 fibers in smooth-muscle of AG bowel. Intense mRNA signals encoding m2 and m3 and to a lesser degree ml were detected in NG bowel, and these signals were weak in AG bowel. Immunoreactivity and mRNA expression of m4 and m5 was not detected in NG or AG bowel. The lack of m3-immunoreactive fibers in the

  17. Sheared-root inocula of vesicular-arbuscular mycorrhizal fungi.

    Science.gov (United States)

    Sylvia, D M; Jarstfer, A G

    1992-01-01

    For efficient handling, vesicular-arbuscular mycorrhizal fungi should be processed into small and uniform inocula; however, processing can reduce the inoculum density. In this article we describe the preparation and use of sheared-root inocula of Glomus spp. in which inoculum densities were increased during processing. Our objectives were to determine inoculum viability and density after shearing and to ascertain if the sheared inocula could be pelletized or used with a gel carrier. Root samples were harvested from aeroponic cultures, blotted dry, cut into 1-cm lengths, and sheared in a food processor for up to 80 s. After shearing, the inoculum was washed over sieves, and the propagule density in each fraction was determined. Sheared inocula were also encapsulated in carrageenan or used in a gel carrier. Shearing aeroponically produced root inocula reduced particle size. Propagule density increased with decreasing size fraction down to a size of 63 mum, after which propagule density decreased. The weighted-average propagule density of the inoculum was 135,380 propagules g (dry weight) of sheared root material. Sheared roots were encapsulated successfully in carrageenan, and the gel served as an effective carrier. Aeroponic root inoculum was stored dry at 4 degrees C for 23 months without significant reduction in propagule density; however, this material was not appropriate for shearing. Moist roots, useful for shearing, began to lose propagule density after 1 month of storage. Shearing proved to be an excellent method to prepare viable root inocula of small and uniform size, allowing for more efficient and effective use of limited inoculum supplies.

  18. Nicotinic Acetylcholine Receptors in the Pathophysiology of Alzheimer's Disease

    DEFF Research Database (Denmark)

    Thomsen, Morten Skøtt; Andreasen T., Jesper; Arvaniti, Maria

    2016-01-01

    Nicotinic acetylcholine receptors (nAChRs) have been pursued for decades as potential molecular targets to treat cognitive dysfunction in Alzheimer's disease (AD) due to their positioning within regions of the brain critical in learning and memory, such as the prefrontal cortex and hippocampus...

  19. Measurement of anti- acetylcholine receptor auto-antibodies in ...

    African Journals Online (AJOL)

    auto-antibodies in myasthenia gravis. K. J. Steenkamp, W. Duim, M. s. Myer,. S. C. K. Malfeld, R. Anderson. Two different acetylcholine receptor (AChR) preparations derived from ... the detection of AChR auto-antibodies in serum specimens from 20 ... 4°C. Thereafter, 1 ml of washing solution (phosphate- buffered saline ...

  20. Cholinergic modulation of dopamine pathways through nicotinic acetylcholine receptors.

    NARCIS (Netherlands)

    de Kloet, S.F.; Mansvelder, H.D.; de Vries, T.J.

    2015-01-01

    Nicotine addiction is highly prevalent in current society and is often comorbid with other diseases. In the central nervous system, nicotine acts as an agonist for nicotinic acetylcholine receptors (nAChRs) and its effects depend on location and receptor composition. Although nicotinic receptors are

  1. The beneficial effect of dual inoculation of vesicular-arbuscular mycorrhizae + rhizobium on growth of white clover

    OpenAIRE

    Lin, XG.; Hao, WY.; Wu, TH.

    1993-01-01

    Investigation on the effect of phosphorus on vesicular-arbuscular mycorrhizal infection, and dual inoculation of vesicular-arbuscular mycorrhizae + rhizobium on growth of white clover under field microplots and pot experiments was conducted on fluvo-aquic soils of semi-arid region in north China. The results showed that 60 kg P205 ha in form of superphosphate was the most favorable phosphorus level for vesicular-arbuscular mycorrhizal infection ; mycorrhizal infection, nodulation, dry weight ...

  2. Transportation

    Science.gov (United States)

    2007-01-01

    Faculty ii INDUSTRY TRAVEL Domestic Assistant Deputy Under Secretary of Defense (Transportation Policy), Washington, DC Department of...developed between the railroad and trucking industries. Railroads: Today’s seven Class I freight railroad systems move 42% of the nation’s intercity ...has been successfully employed in London to reduce congestion and observed by this industry study during its travels . It is currently being

  3. Vesicular stomatitis virus (indiana 2 serotype as experimental model to study acute encephalitis – morphological features Vírus da estomatite vesicular (sorotipo indiana 2 como modelo experimental para o estudo de encefalite aguda – aspectos morfológicos

    Directory of Open Access Journals (Sweden)

    Florêncio Figueiredo Cavalcanti Neto

    2003-10-01

    Full Text Available The Vesicular Stomatitis Virus (VSV is a Vesiculovirus of the Rhabdoviridae family that infects mammals and causes vesicular lesions similar to those of foot-and-mouth disease. VSV experimental encephalitis can be induced in rodents and the symptoms are similar to those observed in rabies. However, the lesions observed in the animals´ encephalon are different. Inclusion bodies are not observed. There is necrosis, particularly in the region of the olfactory bulb, and, in some cases, ventriculitis. It was observed that the time pattern of VSV dissemination and the morphological aspects of the lesions are similar to those described in literature. The virus seems to be disseminated through the brain ventricles, being multiplied in the ependyma cells and in the neurons, besides using retrograde and anterograde transport. It was noticed that, due to the facility of virus manipulation, this experimental model has been used in innumerable research studies in several fields. If, on the one hand there are plenty of reports on the infection pathogenesis, on the other hand there are many gaps involving, for instance, aspects about virus transmission, recovery of infected animals and participation of glial cells in the acute as well as in the recovery phases.   O vírus da estomatite vesicular (VEV é um Vesiculovírus da família Rhabdoviridae que infecta mamíferos e causa lesões vesiculares semelhantes às observadas na febre aftosa. A encefalite experimental pode ser induzida em roedores e os sintomas são semelhantes aos observados na raiva; entretanto, as lesões observadas no encéfalo dos animais são diferentes. Corpúsculos de inclusão não são observados, há necrose especialmente da região do bulbo olfatório e em alguns casos, ventriculite. Observamos que o padrão temporal de disseminação do VEV e os aspectos morfológicos das lesões são similares aos descritos na literatura. O vírus parece se disseminar através dos ventr

  4. EVP-6124, a novel and selective α7 nicotinic acetylcholine receptor partial agonist, improves memory performance by potentiating the acetylcholine response of α7 nicotinic acetylcholine receptors.

    Science.gov (United States)

    Prickaerts, Jos; van Goethem, Nick P; Chesworth, Richard; Shapiro, Gideon; Boess, Frank G; Methfessel, Christoph; Reneerkens, Olga A H; Flood, Dorothy G; Hilt, Dana; Gawryl, Maria; Bertrand, Sonia; Bertrand, Daniel; König, Gerhard

    2012-02-01

    EVP-6124, (R)-7-chloro-N-quinuclidin-3-yl)benzo[b]thiophene-2-carboxamide, is a novel partial agonist of α7 neuronal nicotinic acetylcholine receptors (nAChRs) that was evaluated here in vitro and in vivo. In binding and functional experiments, EVP-6124 showed selectivity for α7 nAChRs and did not activate or inhibit heteromeric α4β2 nAChRs. EVP-6124 had good brain penetration and an adequate exposure time. EVP-6124 (0.3 mg/kg, p.o.) significantly restored memory function in scopolamine-treated rats (0.1 mg/kg, i.p.) in an object recognition task (ORT). Although donepezil at 0.1 mg/kg, p.o. or EVP-6124 at 0.03 mg/kg, p.o. did not improve memory in this task, co-administration of these sub-efficacious doses fully restored memory. In a natural forgetting test, an ORT with a 24 h retention time, EVP-6124 improved memory at 0.3 mg/kg, p.o. This improvement was blocked by the selective α7 nAChR antagonist methyllycaconitine (0.3 mg/kg, i.p. or 10 μg, i.c.v.). In co-application experiments of EVP-6124 with acetylcholine, sustained exposure to EVP-6124 in functional investigations in oocytes caused desensitization at concentrations greater than 3 nM, while lower concentrations (0.3-1 nM) caused an increase in the acetylcholine-evoked response. These actions were interpreted as representing a co-agonist activity of EVP-6124 with acetylcholine on α7 nAChRs. The concentrations of EVP-6124 that resulted in physiological potentiation were consistent with the free drug concentrations in brain that improved memory performance in the ORT. These data suggest that the selective partial agonist EVP-6124 improves memory performance by potentiating the acetylcholine response of α7 nAChRs and support new therapeutic strategies for the treatment of cognitive impairment. This article is part of a Special Issue entitled 'Post-Traumatic Stress Disorder'. Copyright © 2011 Elsevier Ltd. All rights reserved.

  5. Computed tomography of the vesicular glands: anatomical animal model (Oryctolagus cuniculus)

    International Nuclear Information System (INIS)

    Dimitrov, R.; Stamatova-Yovcheva, K.; Hamza, S.; Toneva, Y.

    2014-01-01

    Spiral CT is a non-invasive imaging method of choice for animal anatomical studies. The aim of the study was to establish the imaging anatomical features of the vesicular glands in the rabbit. Eight sexually mature healthy clinically male New Zealand rabbits of 18 months of age with body weight from 2.8 kg to 3.2 kg were used. The animals were anesthetized. As contrast medium Opti-ray350 was administrated. The computed tomography scan was complied with certain bone and soft tissue markers. For this purpose, a whole body multi-slice spiral computed tomography scanner was used. The both soft tissue glands were heterogeneous and relatively hyperdense structures, and defined in detail from the adjacent soft tissues. The urinary bladder neck was ventrally to the glands. Both vesicular glands were better differentiated each other when the rabbit is examined in abdominal recumbence. In dorsal recumbence the shape of the transversal image of the glandular finding was oval. In abdominal recumbence both the left and right soft tissue vesicular gland were defined. Transversal anatomical computed tomographic investigation of the rabbit vesicular gland is a detailed and definitive method, to study the normal morphology of these glands. Key words: Vesicular Gland. Helical Computed Tomography. Anatomy. Rabbit

  6. Modulative effects of acetylcholine and GABA upon homeostatic patterns in nerve tissue under the conditions of low-intensive irradiation

    International Nuclear Information System (INIS)

    Dvorets'kij, A.Yi.; Anan'jeva, T.V.

    1999-01-01

    Investigation of both the acetylcholine and γ-aminobutyric acid modulative effects upon the K + content and Na, K-pump activity in rat brain cortex slices after single prolonged X-irradiation or chronic (fractionated) one with doses of 25 cGy and 50 cGy (1.75 mGy/min) resulted in considerable modification of the metabolic regulation processes. A role of synaptic neuro mediators in supporting the nerve cell functional activity via modulation of the anti gradient K + transport and intracellular potassium homeostasis under any harmful factor influence is discussed

  7. Branched nanotrees with immobilized acetylcholine esterase for nanobiosensor applications

    DEFF Research Database (Denmark)

    Risveden, Klas; Dick, Kimberly A; Bhand, Sunil

    2010-01-01

    A novel lab-on-a-chip nanotree enzyme reactor is demonstrated for the detection of acetylcholine. The reactors are intended for use in the RISFET (regional ion sensitive field effect transistor) nanosensor, and are constructed from gold-tipped branched nanorod structures grown on SiN(x)-covered w......A novel lab-on-a-chip nanotree enzyme reactor is demonstrated for the detection of acetylcholine. The reactors are intended for use in the RISFET (regional ion sensitive field effect transistor) nanosensor, and are constructed from gold-tipped branched nanorod structures grown on Si......N(x)-covered wafers. Two different reactors are shown: one with simple, one-dimensional nanorods and one with branched nanorod structures (nanotrees). Significantly higher enzymatic activity is found for the nanotree reactors than for the nanorod reactors, most likely due to the increased gold surface area...

  8. Subcellular distribution of swine vesicular disease virus proteins and alterations induced in infected cells: A comparative study with foot-and-mouth disease virus and vesicular stomatitis virus

    International Nuclear Information System (INIS)

    Martin-Acebes, Miguel A.; Gonzalez-Magaldi, Monica; Rosas, Maria F.; Borrego, Belen; Brocchi, Emiliana; Armas-Portela, Rosario; Sobrino, Francisco

    2008-01-01

    The intracellular distribution of swine vesicular disease virus (SVDV) proteins and the induced reorganization of endomembranes in IBRS-2 cells were analyzed. Fluorescence to new SVDV capsids appeared first upon infection, concentrated in perinuclear circular structures and colocalized to dsRNA. As in foot-and-mouth disease virus (FMDV)-infected cells, a vesicular pattern was predominantly found in later stages of SVDV capsid morphogenesis that colocalized with those of non-structural proteins 2C, 2BC and 3A. These results suggest that assembly of capsid proteins is associated to the replication complex. Confocal microscopy showed a decreased fluorescence to ER markers (calreticulin and protein disulfide isomerase), and disorganization of cis-Golgi gp74 and trans-Golgi caveolin-1 markers in SVDV- and FMDV-, but not in vesicular stomatitis virus (VSV)-infected cells. Electron microscopy of SVDV-infected cells at an early stage of infection revealed fragmented ER cisternae with expanded lumen and accumulation of large Golgi vesicles, suggesting alterations of vesicle traffic through Golgi compartments. At this early stage, FMDV induced different patterns of ER fragmentation and Golgi alterations. At later stages of SVDV cytopathology, cells showed a completely vacuolated cytoplasm containing vesicles of different sizes. Cell treatment with brefeldin A, which disrupts the Golgi complex, reduced SVDV (∼ 5 log) and VSV (∼ 4 log) titers, but did not affect FMDV growth. Thus, three viruses, which share target tissues and clinical signs in natural hosts, induce different intracellular effects in cultured cells

  9. Long release latencies are increased by acetylcholine at frog endplate

    Czech Academy of Sciences Publication Activity Database

    Samigullin, D.; Bukharaeva, E. A.; Nikolsky, E.; Adámek, S.; Vyskočil, František

    2003-01-01

    Roč. 52, č. 4 (2003), s. 475-480 ISSN 0862-8408 R&D Projects: GA ČR GA305/02/1333; GA ČR GA202/02/1213 Grant - others:RFBR(RU) 02/04/48901 Institutional research plan: CEZ:AV0Z5011922; CEZ:MSM 113100003 Keywords : quantal release * acetylcholine * synaptic latency Subject RIV: ED - Physiology Impact factor: 0.939, year: 2003

  10. The beneficial effect of dual inoculation of vesicular-arbuscular mycorrhizae + rhizobium on growth of white clover

    Directory of Open Access Journals (Sweden)

    Lin, XG.

    1993-01-01

    Full Text Available Investigation on the effect of phosphorus on vesicular-arbuscular mycorrhizal infection, and dual inoculation of vesicular-arbuscular mycorrhizae + rhizobium on growth of white clover under field microplots and pot experiments was conducted on fluvo-aquic soils of semi-arid region in north China. The results showed that 60 kg P205 ha in form of superphosphate was the most favorable phosphorus level for vesicular-arbuscular mycorrhizal infection ; mycorrhizal infection, nodulation, dry weight of shoots and roots, total uptake of nitrogen, phosphorus and other elements, the final yields and recovery of phosphorus of white clover were significantly increased by vesicular-arbuscular mycorrhizal inoculation and dual inoculation with vesicular-arbuscular mycorrhizal fungi and rhizobium. The highest response of inoculation was obtained by adding fertilizer phosphorus at the level of 60 kg P205 ha in form of superphosphate.

  11. Gold nanoparticle–choline complexes can block nicotinic acetylcholine receptors

    Directory of Open Access Journals (Sweden)

    Chur Chin

    2010-04-01

    Full Text Available Chur Chin1, In Kyeom Kim2, Dong Yoon Lim3, Ki Suk Kim4, Hyang Ae Lee4, Eun Joo Kim41Department of Pediatrics, Fatima Hospital, Daegu, Korea; 2Department of Pharmacology, School of Medicine, Kyungpook National University, Daegu, Korea; 3Department of Pharmacology, School of Medicine, Chosun University, Gwangju, Korea; 4Korea Institute of Toxicology, Daejeon, KoreaAbstract: We identified a novel class of direct ion-channel blockers of ligand-gated ion channels called the gold nanoparticle–choline complex. Negatively charged gold nanoparticles (1.4 nm block ion pores by binding to the sulfur group of the cysteine loop of nicotinic acetylcholine receptors (nAChRs, and currents evoked by acetylcholine (Ach can break these bonds. The current evoked by ACh in nAChRs was blocked directly in ion pores by the gold nanoparticle–choline complex. In adrenal-gland perfusion studies, the complex also blocked nAChRs by diminishing catecholamine release by about 75%. An in vivo study showed muscle relaxation in rats after injection of the complex. These results will foster the application of gold nanoparticles as a direct ion-channel blocker. Keywords: negatively charged gold nanoparticle, choline, gold–sulfur bond, nicotinic acetylcholine receptor, direct ion-channel blocker

  12. Branched nanotrees with immobilized acetylcholine esterase for nanobiosensor applications

    Energy Technology Data Exchange (ETDEWEB)

    Risveden, Klas; Bhand, Sunil; Danielsson, Bengt [Department of Pure and Applied Biochemistry, Center for Chemistry and Chemical Engineering, Lund University, PO Box 124, SE-22100 Lund (Sweden); Dick, Kimberly A; Samuelson, Lars [Solid State Physics, Lund University, Box 118, S-22100 Lund (Sweden); Rydberg, Patrik, E-mail: Kimberly.Dick@ftf.lth.se [Department of Medicinal Chemistry, Faculty of Pharmaceutical Sciences, University of Copenhagen, Universitetsparken 2, DK-2100 Copenhagen (Denmark)

    2010-02-05

    A novel lab-on-a-chip nanotree enzyme reactor is demonstrated for the detection of acetylcholine. The reactors are intended for use in the RISFET (regional ion sensitive field effect transistor) nanosensor, and are constructed from gold-tipped branched nanorod structures grown on SiN{sub x}-covered wafers. Two different reactors are shown: one with simple, one-dimensional nanorods and one with branched nanorod structures (nanotrees). Significantly higher enzymatic activity is found for the nanotree reactors than for the nanorod reactors, most likely due to the increased gold surface area and thereby higher enzyme binding capacity. A theoretical calculation is included to show how the enzyme kinetics and hence the sensitivity can be influenced and increased by the control of electrical fields in relation to the active sites of enzymes in an electronic biosensor. The possible effects of electrical fields employed in the RISFET on the function of acetylcholine esterase is investigated using quantum chemical methods, which show that the small electric field strengths used are unlikely to affect enzyme kinetics. Acetylcholine esterase activity is determined using choline oxidase and peroxidase by measuring the amount of choline formed using the chemiluminescent luminol reaction.

  13. Tethering factors as organizers of intracellular vesicular traffic.

    Science.gov (United States)

    Yu, I-Mei; Hughson, Frederick M

    2010-01-01

    Intracellular trafficking entails the budding, transport, tethering, and fusion of transport vesicles and other membrane carriers. Here we review recent progress toward a mechanistic understanding of vesicle tethering. The known tethering factors are large complexes important for one or more intracellular trafficking pathways and are capable of interacting directly with many of the other principal components of the cellular trafficking machinery. Our review emphasizes recent developments in the in vitro reconstitution of vesicle tethering and the structural characterization of multisubunit tethering factors. The combination of these and other approaches has led to exciting progress toward understanding how these essential nanomachines work.

  14. Interactions of calcium homeostasis, acetylcholine metabolism, behavior and 3, 4-diaminopyridine during aging

    International Nuclear Information System (INIS)

    Gibson, G.E.; Peterson, C.

    1986-01-01

    Acetylcholine synthesis declines with aging in both whole brain and in various brain regions. Since neither enzyme activities nor acetylcholine concentrations, accurately reflect the dynamics of the cholinergic system, in vivo acetylcholine formation was measured. Incorporation of U-C 14-glucose of 2 H 4 choline into whole brain acetylcholine decreases from 100% (3 months) in two strains of mice. The diminished synthesis is apparently not due to a lack of precursor availability because U- C 14-glucose and 2 H 4 choline entry into the brain is similar at all ages. It is shown that altered brain calcium homeostasis during aging may underlie the deficits in acetylcholine metabolism, as well as those in behavior. Diminished calcium uptake during aging parallels the decline in the calcium dependent release of acetylcholine

  15. Vesicular Stomatitis Virus Infection Promotes Immune Evasion by Preventing NKG2D-Ligand Surface Expression

    DEFF Research Database (Denmark)

    Jensen, Helle; Andresen, Lars; Nielsen, Jens

    2011-01-01

    Vesicular stomatitis virus (VSV) has recently gained attention for its oncolytic ability in cancer treatment. Initially, we hypothesized that VSV infection could increase immune recognition of cancer cells through induction of the immune stimulatory NKG2D-ligands. Here we show that VSV infection ...

  16. Signifiance of Arginine 20 in the 2A protease for swine vesicular disease virus pathogenicity

    DEFF Research Database (Denmark)

    Inoue, Toru; Zhang, Zhidong; Wang, Leyuan

    2007-01-01

    Pathogenic and attenuated strains of swine vesicular disease virus (SVDV), an enterovirus, have been characterized previously and, by using chimeric infectious cDNA clones, the key determinants of pathogenicity in pigs have been mapped to the coding region for 1D–2A. Within this region, residue 20...

  17. Nanoscale distribution of presynaptic Ca(2+) channels and its impact on vesicular release during development.

    Science.gov (United States)

    Nakamura, Yukihiro; Harada, Harumi; Kamasawa, Naomi; Matsui, Ko; Rothman, Jason S; Shigemoto, Ryuichi; Silver, R Angus; DiGregorio, David A; Takahashi, Tomoyuki

    2015-01-07

    Synaptic efficacy and precision are influenced by the coupling of voltage-gated Ca(2+) channels (VGCCs) to vesicles. But because the topography of VGCCs and their proximity to vesicles is unknown, a quantitative understanding of the determinants of vesicular release at nanometer scale is lacking. To investigate this, we combined freeze-fracture replica immunogold labeling of Cav2.1 channels, local [Ca(2+)] imaging, and patch pipette perfusion of EGTA at the calyx of Held. Between postnatal day 7 and 21, VGCCs formed variable sized clusters and vesicular release became less sensitive to EGTA, whereas fixed Ca(2+) buffer properties remained constant. Experimentally constrained reaction-diffusion simulations suggest that Ca(2+) sensors for vesicular release are located at the perimeter of VGCC clusters (<30 nm) and predict that VGCC number per cluster determines vesicular release probability without altering release time course. This "perimeter release model" provides a unifying framework accounting for developmental changes in both synaptic efficacy and time course. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.

  18. Toluene-induced, Ca2+-dependent vesicular catecholamine release in rat PC12 cells

    NARCIS (Netherlands)

    Westerink, R.H.S.|info:eu-repo/dai/nl/239425952; Vijverberg, H.P.M.|info:eu-repo/dai/nl/068856474

    2002-01-01

    Acute effects of toluene on vesicular catecholamine release from intact PC12 phaeochromocytoma cells have been investigated using carbon fiber microelectrode amperometry. The frequency of vesicles released is low under basal conditions and is enhanced by depolarization. Toluene causes an increase in

  19. The development and significance of vesicular-arbuscular mycorrhizas as influenced by agricultural practices

    NARCIS (Netherlands)

    Ruissen, M.A.

    1982-01-01

    The development and significance of vesicular- arbuscular mycorrhizas (VAM) in wheat and potatoes have been studied in relation to various farming systems and agricultural practices. The effects of farming systems on VAM have been observed on three neighbouring experimental farms in the vicinity of

  20. Stimulation of vesicular-arbuscular mycorrhizal fungi by mycotrophic and nonmycotrophic plant root systems.

    Science.gov (United States)

    Schreiner, R P; Koide, R T

    1993-08-01

    Transformed root cultures of three nonmycotrophic and one mycotrophic plant species stimulated germination and hyphal growth of the vesicular-arbuscular mycorrhizal fungus Glomus etunicatum (Becker & Gerd.) in a gel medium. However, only roots of the mycotrophic species (carrot) supported continued hyphal exploration after 3 to 4 weeks and promoted appressoria formation by G. etunicatum.

  1. Stimulation of Vesicular-Arbuscular Mycorrhizal Fungi by Mycotrophic and Nonmycotrophic Plant Root Systems

    OpenAIRE

    Schreiner, R. Paul; Koide, Roger T.

    1993-01-01

    Transformed root cultures of three nonmycotrophic and one mycotrophic plant species stimulated germination and hyphal growth of the vesicular-arbuscular mycorrhizal fungus Glomus etunicatum (Becker & Gerd.) in a gel medium. However, only roots of the mycotrophic species (carrot) supported continued hyphal exploration after 3 to 4 weeks and promoted appressoria formation by G. etunicatum.

  2. Detection of three porcine vesicular viruses using multiplex real-time primer-probe energy transfer

    DEFF Research Database (Denmark)

    Rasmussen, Thomas Bruun; Uttenthal, Åse; Aguero, M.

    2006-01-01

    Rapid identification of the etiologic agent in infected animals is important for the control of an outbreak of vesicular disease in livestock. We have in the present study developed a multiplex real-time reverse transcription-PCR, based on primer-probe energy transfer (PriProET), for simultaneous...

  3. Influence of the preparation route on the supramolecular organization of lipids in a vesicular system

    DEFF Research Database (Denmark)

    Elizondo, Elisa; Larsen, Jannik; Hatzakis, Nikos

    2012-01-01

    A confocal fluorescence microscopy-based assay was used for studying the influence of the preparation route on the supramolecular organization of lipids in a vesicular system. In this work, vesicles composed of cholesterol and CTAB (1/1 mol %) or cholesterol and DOPC (2/8 mol %) and incorporating...

  4. Why we forget our dreams: Acetylcholine and norepinephrine in wakefulness and REM sleep.

    Science.gov (United States)

    Becchetti, Andrea; Amadeo, Alida

    2016-01-01

    The ascending fibers releasing norepinephrine and acetylcholine are highly active during wakefulness. In contrast, during rapid-eye-movement sleep, the neocortical tone is sustained mainly by acetylcholine. By comparing the different physiological features of the norepinephrine and acetylcholine systems in the light of the GANE (glutamate amplifies noradrenergic effects) model, we suggest how to interpret some functional differences between waking and rapid-eye-movement sleep.

  5. Porters and neurotransmitter transporters.

    Science.gov (United States)

    Nelson, N; Lill, H

    1994-11-01

    Uptake of neurotransmitters involves multiple transporters acting in different brain locations under different physiological conditions. The vesicular transporters are driven by a proton-motive force generated by a V-ATPase and their substrates are taken up via proton/substrate exchange. The plasma membrane transporters are driven by an electrochemical gradient of sodium generated by a Na+/K(+)-ATPase. Two distinct families of transporters were identified in this group. One cotransports sodium with glutamate and other amino acids and requires additionally an outwardly directed potassium gradient. The second cotransports sodium, chloride and a variety of neurotransmitters, including gamma-aminobutyric acid (GABA), glycine and monoamines. Genes and cDNA encoding several members of the latter family have been cloned and studied in detail. The structure and function as well as the evolutionary relationships among these neurotransmitter transporters are discussed.

  6. Contrasting actions of philanthotoxin-343 and philanthotoxin-(12) on human muscle nicotinic acetylcholine receptors

    DEFF Research Database (Denmark)

    Brier, Tim J; Mellor, Ian R; Tikhonov, Denis B

    2003-01-01

    Whole-cell recordings and outside-out patch recordings from TE671 cells were made to investigate antagonism of human muscle nicotinic acetylcholine receptors (nAChR) by the philanthotoxins, PhTX-343 and PhTX-(12). When coapplied with acetylcholine (ACh), PhTX-343 caused activation-dependent, nonc......Whole-cell recordings and outside-out patch recordings from TE671 cells were made to investigate antagonism of human muscle nicotinic acetylcholine receptors (nAChR) by the philanthotoxins, PhTX-343 and PhTX-(12). When coapplied with acetylcholine (ACh), PhTX-343 caused activation...

  7. Back to the future: Rational maps for exploring acetylcholine receptor space and time.

    Science.gov (United States)

    Tessier, Christian J G; Emlaw, Johnathon R; Cao, Zhuo Qian; Pérez-Areales, F Javier; Salameh, Jean-Paul J; Prinston, Jethro E; McNulty, Melissa S; daCosta, Corrie J B

    2017-11-01

    Global functions of nicotinic acetylcholine receptors, such as subunit cooperativity and compatibility, likely emerge from a network of amino acid residues distributed across the entire pentameric complex. Identification of such networks has stymied traditional approaches to acetylcholine receptor structure and function, likely due to the cryptic interdependency of their underlying amino acid residues. An emerging evolutionary biochemistry approach, which traces the evolutionary history of acetylcholine receptor subunits, allows for rational mapping of acetylcholine receptor sequence space, and offers new hope for uncovering the amino acid origins of these enigmatic properties. Copyright © 2017 Elsevier B.V. All rights reserved.

  8. Vesicular transport protein Arf6 modulates cytoskeleton dynamics for polar body extrusion in mouse oocyte meiosis.

    Science.gov (United States)

    Duan, Xing; Zhang, Hao-Lin; Pan, Meng-Hao; Zhang, Yu; Sun, Shao-Chen

    2018-02-01

    Arf6 (ADP-ribosylation factor 6) is known to play important roles in membrane dynamics through the regulation of actin filament reorganization for multiple cellular processes such as cytokinesis, phagocytosis, cell migration and tumor cell invasion. However, the functions of Arf6 in mammalian oocyte meiosis have not been clarified. In present study we showed that Arf6 expressed in mouse oocytes and was mainly distributed around the spindle during meiosis. Depletion of Arf6 by morpholino microinjection caused oocytes failing to extrude first polar body. Further analysis indicated that Arf6 knock down caused the aberrant actin distribution, which further induced the failure of meiotic spindle movement. And the loss of oocyte polarity also confirmed this. The regulation of Arf6 on actin filaments in mouse oocytes might be due to its effects on the phosphorylation level of cofilin and the expression of Arp2/3 complex. Moreover, we found that the decrease of Arf6 caused the disruption of spindle formation, indicating the multiple roles of Arf6 on cytoskeleton dynamics in meiosis. In summary, our results indicated that Arf6 was involved in mouse oocyte meiosis through its functional roles in actin-mediated spindle movement and spindle organization. Copyright © 2017 Elsevier B.V. All rights reserved.

  9. Subcellular localization analysis of the closely related Fps/Fes and Fer protein-tyrosine kinases suggests a distinct role for Fps/Fes in vesicular trafficking.

    Science.gov (United States)

    Zirngibl, R; Schulze, D; Mirski, S E; Cole, S P; Greer, P A

    2001-05-15

    The subcellular localizations of the Fps/Fes and closely related Fer cytoplasmic tyrosine kinases were studied using green fluorescent protein (GFP) fusions and confocal fluorescence microscopy. In contrast to previous reports, neither kinase localized to the nucleus. Fer was diffusely cytoplasmic throughout the cell cycle. Fps/Fes also displayed a diffuse cytoplasmic localization, but in addition it showed distinct accumulations in cytoplasmic vesicles as well as in a perinuclear region consistent with the Golgi. This localization was very similar to that of TGN38, a known marker of the trans Golgi. The localization of Fps/Fes and TGN38 were both perturbed by brefeldin A, a fungal metabolite that disrupts the Golgi apparatus. Fps/Fes was also found to colocalize to various extents with several Rab proteins, which are members of the monomeric G-protein superfamily involved in vesicular transport between specific subcellular compartments. Using Rabs that are involved in endocytosis (Rab5B and Rab7) or exocytosis (Rab1A and Rab3A), we showed that Fps/Fes is localized in both pathways. These results suggest that Fps/Fes may play a general role in the regulation of vesicular trafficking. Copyright 2001 Academic Press.

  10. Acetylcholine synthesis and possible functions during sea urchin development

    Directory of Open Access Journals (Sweden)

    C Angelini

    2009-06-01

    Full Text Available Cholinergic neurotransmitter system molecules were found to play a role during fertilisation and early cell cycles of a large number of invertebrate and vertebrate organisms. In this study, we investigated the presence and possible function of choline acetyltransferase (ChAT, the biosynthetic enzyme of acetylcholine in gametes of the sea urchin, Paracentrotus lividus, through localisation and functional studies. ChAT-like molecules were detected in oocytes, mature eggs and zygotes with indirect immunofluorescence methods. Positive immunoreactivity was found in the ovarian egg cytoplasm and surface as well as at the zygote surface. This suggests the eggs' capacity to autonomously synthesise acetylcholine (ACh, the signal molecule of the cholinergic system. Acetylcholinesterase (AChE, the lytic enzyme of acetylcholine was also found in ovarian eggs, with a similar distribution; however, it disappeared after fertilisation. Ultrastructural ChAT localisation in sperms, which was carried out with the immuno-gold method, showed immunoreactivity in the acrosome of unreacted sperms and at the head surface of reacted sperms. In order to verify a functional role of ACh during fertilization and sea urchin development, in vivo experiments were performed. Exposure of the eggs before fertilisation to 1 mM ACh + 1 ?M eserine caused an incomplete membrane depolarisation and consequently enhanced polyspermy, while lower concentrations of ACh caused developmental anomalies. The exposure of zygotes to 0,045 AChE Units/mL of sea water caused developmental anomalies as well, in 50% of the embryos. Altogether, these findings and other previously obtained results, suggest that the cholinergic system may subserve two different tasks during development, according to which particular type of ACh receptor is active during each temporal window. The first function, taking place in the course of fertilisation is a result of autonomously synthesised ACh in sperms, while the

  11. Impact of acetylcholine and nicotine on human osteoclastogenesis in vitro.

    Science.gov (United States)

    Ternes, Sebastian; Trinkaus, Katja; Bergen, Ivonne; Knaack, Sven; Gelinsky, Michael; Kilian, Olaf; Heiss, Christian; Lips, Katrin Susanne

    2015-11-01

    Recent studies showed that the non-neuronal cholinergic system (NNCS) is taking part in bone metabolism. Most studies investigated its role in osteoblasts, but up to now, the involvement of the NNCS in human osteoclastogenesis remains relatively unclear. Thus, aim of the present study was to determine whether the application of acetylcholine (ACh, 10(−4) M), nicotine (10(−6) M), mineralized collagen membranes or brain derived neurotrophic factor (BDNF, 40 ng/mL) influences the mRNA regulation of molecular components of the NNCS and the neurotrophin family during osteoclastogenesis. Peripheral blood mononuclear cells (PBMCs) were isolated from the blood of young healthy donors (n = 8) and incubated with bone fragments and osteoclast differentiation media for 21 days. All the results are based on the measurement of RNA. Real-time RT-PCR analysis demonstrated a down-regulation of nicotinic acetylcholine receptor (nAChR) subunit α2 and muscarinic acetylcholine receptor (mAChR) M3by osteoclastogenesis while BDNF mRNA expression was not regulated. Application of ACh, nicotine, BDNF or collagen membranes did not affect osteoclastic differentiation.No regulation was detected for nAChR subunit α7, tropomyosin-related kinase receptor B (TrkB), and cholineacetyl transferase (ChAT). Taken together, we assume that the transcriptional level of osteoclastogenesis of healthy young humans is not regulated by BDNF, ACh, and nicotine. Thus, these drugs do not seem to worsen bone degradation and might therefore be suitable as modulators of bone substitution materials if having a positive effect on bone formation.

  12. Immuno-detection of OCTN1 (SLC22A4) in HeLa cells and characterization of transport function.

    Science.gov (United States)

    Pochini, Lorena; Scalise, Mariafrancesca; Indiveri, Cesare

    2015-11-01

    OCTN1 was immuno-detected in the cervical cancer cell HeLa, in which the complete pattern of acetylcholine metabolizing enzymes is expressed. Comparison of immuno-staining intensity of HeLa OCTN1 with the purified recombinant human OCTN1 allowed measuring the specific OCTN1 concentration in the HeLa cell extract and, hence calculating the HeLa OCTN1 specific transport activity that was about 10 nmol×min(-1)×mg protein(-1), measured as uptake of [(3)H]acetylcholine in proteoliposomes reconstituted with HeLa extract. This value was very similar to the specific activity of the recombinant protein. Acetylcholine transport was suppressed by incubation of the protein or proteoliposomes with the anti-OCTN1 antibody and was strongly inhibited by PLP and MTSEA, known inhibitors of OCTN1. The absence of ATP in the internal side of proteoliposomes strongly impaired transport function of both the HeLa and, as expected, the recombinant OCTN1. HeLa OCTN1 was inhibited by spermine, NaCl (Na(+)), TEA, γ-butyrobetaine, choline, acetylcarnitine and ipratropium but not by neostigmine. Besides acetylcholine, choline was taken up by HeLa OCTN1 proteoliposomes. The transporter catalyzed also acetylcholine and choline efflux which, differently from uptake, was not inhibited by MTSEA. Time course of [(3)H]acetylcholine uptake in intact HeLa cells was measured. As in proteoliposomes, acetylcholine transport in intact cells was inhibited by TEA and NaCl. Efflux of [(3)H]acetylcholine occurred in intact cells, as well. The experimental data concur in demonstrating a role of OCTN1 in transporting acetylcholine and choline in HeLa cells. Copyright © 2015 Elsevier B.V. All rights reserved.

  13. Unraveling the high- and low-sensitivity agonist responses of nicotinic acetylcholine receptors

    DEFF Research Database (Denmark)

    Harpsøe, Kasper; Ahring, Philip K; Christensen, Jeppe K

    2011-01-01

    The neuronal a4ß2 nicotinic acetylcholine receptors exist as two distinct subtypes, (a4)(2)(ß2)(3) and (a4)(3)(ß2)(2), and biphasic responses to acetylcholine and other agonists have been ascribed previously to coexistence of these two receptor subtypes. We offer a novel and radical explanation...

  14. Conformationally restrained carbamoylcholine homologues. Synthesis, pharmacology at neuronal nicotinic acetylcholine receptors and biostructural considerations

    DEFF Research Database (Denmark)

    de la Fuente Revenga, M; Balle, Thomas; Jensen, Anders A.

    2015-01-01

    Exploration of small selective ligands for the nicotinic acetylcholine receptors (nAChRs) based on acetylcholine (ACh) has led to the development of potent agonists with clear preference for the α4β2 nAChR, the most prevalent nAChR subtype in the central nervous system. In this work we present...

  15. Characterization of an acetylcholine receptor gene of Haemonchus contortus in relation to levamisole resistance

    NARCIS (Netherlands)

    Hoekstra, R.; Visser, A.; Wiley, L. J.; Weiss, A. S.; Sangster, N. C.; Roos, M. H.

    1997-01-01

    The anthelminitic drug levamisole is thought to bind to nicotinic acetylcholine receptors of nematodes. It is possible that resistance to this drug is associated with either a change in binding characteristics or a reduction in the number of nicotinic acetylcholine receptors. Therefore, the

  16. Characterisation of an acetylcholine receptor gene of Haemonchus contortus in relation to levamisole resistance

    NARCIS (Netherlands)

    Hoekstra, R.; Visser, A.; Wiley, L.; Weiss, A.S.; Sangster, N.C.; Roos, M.H.

    1997-01-01

    The anthelmintic drug levamisole is thought to bind to nicotinic acetylcholine receptors of nematodes. It is possible that resistance to this drug is associated with either a change in binding characteristics or a reduction in the number of nicotinic acetylcholine receptors. Therefore, the molecular

  17. Stochastic Model of Vesicular Sorting in Cellular Organelles

    Science.gov (United States)

    Vagne, Quentin; Sens, Pierre

    2018-02-01

    The proper sorting of membrane components by regulated exchange between cellular organelles is crucial to intracellular organization. This process relies on the budding and fusion of transport vesicles, and should be strongly influenced by stochastic fluctuations, considering the relatively small size of many organelles. We identify the perfect sorting of two membrane components initially mixed in a single compartment as a first passage process, and we show that the mean sorting time exhibits two distinct regimes as a function of the ratio of vesicle fusion to budding rates. Low ratio values lead to fast sorting but result in a broad size distribution of sorted compartments dominated by small entities. High ratio values result in two well-defined sorted compartments but sorting is exponentially slow. Our results suggest an optimal balance between vesicle budding and fusion for the rapid and efficient sorting of membrane components and highlight the importance of stochastic effects for the steady-state organization of intracellular compartments.

  18. Acetylcholine receptors and cholinergic ligands: biochemical and genetic aspects in Torpedo californica and Drosophila melanogaster

    International Nuclear Information System (INIS)

    Rosenthal, L.S.

    1987-01-01

    This study evaluates the biochemical and genetic aspects of the acetylcholine receptor proteins and cholinergic ligands in Drosophila melanogaster and Torpedo californica. Included are (1) a comparative study of nicotinic ligand-induced cation release from acetylcholine receptors isolated from Torpedo californica and from Drosophila melanogaster, (2) solution studies of the cholinergic ligands, nikethamide and ethamivan, aimed at measuring internal molecular rotational barriers in solvents of different polarity; and (3) the isolation and characterization of the gene(s) for the acetylcholine receptor in Drosophila melasogaster. Acetylcholine receptor proteins isolated from Drosphila melanogaster heads were found to behave kinetically similar (with regards to cholinergic ligand-induced 155 Eu: 3+ displacement from prelabeled proteins) to receptor proteins isolated from Torpedo californica electric tissue, providing additional biochemical evidence for the existence of a Drosophila acetylcholine receptor

  19. Organophosphate acetylcholine esterase inhibitor poisoning from a home-made shampoo.

    Science.gov (United States)

    Sadaka, Yair; Broides, Arnon; Tzion, Raffi Lev; Lifshitz, Matitiahu

    2011-07-01

    Organophosphate acetylcholine esterase inhibitor poisoning is a major health problem in children. We report an unusual cause of organophosphate acetylcholine esterase inhibitor poisoning. Two children were admitted to the pediatric intensive care unit due to organophosphate acetylcholine esterase inhibitor poisoning after exposure from a home-made shampoo that was used for the treatment of head lice. Owing to no obvious source of poisoning, the diagnosis of organophosphate acetylcholine esterase inhibitor poisoning in one of these patients was delayed. Both patients had an uneventful recovery. Organophosphate acetylcholine esterase inhibitor poisoning from home-made shampoo is possible. In cases where the mode of poisoning is unclear, direct questioning about the use of home-made shampoo is warranted, in these cases the skin and particularly the scalp should be rinsed thoroughly as soon as possible.

  20. Analysis of micellar and vesicular lecithin and cholesterol in model bile using 1H- and 31P-NMR

    NARCIS (Netherlands)

    de Graaf, M. P.; Groen, A. K.; Bovée, W. M.

    1995-01-01

    The distribution of phospholipid and cholesterol between the vesicular and micellar phases in bile plays an important role in the formation of cholesterol gallstones. Conventional analytical procedures to determine the distribution are potentially unreliable because they disturb the distribution of

  1. Adult celiac disease with acetylcholine receptor antibody positive myasthenia gravis

    Institute of Scientific and Technical Information of China (English)

    Hugh J Freeman; Helen R Gillett; Peter M Gillett; Joel Oger

    2009-01-01

    Celiac disease has been associated with some autoimmune disorders. A 40-year-old competitive strongman with celiac disease responded to a glutenfree diet, but developed profound and generalized motor weakness with acetylcholine receptor antibody positive myasthenia gravis, a disorder reported to occur in about 1 in 5000. This possible relationship between myasthenia gravis and celiac disease was further explored in serological studies. Frozen stored serum samples from 23 acetylcholine receptor antibody positive myasthenia gravis patients with no intestinal symptoms were used to screen for celiac disease. Both endomysial and tissue transglutaminase antibodies were examined. One of 23 (or, about 4.3%) was positive for both IgA-endomysial and IgA tissue transglutaminase antibodies. Endoscopic studies subsequently showed duodenal mucosal scalloping and biopsies confirmed the histopathological changes of celiac disease. Celiac disease and myasthenia gravis may occur together more often than is currently appreciated. The presence of motor weakness in celiac disease may be a clue to occult myasthenia gravis, even in the absence of intestinal symptoms.

  2. Nucleus Accumbens Acetylcholine Receptors Modulate Dopamine and Motivation.

    Science.gov (United States)

    Collins, Anne L; Aitken, Tara J; Greenfield, Venuz Y; Ostlund, Sean B; Wassum, Kate M

    2016-11-01

    Environmental reward-predictive cues can motivate reward-seeking behaviors. Although this influence is normally adaptive, it can become maladaptive in disordered states, such as addiction. Dopamine release in the nucleus accumbens core (NAc) is known to mediate the motivational impact of reward-predictive cues, but little is known about how other neuromodulatory systems contribute to cue-motivated behavior. Here, we examined the role of the NAc cholinergic receptor system in cue-motivated behavior using a Pavlovian-to-instrumental transfer task designed to assess the motivating influence of a reward-predictive cue over an independently-trained instrumental action. Disruption of NAc muscarinic acetylcholine receptor activity attenuated, whereas blockade of nicotinic receptors augmented cue-induced invigoration of reward seeking. We next examined a potential dopaminergic mechanism for this behavioral effect by combining fast-scan cyclic voltammetry with local pharmacological acetylcholine receptor manipulation. The data show evidence of opposing modulation of cue-evoked dopamine release, with muscarinic and nicotinic receptor antagonists causing suppression and augmentation, respectively, consistent with the behavioral effects of these manipulations. In addition to demonstrating cholinergic modulation of naturally-evoked and behaviorally-relevant dopamine signaling, these data suggest that NAc cholinergic receptors may gate the expression of cue-motivated behavior through modulation of phasic dopamine release.

  3. Caenorhabditis elegans nicotinic acetylcholine receptors are required for nociception

    Science.gov (United States)

    Cohen, Emiliano; Chatzigeorgiou, Marios; Husson, Steven J.; Steuer-Costa, Wagner; Gottschalk, Alexander; Schafer, William R.; Treinin, Millet

    2014-01-01

    Polymodal nociceptors sense and integrate information on injurious mechanical, thermal, and chemical stimuli. Chemical signals either activate nociceptors or modulate their responses to other stimuli. One chemical known to activate or modulate responses of nociceptors is acetylcholine (ACh). Across evolution nociceptors express subunits of the nicotinic acetylcholine receptor (nAChR) family, a family of ACh-gated ion channels. The roles of ACh and nAChRs in nociceptor function are, however, poorly understood. Caenorhabditis elegans polymodal nociceptors, PVD, express nAChR subunits on their sensory arbor. Here we show that mutations reducing ACh synthesis and mutations in nAChR subunits lead to defects in PVD function and morphology. A likely cause for these defects is a reduction in cytosolic calcium measured in ACh and nAChR mutants. Indeed, overexpression of a calcium pump in PVD mimics defects in PVD function and morphology found in nAChR mutants. Our results demonstrate, for the first time, a central role for nAChRs and ACh in nociceptor function and suggest that calcium permeating via nAChRs facilitates activity of several signaling pathways within this neuron. PMID:24518198

  4. Increased Sporulation of Vesicular-Arbuscular Mycorrhizal Fungi by Manipulation of Nutrient Regimens †

    OpenAIRE

    Douds, David D.; Schenck, N. C.

    1990-01-01

    Adjustment of pot culture nutrient solutions increased root colonization and sporulation of vesicular-arbuscular mycorrhizal (VAM) fungi. Paspalum notatum Flugge and VAM fungi were grown in a sandy soil low in N and available P. Hoagland nutrient solution without P enhanced sporulation in soil and root colonization of Acaulospora longula, Scutellospora heterogama, Gigaspora margarita, and a wide range of other VAM fungi over levels produced by a tap water control or nutrient solutions contain...

  5. JST Thesaurus Headwords and Synonyms: vesicular stomatitis virus [MeCab user dictionary for science technology term[Archive

    Lifescience Database Archive (English)

    Full Text Available MeCab user dictionary for science technology term vesicular stomatitis virus 名詞 一般 ...* * * * 水疱性口内炎ウイルス スイホウセイコウナイエンウイルス スイホーセイコーナイエンウイルス Thesaurus2015 200906056003651861 C LS07 UNKNOWN_2 vesicular stomatitis virus

  6. Identification of a Vesicular-Arbuscular Mycorrhizal Fungus by Using Monoclonal Antibodies in an Enzyme-Linked Immunosorbent Assay †

    OpenAIRE

    Wright, Sara F.; Morton, Joseph B.; Sworobuk, Janis E.

    1987-01-01

    Spore morphology is currently used to identify species of vesicular-arbuscular mycorrhizal fungi. We report the first use of a highly specific immunological method for identification of a vesicular-arbuscular mycorrhizal fungus. Two monoclonal antibodies were produced against Glomus occultum. Monoclonal antibodies reacted strongly with both spores and hyphae in an indirect enzyme-linked immunosorbent assay. All other mycorrhizal (29 species) and nonmycorrhizal (5 species) fungi tested were no...

  7. Di/tri-peptide transporters as drug delivery targets

    DEFF Research Database (Denmark)

    Nielsen, C U; Brodin, Birger

    2003-01-01

    -dependent, and the transporters thus belong to the Proton-dependent Oligopeptide Transporter (POT)-family. The transporters are not drug targets per se, however due to their uniquely broad substrate specificity; they have proved to be relevant drug targets at the level of drug transport. Drug molecules such as oral active beta....../tri-peptide transporters from vesicular storages 3) changes in gene transcription/mRNA stability. The aim of the present review is to discuss physiological, patho-physiological and drug-induced regulation of di/tri-peptide transporter mediated transport....

  8. Acetylcholine content and viability of cholinergic neurons are influenced by the activity of protein histidine phosphatase

    Science.gov (United States)

    2012-01-01

    Background The first mammalian protein histidine phosphatase (PHP) was discovered in the late 90s of the last century. One of the known substrates of PHP is ATP-citrate lyase (ACL), which is responsible - amongst other functions - for providing acetyl-CoA for acetylcholine synthesis in neuronal tissues. It has been shown in previous studies that PHP downregulates the activity of ACL by dephosphorylation. According to this our present work focused on the influence of PHP activity on the acetylcholine level in cholinergic neurons. Results The amount of PHP in SN56 cholinergic neuroblastoma cells was increased after overexpression of PHP by using pIRES2-AcGFP1-PHP as a vector. We demonstrated that PHP overexpression reduced the acetylcholine level and induced cell death. The acetylcholine content of SN56 cells was measured by fast liquid chromatography-tandem mass spectrometry method. Overexpression of the inactive H53A-PHP mutant also induced cell damage, but in a significantly reduced manner. However, this overexpression of the inactive PHP mutant did not change the acetylcholine content of SN56 cells significantly. In contrast, PHP downregulation, performed by RNAi-technique, did not induce cell death, but significantly increased the acetylcholine content in SN56 cells. Conclusions We could show for the first time that PHP downregulation increased the acetylcholine level in SN56 cells. This might be a potential therapeutic strategy for diseases involving cholinergic deficits like Alzheimer's disease. PMID:22436051

  9. Dietary restriction of choline reduces hippocampal acetylcholine release in rats: in vivo microdialysis study.

    Science.gov (United States)

    Nakamura, A; Suzuki, Y; Umegaki, H; Ikari, H; Tajima, T; Endo, H; Iguchi, A

    2001-12-01

    We fed rats with a diet deficient in choline for 12 weeks and studied how dietary choline deficiency affected their behavior and their ability to release acetylcholine in discrete regions of rat brain using step-through passive avoidance task and in vivo microdialysis. In comparison with the control, rats fed the choline-deficient diet showed poorer retention of nociceptive memory in the passive avoidance task. Average choline level in cerebrospinal fluid in the choline-deficient group was significantly less (33.1%) than that of control rats. In vivo microdialysis showed no difference in the pattern of acetylcholine release enhanced by intraperitoneal administration of scopolamine hydrochloride (2 mg/kg) in the striatum between the two groups, whereas in the hippocampus, the maximum and subsequent increase of acetylcholine from the baseline by scopolamine injection was significantly lower in the choline-deficient group than in the control. From the results of our study, we speculate that long-term dietary restriction of choline can affect extra- and intracellular sources of substrates required for acetylcholine synthesis, and eventually limit the ability to release acetylcholine in the hippocampus. Reduced capacity to release acetylcholine in the hippocampus implies that the mechanism, maintaining acetylcholine synthesis on increased neuronal demand, may vary in discrete regions of the brain in response to dietary manipulation. The vulnerability of the mechanism in the hippocampus to dietary choline restriction is indicated by impaired mnemonic performance we observed.

  10. Modulation of the effect of acetylcholine on insulin release by the membrane potential of B cells

    International Nuclear Information System (INIS)

    Hermans, M.P.; Schmeer, W.; Henquin, J.C.

    1987-01-01

    Mouse islets were used to test the hypothesis that the B cell membrane must be depolarized for acetylcholine to increase insulin release. The resting membrane potential of B cells (at 3 mM glucose) was slightly decreased (5 mV) by acetylcholine, but no electrical activity appeared. This depolarization was accompanied by a Ca-independent acceleration of 86 Rb and 45 Ca efflux but no insulin release. When the B cell membrane was depolarized by a stimulatory concentration of glucose (10 mM), acetylcholine potentiated electrical activity, accelerated 86 Rb and 45 Ca efflux, and increased insulin release. This latter effect, but not the acceleration of 45 Ca efflux, was totally dependent on extracellular Ca. If glucose-induced depolarization of the B cell membrane was prevented by diazoxide, acetylcholine lost all effects but those produced at low glucose. In contrast, when the B cell membrane was depolarized by leucine or tolbutamide (at 3 mM glucose), acetylcholine triggered a further depolarization with appearance of electrical activity, accelerated 86 Rb and 45 Ca efflux, and stimulated insulin release. Acetylcholine produced similar effects (except for electrical activity) in the presence of high K or arginine which, unlike the above test agents, depolarize the B cell membrane by a mechanism other than a decrease in K+ permeability. Omission of extracellular Ca abolished the releasing effect of acetylcholine under all conditions but only partially decreased the stimulation of 45 Ca efflux. The results show thus that acetylcholine stimulation of insulin release does not result from mobilization of cellular Ca but requires that the B cell membrane be sufficiently depolarized to reach the threshold potential where Ca channels are activated. This may explain why acetylcholine alone does not initiate release but becomes active in the presence of a variety of agents

  11. Functional interaction between Lypd6 and nicotinic acetylcholine receptors

    DEFF Research Database (Denmark)

    Arvaniti, Maria; Jensen, Majbrit M; Soni, Neeraj

    2016-01-01

    Nicotinic acetylcholine receptors (nAChRs) affect multiple physiological functions in the brain and their functions are modulated by regulatory proteins of the Lynx family. Here, we report for the first time a direct interaction of the Lynx protein LY6/PLAUR domain-containing 6 (Lypd6) with n...... brain. Additionally, soluble recombinant Lypd6 protein attenuates nicotine-induced hippocampal inward currents in rat brain slices and decreases nicotine-induced extracellular signal-regulated kinase phosphorylation in PC12 cells, suggesting that binding of Lypd6 is sufficient to inhibit n......AChR-mediated intracellular signaling. We further show that perinatal nicotine exposure in rats (4 mg/kg/day through minipumps to dams from embryonic day 7 to post-natal day 21) significantly increases Lypd6 protein levels in the hippocampus in adulthood, which did not occur after exposure to nicotine in adulthood only. Our...

  12. Valence of acetylcholine-receptor-antibody-titers in myasthenia gravis

    International Nuclear Information System (INIS)

    Zeitlhofer, J.; Maida, E.M.; Mamoli, B.; Mayr, N.

    1986-01-01

    In a retrospective study in 47 patients with myasthenia gravis acetylcholine-receptor-antibody-titers (AChR-AB) were correlated with the severity of the disease. In 18 patients the course of titers was studied and two groups of patients could be differentiated: patients with relative constant and patients with fluctuating titers. Age, age of begin of myasthenia and sex did not influence the titers. Also the duration of the disease and the severity of symptoms did not influence the level of AChR-AB-titers. In this retrospective study the influence of immunsuppressive therapy on the intra-individual course of AB-titers and their correlation with the clinical symptoms could not be judged. Measurement of AChR-AB is of value for the diagnosis of myasthenia gravis and important for judging the clinical course and the effect of therapy. (Author)

  13. Eficacia del microsistema de oreja en el diagnóstico de la litiasis vesicular Effectiveness of the ear microsystem in the diagnosis of the vesicular lithiasis

    Directory of Open Access Journals (Sweden)

    Juan Luis Cobas Pérez

    Full Text Available Se llevó a cabo un ensayo clínico para el diagnóstico de la litiasis vesicular mediante el microsistema de oreja en 110 pacientes ingresados en los Servicios de Cirugía y Medicina Interna del Hospital General Docente "Dr. Juan Bruno Zayas Alfonso" de Santiago de Cuba, a fin de evaluar la eficacia del método tradicional. Los principales signos estuvieron dados por manchas blancas en el punto de vesícula biliar en ambas orejas y nódulos en la parte posterior de la concha de la oreja derecha. Se concluyó que este es un procedimiento eficaz, por lo cual es necesario generalizarlo, para de esta manera favorecer la calidad de atención a los pacientes y garantizar la base científica de su aplicación.A clinical trial for the diagnosis of the vesicular lithiasis was carried out by means of the ear microsystem in 110 patients admitted in the Surgery and Internal Medicine Services of "Dr. Juan Bruno Zayas Alfonso" Teaching General Hospital in Santiago de Cuba, in order to evaluate the effectiveness of the traditional method. The main signs were white stains in the billiary vesicle site in both ears and nodules, in the back part of the right ear shell. The conclusion was that this it is an effective procedure, thus it is necessary to generalize it, so as to favor the quality of care to the patients and to guarantee the scientific base of its application.

  14. Visualization of cholinoceptive neurons in the rat neocortex : colocalization of muscarinic and nicotinic acetylcholine receptors

    NARCIS (Netherlands)

    Zee, E.A. van der; Streefland, C.; Strosberg, A.D.; Schröder, H.; Luiten, P.G.M.

    The present investigation analyzes the cellular distribution of muscarinic and nicotinic acetylcholine receptors in rat neocortex, by use of monoclonal antibodies raised against purified receptor proteins. The degree of colocalization of both types of receptors was determined by way of

  15. Electrolyte and protein secretion by the perfused rabbit mandibular gland stimulated with acetylcholine or catecholamines

    DEFF Research Database (Denmark)

    Case, R M; Conigrave, A D; Novak, I

    1980-01-01

    stimulation, the rate of protein secretion fell off much faster than the rate of fluid secretion.7. The beta-adrenergic agonist isoproterenol evoked a fluid secretory response only equal to about 5% of that evoked by acetylcholine, but still the response declined during continued stimulation. The electrolyte...... composition of isoproterenol-evoked saliva was vastly different from that evoked by acetylcholine, being particularly rich in K and HCO(3). The isoproterenol-evoked saliva was also extremely rich in protein so that the total protein secretion evoked by isoproterenol was much greater than that evoked...... unstimulated or evoked by acetylcholine or eserine, could be blocked completely by atropine.4. During prolonged stimulation with acetylcholine, the fluid secretory response declined rapidly over a period of about 15 min from an initial high value to a much lower plateau value. After 3 or more hours...

  16. Enhanced Gene Transfer with Fusogenic Liposomes Containing Vesicular Stomatitis Virus G Glycoprotein

    Science.gov (United States)

    Abe, Akihiro; Miyanohara, Atsushi; Friedmann, Theodore

    1998-01-01

    Exposure of Lipofectin-DNA complexes to the partially purified G glycoprotein of the vesicular stomatitis virus envelope (VSV-G) results in loss of serum-mediated inhibition and in enhanced efficiency of gene transfer. Sucrose density gradient sedimentation analysis indicated that the VSV-G associates physically with the DNA-lipid complex to produce a VSV-G liposome. The ability to incorporate surrogate viral or cellular envelope components such as VSV-G into liposomes may allow more-efficient and possibly targeted gene delivery by lipofection, both in vitro and in vivo. PMID:9621082

  17. Analysis of the RNA species isolated from defective particles of vesicular stomatitis virus.

    Science.gov (United States)

    Adler, R; Banerjee, A K

    1976-10-01

    Serial high multiplicity passage of a cloned stock of vesicular stomatitis virus was found to generate defective interfering particles containing three size classes of RNA, with sedimentaiton coefficients of 31 S, 23 S and 19 S. The 31 S and 23 S RNA species were found to be complementary to both the 12 to 18 S and 31 S size classes of VSV mRNAs. The 19 S class of RNA was found to be partially base-paired. All three RNA species were found to contain ppAp at their 5' termini.

  18. Conditional expression of the vesicular stomatitis virus glycoprotein gene in Escherichia coli.

    OpenAIRE

    Rose, J K; Shafferman, A

    1981-01-01

    Bacterial plasmids that directed expression of the vesicular stomatitis virus glycoprotein (G-protein) gene under control of the tryptophan operon regulatory region were constructed. A plasmid directing the synthesis of a G-protein-like protein (containing the NH2-terminal segment of seven amino acids encoded by the trpE gene fused to the complete G-protein sequence lacking only its NH2-terminal methionine) could be transformed into trpR+ (repressed) but not into trpR- (derepressed) cells. Th...

  19. Natural re-establishment of vesicular-arbuscular mycorrhizae following stripmine reclamation in Wyoming

    Energy Technology Data Exchange (ETDEWEB)

    Allen, E.B.; Allen, M.F. (University of Wyoming, Laramie, WY (USA))

    1980-01-01

    The % root infection of {ital Agropyron smithii} and {ital A. intermedium} by vesicular-arbuscular mycorrhizae was measured and spoil spores were counted in six reclaimed stripmine sites in Wyoming. On 2- and 3-yr old sites % infection and spore counts were c. 50% or less than native prairie levels. Spore counts of a 3-yr old disked prairie site were not different from the undisturbed prairie level, but infection was significantly lower. Spore counts of the reclimed sites were not highly correlated with % root infection. Five of seven annuals which colonized the reclaimed and disked sites were non-mycorrhizal. 43 refs., 3 tabs.

  20. Role of dopamine receptor and muscarinic acetylcholine receptor blockade in the antiapomorphine action of neuroleptics

    Energy Technology Data Exchange (ETDEWEB)

    Zharkovskii, A.M.; Langel, Yu.L.; Chereshka, K.S.; Zharkovskaya, T.A.

    1987-08-01

    The authors analyze the role of dopamine and muscarinic acetylcholine receptor blocking components in the antistereotypic action of neuroleptics with different chemical structure. To determine dopamine-blocking activity in vitro, binding of /sup 3/H-spiperone with membranes of the rat striatum was measured. To study the blocking action of the substances on muscarinic acetylcholine receptors, binding of /sup 3/H-quinuclidinyl benzylate with brain membranes was chosen.

  1. Activation of muscarinic acetylcholine receptors elicits pigment granule dispersion in retinal pigment epithelium isolated from bluegill

    OpenAIRE

    González, Alfredo; Crittenden, Elizabeth L; García, Dana M

    2004-01-01

    Abstract Background In fish, melanin pigment granules in the retinal pigment epithelium disperse into apical projections as part of the suite of responses the eye makes to bright light conditions. This pigment granule dispersion serves to reduce photobleaching and occurs in response to neurochemicals secreted by the retina. Previous work has shown that acetylcholine may be involved in inducing light-adaptive pigment dispersion. Acetylcholine receptors are of two main types, nicotinic and musc...

  2. [Regulation of acetylcholine synthesis in presynaptic endings of cholinergic neurons of the central nervous system].

    Science.gov (United States)

    Tuchek, S; Dolezhal, V; Richny, Ia

    1984-01-01

    Data on the acetylcholine (ACh) synthesis in nerve cells with special attention to its control are summarized in the paper. At rest or during moderate synaptic activity, the concentration of ACh in the compartment of its synthesis probably corresponds to the equilibrium between the substrates and products in the reaction catalysed by choline acetyltransferase. The release of ACh is followed by a transfer of ACh from the compartment of its synthesis to the compartment of release, and, automatically, by the synthesis of new ACh until a new equilibrium is reached in the compartment of synthesis. In addition, synaptic activity and the release of ACh support the synthesis of new ACh in the following ways: choline carriers are disinhibited by lowering the concentration of ACh in the nerve endings, and the transport of choline from the extracellular fluid to the cell interior according to its electro-chemical gradient is thus facilitated; the concentration of choline in the extracellular fluid is increased in the vicinity of the nerve endings as a consequence of the hydrolysis of the released ACh; postactivation hyperpolarization of the nerve endings brings about an increase of the choline transport and concentration in the nerve endings; presumably, the stimulation of muscarinic receptors brings about a further increase in the choline concentration in the vicinity of the nerve endings by the phosphatidylcholine hydrolysis intensification in postsynaptic cells; the decrease in the concentration of acetyl-CoA (as a consequence of the resynthesis of ACh) increases the activity of pyruvate dehydrogenase and the production of acetyl-CoA; conceivably, the increase in the concentration of Ca2+ ions in the nerve endings assists direct passage of acetyl-CoA from the mitochondria to the cytosol of the nerve endings, where the synthesis of ACh occurs.(ABSTRACT TRUNCATED AT 250 WORDS)

  3. Acetylcholine release by human colon cancer cells mediates autocrine stimulation of cell proliferation.

    Science.gov (United States)

    Cheng, Kunrong; Samimi, Roxana; Xie, Guofeng; Shant, Jasleen; Drachenberg, Cinthia; Wade, Mark; Davis, Richard J; Nomikos, George; Raufman, Jean-Pierre

    2008-09-01

    Most colon cancers overexpress M3 muscarinic receptors (M3R), and post-M3R signaling stimulates human colon cancer cell proliferation. Acetylcholine (ACh), a muscarinic receptor ligand traditionally regarded as a neurotransmitter, may be produced by nonneuronal cells. We hypothesized that ACh release by human colon cancer cells results in autocrine stimulation of proliferation. H508 human colon cancer cells, which have robust M3R expression, were used to examine effects of muscarinic receptor antagonists, acetylcholinesterase inhibitors, and choline transport inhibitors on cell proliferation. A nonselective muscarinic receptor antagonist (atropine), a selective M3R antagonist (p-fluorohexahydro-sila-difenidol hydrochloride), and a choline transport inhibitor (hemicholinum-3) all inhibited unstimulated H508 colon cancer cell proliferation by approximately 40% (P<0.005). In contrast, two acetylcholinesterase inhibitors (eserine-hemisulfate and bis-9-amino-1,2,3,4-tetrahydroacridine) increased proliferation by 2.5- and 2-fold, respectively (P<0.005). By using quantitative real-time PCR, expression of choline acetyltransferase (ChAT), a critical enzyme for ACh synthesis, was identified in H508, WiDr, and Caco-2 colon cancer cells. By using high-performance liquid chromatography-electrochemical detection, released ACh was detected in H508 and Caco-2 cell culture media. Immunohistochemistry in surgical specimens revealed weak or no cytoplasmic staining for ChAT in normal colon enterocytes (n=25) whereas half of colon cancer specimens (n=24) exhibited moderate to strong staining (P<0.005). We conclude that ACh is an autocrine growth factor in colon cancer. Mechanisms that regulate colon epithelial cell production and release of ACh warrant further investigation.

  4. Mechanics of channel gating of the nicotinic acetylcholine receptor.

    Directory of Open Access Journals (Sweden)

    Xinli Liu

    2008-01-01

    Full Text Available The nicotinic acetylcholine receptor (nAChR is a key molecule involved in the propagation of signals in the central nervous system and peripheral synapses. Although numerous computational and experimental studies have been performed on this receptor, the structural dynamics of the receptor underlying the gating mechanism is still unclear. To address the mechanical fundamentals of nAChR gating, both conventional molecular dynamics (CMD and steered rotation molecular dynamics (SRMD simulations have been conducted on the cryo-electron microscopy (cryo-EM structure of nAChR embedded in a dipalmitoylphosphatidylcholine (DPPC bilayer and water molecules. A 30-ns CMD simulation revealed a collective motion amongst C-loops, M1, and M2 helices. The inward movement of C-loops accompanying the shrinking of acetylcholine (ACh binding pockets induced an inward and upward motion of the outer beta-sheet composed of beta9 and beta10 strands, which in turn causes M1 and M2 to undergo anticlockwise motions around the pore axis. Rotational motion of the entire receptor around the pore axis and twisting motions among extracellular (EC, transmembrane (TM, and intracellular MA domains were also detected by the CMD simulation. Moreover, M2 helices undergo a local twisting motion synthesized by their bending vibration and rotation. The hinge of either twisting motion or bending vibration is located at the middle of M2, possibly the gate of the receptor. A complementary twisting-to-open motion throughout the receptor was detected by a normal mode analysis (NMA. To mimic the pulsive action of ACh binding, nonequilibrium MD simulations were performed by using the SRMD method developed in one of our laboratories. The result confirmed all the motions derived from the CMD simulation and NMA. In addition, the SRMD simulation indicated that the channel may undergo an open-close (O C motion. The present MD simulations explore the structural dynamics of the receptor under its

  5. Identification of a vesicular-arbuscular mycorrhizal fungus by using monoclonal antibodies in an enzyme-linked immunosorbent assay.

    Science.gov (United States)

    Wright, S F; Morton, J B; Sworobuk, J E

    1987-09-01

    Spore morphology is currently used to identify species of vesicular-arbuscular mycorrhizal fungi. We report the first use of a highly specific immunological method for identification of a vesicular-arbuscular mycorrhizal fungus. Two monoclonal antibodies were produced against Glomus occultum. Monoclonal antibodies reacted strongly with both spores and hyphae in an indirect enzyme-linked immunosorbent assay. All other mycorrhizal (29 species) and nonmycorrhizal (5 species) fungi tested were nonreactive with the monoclonal antibodies. A single spore of G. occultum was detectable in the presence of high numbers of spores of other vesicular-arbuscular mycorrhizal fungi. Variation in the reaction of G. occultum isolates from West Virginia, Florida, and Colombia suggests that monoclonal antibodies may differentiate strains.

  6. Membrane permeable C-terminal dopamine transporter peptides attenuate amphetamine-evoked dopamine release

    DEFF Research Database (Denmark)

    Rickhag, Karl Mattias; Owens, WA; Winkler, Marie-Therese

    2013-01-01

    The dopamine transporter (DAT) is responsible for sequestration of extracellular dopamine (DA). The psychostimulant amphetamine (AMPH) is a DAT substrate, which is actively transported into the nerve terminal, eliciting vesicular depletion and reversal of DA transport via DAT. Here, we investigate......-terminal protein-protein interactions are critical for AMPH-evoked DA efflux and suggest that it may be possible to target protein-protein interactions to modulate transporter function and interfere with psychostimulant effects....

  7. The H2O/D2O exchange across vesicular lipid bilayers

    International Nuclear Information System (INIS)

    Engelbert, H.P.; Lawaczek, R.

    1985-01-01

    A new method to measure the water (D 2 O/H 2 O) permeation across vesicular lipid bilayers is described. The method is based on the solvent isotope effect of the light scattering which is a consequence of the different indices of refraction of D 2 O and H 2 O. Unilamellar lipid vesicles in excess of H 2 O are rapidly mixed with D 2 O or vice versa. As result of the H 2 O/D 2 O exchange across the vesicular bilayer the light scattering signal has a time dependent, almost single exponential component allowing the deduction of the exchange relaxation rate and, at known size, of the permeability coefficient. The experimental results are in accord with calculations from the Mie theory of light scattering for coated spheres. The method is applicable for large vesicles where the permeation is the rate-limiting step. Size separations are performed by a flow dialysis through a sequence of pore-membrane-filters. For dimyristoyl-lecithin bilayers the water permeability-coefficient is 1.9 . 10 -5 cm/s in the crystalline phase and increases by a factor of 10-100 in the liquid-crystalline state. The temperature dependence of the permeation exhibits a sharp change at the phase transition. For binary mixtures of lecithins this sharp change follows the solidus curve of the non-ideal phase diagram determined by spectroscopic techniques. (orig.)

  8. Two transcription products of the vesicular stomatitis virus genome may control L-cell protein synthesis

    International Nuclear Information System (INIS)

    Dunigan, D.D.; Lucas-Lenard, J.M.

    1983-01-01

    When mouse L-cells are infected with vesicular stomatitis virus, there is a decrease in the rate of protein synthesis ranging from 20 to 85% of that in mock-infected cells. Vesicular stomatitis virus, irradiated with increasing doses of UV light, eventually loses this capacity to inhibit protein synthesis. The UV inactivation curve was biphasic, suggesting that transcription of two regions of the viral genome is necessary for the virus to become inactivated in this capacity. The first transcription produced corresponded to about 373 nucleotides, and the second corresponded to about 42 nucleotides. Inhibition of transcription of the larger product by irradiating the virus with low doses of UV light left a residual inhibition of protein synthesis consisting of approximately 60 to 65% of the total inhibition. This residual inhibition could be obviated by irradiating the virus with a UV dose of greater than 20,000 ergs/mm 2 and was thus considered to represent the effect of the smaller transcription product. In the R1 mutant of another author, the inhibition of transcription of the larger product sufficed to restore protein synthesis to the mock-infected level, suggesting that the smaller transcription product is nonfunctional with respect to protein synthesis inhibition. Extracts from cells infected with virus irradiated with low doses of UV light showed a protein synthesis capacity quite similar to that of their in vivo counterparts, indicating that these extracts closely reflect the in vivo effects of virus infection

  9. Black fly involvement in the epidemic transmission of vesicular stomatitis New Jersey virus (Rhabdoviridae: Vesiculovirus).

    Science.gov (United States)

    Mead, Daniel G; Howerth, Elizabeth W; Murphy, Molly D; Gray, Elmer W; Noblet, Raymond; Stallknecht, David E

    2004-01-01

    The transmission routes of Vesicular stomatitis New Jersey virus (VSNJV), a causative agent of vesicular stomatitis, an Office International des Epizooties List-A disease, are not completely understood. Epidemiological and entomological studies conducted during the sporadic epidemics in the western United States have identified potential virus transmission routes involving insect vectors and animal-to-animal contact. In the present study we experimentally tested the previously proposed transmission routes which were primarily based on field observations. Results obtained provide strong evidence for the following: (1) hematophagous insects acquire VSNJV by unconventional routes while blood feeding on livestock, (2) clinical course of VSNJV infection in livestock following transmission by an infected insect is related to insect bite site, (3) infection of livestock via insect bite can result in multiple transmission possibilities, including animal-to-animal contact. Taken together, these data significantly add to our understanding of the transmission routes of a causative agent of one of the oldest known infectious diseases of livestock, for which the details have remained largely unknown despite decades of research.

  10. Vesicular GABA Uptake Can Be Rate Limiting for Recovery of IPSCs from Synaptic Depression

    Directory of Open Access Journals (Sweden)

    Manami Yamashita

    2018-03-01

    Full Text Available Summary: Synaptic efficacy plays crucial roles in neuronal circuit operation and synaptic plasticity. Presynaptic determinants of synaptic efficacy are neurotransmitter content in synaptic vesicles and the number of vesicles undergoing exocytosis at a time. Bursts of presynaptic firings depress synaptic efficacy, mainly due to depletion of releasable vesicles, whereas recovery from strong depression is initiated by endocytic vesicle retrieval followed by refilling of vesicles with neurotransmitter. We washed out presynaptic cytosolic GABA to induce a rundown of IPSCs at cerebellar inhibitory cell pairs in slices from rats and then allowed fast recovery by elevating GABA concentration using photo-uncaging. The time course of this recovery coincided with that of IPSCs from activity-dependent depression induced by a train of high-frequency stimulation. We conclude that vesicular GABA uptake can be a limiting step for the recovery of inhibitory neurotransmission from synaptic depression. : Recovery of inhibitory synaptic transmission from activity-dependent depression requires refilling of vesicles with GABA. Yamashita et al. find that vesicular uptake rate of GABA is a slow process, limiting the recovery rate of IPSCs from depression.

  11. Presynaptic mechanisms of lead neurotoxicity: effects on vesicular release, vesicle clustering and mitochondria number.

    Science.gov (United States)

    Zhang, Xiao-Lei; Guariglia, Sara R; McGlothan, Jennifer L; Stansfield, Kirstie H; Stanton, Patric K; Guilarte, Tomás R

    2015-01-01

    Childhood lead (Pb2+) intoxication is a global public health problem and accounts for 0.6% of the global burden of disease associated with intellectual disabilities. Despite the recognition that childhood Pb2+ intoxication contributes significantly to intellectual disabilities, there is a fundamental lack of knowledge on presynaptic mechanisms by which Pb2+ disrupts synaptic function. In this study, using a well-characterized rodent model of developmental Pb2+ neurotoxicity, we show that Pb2+ exposure markedly inhibits presynaptic vesicular release in hippocampal Schaffer collateral-CA1 synapses in young adult rats. This effect was associated with ultrastructural changes which revealed a reduction in vesicle number in the readily releasable/docked vesicle pool, disperse vesicle clusters in the resting pool, and a reduced number of presynaptic terminals with multiple mitochondria with no change in presynaptic calcium influx. These studies provide fundamental knowledge on mechanisms by which Pb2+ produces profound inhibition of presynaptic vesicular release that contribute to deficits in synaptic plasticity and intellectual development.

  12. Partitioning of Intermediary Carbon Metabolism in Vesicular-Arbuscular Mycorrhizal Leek.

    Science.gov (United States)

    Shachar-Hill, Y.; Pfeffer, P. E.; Douds, D.; Osman, S. F.; Doner, L. W.; Ratcliffe, R. G.

    1995-05-01

    Vesicular-arbuscular mycorrhizal fungi are symbionts for a large variety of crop plants; however, the form in which they take up carbon from the host is not established. To trace the course of carbon metabolism, we have used nuclear magnetic resonance spectroscopy with [13C]glucose labeling in vivo and in extracts to examine leek (Allium porrum) roots colonized by Glomus etunicatum (and uncolonized controls) as well as germinating spores. These studies implicate glucose as a likely substrate for vesicular-arbuscular mycorrhizal fungi in the symbiotic state. Root feeding of 0.6 mM 1-[13C]glucose labeled only the fungal metabolites trehalose and glycogen. The time course of this labeling was dependent on the status of the host. Incubation with 50 mM 1-[13C]glucose caused labeling of sucrose (in addition to fungal metabolites) with twice as much labeling in uncolonized plants. There was no detectable scrambling of the label from C1 glucose to the C6 position of glucose moieties in trehalose or glycogen. Labeling of mannitol C1,6 in the colonized root tissue was much less than in axenically germinating spores. Thus, carbohydrate metabolism of host and fungus are significantly altered in the symbiotic state.

  13. Interaction of rhizosphere bacteria, fertilizer, and vesicular-arbuscular mycorrhizal fungi with sea oats.

    Science.gov (United States)

    Will, M E; Sylvia, D M

    1990-07-01

    Plants must be established quickly on replenished beaches in order to stabilize the sand and begin the dune-building process. The objective of this research was to determine whether inoculation of sea oats (Uniola paniculata L.) with bacteria (indigenous rhizosphere bacteria and N(2) fixers) alone or in combination with vesicular-arbuscular mycorrhizal fungi would enhance plant growth in beach sand. At two fertilizer-N levels, Klebsiella pneumoniae and two Azospirillum spp. did not provide the plants with fixed atmospheric N; however, K. pneumoniae increased root and shoot growth. When a sparingly soluble P source (CaHPO(4)) was added to two sands, K. pneumoniae increased plant growth in sand with a high P content. The phosphorus content of shoots was not affected by bacterial inoculation, indicating that a mechanism other than bacterially enhanced P availability to plants was responsible for the growth increases. When sea oats were inoculated with either K. pneumoniae or Acaligenes denitrificans and a mixed Glomus inoculum, there was no consistent evidence of a synergistic effect on plant growth. Nonetheless, bacterial inoculation increased root colonization by vesicular-arbuscular mycorrhizal fungi when the fungal inoculum consisted of colonized roots but had no effect on colonization when the inoculum consisted of spores alone. K. pneumoniae was found to increase spore germination and hyphal growth of Glomus deserticola compared with the control. The use of bacterial inoculants to enhance establishment of pioneer dune plants warrants further study.

  14. Partial neuromuscular blockade in humans enhances muscle blood flow during exercise independently of muscle oxygen uptake and acetylcholine receptor blockade

    DEFF Research Database (Denmark)

    Hellsten, Ylva; Krustrup, Peter; Iaia, F Marcello

    2009-01-01

    This study examined the role of acetylcholine for skeletal muscle blood flow during exercise by use of the competitive neuromuscular blocking agent cisatracurium in combination with the acetylcholine receptor blocker glycopyrrone. Nine healthy male subjects performed a 10-min bout of one-legged k......This study examined the role of acetylcholine for skeletal muscle blood flow during exercise by use of the competitive neuromuscular blocking agent cisatracurium in combination with the acetylcholine receptor blocker glycopyrrone. Nine healthy male subjects performed a 10-min bout of one...... conductance during exercise, events that are not associated with either acetylcholine or an increased oxygen demand. The results do not support an essential role for acetylcholine, released form the neuromuscular junction, in exercise hyperaemia or for the enhanced blood flow during neuromuscular blockade....... The enhanced exercise hyperemia during partial neuromuscular blockade may be related to a greater recruitment of fast-twitch muscle fibres. Key words: blood flow, neuromuscular blockade, exercise, skeletal muscle....

  15. Near-complete genome sequencing of swine vesicular disease virus using the Roche GS FLX sequencing platform

    DEFF Research Database (Denmark)

    Nielsen, Sandra Cathrine Abel; Bruhn, Christian Anders Wathne; Samaniego Castruita, Jose Alfredo

    2014-01-01

    Swine vesicular disease virus (SVDV) is an enterovirus that is both genetically and antigenically closely related to human coxsackievirus B5 within the Picornaviridae family. SVDV is the causative agent of a highly contagious (though rarely fatal) vesicular disease in pigs. We report a rapid method...... with significant genetic distances within the same species of viruses. All reference mappings used an iterative method to avoid bias. Further verification was achieved through phylogenetic analysis against published SVDV genomes and additional Enterovirus B sequences. This approach allows high confidence...

  16. Dopamine D(1) receptor-mediated control of striatal acetylcholine release by endogenous dopamine.

    Science.gov (United States)

    Acquas, E; Di Chiara, G

    1999-10-27

    The role of dopamine D(1) and D(2) receptors in the control of acetylcholine release in the dorsal striatum by endogenous dopamine was investigated by monitoring with microdialysis the effect of the separate or combined administration of the dopamine D(1) receptor antagonist, SCH 39166 ¿(-)-trans-6,7,7a,8,9, 13b-exahydro-3-chloro-2-hydroxy-N-methyl-5H-benzo-[d]-nap hto-[2, 1b]-azepine hydrochloride¿ (50 microg/kg subcutaneous (s.c.)), of the dopamine D(2)/D(3) receptor agonist, quinpirole (trans-(-)-4aR, 4a,5,6,7,8,8a,9-octahydro-5-propyl-1H-pyrazolo-(3,4-g)-quinoline hydrochloride) (5 and 10 microg/kg s.c.), and of the D(3) receptor selective agonist, PD 128,907 [S(+)-(4aR,10bR)-3,4,4a, 10b-tetrahydro-4-propyl-2H,5H-[1]benzopyrano-[4,3-b]-1,4-oxazin -9-ol hydrochloride] (50 microg/kg s.c.), on in vivo dopamine and acetylcholine release. Microdialysis was performed with a Ringer containing low concentrations (0.01 microM) of the acetylcholinesterase inhibitor, neostigmine. Quinpirole (10 microg/kg s.c.) decreased striatal dopamine and acetylcholine release. Administration of PD 128,907 (50 microg/kg) decreased dopamine but failed to affect acetylcholine release. SCH 39166 (50 microg/kg s.c.) stimulated dopamine release and reduced acetylcholine release. Pretreatment with quinpirole reduced (5 microg/kg s.c.) or completely prevented (10 microg/kg s.c.) the stimulation of dopamine release elicited by SCH 39166 (50 microg/kg s.c.); on the other hand, pretreatment with quinpirole (5 and 10 microg/kg) potentiated the reduction of striatal acetylcholine release induced by SCH 39166 (50 microg/kg s.c.). Similarly, pretreatment with PD 128,907 (50 microg/kg) which prevented the increase of dopamine release induced by SCH 39166 (50 microg/kg), potentiated the reduction of striatal acetylcholine transmission elicited by SCH 39166. Thus, pretreatment with low doses of quinpirole or PD 128,907 influences in opposite manner the effect of SCH 39166 on striatal dopamine and

  17. Menthol binding and inhibition of α7-nicotinic acetylcholine receptors.

    Directory of Open Access Journals (Sweden)

    Abrar Ashoor

    Full Text Available Menthol is a common compound in pharmaceutical and commercial products and a popular additive to cigarettes. The molecular targets of menthol remain poorly defined. In this study we show an effect of menthol on the α7 subunit of the nicotinic acetylcholine (nACh receptor function. Using a two-electrode voltage-clamp technique, menthol was found to reversibly inhibit α7-nACh receptors heterologously expressed in Xenopus oocytes. Inhibition by menthol was not dependent on the membrane potential and did not involve endogenous Ca(2+-dependent Cl(- channels, since menthol inhibition remained unchanged by intracellular injection of the Ca(2+ chelator BAPTA and perfusion with Ca(2+-free bathing solution containing Ba(2+. Furthermore, increasing ACh concentrations did not reverse menthol inhibition and the specific binding of [(125I] α-bungarotoxin was not attenuated by menthol. Studies of α7- nACh receptors endogenously expressed in neural cells demonstrate that menthol attenuates α7 mediated Ca(2+ transients in the cell body and neurite. In conclusion, our results suggest that menthol inhibits α7-nACh receptors in a noncompetitive manner.

  18. Electrically induced release of acetylcholine from denervated Schwann cells.

    Science.gov (United States)

    Dennis, M J; Miledi, R

    1974-03-01

    1. Focal electrical stimulation of Schwann cells at the end-plates of denervated frog muscles elicited slow depolarizations of up to 30 mV in the muscle fibres. This response is referred to as a Schwann-cell end-plate potential (Schwann-e.p.p.).2. Repeated stimulation sometimes evoked further Schwann-e.p.p.s, but they were never sustained for more than 30 pulses. Successive e.p.p.s varied in amplitude and time course independently of the stimulus.3. The Schwann-e.p.p.s were reversibly blocked by curare, suggesting that they result from a release of acetylcholine (ACh) by the Schwann cells.4. ACh release by electrical stimulation did not seem to occur in quantal form and was not dependent on the presence of calcium ions in the external medium; nor was it blocked by tetrodotoxin.5. Stimulation which caused release of ACh also resulted in extensive morphological disruption of the Schwann cells, as seen with both light and electron microscopy.6. It is concluded that electrical stimulation of denervated Schwann cells causes break-down of the cell membrane and releases ACh, presumably in molecular form.

  19. Cost-benefit decision circuitry: proposed modulatory role for acetylcholine.

    Science.gov (United States)

    Fobbs, Wambura C; Mizumori, Sheri J Y

    2014-01-01

    In order to select which action should be taken, an animal must weigh the costs and benefits of possible outcomes associate with each action. Such decisions, called cost-benefit decisions, likely involve several cognitive processes (including memory) and a vast neural circuitry. Rodent models have allowed research to begin to probe the neural basis of three forms of cost-benefit decision making: effort-, delay-, and risk-based decision making. In this review, we detail the current understanding of the functional circuits that subserve each form of decision making. We highlight the extensive literature by detailing the ability of dopamine to influence decisions by modulating structures within these circuits. Since acetylcholine projects to all of the same important structures, we propose several ways in which the cholinergic system may play a local modulatory role that will allow it to shape these behaviors. A greater understanding of the contribution of the cholinergic system to cost-benefit decisions will permit us to better link the decision and memory processes, and this will help us to better understand and/or treat individuals with deficits in a number of higher cognitive functions including decision making, learning, memory, and language. © 2014 Elsevier Inc. All rights reserved.

  20. Molecular alteration of a muscarinic acetylcholine receptor system during synaptogenesis

    International Nuclear Information System (INIS)

    Large, T.H.; Cho, N.J.; De Mello, F.G.; Klein, W.L.

    1985-01-01

    Biochemical properties of the muscarinic acetylcholine receptor system of the avian retina were found to change during the period when synapses form in ovo. Comparison of ligand binding to membranes obtained before and after synaptogenesis showed a significant increase in the affinity, but not proportion, of the high affinity agonist-binding state. There was no change in receptor sensitivity to antagonists during this period. Pirenzepine binding, which can discriminate muscarinic receptor subtypes, showed the presence of a single population of low affinity sites (M2) before and after synaptogenesis. The change in agonist binding was not due to the late development of receptor function. However, detergent-solubilization of membranes eliminated differences in agonist binding between receptors from embryos and hatched chicks, suggesting a developmental change in interactions of the receptor with functionally related membrane components. A possible basis for altered interactions was obtained from isoelectric point data showing that the muscarinic receptor population underwent a transition from a predominantly low pI form (4.25) in 13 day embryos to a predominantly high pI form (4.50) in newly hatched chicks. The possibility that biochemical changes in the muscarinic receptor play a role in differentiation of the system by controlling receptor position on the surface of nerve cells is discussed

  1. Actions of piperidine alkaloid teratogens at fetal nicotinic acetylcholine receptors.

    Science.gov (United States)

    Green, Benedict T; Lee, Stephen T; Panter, Kip E; Welch, Kevin D; Cook, Daniel; Pfister, James A; Kem, William R

    2010-01-01

    Teratogenic alkaloids are found in many species of plants including Conium maculatum L., Nicotiana glauca, Nicotiana tabaccum, and multiple Lupinus spp. Fetal musculoskeletal defects produced by alkaloids from these plants include arthrogyropisis, scoliosis, torticollis, kyposis, lordosis, and cleft palate. A pharmacodynamic comparison of the alkaloids ammodendrine, anabasine, anabaseine, anagyrine, and coniine in SH-SY5Y cells and TE-671 cells was made. These alkaloids and their enantiomers were more effective in depolarizing TE-671 cells which express the human fetal-muscle type nicotinic acetylcholine receptor (nAChR) relative to SH-SY5Y cells which predominately express autonomic nAChRs. The rank order of potency in TE-671 cells was: anabaseine>(+)-anabasine>(-)-anabasine > (+/-)-anabasine>anagyrine>(-)-coniine > (+/-)-coniine>(+)-coniine>(+/-)-ammodendrine>(+)-ammodendrine. The rank order potency in SH-SY5Y cells was: anabaseine>(+)-anabasine>(-)-coniine>(+)-coniine>(+)-ammodendrine>anagyrine>(-)-anabasine>(+/-)-coniine>(+/-)-anabasine>(-)-ammodendrine. The actions of these alkaloids at nAChRs in both cell lines could be distinguished by their maximum effects in depolarizing cell membrane potential. The teratogenic action of these compounds may be related to their ability to activate and subsequently desensitize nAChRs.

  2. Effects of two oxadiazolidinones on cholinesterases and acetylcholine receptors

    International Nuclear Information System (INIS)

    Bakry, N.; Lockyer, S.; Sherby, S.; Eldefrawi, A.; Eldefrawi, M.

    1986-01-01

    Inhibition of acetylcholinesterase (AChE) and butyryl cholinesterase (BuChE) by 3-(2,3-dihydro-2,2-dimethyl-benzofuran-'7-yl)-5-methoxy-1,3,4-oxadiazol-2( 3 H)-one (DBOX) and 3-(2-methoxyphenyl)-5-methoxy-1,3,4-oxadiazol-2( 3 H)-one (MPOX) was measured by the Ellmann spectrophotometric method. Inhibition was quasi first order and irreversible. DBOX was 2-3 orders of magnitude more potent than MPOX. Housefly brain AChE and horse serum BuChE were more sensitive than AChEs of red blood cells or eel and Torpedo electric organs. It is suggested that the nonesteratic oxadiazolidinones are activated to carbanillates on the surface of the enzyme and produce a carbanillated enzyme which ages rapidly. Carbamate anticholinesterases protected AChE against carbanillation as they did against phosphorylation. At higher concentrations, the two oxadiazolidinones also affected binding of [ 125 I] α bungarotoxin and [ 3 H]perhydrohistrionicotoxin to Torpedo nicotinic acetylcholine receptors, but did not affect binding of [ 3 H]quinuclidinyl benzilate to rat brain muscarinic receptors

  3. Acetylcholine esterase activity in mild cognitive impairment and Alzheimer's disease

    International Nuclear Information System (INIS)

    Herholz, Karl

    2008-01-01

    Impairment of cholinergic neurotransmission is a well-established fact in Alzheimer's disease (AD), but there is controversy about its relevance at the early stages of the disease and in mild cognitive impairment (MCI). In vivo positron emission tomography imaging of cortical acetylcholine esterase (AChE) activity as a marker of cholinergic innervation that is expressed by cholinergic axons and cholinoceptive neurons has demonstrated a reduction of this enzyme activity in manifest AD. The technique is also useful to measure the inhibition of cerebral AChE induced by cholinesterase inhibitors for treatment of dementia symptoms. A reduction of cortical AchE activity was found consistently in all studies of AD and in few cases of MCI who later concerted to AD. The in vivo findings in MCI and very mild AD are still preliminary, and studies seem to suggest that cholinergic innervation and AChE as the main degrading enzyme are both reduced, which might result in partial compensation of their effect. (orig.)

  4. Muscarinic acetylcholine receptors: location of the ligand binding site

    International Nuclear Information System (INIS)

    Hulme, E.; Wheatley, M.; Curtis, C.; Birdsall, N.

    1987-01-01

    The key to understanding the pharmacological specificity of muscarinic acetylcholine receptors (mAChR's) is the location within the receptor sequence of the amino acid residues responsible for ligand binding. To approach this problem, they have purified mAChR's from rat brain to homogeneity by sequential ion-exchange chromatography, affinity chromatography and molecular weight fractionation. Following labelling of the binding site with an alkylating affinity label, 3 H-propylbenzilycholine mustard aziridinium ion ( 3 H-PrBCM), the mAChR was digested with a lysine-specific endoproteinase, and a ladder of peptides of increasing molecular weight, each containing the glycosylated N-terminus, isolated by chromatography on wheat-germ agglutinin sepharose. The pattern of labelling showed that a residue in the peptides containing transmembrane helices 2 and/or 3 of the mAChR was alkylated. The linkage was cleaved by 1 M hydroxylamine, showing that 3 H-PrBCM was attached to an acidic residue, whose properties strongly suggested it to be embedded in a hydrophobic intramembrane region of the mAChR. Examination of the cloned sequence of the mAChR reveals several candidate residues, the most likely of which is homologous to an aspartic acid residue thought to protonate the retinal Schiff's base in the congeneric protein rhodopsin

  5. Transmembrane topology of the acetylcholine receptor examined in reconstituted vesicles

    International Nuclear Information System (INIS)

    McCrea, P.D.

    1987-01-01

    Each of the five acetylcholine receptor (AChR) subunits, α 2 β-γδ, is believed to have the same number of transmembrane crossing and to share the same general folding pattern. AChR isolated from the electric organ of electric fish is predominantly dimeric. We have used this bridge as a marker for the C-terminus of the δ subunit, and presumably that of the other subunits in addition. The disulfide's accessibility to hydrophilic reductants, principally glutathione (GSH), was tested in a reconstituted vesicle system. The reduction of the δ-δ desulfide, as evidenced by the transition of AChrR dimers to monomers, was quantitatively monitored on velocity sedimentation sucrose gradients. Alternatively, the reduction of δ 2 to δ was followed by employing non-reducing SDS-PAGE. Reductants such as GSH were able to access the bridge in intact right-side-out vesicles. No acceleration of this process was evident when the vesicles were disrupted by freeze-thaw or by detergents. Control experiments which determined the rate of reduction of entrapped diphtheria toxin, or that of 3 H-GSH efflux, demonstrated that intact reconstituted vesicles provide an adequate permeability barrier to GSH access of their intravesicular space

  6. Biochemical and immunological studies of the Muscarinic acetylcholine receptor

    International Nuclear Information System (INIS)

    Gainer, M.W.

    1985-01-01

    Muscarinic acetylcholine receptors were solubilized from bovine brain membranes with 3[3-cholamidopropyl)dimethylammonio]propanesulfonate (CHAPS). A combination of 10 mM CHAPS and 1 M NaCl solubilized 15-40% of the specific receptor binding sites from these membranes. The solubilized receptors displayed high affinity binding of the muscarinic antagonist, [ 3 H]quinuclidinyl benzilate with a K/sub D/ = 300 pM. In addition, the solubilized and retained guanyl nucleotide regulation of agonist binding characteristic of membrane bound receptors. Gel filtration experiments showed that solubilized receptors from cortex and cerebellum had different elution profiles. Analysis by sucrose density gradient centrifugation showed that receptors in the lower molecular weight peak sedimented with a coefficient of 5S. Receptors in the larger molecular weight peak sedimented to the bottom of the gradient. Attempts to purify receptors by chromatography on propylbenzilycholine Sepharose were unsuccessful. The technique used to attach the ligand to the solid support, however, was used to synthesize a PrBCM-BSA conjugate and the conjugate used as an antigen in the production of anti-ligand antibodies. Two anti-PrBCM monoclonal antibodies were isolated that recognize muscarinic but not nicotinic cholinergic ligands. The abilities of the antibodies to recognize other muscarinic ligands indicated the antibodies recognized a portion of PrBCM involved in binding to the receptor. Construction of an antibody affinity resin resulted in the purification of this fragment a minimum of 170 fold

  7. Early Life Stress, Nicotinic Acetylcholine Receptors and Alcohol Use Disorders

    Directory of Open Access Journals (Sweden)

    Joan Y. Holgate

    2015-06-01

    Full Text Available Stress is a major driving force in alcohol use disorders (AUDs. It influences how much one consumes, craving intensity and whether an abstinent individual will return to harmful alcohol consumption. We are most vulnerable to the effects of stress during early development, and exposure to multiple traumatic early life events dramatically increases the risk for AUDs. However, not everyone exposed to early life stress will develop an AUD. The mechanisms determining whether an individual’s brain adapts and becomes resilient to the effects of stress or succumbs and is unable to cope with stress remain elusive. Emerging evidence suggests that neuroplastic changes in the nucleus accumbens (NAc following early life stress underlie the development of AUDs. This review discusses the impact of early life stress on NAc structure and function, how these changes affect cholinergic signaling within the mesolimbic reward pathway and the role nicotinic acetylcholine receptors (nAChRs play in this process. Understanding the neural pathways and mechanism determining stress resilience or susceptibility will improve our ability to identify individuals susceptible to developing AUDs, formulate cognitive interventions to prevent AUDs in susceptible individuals and to elucidate and enhance potential therapeutic targets, such as the nAChRs, for those struggling to overcome an AUD.

  8. Shear-induced Bubble Coalescence in Rhyolitic Melts with Low Vesicularity

    Science.gov (United States)

    Okumura, S.; Nakamura, M.; Tsuchiyama, A.

    2006-12-01

    Development of bubble structure during magma ascent controls the dynamics of volcanic eruption, because the bubble structure influences the magma rheology and permeability, and hence magma degassing. In the flowing magmas, the bubble structure is expected to be changed by shear, as pointed out by some previous studies based on geological observations. However, the development of bubble structure has been experimentally studied only in the isostatic magmas. We have experimentally demonstrated for the first time, the shear-induced development of number density, size and shape of bubbles in a rhyolitic melt. The deformation experiments were performed by using an externally heated, piston-cylinder type apparatus with a rotational piston. At 975°C, natural obsidian (initial water content of 0.5 wt%) having cylindrical shape (ca. 4.7 mm in diameter and 5 mm in length) was vesiculated in the graphite container (ca. 5 and 9 mm in the inner and the outer diameters, respectively, and 5 mm in length), and the vesiculated samples were twisted at various rotational speeds up to 1 rpm. The number density, size and shape of bubbles in the quenched samples were then measured by using the X-ray computed tomography. The size distribution of bubbles shows that the number of larger bubbles increases with the rotational speed and at the outer zone of the samples at which the shear rate is high. In the high shear rate zone, the magnitude of bubble deformation is large. The 3D images of large bubbles clearly indicate that they were formed by coalescence. These results indicate that the degree of bubble coalescence is enhanced with the shear rate. The experimental results also demonstrated that the coalescence of bubbles occur even at low vesicularity (ca. 20 vol.%). Because the shear rate induced in this study (in the order of 0.01 1/s) seems to be produced for magmas ascending in a volcanic conduit, we propose the possibility that the vesiculated magmas undergo bubble coalescence at a

  9. Bovine Vaccinia in dairy cattle and suspicion of vesicular disease on milkers in Brazil

    Directory of Open Access Journals (Sweden)

    Thaís Garcia da Silva

    2018-05-01

    Full Text Available ABSTRACT: Bovine vaccinia (BV is a vesicular disease induced by the Vaccinia virus (VACV that affects milk production and is an occupational zoonosis. This research had the following objectives: (i detection of VACV by qPCR in cattle with clinical suspicion of vesicular disease; (ii symptoms characterization in animals and milkers with clinical suspicion of the disease and virus detection in humans; and (iii identification of risk factors for infections of VACV in herds from several Brazilian states. A total of 471 bovine epithelial samples from dairy farms, in 15 Brazilian states, were evaluated between 2007 and 2012. The samples were tested by quantitative PCR (qPCR using SYBR Green® reagents, validated with a lower limit of detection of 100 TCID50/50µL (1.7x100 viral particles, and 45.1% of VACV positive samples were detected. Using official forms for epidemiological investigation (FORM-IN, the risk factors for VACV infections in cattle were determined to be farms with a lack of technological facilities (P=0.029 and the presence of rodents (P=0.001. There was an effect of seasonality in cattle with a higher occurrence of BV during the dry season. A total of 420 epidemiological questionnaires were applied at public health care centers, where 100% of the milkers had vesicular lesions on their hands (98.1% and on their arms (6.9%. The most frequent clinical symptoms in humans were: local swelling (74.2%, headache (20.7%, fever (10.4% and inguinal lymphadenopathy (74.2%. Only 19.98% of milkers aged between 39 and 58 years were seroreactive to VACV and were immunized with the human anti-smallpox vaccine. There was an increase in the frequency of BV in older individuals due to their natural decrease in specific immunity. It has been shown that the implementation of zootechnical management techniques and health planning are important for the prevention of BV in animals and humans.

  10. Vesicular Trafficking Defects, Developmental Abnormalities, and Alterations in the Cellular Death Process Occur in Cell Lines that Over-Express Dictyostelium GTPase, Rab2, and Rab2 Mutants

    Directory of Open Access Journals (Sweden)

    Katherine Maringer

    2014-08-01

    Full Text Available Small molecular weight GTPase Rab2 has been shown to be a resident of pre-Golgi intermediates and required for protein transport from the ER to the Golgi complex, however, the function of Rab2 in Dictyostelium has yet to be fully characterized. Using cell lines that over-express DdRab2, as well as cell lines over-expressing constitutively active (CA, and dominant negative (DN forms of the GTPase, we report a functional role in vesicular transport specifically phagocytosis, and endocytosis. Furthermore, Rab2 like other GTPases cycles between an active GTP-bound and an inactive GDP-bound state. We found that this GTP/GDP cycle for DdRab2 is crucial for normal Dictyostelium development and cell–cell adhesion. Similar to Rab5 and Rab7 in C. elegans, we found that DdRab2 plays a role in programmed cell death, possibly in the phagocytic removal of apoptotic corpses.

  11. Cognitive effects of dopamine depletion in the context of diminished acetylcholine signaling capacity in mice

    Directory of Open Access Journals (Sweden)

    Lilia Zurkovsky

    2013-01-01

    A subset of patients with Parkinson’s disease acquires a debilitating dementia characterized by severe cognitive impairments (i.e. Parkinson’s disease dementia; PDD. Brains from PDD patients show extensive cholinergic loss as well as dopamine (DA depletion. We used a mutant mouse model to directly test whether combined cholinergic and DA depletion leads to a cognitive profile resembling PDD. Mice carrying heterozygous deletion of the high-affinity, hemicholinium-3-sensitive choline transporter (CHTHET show reduced levels of acetylcholine throughout the brain. We achieved bilateral DA depletion in CHTHET and wild-type (WT littermates via intra-striatal infusion of 6-hydroxydopamine (6-OHDA, or used vehicle as control. Executive function and memory were evaluated using rodent versions of cognitive tasks commonly used with human subjects: the set-shifting task and spatial and novel-object recognition paradigms. Our studies revealed impaired acquisition of attentional set in the set-shifting paradigm in WT-6OHDA and CHTHET-vehicle mice that was exacerbated in the CHTHET-6OHDA mice. The object recognition test following a 24-hour delay was also impaired in CHTHET-6OHDA mice compared with all other groups. Treatment with acetylcholinesterase (AChE inhibitors physostigmine (0.05 or 0.1 mg/kg and donepezil (0.1 and 0.3 mg/kg reversed the impaired object recognition of the CHTHET-6OHDA mice. Our data demonstrate an exacerbated cognitive phenotype with dual ACh and DA depletion as compared with either insult alone, with traits analogous to those observed in PDD patients. The results suggest that combined loss of DA and ACh could be sufficient for pathogenesis of specific cognitive deficits in PDD.

  12. The lipidomes of vesicular stomatitis virus, semliki forest virus, and the host plasma membrane analyzed by quantitative shotgun mass spectrometry

    DEFF Research Database (Denmark)

    Kalvodova, Lucie; Sampaio, Julio L; Cordo, Sandra

    2009-01-01

    kidney cells can be infected by two different viruses, namely, vesicular stomatitis virus and Semliki Forest virus, from the Rhabdoviridae and Togaviridae families, respectively. We purified the host plasma membrane and the two different viruses after exit from the host cells and analyzed the lipid...

  13. Carbon Fiber Ultramicrodic Electrode Electrodeposited with Over-Oxidized Polypyrrole for Amperometric Detection of Vesicular Exocytosis from Pheochromocytoma Cell

    Directory of Open Access Journals (Sweden)

    Li Wang

    2015-01-01

    Full Text Available Vesicular exocytosis is ubiquitous, but it is difficult to detect within the cells’ communication mechanism. For this purpose, a 2 µm ultramicrodic carbon fiber electrode was fabricated in this work based on electrodeposition with over-oxidized polypyrrole nanoparticle (PPyox-CFE, which was applied successfully for real-time monitoring of quantal exocytosis from individual pheochromocytoma (PC12 cells. PPyox-CFE was evaluated by dopamine (DA solutions through cyclic voltammetry and amperometry electrochemical methods, and results revealed that PPyox-CFE improved the detection limit of DA. In particular, the sensitivity of DA was improved to 24.55 µA·µM−1·µm−2 using the PPyox-CFE. The ultramicrodic electrode combined with the patch-clamp system was used to detect vesicular exocytosis of DA from individual PC12 cells with 60 mM K+ stimulation. A total of 287 spikes released from 7 PC12 cells were statistically analyzed. The current amplitude (Imax and the released charge (Q of the amperometric spikes from the DA release by a stimulated PC12 cell is 45.1 ± 12.5 pA and 0.18 ± 0.04 pC, respectively. Furthermore, on average ~562,000 molecules were released in each vesicular exocytosis. PPyox-CFE, with its capability of detecting vesicular exocytosis, has potential application in neuron communication research.

  14. APPARENT LACK OF VESICULAR-ARBUSCULAR MYCORRHIZA (VAM) IN SEAGRASSES ZOSTERA MARINA L. AND THALASSIA TESTUDIUM BANKS EX KONIG

    Science.gov (United States)

    We examined two populations of Zostera marina L. and one of Thalassia testudinum Banks ex Konig for presence of vesicular-arbuscular mycorrhiza (VAM). None of these plants showed any VAM colonization. In addition, we were unable to find any literature references on the presence o...

  15. Loss-of-function mutations in ATP6V0A2 impair vesicular trafficking, tropoelastin secretion and cell survival.

    NARCIS (Netherlands)

    Hucthagowder, V.; Morava, E.; Kornak, U.; Lefeber, D.J.; Fischer, B.; Dimopoulou, A.; Aldinger, A.; Choi, J.; Davis, E.C.; Abuelo, D.N.; Adamowicz, M.; Al-Aama, J.Y.; Basel-Vanagaite, L.; Fernandez, B.; Greally, M.T.; Gillessen-Kaesbach, G.; Kayserili, H.; Lemyre, E.; Tekin, M.; Turkmen, S.; Tuysuz, B.; Yuksel-Konuk, B.; Mundlos, S.; Maldergem, L. van; Wevers, R.A.; Urban, Z.

    2009-01-01

    Autosomal recessive cutis laxa type 2 (ARCL2), a syndrome of growth and developmental delay and redundant, inelastic skin, is caused by mutations in the a2 subunit of the vesicular ATPase H+-pump (ATP6V0A2). The goal of this study was to define the disease mechanisms that lead to connective tissue

  16. Rapid synthesis of acetylcholine receptors at neuromuscular junctions.

    Science.gov (United States)

    Ramsay, D A; Drachman, D B; Pestronk, A

    1988-10-11

    The rate of acetylcholine receptor (AChR) degradation in mature, innervated mammalian neuromuscular junctions has recently been shown to be biphasic; up to 20% are rapidly turned over (RTOs; half life less than 1 day) whereas the remainder are lost more slowly ('stable' AChRs; half life 10-12 days). In order to maintain normal junctional receptor density, synthesis and insertion of AChRs should presumably be sufficiently rapid to replace both the RTOs and the stable receptors. We have tested this prediction by blocking pre-existing AChRs in the mouse sternomastoid muscle with alpha-bungarotoxin (alpha-BuTx), and monitoring the subsequent appearance of 'new' junctional AChRs at intervals of 3 h to 20 days by labeling them with 125I-alpha-BuTx. The results show that new receptors were initially inserted rapidly (16% at 24 h and 28% at 48 h). The rate of increase of 'new' 125I-alpha-BuTx binding sites gradually slowed down during the remainder of the time period studied. Control observations excluded possible artifacts of the experimental procedure including incomplete blockade of AChRs, dissociation of toxin-receptor complexes, or experimentally induced alteration of receptor synthesis. The present demonstration of rapid synthesis and incorporation of AChRs at innervated neuromuscular junctions provides support for the concept of a subpopulation of rapidly turned over AChRs. The RTOs may serve as precursors for the larger population of stable receptors and have an important role in the metabolism of the neuromuscular synapse.

  17. Acetylcholine receptors in dementia and mild cognitive impairment

    Energy Technology Data Exchange (ETDEWEB)

    Sabri, Osama; Kendziorra, Kai [University of Leipzig, Department of Nuclear Medicine, Leipzig (Germany); Wolf, Henrike; Gertz, Hermann-Josef [University of Leipzig, Department of Psychiatry, Leipzig (Germany); Brust, Peter [Institute of Interdisciplinary Isotope Research, Leipzig (Germany)

    2008-03-15

    To clarify whether changes in the cholinergic transmission occur early in the course of Alzheimer's disease (AD), we carried out positron emission tomography (PET) with the radioligand 2-[{sup 18}F]F-A-85380, which is supposed to be specific for {alpha}4{beta}2 nicotinic acetylcholine receptors (nAChRs). We included patients with moderate to severe AD and patients with amnestic mild cognitive impairment (MCI), presumed to present preclinical AD. Both patients with AD and MCI showed significant reductions in {alpha}4{beta}2 nAChRs in brain regions typically affected by AD pathology. These findings indicate that a reduction in {alpha}4{beta}2 nAChRs occurs during early symptomatic stages of AD. The {alpha}4{beta}2 nAChR availability in these regions correlated with the severity of cognitive impairment, indicating a stage sensitivity of the {alpha}4{beta}2 nAChR status. Together, our results provide evidence for the potential of 2-[{sup 18}]F-A-85380 nAChR PET in the diagnosis of patients at risk for AD. Because of the extraordinary long acquisition time with 2-[{sup 18}F]F-A-85380, we developed the new {alpha}4{beta}2 nAChR-specific radioligands (+)- and (-)-[{sup 18}F]norchloro-fluoro-homoepibatidine (NCFHEB) and evaluated them preclinically. (-)-[{sup 18}F]NCFHEB shows twofold higher brain uptake and significantly shorter acquisition times. Therefore, (-)-[{sup 18}F]NCFHEB should be a suitable radioligand for larger clinical investigations. (orig.)

  18. The formation of acetylcholine receptor clusters visualized with quantum dots

    Directory of Open Access Journals (Sweden)

    Peng H Benjamin

    2009-07-01

    Full Text Available Abstract Background Motor innervation of skeletal muscle leads to the assembly of acetylcholine receptor (AChR clusters in the postsynaptic membrane at the vertebrate neuromuscular junction (NMJ. Synaptic AChR aggregation, according to the diffusion-mediated trapping hypothesis, involves the establishment of a postsynaptic scaffold that "traps" freely diffusing receptors into forming high-density clusters. Although this hypothesis is widely cited to explain the formation of postsynaptic AChR clusters, direct evidence at molecular level is lacking. Results Using quantum dots (QDs and live cell imaging, we provide new measurements supporting the diffusion-trap hypothesis as applied to AChR cluster formation. Consistent with published works, experiments on cultured Xenopus myotomal muscle cells revealed that AChRs at clusters that formed spontaneously (pre-patterned clusters, also called hot spots and at those induced by nerve-innervation or by growth factor-coated latex beads were very stable whereas diffuse receptors outside these regions were mobile. Moreover, despite the restriction of AChR movement at sites of synaptogenic stimulation, individual receptors away from these domains continued to exhibit free diffusion, indicating that AChR clustering at NMJ does not involve an active attraction of receptors but is passive and diffusion-driven. Conclusion Single-molecular tracking using QDs has provided direct evidence that the clustering of AChRs in muscle cells in response to synaptogenic stimuli is achieved by two distinct cellular processes: the Brownian motion of receptors in the membrane and their trapping and immobilization at the synaptic specialization. This study also provides a clearer picture of the "trap" that it is not a uniformly sticky area but consists of discrete foci at which AChRs are immobilized.

  19. Evaluation of PET Radioligands for the neuronal nicotinic acetylcholine receptor

    International Nuclear Information System (INIS)

    Schoenbaechler, R.; Westera, G.; Nan-Horng Lin

    2002-01-01

    Full text: A-186253.1, a compound made by Abbott laboratories, was labelled with carbon-11 and evaluated as a PET ligand for the neuronal nicotinic acetylcholine receptor (nAChR). The compound was labelled with C-11 by methylation with 11C-MeI of the desmethyl precursor A-183828.1. The affinity of A-186253.1 for the α4β2 and the α7 subtype of the nAChR was determined in displacement studies. PET-studies were performed in rats and pigs Inhibitory constants (K i ) versus cytsine were 461 ± 99 pM for A-186253.1 and versus α-Bungarotoxin >100 μM. which means a very high selectivity for the α4β2-receptor (>227,000). Highest uptake of [ 11 C]-A-186253.1 was observed in the thalamus where an increase in radiotracer uptake was seen until 45 min p.i.. Thereafter, the radiotracer concentration remained constant until the end of the scan indicating slow washout of [ 11 C]-A-186253.1. Application of cold A-186253.1 (0.5 mg/kg) 40 min p.i. resulted in a decrease in radiotracer concentration in the thalamus and the cortex indicating displacement of [ 11 C]-A-186253.1. Blockade studies with cytisine (0.5 mg/kg), a selective ligand for the α4β2 nicotinic receptor, showed just a slight reduction of the radioligand uptake in the thalamus and in the cortex whereas the blockade with cold A-186253.1 (1 mg/kg) resulted in a 50 % reduction. These results suggest, that 50 % of the [ 11 C]-A-186253.1 in the brain corresponds to specifically bound radioligand, but not to the α4β2 subtype of the nicotinic receptor. (author)

  20. Loss of Acetylcholine Signaling Reduces Cell Clearance Deficiencies in Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Sérgio M Pinto

    Full Text Available The ability to eliminate undesired cells by apoptosis is a key mechanism to maintain organismal health and homeostasis. Failure to clear apoptotic cells efficiently can cause autoimmune diseases in mammals. Genetic studies in Caenorhabditis elegans have greatly helped to decipher the regulation of apoptotic cell clearance. In this study, we show that the loss of levamisole-sensitive acetylcholine receptor, but not of a typical neuronal acetylcholine receptor causes a reduction in the number of persistent cell corpses in worms suffering from an engulfment deficiency. This reduction is not caused by impaired or delayed cell death but rather by a partial restoration of the cell clearance capacity. Mutants in acetylcholine turn-over elicit a similar phenotype, implying that acetylcholine signaling is the process responsible for these observations. Surprisingly, tissue specific RNAi suggests that UNC-38, a major component of the levamisole-sensitive receptor, functions in the dying germ cell to influence engulfment efficiency. Animals with loss of acetylcholine receptor exhibit a higher fraction of cell corpses positive for the "eat-me" signal phosphatidylserine. Our results suggest that modulation by ion channels of ion flow across plasma membrane in dying cells can influence the dynamics of phosphatidylserine exposure and thus clearance efficiency.

  1. Two types of muscarinic acetylcholine receptors in Drosophila and other arthropods

    DEFF Research Database (Denmark)

    Collin, Caitlin Alexis; Hauser, Frank; Gonzalez de Valdivia, Ernesto I

    2013-01-01

    Muscarinic acetylcholine receptors (mAChRs) play a central role in the mammalian nervous system. These receptors are G protein-coupled receptors (GPCRs), which are activated by the agonists acetylcholine and muscarine, and blocked by a variety of antagonists. Mammals have five mAChRs (m1-m5......). In this study, we cloned two structurally related GPCRs from the fruit fly Drosophila melanogaster, which, after expression in Chinese hamster ovary cells, proved to be muscarinic acetylcholine receptors. One mAChR (the A-type; encoded by gene CG4356) is activated by acetylcholine (EC50, 5 × 10(-8) M......) and muscarine (EC50, 6 × 10(-8) M) and blocked by the classical mAChR antagonists atropine, scopolamine, and 3-quinuclidinyl-benzilate (QNB), while the other (the B-type; encoded by gene CG7918) is also activated by acetylcholine, but has a 1,000-fold lower sensitivity to muscarine, and is not blocked...

  2. Non-Neuronal Functions of the M2 Muscarinic Acetylcholine Receptor

    Directory of Open Access Journals (Sweden)

    Ritva Tikkanen

    2013-04-01

    Full Text Available Acetylcholine is an important neurotransmitter whose effects are mediated by two classes of receptors. The nicotinic acetylcholine receptors are ion channels, whereas the muscarinic receptors belong to the large family of G protein coupled seven transmembrane helix receptors. Beyond its function in neuronal systems, it has become evident that acetylcholine also plays an important role in non-neuronal cells such as epithelial and immune cells. Furthermore, many cell types in the periphery are capable of synthesizing acetylcholine and express at least some of the receptors. In this review, we summarize the non-neuronal functions of the muscarinic acetylcholine receptors, especially those of the M2 muscarinic receptor in epithelial cells. We will review the mechanisms of signaling by the M2 receptor but also the cellular trafficking and ARF6 mediated endocytosis of this receptor, which play an important role in the regulation of signaling events. In addition, we provide an overview of the M2 receptor in human pathological conditions such as autoimmune diseases and cancer.

  3. The novel protein kinase C epsilon isoform modulates acetylcholine release in the rat neuromuscular junction.

    Science.gov (United States)

    Obis, Teresa; Hurtado, Erica; Nadal, Laura; Tomàs, Marta; Priego, Mercedes; Simon, Anna; Garcia, Neus; Santafe, Manel M; Lanuza, Maria A; Tomàs, Josep

    2015-12-01

    Various protein kinase C (PKC) isoforms contribute to the phosphorylating activity that modulates neurotransmitter release. In previous studies we showed that nPKCε is confined in the presynaptic site of the neuromuscular junction and its presynaptic function is activity-dependent. Furthermore, nPKCε regulates phorbol ester-induced acetylcholine release potentiation, which further indicates that nPKCε is involved in neurotransmission. The present study is designed to examine the nPKCε involvement in transmitter release at the neuromuscular junction. We use the specific nPKCε translocation inhibitor peptide εV1-2 and electrophysiological experiments to investigate the involvement of this isoform in acetylcholine release. We observed that nPKCε membrane translocation is key to the synaptic potentiation of NMJ, being involved in several conditions that upregulate PKC isoforms coupling to acetylcholine (ACh) release (incubation with high Ca(2+), stimulation with phorbol esters and protein kinase A, stimulation with adenosine 3',5'-cyclic monophosphorothioate, 8-Bromo-, Rp-isomer, sodium salt -Sp-8-BrcAMP-). In all these conditions, preincubation with the nPKCε translocation inhibitor peptide (εV1-2) impairs PKC coupling to acetylcholine release potentiation. In addition, the inhibition of nPKCε translocation and therefore its activity impedes that presynaptic muscarinic autoreceptors and adenosine autoreceptors modulate transmitter secretion. Together, these results point to the importance of nPKCε isoform in the control of acetylcholine release in the neuromuscular junction.

  4. Interactions of macrophages with probiotic bacteria lead to increased antiviral response against vesicular stomatitis virus

    DEFF Research Database (Denmark)

    Ivec, Martin; Botic, Tanja; Koren, Srecko

    2007-01-01

    and by producing chemokines and immunoregulatory cytokines that enable the adaptive immune response to recognize infected cells and perform antiviral effector functions. Probiotics, as a part of the normal gut intestinal flora, are important in supporting a functional yet balanced immune system. Improving our...... understanding of their role in the activation of macrophages and their stimulation of proinflammatory cytokine production in early viral infection was the main goal of this study. Our in vitro model study showed that probiotic bacteria, either from the species Lactobacillus or Bifidobacteria have the ability...... dehydrogenases activity could be implied as the first indicator of potential inhibitory effects of the probiotics on virus replication. The interactions between probiotic bacteria, macrophages and vesicular stomatitis virus (VSV), markedly depended on the bacterial strain studied....

  5. Viral meningitis epidemics and a single, recent, recombinant and anthroponotic origin of swine vesicular disease virus

    DEFF Research Database (Denmark)

    Bruhn, Christian Anders Wathne; Nielsen, Sandra Cathrine Abel; Samaniego Castruita, Jose Alfredo

    2015-01-01

    BACKGROUND AND OBJECTIVES: Swine vesicular disease virus (SVDV) is a close relative of the human Enterovirus B serotype, coxsackievirus B5. As the etiological agent of a significant emergent veterinary disease, several studies have attempted to explain its origin. However, several key questions...... and non-coding regions supports that SVDV has a recombinant origin between coxsackievirus B5 and another Enterovirus B serotype, most likely coxsackievirus A9. Extensive Bayesian sequence-based analysis of the time of the most recent common ancestor of all analysed sequences places this within a few years...... around 1961. Epidemiological evidence points to China as an origin, but there are no available samples to test this conclusively. CONCLUSIONS AND IMPLICATIONS: Historical investigation and the clinical aspects of the involved Enterovirus B serotypes, makes the current results consistent with a hypothesis...

  6. `Full fusion' is not ineluctable during vesicular exocytosis of neurotransmitters by endocrine cells

    Science.gov (United States)

    Oleinick, Alexander; Svir, Irina; Amatore, Christian

    2017-01-01

    Vesicular exocytosis is an essential and ubiquitous process in neurons and endocrine cells by which neurotransmitters are released in synaptic clefts or extracellular fluids. It involves the fusion of a vesicle loaded with chemical messengers with the cell membrane through a nanometric fusion pore. In endocrine cells, unless it closes after some flickering (`Kiss-and-Run' events), this initial pore is supposed to expand exponentially, leading to a full integration of the vesicle membrane into the cell membrane-a stage called `full fusion'. We report here a compact analytical formulation that allows precise measurements of the fusion pore expansion extent and rate to be extracted from individual amperometric spike time courses. These data definitively establish that, during release of catecholamines, fusion pores enlarge at most to approximately one-fifth of the radius of their parent vesicle, hence ruling out the ineluctability of `full fusion'.

  7. Pharmacological factors in the saliva of blood-feeding insects. Implications for vesicular stomatitis epidemiology.

    Science.gov (United States)

    Tabachnick, W J

    2000-01-01

    Vesicular stomatitis (VS) epizootics in the Western United States have caused substantial economic losses to U.S. livestock industries in 1995, 1997, and 1998. The role of arthropods in transmitting VS to U.S. livestock is unclear. In particular, the impact of arthropod salivary gland factors in VS infections in livestock needs study. Pharmacological effects of arthropod salivary gland factors on animals are reviewed. The potential effects of arthropod saliva on the transmission and spread of VS virus to livestock in the Western U.S. is presented with emphasis on the biting midge, Culicoides sonorensis. Information is discussed with attention to vector potential of C. sonorensis, and its use as a model for evaluating insect salivary gland pharmacology on livestock response to VS.

  8. Phosphorus use efficiency of tomato as influenced by phosphorus and vesicular arbuscular mycorrhizal (VAM) fungi inoculation

    International Nuclear Information System (INIS)

    Dhinakaran, R.; Savithri, P.

    1997-01-01

    A pot experiment was conducted on tomato (Lycopersicon esculentum L.var. CO3) grown in red non-calcareous soil (Paralythic Ustochrept) to study the effect of different P treatments involving single superphosphate (SSP) and Mussoorie rock phosphate (MRP) added at different levels, viz. 100 and 75 kg P 2 O 5 /ha along with and without vesicular arbuscular mycorrhizal (VAM) fungi inoculation. The results revealed that the P application as superphosphate at 100 kg P 2 O 5 /ha significantly increased the yield of tomato but the application of VAM fungi did not have any pronounced effect on tomato yield. The 32 P studies confirmed the increased uptake of P by the plants at higher level of P application. P content and its uptake by tomato fruit increased with the increasing levels of P application and VAM inoculation. The VAM fungi inoculation was also helpful in increasing the fertilizer use efficiency and also per cent P derived from fertilizer. (author)

  9. Increased sporulation of vesicular-arbuscular mycorrhizal fungi by manipulation of nutrient regimens.

    Science.gov (United States)

    Douds, D D; Schenck, N C

    1990-02-01

    Adjustment of pot culture nutrient solutions increased root colonization and sporulation of vesicular-arbuscular mycorrhizal (VAM) fungi. Paspalum notatum Flugge and VAM fungi were grown in a sandy soil low in N and available P. Hoagland nutrient solution without P enhanced sporulation in soil and root colonization of Acaulospora longula, Scutellospora heterogama, Gigaspora margarita, and a wide range of other VAM fungi over levels produced by a tap water control or nutrient solutions containing P. However, Glomus intraradices produced significantly more spores in plant roots in the tap water control treatment. The effect of the nutrient solutions was not due solely to N nutrition, because the addition of NH(4)NO(3) decreased both colonization and sporulation by G. margarita relative to levels produced by Hoagland solution without P.

  10. Effects of vesicular-arbuscular mycorrhizal (VAM) fungi on the seedling growth of three Pistacia species.

    Science.gov (United States)

    Caglar, S; Akgun, A

    2006-07-01

    The experiment was undertaken to test the efficiency of inoculation of vesicular-arbuscular mycorrhizal (VAM) fungi on the seedling growth of three Pistacia species used as rootstocks. The stratified Pistacia seeds were inoculated with VAM fungi. The highest rate of inoculated roots was 96.7% in P. khinjuck seedlings with G. clarum and G. etunicatum, 83.3% in P. vera seedlings with G. caledonium and 73.3% in P. terebinthus seedlings with G. caledonium. Mycorrhizal inoculations improved seedling height only in P. terebinthus. Certain mycorrhizal inoculations increased the leaf N, but not P and K contents. Seedlings inoculated with G. caledonium had higher reducing sugar contents. It was concluded that pre-inoculated Pistacia seedlings could have a better growth in the harsh field conditions.

  11. Inhibition of synaptically evoked cortical acetylcholine release by adenosine: an in vivo microdialysis study in the rat.

    Science.gov (United States)

    Materi, L M; Rasmusson, D D; Semba, K

    2000-01-01

    The release of cortical acetylcholine from the intracortical axonal terminals of cholinergic basal forebrain neurons is closely associated with electroencephalographic activity. One factor which may act to reduce cortical acetylcholine release and promote sleep is adenosine. Using in vivo microdialysis, we examined the effect of adenosine and selective adenosine receptor agonists and antagonists on cortical acetylcholine release evoked by electrical stimulation of the pedunculopontine tegmental nucleus in urethane anesthetized rats. All drugs were administered locally within the cortex by reverse dialysis. None of the drugs tested altered basal release of acetylcholine in the cortex. Adenosine significantly reduced evoked cortical acetylcholine efflux in a concentration-dependent manner. This was mimicked by the adenosine A(1) receptor selective agonist N(6)-cyclopentyladenosine and blocked by the selective A(1) receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX). The A(2A) receptor agonist 2-[p-(2-carboxyethyl)-phenethylamino]-5'-N-ethylcarboxamidoadenosi ne hydrochloride (CGS 21680) did not alter evoked cortical acetylcholine release even in the presence of DPCPX. Administered alone, neither DPCPX nor the non-selective adenosine receptor antagonist caffeine affected evoked cortical acetylcholine efflux. Simultaneous delivery of the adenosine uptake inhibitors dipyridamole and S-(4-nitrobenzyl)-6-thioinosine significantly reduced evoked cortical acetylcholine release, and this effect was blocked by the simultaneous administration of caffeine. These data indicate that activation of the A(1) adenosine receptor inhibits acetylcholine release in the cortex in vivo while the A(2A) receptor does not influence acetylcholine efflux. Such inhibition of cortical acetylcholine release by adenosine may contribute to an increased propensity to sleep during prolonged wakefulness.

  12. Antibodies against vesicular stomatitis virus in horses from southern, midwestern and northeastern Brazilian States

    Directory of Open Access Journals (Sweden)

    Vinícius Leobet Lunkes

    2016-08-01

    Full Text Available ABSTRACT: Vesicular stomatitis virus (VSV is the agent of a vesicular disease that affects many animal species and may be clinically confounded with foot-and-mouth disease in ruminant and swine. Horses are especially susceptible to VSV and may serve as sentinels for virus circulation. The present study investigated the presence of neutralizing antibodies against VSV Indiana III (VSIV-3 in serum samples of 3,626 horses from six states in three Brazilian regions: Southern (RS, n = 1,011, Midwest (GO/DF, n = 1,767 and Northeast (PB, PE, RN and CE, n = 848 collected between 2013 and 2014. Neutralizing antibodies against VSIV-3 (titers ≥40 were detected in 641 samples (positivity of 17.7%; CI95%:16.5-19.0%, being 317 samples from CE (87.3%; CI95%: 83.4-90.5 %; 109 from RN (65.7%; CI95%: 57.8 -72.7%; 124 from PB (45.4%; CI95%: 39.4-51.5%; 78 from GO/DF (4.4%; CI95%: 3.5-5.5% and nine samples of RS (0.9%; CI95%: 0.4-1.7%. Several samples from the Northeast and Midwest harbored high neutralizing titers, indicating a recent exposure to the virus. In contrast, samples from RS had low titers, possibly due to a past remote exposure. Several positive samples presented neutralizing activity against other VSV serotypes (Indiana I and New Jersey, yet in lower titers, indicating the specificity of the response to VSIV-3. These results demonstrated a relatively recent circulation of VSIV-3 in northeastern Brazilian States, confirming clinical findings and demonstrating the sanitary importance of this infection.

  13. Topical vesicular formulations of Curcuma longa extract on recuperating the ultraviolet radiation-damaged skin.

    Science.gov (United States)

    Kaur, Chanchal Deep; Saraf, Swarnlata

    2011-12-01

      Ultraviolet radiations generate reactive oxygen species, leading to adverse effects on skin properties. Botanical extracts are multifunctional in nature having various properties like photoprotection, anti-aging, moisturizing, antioxidant, astringent, anti-irritant, and antimicrobial activity.   The aim of this study was to formulate creams having Curcuma longa extract loaded novel vesicular systems (liposomes, ethosomes, and transfersomes) and study their photoprotective effect by assessment of skin hydration (Cutometer) and sebum content (Sebumeter).   The alcoholic C. longa extract loaded liposomes, ethosomes, and transfersomes having 0.5-2.0% w/w extract were prepared, evaluated for size, entrapment efficiency, and incorporated into the cream. Their long-term interaction with skin (6 weeks) was compared in terms of their effects on skin hydration and sebum content.   Vesicular size obtained was in the range 167.3 ± 3.0 to 262.4 ± 2.4 nm with low polydispersity index (0.2-0.3) and high entrapment efficiency. The efficacy was in the order C. longa extract loaded transfersomal creams > C. longa extract loaded ethosomal creams > C. longa extract loaded liposomal creams > C. longa extract loaded creams > Empty transfersome loaded cream > Empty ethosome loaded cream > Empty liposome loaded cream > Base cream.   The photoprotective properties of the constituents of C. longa extract and hydrant, moisturizing lipid components of nano vesicles with better skin penetration resulted in improvement in skin properties like skin hydration and sebum content. The herbal extract loaded nano vesicles incorporated in cream could be used as photoprotective formulations. © 2011 Wiley Periodicals, Inc.

  14. Leucine-rich repeat-containing G protein-coupled receptor 4 facilitates vesicular stomatitis virus infection by binding vesicular stomatitis virus glycoprotein.

    Science.gov (United States)

    Zhang, Na; Huang, Hongjun; Tan, Binghe; Wei, Yinglei; Xiong, Qingqing; Yan, Yan; Hou, Lili; Wu, Nannan; Siwko, Stefan; Cimarelli, Andrea; Xu, Jianrong; Han, Honghui; Qian, Min; Liu, Mingyao; Du, Bing

    2017-10-06

    Vesicular stomatitis virus (VSV) and rabies and Chandipura viruses belong to the Rhabdovirus family. VSV is a common laboratory virus to study viral evolution and host immune responses to viral infection, and recombinant VSV-based vectors have been widely used for viral oncolysis, vaccination, and gene therapy. Although the tropism of VSV is broad, and its envelope glycoprotein G is often used for pseudotyping other viruses, the host cellular components involved in VSV infection remain unclear. Here, we demonstrate that the host protein leucine-rich repeat-containing G protein-coupled receptor 4 (Lgr4) is essential for VSV and VSV-G pseudotyped lentivirus (VSVG-LV) to infect susceptible cells. Accordingly, Lgr4-deficient mice had dramatically decreased VSV levels in the olfactory bulb. Furthermore, Lgr4 knockdown in RAW 264.7 cells also significantly suppressed VSV infection, and Lgr4 overexpression in RAW 264.7 cells enhanced VSV infection. Interestingly, only VSV infection relied on Lgr4, whereas infections with Newcastle disease virus, influenza A virus (A/WSN/33), and herpes simplex virus were unaffected by Lgr4 status. Of note, assays of virus entry, cell ELISA, immunoprecipitation, and surface plasmon resonance indicated that VSV bound susceptible cells via the Lgr4 extracellular domain. Pretreating cells with an Lgr4 antibody, soluble LGR4 extracellular domain, or R-spondin 1 blocked VSV infection by competitively inhibiting VSV binding to Lgr4. Taken together, the identification of Lgr4 as a VSV-specific host factor provides important insights into understanding VSV entry and its pathogenesis and lays the foundation for VSV-based gene therapy and viral oncolytic therapeutics. © 2017 by The American Society for Biochemistry and Molecular Biology, Inc.

  15. SAD-B kinase regulates pre-synaptic vesicular dynamics at hippocampal Schaffer collateral synapses and affects contextual fear memory.

    Science.gov (United States)

    Watabe, Ayako M; Nagase, Masashi; Hagiwara, Akari; Hida, Yamato; Tsuji, Megumi; Ochiai, Toshitaka; Kato, Fusao; Ohtsuka, Toshihisa

    2016-01-01

    Synapses of amphids defective (SAD)-A/B kinases control various steps in neuronal development and differentiation, such as axon specifications and maturation in central and peripheral nervous systems. At mature pre-synaptic terminals, SAD-B is associated with synaptic vesicles and the active zone cytomatrix; however, how SAD-B regulates neurotransmission and synaptic plasticity in vivo remains unclear. Thus, we used SAD-B knockout (KO) mice to study the function of this pre-synaptic kinase in the brain. We found that the paired-pulse ratio was significantly enhanced at Shaffer collateral synapses in the hippocampal CA1 region in SAD-B KO mice compared with wild-type littermates. We also found that the frequency of the miniature excitatory post-synaptic current was decreased in SAD-B KO mice. Moreover, synaptic depression following prolonged low-frequency synaptic stimulation was significantly enhanced in SAD-B KO mice. These results suggest that SAD-B kinase regulates vesicular release probability at pre-synaptic terminals and is involved in vesicular trafficking and/or regulation of the readily releasable pool size. Finally, we found that hippocampus-dependent contextual fear learning was significantly impaired in SAD-B KO mice. These observations suggest that SAD-B kinase plays pivotal roles in controlling vesicular release properties and regulating hippocampal function in the mature brain. Synapses of amphids defective (SAD)-A/B kinases control various steps in neuronal development and differentiation, but their roles in mature brains were only partially known. Here, we demonstrated, at mature pre-synaptic terminals, that SAD-B regulates vesicular release probability and synaptic plasticity. Moreover, hippocampus-dependent contextual fear learning was significantly impaired in SAD-B KO mice, suggesting that SAD-B kinase plays pivotal roles in controlling vesicular release properties and regulating hippocampal function in the mature brain. © 2015 International

  16. Monte carlo simulation of vesicular release, spatiotemporal distribution of glutamate in synaptic cleft and generation of postsynaptic currents.

    Science.gov (United States)

    Glavinovíc, M I

    1999-02-01

    The release of vesicular glutamate, spatiotemporal changes in glutamate concentration in the synaptic cleft and the subsequent generation of fast excitatory postsynaptic currents at a hippocampal synapse were modeled using the Monte Carlo method. It is assumed that glutamate is released from a spherical vesicle through a cylindrical fusion pore into the synaptic cleft and that S-alpha-amino-3-hydroxy -5-methyl-4-isoxazolepropionic acid (AMPA) receptors are uniformly distributed postsynaptically. The time course of change in vesicular concentration can be described by a single exponential, but a slow tail is also observed though only following the release of most of the glutamate. The time constant of decay increases with vesicular size and a lower diffusion constant, and is independent of the initial concentration, becoming markedly shorter for wider fusion pores. The cleft concentration at the fusion pore mouth is not negligible compared to vesicular concentration, especially for wider fusion pores. Lateral equilibration of glutamate is rapid, and within approximately 50 micros all AMPA receptors on average see the same concentration of glutamate. Nevertheless the single-channel current and the number of channels estimated from mean-variance plots are unreliable and different when estimated from rise- and decay-current segments. Greater saturation of AMPA receptor channels provides higher but not more accurate estimates. Two factors contribute to the variability of postsynaptic currents and render the mean-variance nonstationary analysis unreliable, even when all receptors see on average the same glutamate concentration. Firstly, the variability of the instantaneous cleft concentration of glutamate, unlike the mean concentration, first rapidly decreases before slowly increasing; the variability is greater for fewer molecules in the cleft and is spatially nonuniform. Secondly, the efficacy with which glutamate produces a response changes with time. Understanding

  17. Characterization of pH-sensitive molecular switches that trigger the structural transition of vesicular stomatitis virus glycoprotein from the postfusion state toward the prefusion state.

    Science.gov (United States)

    Ferlin, Anna; Raux, Hélène; Baquero, Eduard; Lepault, Jean; Gaudin, Yves

    2014-11-01

    Vesicular stomatitis virus (VSV; the prototype rhabdovirus) fusion is triggered at low pH and mediated by glycoprotein G, which undergoes a low-pH-induced structural transition. A unique feature of rhabdovirus G is that its conformational change is reversible. This allows G to recover its native prefusion state at the viral surface after its transport through the acidic Golgi compartments. The crystal structures of G pre- and postfusion states have been elucidated, leading to the identification of several acidic amino acid residues, clustered in the postfusion trimer, as potential pH-sensitive switches controlling the transition back toward the prefusion state. We mutated these residues and produced a panel of single and double mutants whose fusion properties, conformational change characteristics, and ability to pseudotype a virus lacking the glycoprotein gene were assayed. Some of these mutations were also introduced in the genome of recombinant viruses which were further characterized. We show that D268, located in the segment consisting of residues 264 to 273, which refolds into postfusion helix F during G structural transition, is the major pH sensor while D274, D395, and D393 have additional contributions. Furthermore, a single passage of recombinant virus bearing the mutation D268L (which was demonstrated to stabilize the G postfusion state) resulted in a pseudorevertant with a compensatory second mutation, L271P. This revealed that the propensity of the segment of residues 264 to 273 to refold into helix F has to be finely tuned since either an increase (mutation D268L alone) or a decrease (mutation L271P alone) of this propensity is detrimental to the virus. Vesicular stomatitis virus enters cells via endocytosis. Endosome acidification induces a structural transition of its unique glycoprotein (G), which mediates fusion between viral and endosomal membranes. G conformational change is reversible upon increases in pH. This allows G to recover its native

  18. Prejunctional inhibition of norepinephrine release caused by acetylcholine in the human saphenous vein

    International Nuclear Information System (INIS)

    Rorie, D.K.; Rusch, N.J.; Shepherd, J.T.; Vanhoutte, P.M.; Tyce, G.M.

    1981-01-01

    We performed experiments to determine whether or not acetylcholine exerts a prejunctional inhibitory effect on adrenergic neurotransmission in the human blood vessel wall. Rings of human greater saphenous veins were prepared 2 to 15 hours after death and mounted for isometric tension recording in organ chambers filled with Krebs-Ringer solution. Acetylcholine depressed contractile responses to electric activation of the sympathetic nerve endings significantly more than those to exogenous norepinephrine; the relaxations caused by the cholinergic transmitter were antagonized by atropine. Helical strips were incubated with [/sub 3/H]norepinephrine and mounted for superfusion. Electric stimulation augmented the fractional release of labeled norepinephrine. Acetylcholine caused a depression of the evoked /sub 3/H release which was antagonized by atropine but not by hexamethonium. These experiments demonstrate that, as in animal cutaneous veins, there are prejunctional inhibitory muscarinic receptors on the adrenergic nerve endings in the human saphenous vein. By contrast, the human vein also contains postjunctional inhibitory muscarinic receptors

  19. Covalent attachment of antagonists to the a7 nicotinic acetylcholine receptor: synthesis and reactivity of substituted maleimides

    DEFF Research Database (Denmark)

    Ambrus, Joseph I; Halliday, Jill I; Kanizaj, Nicholas

    2012-01-01

    The 3-methylmaleimide congeners of the natural product methyllycaconitine (MLA) and an analogue covalently attach to functional cysteine mutants of the a7 nicotinic acetylcholine receptor (nAChR).......The 3-methylmaleimide congeners of the natural product methyllycaconitine (MLA) and an analogue covalently attach to functional cysteine mutants of the a7 nicotinic acetylcholine receptor (nAChR)....

  20. Involvement of spinal serotonin receptors in the regulation of intraspinal acetylcholine release.

    Science.gov (United States)

    Kommalage, Mahinda; Höglund, A Urban

    2005-02-21

    Stimulation of spinal serotonin (5-HT) receptors results in analgesia and release of acetylcholine. We investigated the involvement of 5-HT1, 5-HT2, and 5-HT3 receptor subtypes in the regulation of spinal acetylcholine release. A spinal microdialysis probe was placed dorsally at about the C5 level in anaesthetized rats. The selective serotonin reuptake inhibitor citalopram was found to increase acetylcholine release when infused via the microdialysis probe. Several doses of the 5-HT receptor agonists 8-hydroxy-2-(di-n-propylamino)tetraline (8-OH-DPAT, 5-HT1A), 1,4-dihydro-3-(1,2,3,6-tetrahydro-4-pyridinyl)-5H-pyrrolo[3,2-b]pyridin-5-one dihydrochloride (CP93129, 5-HT1B), alpha-methyl-5-hydroxytryptamine maleate (m5-HT, 5-HT2), 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI, 5-HT2C), and 1-(m-chlorophenyl)-biguanide (5-HT3) were subsequently infused via the microdialysis probe. Only 8-OH-DPAT, CP93129, and m5-HT increased acetylcholine release dose dependently. The 5-HT1A receptor selective antagonist (S)-N-tert-butyl-3-(4-(2-methoxyphenyl)piperazine-1-yl)-2-phenylpropanamide hydrochloride and the 5-HT2A receptor selective antagonist ketanserin tartrate inhibited the 8-OH-DPAT and the m5-HT induced acetylcholine release. The results suggest that 5-HT1A and the 5-HT2A receptors are involved in the regulation of acetylcholine release in the spinal cord.

  1. Pharmacological and biochemical characterization of the D-1 dopamine receptor mediating acetylcholine release in rabbit retina

    International Nuclear Information System (INIS)

    Hensler, J.G.; Cotterell, D.J.; Dubocovich, M.L.

    1987-01-01

    Superfusion with dopamine (0.1 microM-10 mM) evokes calcium-dependent [ 3 H]acetylcholine release from rabbit retina labeled in vitro with [ 3 H]choline. This effect is antagonized by the D-1 dopamine receptor antagonist SCH 23390. Activation or blockade of D-2 dopamine, alpha-2 or beta receptors did not stimulate or attenuate the release of [ 3 H]acetylcholine from rabbit retina. Dopamine receptor agonists evoke the release of [ 3 H]acetylcholine with the following order of potency: apomorphine ≤ SKF(R)82526 3 H]acetylcholine: SCH 23390 (IC50 = 1 nM) 3 H]acetylcholine release is characteristic of the D-1 dopamine receptor. These potencies were correlated with the potencies of dopamine receptor agonists and antagonists at the D-1 dopamine receptor in rabbit retina as labeled by [ 3 H]SCH 23390, or as determined by adenylate cyclase activity. [ 3 H]SCH 23390 binding in rabbit retinal membranes was stable, saturable and reversible. Scatchard analysis of [ 3 H]SCH 23390 saturation data revealed a single high affinity binding site (Kd = 0.175 +/- 0.002 nM) with a maximum binding of 482 +/- 12 fmol/mg of protein. The potencies of dopamine receptor agonists to stimulate [ 3 H]acetylcholine release were correlated with their potencies to stimulate adenylate cyclase (r = 0.784, P less than .05, n = 7) and with their affinities at [ 3 H]SCH 23390 binding sites (r = 0.755, P < .05, n = 8)

  2. Comparative study of muscarinic acetylcholine receptors of human and rat cortical glial cells

    International Nuclear Information System (INIS)

    Demushkin, V.P.; Burbaeva, G.S.; Dzhaliashvili, T.A.; Plyashkevich, Y.G.

    1985-01-01

    The aim of the present investigation was a comparative studyof muscarinic acetylcholine receptors in human and rat glial cells. ( 3 H)Quinuclidinyl-benzylate (( 3 H)-QB), atropine, platiphylline, decamethonium, carbamylcholine, tubocurarine, and nicotine were used. The glial cell fraction was obtained from the cerebral cortex of rats weighing 130-140 g and from the frontal pole of the postmortem brain from men aged 60-70 years. The use of the method of radioimmune binding of ( 3 H)-QB with human and rat glial cell membranes demonstrated the presence of a muscarinic acetylcholine receptor in the glial cells

  3. Orthosteric and allosteric potentiation of heteromeric neuronal nicotinic acetylcholine receptors.

    Science.gov (United States)

    Wang, Jingyi; Lindstrom, Jon

    2018-06-01

    Heteromeric nicotinic ACh receptors (nAChRs) were thought to have two orthodox agonist-binding sites at two α/β subunit interfaces. Highly selective ligands are hard to develop by targeting orthodox agonist sites because of high sequence similarity of this binding pocket among different subunits. Recently, unorthodox ACh-binding sites have been discovered at some α/α and β/α subunit interfaces, such as α4/α4, α5/α4 and β3/α4. Targeting unorthodox sites may yield subtype-selective ligands, such as those for (α4β2) 2 α5, (α4β2) 2 β3 and (α6β2) 2 β3 nAChRs. The unorthodox sites have unique pharmacology. Agonist binding at one unorthodox site is not sufficient to activate nAChRs, but it increases activation from the orthodox sites. NS9283, a selective agonist for the unorthodox α4/α4 site, was initially thought to be a positive allosteric modulator (PAM). NS9283 activates nAChRs with three engineered α4/α4 sites. PAMs, on the other hand, act at allosteric sites where ACh cannot bind. Known PAM sites include the ACh-homologous non-canonical site (e.g. morantel at β/α), the C-terminus (e.g. Br-PBTC and 17β-estradiol), a transmembrane domain (e.g. LY2087101) or extracellular and transmembrane domain interfaces (e.g. NS206). Some of these PAMs, such as Br-PBTC and 17β-estradiol, require only one subunit to potentiate activation of nAChRs. In this review, we will discuss differences between activation from orthosteric and allosteric sites, their selective ligands and clinical implications. These studies have advanced understanding of the structure, assembly and pharmacology of heteromeric neuronal nAChRs. This article is part of a themed section on Nicotinic Acetylcholine Receptors. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.11/issuetoc. © 2017 The British Pharmacological Society.

  4. Central nervous system promotes thermotolerance via FoxO/DAF-16 activation through octopamine and acetylcholine signaling in Caenorhabditis elegans.

    Science.gov (United States)

    Furuhashi, Tsubasa; Sakamoto, Kazuichi

    2016-03-25

    The autonomic nervous system (ANS) responds to many kinds of stressors to maintain homeostasis. Although the ANS is believed to regulate stress tolerance, the exact mechanism underlying this is not well understood. To understand this, we focused on longevity genes, which have functions such as lifespan extension and promotion of stress tolerance. To understand the relationship between ANS and longevity genes, we analyzed stress tolerance of Caenorhabditis elegans treated with octopamine, which has an affinity to noradrenaline in insects, and acetylcholine. Octopamine and acetylcholine did not show resistance against H2O2, but the neurotransmitters promoted thermotolerance via DAF-16. However, chronic treatment with octopamine and acetylcholine did not extend the lifespan, although DAF-16 plays an important role in longevity. In conclusion, our results show that octopamine and acetylcholine activate DAF-16 in response to stress, but chronic induction of octopamine and acetylcholine is not beneficial for increasing longevity. Copyright © 2016 Elsevier Inc. All rights reserved.

  5. Cre-expressing neurons in visual cortex of Ntsr1-Cre GN220 mice are corticothalamic and are depolarized by acetylcholine.

    Science.gov (United States)

    Sundberg, Sofie Charlotte; Lindström, Sarah Helen; Sanchez, Gonzalo Manuel; Granseth, Björn

    2018-01-01

    The Ntsr1-Cre GN220 mouse expresses Cre-recombinase in corticothalamic (CT) neurons in neocortical layer 6. It is not known if the other major types of pyramidal neurons in this layer also express this enzyme. By electrophysiological recordings in slices and histological analysis of the uptake of retrogradely transported beads we show that Cre-positive neurons are CT and not corticocortical or corticoclaustral types. Furthermore, we show that Ntsr1-Cre-positive cells are immuno-positive for the nuclear transcription factor Forkhead box protein P2 (FoxP2). We conclude that Cre-expression is limited to a specific type of pyramidal neuron: CT. However, it appears as not all CT neurons are Cre-expressing; there are indications that the penetrance of the gene is about 90%. We demonstrate the utility of assigning a specific identity to individual neurons by determining that the CT neurons are potently modulated by acetylcholine acting on both nicotinic and muscarinic acetylcholine receptors. These results corroborate the suggested function of these neurons in regulating the gain of thalamocortical transfer of sensory information depending on attentional demand and state of arousal. © 2017 Wiley Periodicals, Inc.

  6. Historia natural del virus de la estomatitis vesicular en zonas enzoóticas de Antioquia

    Directory of Open Access Journals (Sweden)

    John Arboleda

    2003-01-01

    Full Text Available

    La Estomatitis Vesicular (EV es una enfermedad producida
    por el virus de la Estomatitis Vesicular, serotipos New Jersey (VSV-NJ e Indiana (VSV-IN, afecta bovinos y equinos, porcinos y causa infección natural en humanos, principalmente granjeros, ordeñadores y personal de laboratorio.
    Se caracteriza por producir vesículas en las membranas mucosas
    de la boca (epitelio de la lengua y el paladar, bandas coronarias,
    pezones y tejidos blandos de los cascos; hay pérdida de peso y decrecimiento en la producción de leche. Está clasificada en la Lista A de la Organización Internacional de Epizootias, debido a su gran poder de difusión, a las graves consecuencias socioeconómicas y a las restricciones comerciales. Además, clínicamente la EV es indistinguible de la Fiebre Aftosa (FA (1.
    La enfermedad se presenta por ciclos estacionales; la mayoría
    de ellos ocurre en las épocas de transición de los períodos de lluvias a los de verano y viceversa (2. Estudios serológicos realizados en áreas endémicas han demostrado que VSV-NJ y VSV-IN infectan en forma natural una amplia variedad de animales silvestres, los cuales están posiblemente implicados en la ecozootiología de la EV, bien como hospederos portadores, amplificadores o reservorios. Igualmente, dos especies de artrópodos, Lutzomyia shannoni y Simulium vittatum son infectados naturalmente, replican y transmiten experimentalmente
    el VSV, convirtiéndolos en posibles vectores y/o reservorios.
    Sin embargo, en ningún animal se produce la viremia necesaria para infectar los artrópodos hematófagos. El reservorio natural nunca ha sido encontrado entre los animales domésticos y silvestres investigados (3.

    El objetivo es identificar los factores ecológicos (cobertura
    vegetal, temperatura promedio, pluviosidad y humedad relativa, los vectores artrópodos y los mamíferos reservorios asociados con el antenimiento y transmisión de la VSV en

  7. A Polyamide Inhibits Replication of Vesicular Stomatitis Virus by Targeting RNA in the Nucleocapsid

    Energy Technology Data Exchange (ETDEWEB)

    Gumpper, Ryan H.; Li, Weike; Castañeda, Carlos H.; Scuderi, M. José; Bashkin, James K.; Luo, Ming; Dutch, Rebecca Ellis

    2018-02-07

    Polyamides have been shown to bind double-stranded DNA by complementing the curvature of the minor groove and forming various hydrogen bonds with DNA. Several polyamide molecules have been found to have potent antiviral activities against papillomavirus, a double-stranded DNA virus. By analogy, we reason that polyamides may also interact with the structured RNA bound in the nucleocapsid of a negative-strand RNA virus. Vesicular stomatitis virus (VSV) was selected as a prototype virus to test this possibility since its genomic RNA encapsidated in the nucleocapsid forms a structure resembling one strand of an A-form RNA duplex. One polyamide molecule, UMSL1011, was found to inhibit infection of VSV. To confirm that the polyamide targeted the nucleocapsid, a nucleocapsid-like particle (NLP) was incubated with UMSL1011. The encapsidated RNA in the polyamide-treated NLP was protected from thermo-release and digestion by RNase A. UMSL1011 also inhibits viral RNA synthesis in the intracellular activity assay for the viral RNA-dependent RNA polymerase. The crystal structure revealed that UMSL1011 binds the structured RNA in the nucleocapsid. The conclusion of our studies is that the RNA in the nucleocapsid is a viable antiviral target of polyamides. Since the RNA structure in the nucleocapsid is similar in all negative-strand RNA viruses, polyamides may be optimized to target the specific RNA genome of a negative-strand RNA virus, such as respiratory syncytial virus and Ebola virus.

    IMPORTANCENegative-strand RNA viruses (NSVs) include several life-threatening pathogens, such as rabies virus, respiratory syncytial virus, and Ebola virus. There are no effective antiviral drugs against these viruses. Polyamides offer an exceptional opportunity because they may be optimized to target each NSV. Our studies on vesicular stomatitis virus, an NSV, demonstrated that a polyamide molecule could specifically target the viral RNA in the nucleocapsid and inhibit

  8. Functional Characterization of a Novel Class of Morantel-Sensitive Acetylcholine Receptors in Nematodes.

    Directory of Open Access Journals (Sweden)

    Elise Courtot

    2015-12-01

    Full Text Available Acetylcholine receptors are pentameric ligand-gated channels involved in excitatory neuro-transmission in both vertebrates and invertebrates. In nematodes, they represent major targets for cholinergic agonist or antagonist anthelmintic drugs. Despite the large diversity of acetylcholine-receptor subunit genes present in nematodes, only a few receptor subtypes have been characterized so far. Interestingly, parasitic nematodes affecting human or animal health possess two closely related members of this gene family, acr-26 and acr-27 that are essentially absent in free-living or plant parasitic species. Using the pathogenic parasitic nematode of ruminants, Haemonchus contortus, as a model, we found that Hco-ACR-26 and Hco-ACR-27 are co-expressed in body muscle cells. We demonstrated that co-expression of Hco-ACR-26 and Hco-ACR-27 in Xenopus laevis oocytes led to the functional expression of an acetylcholine-receptor highly sensitive to the anthelmintics morantel and pyrantel. Importantly we also reported that ACR-26 and ACR-27, from the distantly related parasitic nematode of horses, Parascaris equorum, also formed a functional acetylcholine-receptor highly sensitive to these two drugs. In Caenorhabditis elegans, a free-living model nematode, we demonstrated that heterologous expression of the H. contortus and P. equorum receptors drastically increased its sensitivity to morantel and pyrantel, mirroring the pharmacological properties observed in Xenopus oocytes. Our results are the first to describe significant molecular determinants of a novel class of nematode body wall muscle AChR.

  9. Design, synthesis and biological evaluation of Erythrina alkaloid analogues as neuronal nicotinic acetylcholine receptor antagonists

    DEFF Research Database (Denmark)

    Crestey, François; Jensen, Anders A.; Borch, Morten

    2013-01-01

    The synthesis of a new series of Erythrina alkaloid analogues and their pharmacological characterization at various nicotine acetylcholine receptor (nAChR) subtypes are described. The compounds were designed to be simplified analogues of aromatic erythrinanes with the aim of obtaining subtype...

  10. Tying up Nicotine: New Selective Competitive Antagonist of the Neuronal Nicotinic Acetylcholine Receptors

    DEFF Research Database (Denmark)

    Petersen, Ida Nymann; Crestey, François; Jensen, Anders A

    2015-01-01

    Conformational restriction of the pyrrolidine nitrogen in nicotine by the introduction of an ethylene bridge provided a potent and selective antagonist of the α4β2-subtype of the nicotinic acetylcholine receptors. Resolution by chiral SFC, pharmacological characterization of the two enantiomers...

  11. Block of nicotinic acetylcholine receptors by philanthotoxins is strongly dependent on their subunit composition

    DEFF Research Database (Denmark)

    Kachel, Hamid S; Patel, Rohit N; Franzyk, Henrik

    2016-01-01

    -fold selectivity of PhTX-12 over PhTX-343 for embryonic muscle-type nicotinic acetylcholine receptors (nAChRs) in TE671 cells. We investigated their inhibition of different neuronal nAChR subunit combinations as well as of embryonic muscle receptors expressed in Xenopus oocytes. Whole-cell currents...

  12. Functional aspects of dexamethasone upregulated nicotinic acetylcholine receptors in C2C12 myotubes

    NARCIS (Netherlands)

    Maestrone, E; Lagostena, L; Henning, RH; DenHertog, A; Nobile, M

    Three days of treatment with the glucocorticoid dexamethasone (1 nM-mu M) induced a concentration-dependent up-regulation of muscle nicotinic acetylcholine receptor (nAChR) in C2C12 mouse myotubes (EC(50)=10+/-7.3 nM), as assessed by [H-3]alpha-BuTx binding. The maximum increase in binding amounted

  13. Intersubunit bridge formation governs agonist efficacy at nicotinic acetylcholine alpha4beta2 receptors

    DEFF Research Database (Denmark)

    Rohde, Line Aagot Hede; Ahring, Philip Kiær; Jensen, Marianne Lerbech

    2012-01-01

    The a4ß2 subtype of the nicotinic acetylcholine receptor (nAChR) has been pursued as a drug target for treatment of psychiatric and neurodegenerative disorders and smoking cessation aids for decades. Still, a thorough understanding of structure-function relationships of a4ß2 agonists is lacking...

  14. Synthesis and pharmacological evaluation of DHβE analogs as neuronal nicotinic acetylcholine receptor antagonists

    DEFF Research Database (Denmark)

    Jepsen, Tue H.; Jensen, Anders A.; Lund, Mads Henrik

    2014-01-01

    Dihydro-β-erythroidine (DHβE) is a member of the Erythrina family of alkaloids and a potent competitive antagonist of the α4β2-subtype of the nicotinic acetylcholine receptors (nAChRs). Guided by an X-ray structure of DHβE in complex with an ACh binding protein, we detail the design, synthesis...

  15. alpha(7) Nicotinic acetylcholine receptor activation prevents behavioral and molecular changes induced by repeated phencyclidine treatment

    DEFF Research Database (Denmark)

    Thomsen, Morten Skøtt; Christensen, Ditte Z; Hansen, Henrik H

    2009-01-01

    in a modified Y-maze test. Polymorphisms in the alpha(7) nicotinic acetylcholine receptor (nAChR) gene have been linked to schizophrenia. Here we demonstrate that acute administration of the selective alpha(7) nAChR partial agonist SSR180711 dose-dependently reversed the behavioral impairment induced by PCP...

  16. Cognitive improvement by activation of alpha7 nicotinic acetylcholine receptors: from animal models to human pathophysiology

    DEFF Research Database (Denmark)

    Thomsen, Morten S; Hansen, Henrik H; Timmerman, Daniel B

    2010-01-01

    Agonists and positive allosteric modulators of the alpha(7) nicotinic acetylcholine receptor (nAChR) are currently being developed for the treatment of cognitive disturbances in patients with schizophrenia or Alzheimer's disease. This review describes the neurobiological properties of the alpha n...

  17. Nicotinic Acetylcholine Receptor Expression and Susceptibility to Cholinergic Immunomodulation in Human Monocytes of Smoking Individuals

    NARCIS (Netherlands)

    van der Zanden, Esmerij P.; Hilbers, Francisca W.; Verseijden, Caroline; van den Wijngaard, Rene M.; Skynner, Mike; Lee, Kevin; Ulloa, Luis; Boeckxstaens, Guy E.; de Jonge, Wouter J.

    2012-01-01

    Objective: Smoking is generally accepted as a factor that affects the disease course in inflammatory bowel disease patients. Whether these effects can be contributed to the immunomodulatory effects of nicotine via nicotinic acetylcholine receptor (nAChR) activation is unclear. As previous data

  18. Distinct neural pathways mediate alpha7 nicotinic acetylcholine receptor-dependent activation of the forebrain

    DEFF Research Database (Denmark)

    Thomsen, Morten S; Hay-Schmidt, Anders; Hansen, Henrik H

    2010-01-01

    alpha(7) nicotinic acetylcholine receptor (nAChR) agonists are candidates for the treatment of cognitive deficits in schizophrenia. Selective alpha(7) nAChR agonists, such as SSR180711, activate neurons in the medial prefrontal cortex (mPFC) and nucleus accumbens shell (ACCshell) in rats, regions...

  19. MUSCARINIC ACETYLCHOLINE RECEPTOR-EXPRESSION IN ASTROCYTES IN THE CORTEX OF YOUNG AND AGED RATS

    NARCIS (Netherlands)

    VANDERZEE, EA; DEJONG, GI; STROSBERG, AD; LUITEN, PGM

    The present report describes the cellular and subcellular distribution pattern of immunoreactivity to M35, a monoclonal antibody raised against purified muscarinic acetylcholine receptor protein, in astrocytes in the cerebral cortex of young and aged rats. Most M35-positive astrocytes were localized

  20. Effect of basal forebrain stimulation on extracellular acetylcholine release and blood flow in the olfactory bulb.

    Science.gov (United States)

    Uchida, Sae; Kagitani, Fusako

    2017-05-12

    The olfactory bulb receives cholinergic basal forebrain input, as does the neocortex; however, the in vivo physiological functions regarding the release of extracellular acetylcholine and regulation of regional blood flow in the olfactory bulb are unclear. We used in vivo microdialysis to measure the extracellular acetylcholine levels in the olfactory bulb of urethane-anesthetized rats. Focal chemical stimulation by microinjection of L-glutamate into the horizontal limb of the diagonal band of Broca (HDB) in the basal forebrain, which is the main source of cholinergic input to the olfactory bulb, increased extracellular acetylcholine release in the ipsilateral olfactory bulb. When the regional cerebral blood flow was measured using laser speckle contrast imaging, the focal chemical stimulation of the HDB did not significantly alter the blood flow in the olfactory bulb, while increases were observed in the neocortex. Our results suggest a functional difference between the olfactory bulb and neocortex regarding cerebral blood flow regulation through the release of acetylcholine by cholinergic basal forebrain input.

  1. Mechanisms of the inhibition of endplate acetylcholine receptors by antiseptic chlorhexidine (experiments and models)

    Czech Academy of Sciences Publication Activity Database

    Shaihutdinova, A.R.; Nikolsky, E. E.; Vyskočil, František; Skorinkin, A.I.

    2009-01-01

    Roč. 380, č. 6 (2009), s. 551-560 ISSN 0028-1298 R&D Projects: GA AV ČR(CZ) IAA500110905 Institutional research plan: CEZ:AV0Z50110509 Keywords : acetylcholine * endplate currents Subject RIV: ED - Physiology Impact factor: 2.631, year: 2009

  2. Cloning and structural analysis of partial acetylcholine receptor subunit genes from the parasitic nematode Teladorsagia circumcincta

    NARCIS (Netherlands)

    Walker, J.; Hoekstra, R.; Roos, M. H.; Wiley, L. J.; Weiss, A. S.; Sangster, N. C.; Tait, A.

    2001-01-01

    Nematode nicotinic acetylcholine receptors (nAChRs) are the sites of action for the anthelmintic drug levamisole. Recent findings indicate that the molecular mechanism of levamisole resistance may involve changes in the number and/or functions of target nAChRs. Accordingly, we have used an RT-PCR

  3. Characterization of the positive and negative inotropic effects of acetylcholine in the human myocardium

    NARCIS (Netherlands)

    X.Y. Du (Xiaoyi); R.G. Schoemaker (Regien); E. Bos (Egbert); P.R. Saxena (Pramod Ranjan)

    1995-01-01

    textabstractIn the human isolated myocardium, acetylcholine (10−9 to 10−3 M) elicited a biphasic inotropic effect (a decrease in the lower and an increase in the higher concentration range) in atrial and a positive inotropic effect in ventricular trabeculae. However, under conditions of raised

  4. Layer-specific modulation of the prefrontal cortex by nicotinic acetylcholine receptors

    NARCIS (Netherlands)

    Poorthuis, R.B.; Bloem, B.; Schak, B.; Wester, J.; de Kock, C.P.J.; Mansvelder, H.D.

    2013-01-01

    Acetylcholine signaling through nicotinic receptors (nAChRs) in the prefrontal cortex (PFC) is crucial for attention. Nicotinic AChRs are expressed on glutamatergic inputs to layer V (LV) cells and on LV interneurons and LVI pyramidal neurons. Whether PFC layers are activated by nAChRs to a similar

  5. Weak toxin WTX from Naja kaouthia cobra venom interacts with both nicotinic and muscarinic acetylcholine receptors

    Czech Academy of Sciences Publication Activity Database

    Mordvintsev, D.Y.; Polyak, Y.L.; Rodionov, D.I.; Jakubík, Jan; Doležal, Vladimír; Karlsson, E.; Tsetlin, V.I.; Utkin, Y.N.

    2009-01-01

    Roč. 276, č. 18 (2009), s. 5065-5075 ISSN 1742-464X R&D Projects: GA MŠk(CZ) LC554; GA AV ČR(CZ) IAA500110703 Institutional research plan: CEZ:AV0Z50110509 Keywords : acetylcholine receptors * allosteric interaction * toxins Subject RIV: ED - Physiology Impact factor: 3.042, year: 2009

  6. Muscarinic Acetylcholine Receptors Act in Synergy to Facilitate Learning and Memory

    Science.gov (United States)

    Leaderbrand, Katherine; Chen, Helen J.; Corcoran, Kevin A.; Guedea, Anita L.; Jovasevic, Vladimir; Wess, Jurgen; Radulovic, Jelena

    2016-01-01

    Understanding how episodic memories are formed and retrieved is necessary if we are to treat disorders in which they malfunction. Muscarinic acetylcholine receptors (mAChR) in the hippocampus and cortex underlie memory formation, but there is conflicting evidence regarding their role in memory retrieval. Additionally, there is no consensus on…

  7. Apolipoprotein E4 reduces evoked hippocampal acetylcholine release in adult mice

    Czech Academy of Sciences Publication Activity Database

    Dolejší, Eva; Liraz, O.; Rudajev, Vladimír; Zimčík, Pavel; Doležal, Vladimír; Michaelson, D. M.

    2016-01-01

    Roč. 136, č. 3 (2016), s. 503-509 ISSN 0022-3042 R&D Projects: GA MŠk(CZ) LH13269 Institutional support: RVO:67985823 Keywords : acetylcholine release * Alzheimer's disease (AD) * apolipoprotein E4 (apoE4) * hippocampus Subject RIV: FH - Neurology Impact factor: 4.083, year: 2016

  8. Binding of N-methylscopolamine to the extracellular domain of muscarinic acetylcholine receptors

    Czech Academy of Sciences Publication Activity Database

    Jakubík, Jan; Randáková, Alena; Zimčík, Pavel; El-Fakahany, E. E.; Doležal, Vladimír

    2017-01-01

    Roč. 7, Jan 16 (2017), č. článku 40381. ISSN 2045-2322 R&D Projects: GA ČR(CZ) GBP304/12/G069 Institutional support: RVO:67985823 Keywords : muscarinic acetylcholine receptors * N-methylscopolamine * ligand binding * molecular dynamics Subject RIV: ED - Physiology OBOR OECD: Physiology (including cytology) Impact factor: 4.259, year: 2016

  9. Fragment growing induces conformational changes in acetylcholine-binding protein: A structural and thermodynamic analysis

    NARCIS (Netherlands)

    Edink, E.S.; Rucktooa, P.; Retra, K.; Akdemir, A.; Nahar, T.T.; Zuiderveld, O.P.; van Elk, R.; Janssen, E.; van Nierop, P.; van Muijlwijk-Koezen, J.E.; Smit, A.B.; Sixma, T.K.; Leurs, R.; de Esch, I.J.P.

    2011-01-01

    Optimization of fragment hits toward high-affinity lead compounds is a crucial aspect of fragment-based drug discovery (FBDD). In the current study, we have successfully optimized a fragment by growing into a ligand-inducible subpocket of the binding site of acetylcholine-binding protein (AChBP).

  10. Nicotinic acetylcholine receptor β2-subunits in the medial prefrontal cortex control attention

    NARCIS (Netherlands)

    Guillem, K.; Bloem, B.; Poorthuis, R.B.; Loos, M.; Smit, A.B.; Maskos, U.; Spijker, S.; Mansvelder, H.D.

    2011-01-01

    More than one-third of all people are estimated to experience mild to severe cognitive impairment as they age. Acetylcholine (ACh) levels in the brain diminish with aging, and nicotinic ACh receptor (nAChR) stimulation is known to enhance cognitive performance. The prefrontal cortex (PFC) is

  11. Functional Characterization of a Novel Class of Morantel-Sensitive Acetylcholine Receptors in Nematodes

    Science.gov (United States)

    Courtot, Elise; Charvet, Claude L.; Beech, Robin N.; Harmache, Abdallah; Wolstenholme, Adrian J.; Holden-Dye, Lindy; O’Connor, Vincent; Peineau, Nicolas; Woods, Debra J.; Neveu, Cedric

    2015-01-01

    Acetylcholine receptors are pentameric ligand–gated channels involved in excitatory neuro-transmission in both vertebrates and invertebrates. In nematodes, they represent major targets for cholinergic agonist or antagonist anthelmintic drugs. Despite the large diversity of acetylcholine-receptor subunit genes present in nematodes, only a few receptor subtypes have been characterized so far. Interestingly, parasitic nematodes affecting human or animal health possess two closely related members of this gene family, acr-26 and acr-27 that are essentially absent in free-living or plant parasitic species. Using the pathogenic parasitic nematode of ruminants, Haemonchus contortus, as a model, we found that Hco-ACR-26 and Hco-ACR-27 are co-expressed in body muscle cells. We demonstrated that co-expression of Hco-ACR-26 and Hco-ACR-27 in Xenopus laevis oocytes led to the functional expression of an acetylcholine-receptor highly sensitive to the anthelmintics morantel and pyrantel. Importantly we also reported that ACR-26 and ACR-27, from the distantly related parasitic nematode of horses, Parascaris equorum, also formed a functional acetylcholine-receptor highly sensitive to these two drugs. In Caenorhabditis elegans, a free-living model nematode, we demonstrated that heterologous expression of the H. contortus and P. equorum receptors drastically increased its sensitivity to morantel and pyrantel, mirroring the pharmacological properties observed in Xenopus oocytes. Our results are the first to describe significant molecular determinants of a novel class of nematode body wall muscle AChR. PMID:26625142

  12. Bradykinin or acetylcholine as vasodilators to test endothelial venous function in healthy subjects

    Directory of Open Access Journals (Sweden)

    Eneida R. Rabelo

    2008-01-01

    Full Text Available INTRODUCTION: The evaluation of endothelial function has been performed in the arterial bed, but recently evaluation within the venous system has also been explored. Endothelial function studies employ different drugs that act as endothelium-dependent vasodilatory response inductors. OBJECTIVES: The aim of this study is to compare the endothelium-dependent venous vasodilator response mediated by either acetylcholine or bradykinin in healthy volunteers. METHODS AND RESULTS: Changes in vein diameter after phenylephrine-induced venoconstriction were measured to compare venodilation induced by acetylcholine or bradykinin (linear variable differential transformer dorsal hand vein technique. We studied 23 healthy volunteers; 31% were male, and the subject had a mean age of 33 ± 8 years and a mean body mass index of 23 ± 2 kg/m². The maximum endothelium-dependent venodilation was similar for both drugs (p = 0.13, as well as the mean responses for each dose of both drugs (r = 0.96. The maximum responses to acetylcholine and bradykinin also had good agreement. CONCLUSION: There were no differences between acetylcholine and bradykinin as venodilators in this endothelial venous function investigation.

  13. GABAergic Neurons of the Rat Dorsal Hippocampus Express Muscarinic Acetylcholine Receptors

    NARCIS (Netherlands)

    van der Zee, E.A.; Luiten, P.G.M.

    1993-01-01

    The expression of muscarinic acetylcholine receptors (mAChRs) in glutamic acid decarboxylase (GAD)-positive cells in the different strata of CA1, CA3, and the dentate gyrus (DG) of the dorsal hippocampus is examined by way of quantitative immunofluorescent double labeling employing M35, the

  14. Effects of vesicular-arbuscular mycorrhizae on survival and growth of perennial grasses in lignite overburden in Texas

    Energy Technology Data Exchange (ETDEWEB)

    Call, C.A.; Davies, F.T.

    1988-12-01

    Seedlings of sideoats grama (Bouteloua curtipendula), Indiangrass (Sorghastrum nutans), and kleingrass (Panicum coloratum) were inoculated with vesicular-arbuscular mycorrhizal (VAM) fungi (Glomus fasciculatum and Gigaspora margarita) in a containerized system and transplanted into lignite overburden in the Post Oak Savannah region of Texas, U.S.A. After three growing seasons without cultural inputs, plants inoculated with VAM fungi had greater survival percentages, basal diameters, and above-ground biomass than noninoculated plants. Inoculated plants had higher levels of nitrogen and phosphorus in above-ground biomass than noninoculated plants. Root colonization percentages of inoculated plants remained fairly stable while noninoculated plants showed low levels of colonization over the 3-year study period. Vesicular-arbuscular mycorrhizae enhanced the survival and growth of the 3 grass species by making effective use of limited resources in the lignite overburden. 31 refs., 3 tabs.

  15. Extensive In Vitro Hyphal Growth of Vesicular-Arbuscular Mycorrhizal Fungi in the Presence of CO(2) and Flavonols.

    Science.gov (United States)

    Bécard, G; Douds, D D; Pfeffer, P E

    1992-03-01

    Various flavonoids were tested for their ability to stimulate in vitro growth of germinated spores of vesicular-arbuscular mycorrhizal fungi. Experiments were performed in the presence of 2% CO(2), previously demonstrated to be required for growth of Gigaspora margarita (G. Bécard and Y. Piché, Appl. Environ. Microbiol. 55:2320-2325, 1989). Only the flavonols stimulated fungal growth. The flavones, flavanones, and isoflavones tested were generally inhibitory. Quercetin (10 muM) prolonged hyphal growth from germinated spores of G. margarita from 10 to 42 days. An average of more than 500 mm of hyphal growth and 13 auxiliary cells per spore were obtained. Quercetin also stimulated the growth of Glomus etunicatum. The glycosides of quercetin, rutin, and quercitrin were not stimulatory. The axenic growth of G. margarita achieved here under rigorously defined conditions is the most ever reported for a vesicular-arbuscular mycorrhizal fungus.

  16. Effects of acute chlorpyrifos exposure on in vivo acetylcholine accumulation in rat striatum

    International Nuclear Information System (INIS)

    Karanth, Subramanya; Liu, Jing; Mirajkar, Nikita; Pope, Carey

    2006-01-01

    This study examined the acute effects of chlorpyrifos (CPF) on cholinesterase inhibition and acetylcholine levels in the striatum of freely moving rats using in vivo microdialysis. Adult, male Sprague-Dawley rats were treated with vehicle (peanut oil, 2 ml/kg) or CPF (84, 156 or 279 mg/kg, sc) and functional signs of toxicity, body weight and motor activity recorded. Microdialysis was conducted at 1, 4 and 7 days after CPF exposure for measurement of acetylcholine levels in striatum. Rats were then sacrificed and the contralateral striatum and diaphragm were collected for biochemical measurements. Few overt signs of cholinergic toxicity were noted in any rats. Body weight gain was significantly affected in the high-dose (279 mg/kg) group only, while motor activity (nocturnal rearing) was significantly reduced in all CPF-treated groups at one day (84 mg/kg) or from 1-4 days (156 and 279 mg/kg) after dosing. Cholinesterase activities in both diaphragm and striatum were markedly inhibited (50-92%) in a time-dependent manner, but there were relatively minimal dose-related changes. In contrast, time- and dose-dependent changes in striatal acetylcholine levels were noted, with significantly higher levels noted in the high-dose group compared to other groups. Maximal increases in striatal acetylcholine levels were observed at 4-7 days after dosing (84 mg/kg, 7-9-fold; 156 mg/kg, 10-13-fold; 279 mg/kg, 35-57-fold). Substantially higher acetylcholine levels were noted when an exogenous cholinesterase inhibitor was included in the perfusion buffer, but CPF treatment-related differences were substantially lower in magnitude under those conditions. The results suggest that marked differences in acetylcholine accumulation can occur with dosages of CPF eliciting relatively similar degrees of cholinesterase inhibition. Furthermore, the minimal expression of classic signs of cholinergic toxicity in the presence of extensive brain acetylcholine accumulation suggests that some

  17. Tumor Necrosis Factor-Mediated Survival of CD169+ Cells Promotes Immune Activation during Vesicular Stomatitis Virus Infection

    DEFF Research Database (Denmark)

    Shinde, Prashant V; Xu, Haifeng C; Maney, Sathish Kumar

    2018-01-01

    Innate immune activation is essential to mount an effective antiviral response and to prime adaptive immunity. Although a crucial role of CD169(+) cells during vesicular stomatitis virus (VSV) infections is increasingly recognized, factors regulating CD169(+) cells during viral infections remain...... stomatitis virus infection, phagocytes produce tumor necrosis factor (TNF) which signals via TNFR1 and promote "enforced virus replication" in CD169(+) macrophages. Consequently, lack of TNF or TNFR1 resulted in defective immune activation and VSV clearance....

  18. Linhagens de Lentinula edodes inibem fungos fitopatogênicos e o vírus da estomatite vesicular, sorotipo Alagoas

    OpenAIRE

    Sasaki, Selma H.; Linhares, Rosa E.C.; Nozawa, Carlos M.; Montalván, Ricardo; Paccola-Meirelles, Luzia D.

    2001-01-01

    Four Lentinula edodes strains (Le10, 46, K2, Assai) were assessed for their antagonistic effect on four filamentous fungus species of agricultural importance (Helminthosporium euphorbiae, Helminthosporium sp, Fusarium solani and Phomopsis sojae) and on Alagoas serotype of Vesicular Stomatitis Virus (VSA). The L. edodes strains studied had variable effects on the filamentous fungi and on VSA. The K2 and Le10 strains were antagonistic on the fungi assessed and the 46 and K2 strains were efficie...

  19. Vesicular-arbuscular mycorrhizal status of plant species in the peat swamp forest of Setia Alam Jaya, Sebangau, Central Kalimantan

    Directory of Open Access Journals (Sweden)

    Suciatmih Suciatmih

    2003-06-01

    Full Text Available In order to describe the vesicular-arbuscular mycorrhizal (VAM status of plants growing on peat soil, a study was carried out inthe peat swamp forest of Setia Alam Jaya in Sebangau, Central Kalimantan. Out of 146 plant root samples belonging to 48 plantspecies from 25 families examined, all plants colonized by VAM fungi namely 14 (29.2% high level, 32 (66.7% medium level, and 2(4.1% low level respectively.

  20. Vesicular-arbuscular mycorrhizal status of plant species in the peat swamp forest of Setia Alam Jaya, Sebangau, Central Kalimantan

    OpenAIRE

    Suciatmih Suciatmih

    2003-01-01

    In order to describe the vesicular-arbuscular mycorrhizal (VAM) status of plants growing on peat soil, a study was carried out inthe peat swamp forest of Setia Alam Jaya in Sebangau, Central Kalimantan. Out of 146 plant root samples belonging to 48 plantspecies from 25 families examined, all plants colonized by VAM fungi namely 14 (29.2%) high level, 32 (66.7%) medium level, and 2(4.1%) low level respectively.

  1. Topical Non-Iontophoretic Application of Acetylcholine and Nitroglycerin via a Translucent Patch: A New Means for Assessing Microvascular Reactivity

    Science.gov (United States)

    Schonberger, Robert B.; Worden, William S.; Shahmohammadi, Kaveh; Menn, Kirsten; Silverman, Tyler J.; Stout, Robert G.; Shelley, Kirk H.; Silverman, David G.

    2007-01-01

    Objective: Assessments of endothelial cell function with acetylcholine have typically used systemic, regional intra-arterial, or iontophoretic delivery of drug. Each of these techniques induces systemic and/or local changes that compromise their safety or effectiveness. Using translucent drug preparations applied under laser Doppler flowmetry (LDF) probes, we tested whether local vasodilation can be induced with non-iontophoretic transdermal delivery of acetylcholine and how such dilation would compare to the dilation achieved with topical nitroglycerin in healthy volunteers. Methods: Ten subjects without known vascular disease were recruited for LDF monitoring at sites of drug application for this preliminary investigation. Topical acetylcholine chloride, nitroglycerin, and placebo were applied via translucent patches to the forehead directly below LDF probes. Results: LDF readings increased by 406 percent (245 percent to 566 percent) and 36 percent (26 percent to 46 percent), respectively, at the acetylcholine and placebo sites (p = .005 by Wilcoxon Signed Rank Test (WSRT) for acetylcholine vs. placebo); and they increased by 365 percent (179 percent to 550 percent) at the nitroglycerin site (p = .005 by WSRT for nitroglycerin vs. placebo; p = .6 vs. acetylcholine). Conclusion: Transdermal delivery of acetylcholine can induce significant local vasodilatory responses comparable to those achieved with nitroglycerin without requiring iontophoresis. The means of transdermal delivery and monitoring described herein may constitute a new minimally invasive way to interrogate the microvasculature and thereby assess the microcirculatory changes induced by various disorders and therapeutic interventions. PMID:17876370

  2. Concomitant release of ventral tegmental acetylcholine and accumbal dopamine by ghrelin in rats.

    Directory of Open Access Journals (Sweden)

    Elisabet Jerlhag

    Full Text Available Ghrelin, an orexigenic peptide, regulates energy balance specifically via hypothalamic circuits. Growing evidence suggest that ghrelin increases the incentive value of motivated behaviours via activation of the cholinergic-dopaminergic reward link. It encompasses the cholinergic afferent projection from the laterodorsal tegmental area (LDTg to the dopaminergic cells of the ventral tegmental area (VTA and the mesolimbic dopamine system projecting from the VTA to nucleus accumbens (N.Acc.. Ghrelin receptors (GHS-R1A are expressed in these reward nodes and ghrelin administration into the LDTg increases accumbal dopamine, an effect involving nicotinic acetylcholine receptors in the VTA. The present series of experiments were undertaken directly to test this hypothesis. Here we show that ghrelin, administered peripherally or locally into the LDTg concomitantly increases ventral tegmental acetylcholine as well as accumbal dopamine release. A GHS-R1A antagonist blocks this synchronous neurotransmitter release induced by peripheral ghrelin. In addition, local perfusion of the unselective nicotinic antagonist mecamylamine into the VTA blocks the ability of ghrelin (administered into the LDTg to increase N.Acc.-dopamine, but not VTA-acetylcholine. Collectively our data indicate that ghrelin activates the LDTg causing a release of acetylcholine in the VTA, which in turn activates local nicotinic acetylcholine receptors causing a release of accumbal dopamine. Given that a dysfunction in the cholinergic-dopaminergic reward system is involved in addictive behaviours, including compulsive overeating and alcohol use disorder, and that hyperghrelinemia is associated with such addictive behaviours, ghrelin-responsive circuits may serve as a novel pharmacological target for treatment of alcohol use disorder as well as binge eating.

  3. Recent Duplication and Functional Divergence in Parasitic Nematode Levamisole-Sensitive Acetylcholine Receptors.

    Directory of Open Access Journals (Sweden)

    Thomas B Duguet

    2016-07-01

    Full Text Available Helminth parasites rely on fast-synaptic transmission in their neuromusculature to experience the outside world and respond to it. Acetylcholine plays a pivotal role in this and its receptors are targeted by a wide variety of both natural and synthetic compounds used in human health and for the control of parasitic disease. The model, Caenorhabditis elegans is characterized by a large number of acetylcholine receptor subunit genes, a feature shared across the nematodes. This dynamic family is characterized by both gene duplication and loss between species. The pentameric levamisole-sensitive acetylcholine receptor has been characterized from C. elegans, comprised of five different subunits. More recently, cognate receptors have been reconstituted from multiple parasitic nematodes that are found to vary in subunit composition. In order to understand the implications of receptor composition change and the origins of potentially novel drug targets, we investigated a specific example of subunit duplication based on analysis of genome data for 25 species from the 50 helminth genome initiative. We found multiple independent duplications of the unc-29, acetylcholine receptor subunit, where codon substitution rate analysis identified positive, directional selection acting on amino acid positions associated with subunit assembly. Characterization of four gene copies from a model parasitic nematode, Haemonchus contortus, demonstrated that each copy has acquired unique functional characteristics based on phenotype rescue of transgenic C. elegans and electrophysiology of receptors reconstituted in Xenopus oocytes. We found evidence that a specific incompatibility has evolved for two subunits co-expressed in muscle. We demonstrated that functional divergence of acetylcholine receptors, driven by directional selection, can occur more rapidly than previously thought and may be mediated by alteration of receptor assembly. This phenomenon is common among the

  4. Transmission of vesicular stomatitis New Jersey virus to cattle by the biting midge Culicoides sonorensis (Diptera: Ceratopogonidae).

    Science.gov (United States)

    Perez de Leon, Adalberto A; Tabachnick, Walter J

    2006-03-01

    Laboratory-reared Culicoides sonorensis Wirth & Jones were infected with vesicular stomatitis virus serotype New Jersey (family Rhabdoviridae, genus Vesiculovirus, VSNJV) through intrathoracic inoculation. After 10-d incubation at 25 degrees C, these insects were allowed to blood feed on four steers. Two other steers were exposed to VSNJV through intralingual inoculation with 10(8) tissue culture infective dose50 VSNJV. All six steers became seropositive for VSNJV. The results demonstrate the ability of C. sonorensis to transmit VSNJV to livestock. Only the animals intralingually inoculated with VSNJV showed clinical signs in the form of vesicles at the site of inoculation. Uninfected C. sonorensis allowed to feed on the exposed animals did not become infected with VSNJV. Animals infected by C. sonorensis showed a slower antibody response compared with intralingually inoculated animals. This is probably because of different amounts of virus received via insect transmission and syringe inoculation. A significant difference was found in the serum acute-phase protein alpha-1-acid glycoprotein in animals that received VSNJV through C. sonorensis transmission. These animals had previously been exposed to insect attack in the field compared with intralingually inoculated animals and C. sonorensis-infected animals that had been protected from insect attack. The failure to observe clinical signs of vesicular stomatitis through transmission of VSNJV by C. sonorensis may explain widespread subclinical infections during vesicular stomatitis epidemics.

  5. Recombinant vesicular stomatitis virus vaccine vectors expressing filovirus glycoproteins lack neurovirulence in nonhuman primates.

    Directory of Open Access Journals (Sweden)

    Chad E Mire

    Full Text Available The filoviruses, Marburg virus and Ebola virus, cause severe hemorrhagic fever with high mortality in humans and nonhuman primates. Among the most promising filovirus vaccines under development is a system based on recombinant vesicular stomatitis virus (rVSV that expresses an individual filovirus glycoprotein (GP in place of the VSV glycoprotein (G. The main concern with all replication-competent vaccines, including the rVSV filovirus GP vectors, is their safety. To address this concern, we performed a neurovirulence study using 21 cynomolgus macaques where the vaccines were administered intrathalamically. Seven animals received a rVSV vector expressing the Zaire ebolavirus (ZEBOV GP; seven animals received a rVSV vector expressing the Lake Victoria marburgvirus (MARV GP; three animals received rVSV-wild type (wt vector, and four animals received vehicle control. Two of three animals given rVSV-wt showed severe neurological symptoms whereas animals receiving vehicle control, rVSV-ZEBOV-GP, or rVSV-MARV-GP did not develop these symptoms. Histological analysis revealed major lesions in neural tissues of all three rVSV-wt animals; however, no significant lesions were observed in any animals from the filovirus vaccine or vehicle control groups. These data strongly suggest that rVSV filovirus GP vaccine vectors lack the neurovirulence properties associated with the rVSV-wt parent vector and support their further development as a vaccine platform for human use.

  6. Perturbed cholesterol and vesicular trafficking associated with dengue blocking in Wolbachia-infected Aedes aegypti cells.

    Science.gov (United States)

    Geoghegan, Vincent; Stainton, Kirsty; Rainey, Stephanie M; Ant, Thomas H; Dowle, Adam A; Larson, Tony; Hester, Svenja; Charles, Philip D; Thomas, Benjamin; Sinkins, Steven P

    2017-09-13

    Wolbachia are intracellular maternally inherited bacteria that can spread through insect populations and block virus transmission by mosquitoes, providing an important approach to dengue control. To better understand the mechanisms of virus inhibition, we here perform proteomic quantification of the effects of Wolbachia in Aedes aegypti mosquito cells and midgut. Perturbations are observed in vesicular trafficking, lipid metabolism and in the endoplasmic reticulum that could impact viral entry and replication. Wolbachia-infected cells display a differential cholesterol profile, including elevated levels of esterified cholesterol, that is consistent with perturbed intracellular cholesterol trafficking. Cyclodextrins have been shown to reverse lipid accumulation defects in cells with disrupted cholesterol homeostasis. Treatment of Wolbachia-infected Ae. aegypti cells with 2-hydroxypropyl-β-cyclodextrin restores dengue replication in Wolbachia-carrying cells, suggesting dengue is inhibited in Wolbachia-infected cells by localised cholesterol accumulation. These results demonstrate parallels between the cellular Wolbachia viral inhibition phenotype and lipid storage genetic disorders. Wolbachia infection of mosquitoes can block dengue virus infection and is tested in field trials, but the mechanism of action is unclear. Using proteomics, Geoghegan et al. here identify effects of Wolbachia on cholesterol homeostasis and dengue virus replication in Aedes aegypti.

  7. Quantification of Lyssavirus-Neutralizing Antibodies Using Vesicular Stomatitis Virus Pseudotype Particles

    Directory of Open Access Journals (Sweden)

    Sarah Moeschler

    2016-09-01

    Full Text Available Rabies is a highly fatal zoonotic disease which is primarily caused by rabies virus (RABV although other members of the genus Lyssavirus can cause rabies as well. As yet, 14 serologically and genetically diverse lyssaviruses have been identified, mostly in bats. To assess the quality of rabies vaccines and immunoglobulin preparations, virus neutralization tests with live RABV are performed in accordance with enhanced biosafety standards. In the present work, a novel neutralization test is presented which takes advantage of a modified vesicular stomatitis virus (VSV from which the glycoprotein G gene has been deleted and replaced by reporter genes. This single-cycle virus was trans-complemented with RABV envelope glycoprotein. Neutralization of this pseudotype virus with RABV reference serum or immune sera from vaccinated mice showed a strong correlation with the rapid fluorescent focus inhibition test (RFFIT. Importantly, pseudotype viruses containing the envelope glycoproteins of other lyssaviruses were neutralized by reference serum to a significantly lesser extent or were not neutralized at all. Taken together, a pseudotype virus system has been successfully developed which allows the safe, fast, and sensitive detection of neutralizing antibodies directed against different lyssaviruses.

  8. Foot & Mouth Disease & Ulcerative/Vesicular Rule-outs: Challenges Encountered in Recent Outbreaks

    Energy Technology Data Exchange (ETDEWEB)

    Hullinger, P

    2008-01-28

    development and subsequent rupturing of vesicles at the coronary band and in the oral cavity. Vesicles and ulcerations can also occur on the mammary gland. Recovery in adult animals usually occurs in 8-15 days. Clinical signs for most serotypes are less dramatic in sheep and goats. Swine can develop very severe coronary band lesions and high mortality in piglets has been observed. One of the challenges of diagnosing FMD is that it may be clinically similar to several other vesicular or ulcerative diseases. FMD is clinically indistinguishable from Vesicular stomatitis, Swine vesicular disease and Vesicular exanthema of swine. It may also resemble Bovine viral diarrhea, Mucosal disease, Infectious bovine rhinotracheitis, Bluetongue, Bovine papular stomatitis, Bovine mammillitis and Rinderpest.

  9. Quantification of Lyssavirus-Neutralizing Antibodies Using Vesicular Stomatitis Virus Pseudotype Particles.

    Science.gov (United States)

    Moeschler, Sarah; Locher, Samira; Conzelmann, Karl-Klaus; Krämer, Beate; Zimmer, Gert

    2016-09-16

    Rabies is a highly fatal zoonotic disease which is primarily caused by rabies virus (RABV) although other members of the genus Lyssavirus can cause rabies as well. As yet, 14 serologically and genetically diverse lyssaviruses have been identified, mostly in bats. To assess the quality of rabies vaccines and immunoglobulin preparations, virus neutralization tests with live RABV are performed in accordance with enhanced biosafety standards. In the present work, a novel neutralization test is presented which takes advantage of a modified vesicular stomatitis virus (VSV) from which the glycoprotein G gene has been deleted and replaced by reporter genes. This single-cycle virus was trans-complemented with RABV envelope glycoprotein. Neutralization of this pseudotype virus with RABV reference serum or immune sera from vaccinated mice showed a strong correlation with the rapid fluorescent focus inhibition test (RFFIT). Importantly, pseudotype viruses containing the envelope glycoproteins of other lyssaviruses were neutralized by reference serum to a significantly lesser extent or were not neutralized at all. Taken together, a pseudotype virus system has been successfully developed which allows the safe, fast, and sensitive detection of neutralizing antibodies directed against different lyssaviruses.

  10. Initial vesicular-arbuscular mycorrhizal development of slender wheatgrass on two amended mine spoils

    Energy Technology Data Exchange (ETDEWEB)

    Zak, J.C.; Parkinson, D. (University of Calgary, Calgary, AB (Canada). Dept. of Biology)

    1982-01-01

    The initial vesicular-arbuscular (VA) mycorrhizal development of slender wheatgrass on extracted oil-sands and subalpine coal-mine spoils, amended with either fertilizer, peat, or liquid sewage sludge, was examined. Plants were sampled at 2,6 and 10 weeks after plant emergence and the level of infection was expressed as length of mycorrhizal root per plant and length of root which contained arbuscules, vesicles, or only hyphae. Mycorrhizal infection of slender wheatgrass on the oil sands was limited to plants on the peat-amended spoil. Infection of plants on the peat-amended oil-sands spoil was detected by 2 weeks. Plants on the subalpine spoil were infected at 2 weeks only on the peat-amended spoil. While slender wheatgrass on the control and fertilizer-amended spoil developed mycorrhizae by 6 weeks, infection was not observed in plants on the sewage-amended spoil until 10 weeks. At 10 weeks, there were no significant differences in lengths of mycorrhizal root per plant among the amendments. Increased P levels in the fertilizer- and sewage-amended subalpine spoil did not suppress VA mycorrhizal development. 43 refs., 6 tabs.

  11. Vesicular-arbuscular mycorrhizae of Easter lily in the northwestern United States.

    Science.gov (United States)

    Ames, R N; Linderman, R G

    1977-12-01

    The vesicular-arbuscular (VA) mycorrhizal fungi of commercially grown Easter lily (Lilium longiflorum Thunb.) were studied. Soil and root samples were collected monthly from March through September 1975 from five fields in the coastal area of southern Oregon and northern California. Soil seivings were inoculated onto clover, onion, and lily to cause infections resulting in the production of many new mycorrhizal spores facilitating identification. Four VA mycorrhizal species were found: Acaulospora trappei, A. elegans, Glomus monosporus, and G. fasciculatus. All four VA species infected Easter lily, clover, and onion. Acaulospora trappei and G. fasciculatus were the most commonly isolated species from all five fields. Mycorrhizal infections in roots of field-grown lilies were sparse and presumably young in March and gradually increased in size and number until September when bulbs were harvested. Over 75% of each root system became infected with mycorrhizae in fields with all four fungal species, and those levels were reached by July. In fields with only two mycorrhizal species, usually 50% or less of each root system was infected, even by the end of the growing season.

  12. Interactions of Vesicular-Arbuscular Mycorrhizal Fungi, Phosphorus, and Heterodera glycines on Soybean.

    Science.gov (United States)

    Tylka, G L; Hussey, R S; Roncadori, R W

    1991-01-01

    Effects of vesicular-arbuscular mycorrhizal (VAM) fungi and soil phosphorus (P) fertility on parasitism of soybean cultivars Bragg and Wright by soybean cyst nematode (SCN) were investigated in field micropiot and greenhouse experiments. VAM fungi increased height of both cultivars and yield of Wright in microplot studies in 1986 and 1987. Conversely, yield of mycorrhizal and nonmycorrhizal plants of both cultivars was suppressed by SCN. Soil population densities of SCN were unaffected by VAM fungi in 1986 but were greater in microplots infested with VAM fungi than in control microplots in 1987. Growth of Wright soybean was stimulated by VAM fungi and suppressed by SCN in greenhouse experiments. The effect of VAM fungi on SCN varied with time. Numbers of SCN in roots and soil were decreased by VAM fungi by as much as 73% at the highest SCN inoculum level through 49 days after planting. Later, however, SCN numbers were usually comparable on mycorrhizal and nonmycorrhizal plants. Soil P fertility generally had no effect on SCN. Results of a split-root experiment indicated that VAM fungal suppression of SCN was not systemic.

  13. Occurrence of vesicular-arbuscular mycorrhizae in mixed overburden mine spoils of Texas

    Energy Technology Data Exchange (ETDEWEB)

    Mott, J.B.; Zuberer, D.A.

    1987-07-01

    Presently in east Texas, lignite surface mines are reclaimed and revegetated using mixed overburden materials which are equivalent to or better in physical-chemical properties than the poor topsoils removed during mining. Little information is available regarding the biological characteristics of levelled mixed overburden and the re-establishment of endomycorrhizal associations on revegetated mixed overburden sites. Therefore, the authors investigated the occurrence of infection of coastal bermudagrass (Cynodon dactylon), planted vegetatively on reclamation sites (1-10 years post-mining), with vesicular-arbuscular mycorrhizal (VAM) fungi. Numbers of spores were also monitored. For comparison, infection of coastal bermudagrass and spore numbers were determined for an unmined old field succession on soil typical of the region. VAM infection, measured as a percentage of root length infected or as a percentage of root segments exhibiting infection, returned to pre-mining levels by 3-7 years after disturbance. Intensity of infection was not altered by disturbance, age of reclaimed site, or season. Significantly greater numbers of spores (ca. 10-fold) were observed in the unmined soil and no differences were found between numbers of spores from variously aged mine spoil sites. 35 refs., 3 tabs.

  14. Interaction of Vesicular-Arbuscular Mycorrhizae and Cultivars of Alfalfa Susceptible and Resistant to Meloidogyne hapla.

    Science.gov (United States)

    Grandison, G S; Cooper, K M

    1986-04-01

    The interaction between vesicular-arbuscular mycorrhizal (VAM) fungi and the root-knot nematode (Meloidogyne hapla) was investigated using both nematode-susceptible (Grasslands Wairau) and nematode-resistant (Nevada Synthetic XX) cultivars of alfalfa (Medicago sativa) at four levels of applied phosphate. Mycorrhizal inoculation improved plant growth and reduced nematode numbers and adult development in roots in dually infected cultures of the susceptible cultivar. The tolerance of plants to nematode infection and development when preinfected with mycorrhizal fungi was no greater than when they were inoculated with nematodes and mycorrhizal fungi simultaneously. Growth of plants of the resistant cultivar was unaffected by nematode inoculation but was improved by mycorrhizal inoculation. Numbers of nematode juveniles were lower in the roots of the resistant than of the susceptible cultivar and were further reduced by mycorrhizal inoculation, although no adult nematodes developed in any resistant cultivar treatment. Inoculation of alfalfa with VAM fungi increased the tolerance and resistance of a cultivar susceptible to M. hapla and improved the resistance of a resistant cultivar.

  15. Production of vesicular-arbuscular mycorrhizal fungus inoculum in aeroponic culture.

    Science.gov (United States)

    Hung, L L; Sylvia, D M

    1988-02-01

    Bahia grass (Paspalum notatum) and industrial sweet potato (Ipomoea batatas) colonized by Glomus deserticola, G. etunicatum, and G. intraradices were grown in aeroponic cultures. After 12 to 14 weeks, all roots were colonized by the inoculated vesicular-arbuscular mycorrhizal fungi. Abundant vesicles and arbuscules formed in the roots, and profuse sporulation was detected intra-and extraradically. Within each fungal species, industrial sweet potato contained significantly more roots and spores per plant than bahia grass did, although the percent root colonization was similar for both hosts. Mean percent root colonization and sporulation per centimeter of colonized root generally increased with time, although with some treatments colonization declined by week 14. Spore production ranged from 4 spores per cm of colonized root for G. etunicatum to 51 spores per cm for G. intraradices. Infectivity trials with root inocula resulted in a mean of 38, 45, and 28% of bahia grass roots colonized by G. deserticola, G. etunicatum, and G. intraradices, respectively. The germination rate of G. etunicatum spores produced in soil was significantly higher than that produced in aeroponic cultures (64% versus 46%) after a 2-week incubation at 28 degrees C. However, infectivity studies comparing G. etunicatum spores from soil and aeroponic culture indicated no biological differences between the spore sources. Aeroponically produced G. deserticola and G. etunicatum inocula retained their infectivity after cold storage (4 degrees C) in either sterile water or moist vermiculite for at least 4 and 9 months, respectively.

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

    Science.gov (United States)

    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.

  17. Ammonia Assimilation in Zea mays L. Infected with a Vesicular-Arbuscular Mycorrhizal Fungus Glomus fasciculatum.

    Science.gov (United States)

    Cliquet, J. B.; Stewart, G. R.

    1993-03-01

    To investigate nitrogen assimilation and translocation in Zea mays L. colonized by the vesicular-arbuscular mycorrhizal (VAM) fungus Glomus fasciculatum (Thax. sensu Gerd.), we measured key enzyme activities, 15N incorporation into free amino acids, and 15N translocation from roots to shoots. Glutamine synthetase and nitrate reductase activities were increased in both roots and shoots compared with control plants, and glutamate dehydrogenase activity increased in roots only. In the presence of [15N]ammonium, glutamine amide was the most heavily labeled product. More label was incorporated into amino acids in VAM plants. The kinetics of 15N labeling and effects of methionine sulfoximine on distribution of 15N-labeled products were entirely consistent with the operation of the glutamate synthase cycle. No evidence was found for ammonium assimilation via glutamate dehydrogenase. 15N translocation from roots to shoots through the xylem was higher in VAM plants compared with control plants. These results establish that, in maize, VAM fungi increase ammonium assimilation, glutamine production, and xylem nitrogen translocation. Unlike some ectomycorrhizal fungi, VAM fungi do not appear to alter the pathway of ammonium assimilation in roots of their hosts.

  18. Seasonality of vesicular-arbuscular mycorrhizae in sedges in a semi-arid tropical grassland

    Science.gov (United States)

    Muthukumar, T.; Udaiyan, K.

    2002-10-01

    Vesicular-arbuscular mycorrhizal (VAM) colonization and spore numbers in the rhizosphere of Cyperus iria L. and C. rotundus L., growing in a semi-arid tropical grassland, was studied during the 1993 and 1994 monsoons. In addition, climatic and chemical properties of the soils were determined in order to investigate their influence on mycorrhizal variables. VAM fungal association in the sedges was confirmed by plant- and root-trap culture techniques. The soil nutrients exhibited seasonal variations, but were highly variable between years. Intercellular hyphae and vesicles with occasional intraradical spores characterized mycorrhizal association in sedges. Dark septate fungi also colonized roots of sedges. Temporal variations in mycorrhizal colonization and spore numbers occurred, indicating seasonality. However, the patterns of mycorrhizal colonization and spore numbers were different during both the years. The VAM fungal structures observed were intercellular hyphae and vesicles. Changes in the proportion of root length with VAM structures, total colonization levels and spore numbers were related to climatic and edaphic factors. However, the intensity of influence of climatic and soil factors on VAM tended to vary with sedge species.

  19. Arbuscules of vesicular-arbuscular mycorrhizal fungi inhabit an acidic compartment within plant roots.

    Science.gov (United States)

    Guttenberger, M

    2000-08-01

    The most widespread type of mycorrhiza is the so-called vesicular-arbuscular mycorrhiza. In this endomycorrhiza, fungal hyphae penetrate plant cell walls in the root cortex. There they form densely branched arbuscules. Fungus and plant plasma membrane are separated by a common interfacial apoplast. The pH of the compartment between the symbionts is of pivotal importance for nutrient transfer. Histochemical experiments were conducted to check for an acidic nature of the interface in the model system Glomus versiforme (Karst.) Berch-Allium porrum L. Two chemically different acidotropic dyes (neutral red and LysoSensor Green DND-189) stained the arbuscules intensely. The staining of arbuscules could be eliminated by addition of the protonophore carbonylcyanide m-chlorophenylhydrazone (CCCP) or treatments leading to membrane rupture. Therefore, the staining of the arbuscules was based on the ion-trap mechanism, which indicates acidic, membrane-bound compartments. Microscopic examination of stained arbuscules at high optical resolution revealed a peripheral accumulation of the dye. Since plasmolysis rapidly destained the arbuscules, it is concluded that the dyes accumulate in the arbuscular interface, indicating the highly acidic nature of this compartment. The findings are discussed with respect to their relevance for the nutrient transfer in mycorrhizas. In addition, evidence for a discontinuity in the arbuscular interface between the stem and the branches of the arbuscule is given.

  20. Interaction of vesicular-arbuscular mycorrhizal fungi with erosion in an oxisol.

    Science.gov (United States)

    Habte, M; Fox, R L; Aziz, T; El-Swaify, S A

    1988-04-01

    The development of vesicular-arbuscular mycorrhizal (VAM) symbiosis was monitored in Leucaena leucocephala grown in an Oxisol subjected to incremental simulated erosion. The density of VAM infective propagules in the soil diminished as the level of simulated erosion (removal of surface soil) was increased from 0 to 50 cm. The level of infection on L. leucocephala roots observed at harvest was not significantly influenced by simulated erosion unless removal of surface soil exceeded 25 cm. Inoculation of this soil and the uneroded soil with Glomus aggregatum enhanced the early onset of infection but did not significantly influence the level of infection observed at the time of harvest. Simulated erosion in excess of 7.5 cm of surface soil removal significantly delayed the development of VAM effectiveness monitored in terms of the P status of L. leucocephala subleaflets and also curtailed the level of maximum effectiveness observed. Decreases in VAM effectiveness were significantly correlated with decreases in soil chemical constituents. However, VAM effectiveness in a soil subjected to 30 cm of surface soil removal was not restored to a significant extent unless the soil was amended with P, even though other nutrients were restored to sufficiency levels. Our results demonstrate that the development of VAM effectiveness is the phase of the VAM symbiosis that is most adversely influenced by simulated erosion and that this effect appears to be caused primarily by insufficient P in the soil solution.

  1. Factors influencing survival of vesicular-arbuscular mycorrhiza propagules during topsoil storage

    Energy Technology Data Exchange (ETDEWEB)

    Miller, R.M.; Carnes, B.A.; Moorman, T.B.

    1985-01-01

    The survival dynamics of vesicular-arbuscular mycorrhizal fungi were determined, (using a bioassay procedure) for soils stored from 0.5 to 6.0 years in topsoil stockpiles associated with a coal surface-mine in the western United States. Propagule mortality could best be related to in situ soil moisture potential using a piecewise regression model (R/sup 2/ = 0.57; P less than or equal to 0.001) with the breaking point occurring at -2 MPa. The addition of length of storage time was found to contribute significantly to the accuracy of the model (R/sup 2/ = 0.70; P less than or equal to 0.001). In addition, the piece-wise nature of the data suggested two separate populations of VAM fungi - those propagules found in soils with moisture potentials less than -2 MPa and those occurring in soils with moisture potentials greater than -2 MPa. Soil moisture and length of storage time had differing effects on each of these populations. When water potential was less than -2 MPa, moisture was an important predictor of inoculum (P < 0.001), while length of storage had little predictive capability (P = 0.17). However, when water potentials were greater than -2 MPa, the predictive importance of soil moisture (P = 0.86) and length of storage (P = 0.04) were reversed. The significance of these findings to topsoil replacement and subsequent plant community development are discussed. 28 references, 2 figures, 2 tables.

  2. Anterograde or Retrograde Transsynaptic Circuit Tracing in Vertebrates with Vesicular Stomatitis Virus Vectors.

    Science.gov (United States)

    Beier, Kevin T; Mundell, Nathan A; Pan, Y Albert; Cepko, Constance L

    2016-01-04

    Viruses have been used as transsynaptic tracers, allowing one to map the inputs and outputs of neuronal populations, due to their ability to replicate in neurons and transmit in vivo only across synaptically connected cells. To date, their use has been largely restricted to mammals. In order to explore the use of such viruses in an expanded host range, we tested the transsynaptic tracing ability of recombinant vesicular stomatitis virus (rVSV) vectors in a variety of organisms. Successful infection and gene expression were achieved in a wide range of organisms, including vertebrate and invertebrate model organisms. Moreover, rVSV enabled transsynaptic tracing of neural circuitry in predictable directions dictated by the viral envelope glycoprotein (G), derived from either VSV or rabies virus (RABV). Anterograde and retrograde labeling, from initial infection and/or viral replication and transmission, was observed in Old and New World monkeys, seahorses, jellyfish, zebrafish, chickens, and mice. These vectors are widely applicable for gene delivery, afferent tract tracing, and/or directional connectivity mapping. Here, we detail the use of these vectors and provide protocols for propagating virus, changing the surface glycoprotein, and infecting multiple organisms using several injection strategies. Copyright © 2016 John Wiley & Sons, Inc.

  3. Vesicular stomatitis virus enables gene transfer and transsynaptic tracing in a wide range of organisms.

    Science.gov (United States)

    Mundell, Nathan A; Beier, Kevin T; Pan, Y Albert; Lapan, Sylvain W; Göz Aytürk, Didem; Berezovskii, Vladimir K; Wark, Abigail R; Drokhlyansky, Eugene; Bielecki, Jan; Born, Richard T; Schier, Alexander F; Cepko, Constance L

    2015-08-01

    Current limitations in technology have prevented an extensive analysis of the connections among neurons, particularly within nonmammalian organisms. We developed a transsynaptic viral tracer originally for use in mice, and then tested its utility in a broader range of organisms. By engineering the vesicular stomatitis virus (VSV) to encode a fluorophore and either the rabies virus glycoprotein (RABV-G) or its own glycoprotein (VSV-G), we created viruses that can transsynaptically label neuronal circuits in either the retrograde or anterograde direction, respectively. The vectors were investigated for their utility as polysynaptic tracers of chicken and zebrafish visual pathways. They showed patterns of connectivity consistent with previously characterized visual system connections, and revealed several potentially novel connections. Further, these vectors were shown to infect neurons in several other vertebrates, including Old and New World monkeys, seahorses, axolotls, and Xenopus. They were also shown to infect two invertebrates, Drosophila melanogaster, and the box jellyfish, Tripedalia cystophora, a species previously intractable for gene transfer, although no clear evidence of transsynaptic spread was observed in these species. These vectors provide a starting point for transsynaptic tracing in most vertebrates, and are also excellent candidates for gene transfer in organisms that have been refractory to other methods. © 2015 Wiley Periodicals, Inc.

  4. Curcumin and Boswellia serrata gum resin extract inhibit chikungunya and vesicular stomatitis virus infections in vitro.

    Science.gov (United States)

    von Rhein, Christine; Weidner, Tatjana; Henß, Lisa; Martin, Judith; Weber, Christopher; Sliva, Katja; Schnierle, Barbara S

    2016-01-01

    Chikungunya virus (CHIKV) is a mosquito-transmitted alphavirus that causes chikungunya fever and has infected millions of people mainly in developing countries. The associated disease is characterized by rash, high fever, and severe arthritis that can persist for years. CHIKV has adapted to Aedes albopictus, which also inhabits temperate regions including Europe and the United States of America. CHIKV has recently caused large outbreaks in Latin America. No treatment or licensed CHIKV vaccine exists. Traditional medicines are known to have anti-viral effects; therefore, we examined whether curcumin or Boswellia serrata gum resin extract have antiviral activity against CHIKV. Both compounds blocked entry of CHIKV Env-pseudotyped lentiviral vectors and inhibited CHIKV infection in vitro. In addition, vesicular stomatitis virus vector particles and viral infections were also inhibited to the same extent, indicating a broad antiviral activity. Although the bioavailability of these compounds is rather poor, they might be used as a lead structure to develop more effective antiviral drugs or might be used topically to prevent CHIKV spread in the skin after mosquito bites. Copyright © 2015 Elsevier B.V. All rights reserved.

  5. [Antitumor effects of matrix protein of vesicular stomatic virus on EL4 lymphoma mice].

    Science.gov (United States)

    Lin, Shi-jia; Yu, Qin-mei; Meng, Wen-tong; Wen, Yan-jun; Chen, Li-juan; Niu, Ting

    2011-03-01

    To explore antitumor effects of plasmid pcDNA3. 1-MP encoding matrix protein of vesicular stomatitis virus (VSV) complexed with cationic liposome (DOTAP:CHOL) in mice with EL4 lymphoma. C57BL/6 mouse model with EL4 lymphoma was established. Sixty mice bearing EL4 lymphoma were divided randomly into five groups including Lip-MP, Lip-pVAX, Lip, ADM and NS groups, which were intravenously injected with liposome-pcDNA 3. 1-MP complex, liposome-pVAX complex, empty liposome, Adriamycin and normal saline respectively every three days. Tumor volumes and survival time were monitored. Microvessel density and tumor proliferative index in tumor tissues were determined by CD31, Ki-67 immunohistochemistry staining, meanwhile the tumor apoptosis index was measured by TUNEL method. From 6 days after treatments on, the tumor volume in Lip-MP group was much smaller than that in Lip-pVAX, Lip and NS group (P EL4 tumor cells in vivo (P EL4 lymphoma, which may be related to the induction of tumor cell apoptosis, inhibition of tumor angiogenesis, and suppression of tumor cell proliferation.

  6. Chronic nicotine modifies skeletal muscle Na,K-ATPase activity through its interaction with the nicotinic acetylcholine receptor and phospholemman.

    Directory of Open Access Journals (Sweden)

    Alexander V Chibalin

    Full Text Available Our previous finding that the muscle nicotinic acetylcholine receptor (nAChR and the Na,K-ATPase interact as a regulatory complex to modulate Na,K-ATPase activity suggested that chronic, circulating nicotine may alter this interaction, with long-term changes in the membrane potential. To test this hypothesis, we chronically exposed rats to nicotine delivered orally for 21-31 days. Chronic nicotine produced a steady membrane depolarization of ∼3 mV in the diaphragm muscle, which resulted from a net change in electrogenic transport by the Na,K-ATPase α2 and α1 isoforms. Electrogenic transport by the α2 isoform increased (+1.8 mV while the activity of the α1 isoform decreased (-4.4 mV. Protein expression of Na,K-ATPase α1 or α2 isoforms and the nAChR did not change; however, the content of α2 subunit in the plasma membrane decreased by 25%, indicating that its stimulated electrogenic transport is due to an increase in specific activity. The physical association between the nAChR, the Na,K-ATPase α1 or α2 subunits, and the regulatory subunit of the Na,K-ATPase, phospholemman (PLM, measured by co-immuno precipitation, was stable and unchanged. Chronic nicotine treatment activated PKCα/β2 and PKCδ and was accompanied by parallel increases in PLM phosphorylation at Ser(63 and Ser(68. Collectively, these results demonstrate that nicotine at chronic doses, acting through the nAChR-Na,K-ATPase complex, is able to modulate Na,K-ATPase activity in an isoform-specific manner and that the regulatory range includes both stimulation and inhibition of enzyme activity. Cholinergic modulation of Na,K-ATPase activity is achieved, in part, through activation of PKC and phosphorylation of PLM.

  7. Vesicular-arbuscular-/ecto-mycorrhiza succession in seedlings of. Eucalyptus spp. Sucessão de micorrizas vesicular-arbuscular e ectomicorrizas em mudas de Eucalyptus spp.

    Directory of Open Access Journals (Sweden)

    Vera Lúcia dos Santos

    2001-06-01

    Full Text Available The occurrence of vesicular-arbuscular mycorrhizae (AM and ectomycorrhizae (ECM in the same root system was observed when species of Eucalyptus urophylla S.T. Blake, E. citriodora Hook f., E. grandis W. Hill ex Maiden, E. cloeziana F. Muell. and E. camaldulensis Dehnh were simultaneously inoculated with Glomus etunicatum Becker & Gederman and Pisolithus tinctorius (Per. Cocker & Couch, isolate Pt 90A. The succession between the two fungi was observed. In general ectomycorrhizal colonization increased followed by a decrease in AM. Pisolithus tinctorius was favored in simultaneous inoculation with G. etunicatum, and the positive effect of the simultaneous inoculation of both fungi in the percent colonization by the AM fungus occurred up to 60 days after inoculation. After 120 days, colonization of roots by G. etunicatum decreased in the presence of P. tinctorius. When inoculated simultaneously, the proportion of AM and ECM varied with evaluation time, while the combined percentage of mycorrhizal roots approached the maximum and remained more or less constant after 60 days, suggesting that there could be competition between the fungi for limiting substrate. The maximum percent mycorrhizal colonization varied with Eucalyptus species and the highest value was observed for E. camaldulensis, followed in order by E. citriodora, E. urophylla, E. grandis and E. cloeziana.A ocorrência de micorrizas arbusculares (AM e ectomicorrizas (ECM no mesmo sistema radicular foi observada quando Eucalyptus urophylla S.T. Blake, E. citriodora Hook F., E. grandis W. Hill ex Maiden, E. cloeziana F. Muell e E. camaldulensis Dehnh foram inoculadas simultaneamente com Glomus etunicatum Becker & Gederman and Pisolithus tinctorius (Per. Cocker & Couch. A sucessão entre os dois fungos foi observada. De modo geral, o aumento da colonização ECM foi acompanhado de um decréscimo em AM. A inoculação simultânea resultou em percentagens de colonização diferenciadas das

  8. Sigma receptor ligand N,N'-di-(ortho-tolyl)guanidine inhibits release of acetylcholine in the guinea pig ileum.

    Science.gov (United States)

    Cambell, B G; Keana, J F; Weber, E

    1991-11-26

    The inhibition of stimulated contractions of the guinea pig ileum longitudinal muscle/myenteric plexus preparation by sigma receptor ligands has been previously described. In this study, the stimulated release of [3H]acetylcholine from cholinergic nerve terminals in this same preparation was monitored in the presence and absence of sigma receptor ligands. N,N'-Di-(orthotolyl)guanidine (DTG) and other compounds selective for the sigma receptor inhibited stimulated [3H]acetylcholine release. These results suggest that their inhibition of stimulated contractions in this preparation was mediated by inhibition of acetylcholine release.

  9. Hyphal N transport by a vesicular-arbuscular mycorrhizal fungus associated with cucumber grown at three nitrogen levels

    DEFF Research Database (Denmark)

    Johansen, A.; Jakobsen, I.; Jensen, E.S.

    1994-01-01

    colonization at all three levels of N supply, but this effect was strongest in plants of low N status. The results indicated that this increase was due partly to the improved inflow of N via the external hyphae. Root colonization by G. intraradices was unaffected by the amount of N supplied to the RC, while...... hyphal length increased in HC(A) compared to HC(B). Although a considerable N-15 content was detected in mycorrhizal roots adjacent to HC(B), only insignificant amounts of N-15 were found in the external hyphae in HC(B). The external hyphae depleted the soil of inorganic N in both HC(A) and HC(B), while...

  10. On the appearance of acetylcholine receptors in denervated rat diaphragm, and its dependence on nerve stump length

    International Nuclear Information System (INIS)

    Uchitel, O.; Robbins, N.

    1978-01-01

    Acetylcholine (ACh) sensitivity and extrajunctional receptor distribution of the rat diaphragm were closely monitored during the early period following denervation. Both contracture in response to 10 μg/ml of ACh and extrajunctional binding of [ 125 I]alpha-bungarotoxin ([ 125 I]α-BTX) were first detectable 30 h after cutting the phrenic nerve in the thorax. If the nerve were cut more proximally, leaving a 3.5 cm distal nerve stump, the same level of ACh contracture and [ 125 I]α-BTX binding did not appear until 40 h after operation. This 10-h delay was far longer than the 3-h delay in transmission failure reportedly dependent on stump length. The earliest detectable extrajunctional [ 125 I]α-BTX binding appeared throughout the entire muscle fiber, and was not localized to the endplate region as would be expected if degeneration in the nerve terminal induced new receptors. However, later significant increases in [ 125 I]α-BTX binding at the endplate region could have resulted from such degeneration. All these results are consistent with neurotrophic regulation of muscle ACh receptors, working via a mechanism involving axonal transport. (Auth.)

  11. Super-resolution microscopy reveals functional organization of dopamine transporters into cholesterol and neuronal activity-dependent nanodomains

    DEFF Research Database (Denmark)

    Rahbek-Clemmensen, Troels; Lycas, Matthew D.; Erlendsson, Simon

    2017-01-01

    is dynamically sequestrated into cholesterol-dependent nanodomains in the plasma membrane of presynaptic varicosities and neuronal projections of dopaminergic neurons. Stochastic optical reconstruction microscopy reveals irregular dopamine transporter nanodomains (∼70 nm mean diameter) that were highly sensitive...... to cholesterol depletion. Live photoactivated localization microscopy shows a similar dopamine transporter membrane organization in live heterologous cells. In neurons, dual-color dSTORM shows that tyrosine hydroxylase and vesicular monoamine transporter-2 are distinctively localized adjacent to...

  12. Depolarization by K+ and glutamate activates different neurotransmitter release mechanisms in GABAergic neurons: vesicular versus non-vesicular release of GABA

    DEFF Research Database (Denmark)

    Belhage, B; Hansen, G H; Schousboe, A

    1993-01-01

    was also reduced by organic (verapamil) and inorganic (Co++) Ca++ channel blockers but was insensitive to the GABA transport inhibitor SKF 89976A. In contrast, the second phase was less sensitive to nocodazole and Ca++ channel antagonists but could be inhibited by SKF 89976A. The glutamate-induced [3H...

  13. Factores de riesgo en la litiasis vesicular: Estudio en pacientes colecistectomizados

    Directory of Open Access Journals (Sweden)

    Carlos A Romero Díaz

    1999-08-01

    Full Text Available Se realizó un estudio prospectivo desde diciembre de 1991 hasta noviembre de 1997, en 276 pacientes ingresados e intervenidos quirúrgicamente con el diagnóstico de litiasis vesicular. Dichos pacientes se intervinieron por nuestro grupo básico de trabajo y en su mayoría eran remitidos por médicos de la familia graduados en nuestra facultad. Se clasificaron los cálculos en pigmentarios y de colesterol según sus características macroscópicas al corte, y se realizó una encuesta con los posibles factores de riesgos. Predominaron los cálculos de colesterol (76,1 % sobre los pigmentarios, y existió predominio del sexo femenino sobre el masculino en relación de 4:1. El diagnóstico de litiasis vesicular se efectuó con mayor frecuencia en la cuarta y quinta décadas de la vida, mientras los pigmentarios se observaron con mayor frecuencia en edades más avanzadas. La obesidad (39,5 %, la diabetes mellitus (19,5 % y la paridad (31,8 % constituyeron los principales factores de riesgo, por lo que se deberá tomar en cuenta los antecedentes de litiasis en familiares de primera línea y la ingestión de anticonceptivos orales. En los estados hemolíticos predominaron los cálculos pigmentarios y se demostró la relación de las hiperlipoproteinemias de las fracciones IIb y IV con la colelitiasis. Sólo el 34,5 % de los bilicultivos realizados tuvieron crecimiento bacteriano, y fueron la Escherichia coli y el estreptococo los más aisladosA prospective study of 276 patients admitted and operated on with the diagnosis of cholelithiasis was conducted from December, 1991, to November, 1997. These patients were operated on by our basic working group and most of them were referred by family physicians graduated in our Faculty. Gallstones were classified into pigment gallstones and cholesterol gallstones according to their macroscopic characteristics on cutting. A survey was done with the possible risk factors. Cholesterol gallstones (76

  14. Transport Statistics - Transport - UNECE

    Science.gov (United States)

    Sustainable Energy Statistics Trade Transport Themes UNECE and the SDGs Climate Change Gender Ideas 4 Change UNECE Weekly Videos UNECE Transport Areas of Work Transport Statistics Transport Transport Statistics About us Terms of Reference Meetings and Events Meetings Working Party on Transport Statistics (WP.6

  15. Insulin replacement restores the vesicular secretory apparatus in the diabetic rat lacrimal gland

    Directory of Open Access Journals (Sweden)

    Ana Carolina Dias

    2015-06-01

    Full Text Available ABSTRACT Purpose: In the lacrimal gland (LG acinar cells, signaling regulates the release of secretory vesicles through specific Rab and SNARE exocytotic proteins. In diabetes mellitus (DM, the LGs are dysfunctional. The aim of this work was to determine if secretory apparatus changes were associated with any effects on the secretory vesicles (SV in diabetic rats as well as the expression levels of constituent Rab and members of the SNARE family, and if insulin supplementation reversed those changes. Methods: DM was induced in male Wistar rats with an intravenous dose of streptozotocin (60 mg/kg. One of the two diabetic groups was then treated every other day with insulin (1 IU. A third control group was injected with vehicle. After 10 weeks, Western blotting and RT-PCR were used to compared the Rab and SNARE secretory factor levels in the LGs. Transmission electron microscopy evaluated acinar cell SV density and integrity. Results: In the diabetes mellitus group, there were fewer and enlarged SV. The Rab 27b, Rab 3d, and syntaxin-1 protein expression declined in the rats with diabetes mellitus. Insulin treatment restored the SV density and the Rab 27b and syntaxin expression to their control protein levels, whereas the Vamp 2 mRNA expression increased above the control levels. Conclusions: Diabetes mellitus LG changes were associated with the declines in protein expression levels that were involved in supporting exocytosis and vesicular formation. They were partially reversed by insulin replacement therapy. These findings may help to improve therapeutic management of dry eye in diabetes mellitus.

  16. Enhanced immunosurveillance for animal morbilliviruses using vesicular stomatitis virus (VSV) pseudotypes.

    Science.gov (United States)

    Logan, Nicola; Dundon, William G; Diallo, Adama; Baron, Michael D; James Nyarobi, M; Cleaveland, Sarah; Keyyu, Julius; Fyumagwa, Robert; Hosie, Margaret J; Willett, Brian J

    2016-11-11

    The measurement of virus-specific neutralising antibodies represents the "gold-standard" for diagnostic serology. For animal morbilliviruses, such as peste des petits ruminants (PPRV) or rinderpest virus (RPV), live virus-based neutralisation tests require high-level biocontainment to prevent the accidental escape of the infectious agents. In this study, we describe the adaptation of a replication-defective vesicular stomatitis virus (VSVΔG) based pseudotyping system for the measurement of neutralising antibodies against animal morbilliviruses. By expressing the haemagglutinin (H) and fusion (F) proteins of PPRV on VSVΔG pseudotypes bearing a luciferase marker gene, neutralising antibody titres could be measured rapidly and with high sensitivity. Serological responses against the four distinct lineages of PPRV could be measured simultaneously and cross-neutralising responses against other morbilliviruses compared. Using this approach, we observed that titres of neutralising antibodies induced by vaccination with live attenuated PPRV were lower than those induced by wild type virus infection and the level of cross-lineage neutralisation varied between vaccinates. By comparing neutralising responses from animals infected with either PPRV or RPV, we found that responses were highest against the homologous virus, indicating that retrospective analyses of serum samples could be used to confirm the nature of the original pathogen to which an animal had been exposed. Accordingly, when screening sera from domestic livestock and wild ruminants in Tanzania, we detected evidence of cross-species infection with PPRV, canine distemper virus (CDV) and a RPV-related bovine morbillivirus, suggesting that exposure to animal morbilliviruses may be more widespread than indicated previously using existing diagnostic techniques. Copyright © 2016 The Authors. Published by Elsevier Ltd.. All rights reserved.

  17. Enhancement of absorption and hepatoprotective potential through soya-phosphatidylcholine-andrographolide vesicular system.

    Science.gov (United States)

    Jain, Pushpendra Kumar; Khurana, Navneet; Pounikar, Yogesh; Gajbhiye, Asmita; Kharya, Murli Dhar

    2013-06-01

    Andrographis paniculata is a medicinal herb used extensively for various ailments and contains therapeutically active phytoconstituent, andrographolide (AN). Although hepatoprotective activity of AN is established, but their bioavailability is restricted due to its rapid clearance. The aim of this study, therefore, was to formulate AN herbosomes (ANH) through complexation with naturally occurring soya-phosphatidylcholine (SPC), in order to enhance absorption. Prepared andrographolide-soy phosphatidylcholine (AN-SPC) complex prepared was subjected for characterisation of complex and formation of vesicular system known as ANH using rotary evaporation techniques. This complex was subjected to in vitro study using everted small intestine sac technique which showed significantly increased absorption of AN from the ANH as compared to the plain AN. The hepatoprotective potential of ANH and plain AN was evaluated using carbon tetrachloride inducing hepatotoxicity rat model and compared, in which ANH equivalent to 50 mg/kg of plain AN significantly restore serum glutamate oxalacetate transaminase (112.4 ± 9.67 for AN whereas 90.2 ± 4.23 for ANH) and serum glutamate pyruvate transaminase (109.3 ± 7.89 for AN whereas 90.6 ± 4.34 for ANH) level as compared to control group. The ANH showed significantly better absorption than plain AN and this effect of ANH was also comparable to the standard drug (Silymarin). The findings of present study reveal that ANH has better bioavailability as shown by in vitro absorption study and hence improved hepatoprotection as compared to plain AN at equivalent dose.

  18. Reovirus FAST Protein Enhances Vesicular Stomatitis Virus Oncolytic Virotherapy in Primary and Metastatic Tumor Models

    Directory of Open Access Journals (Sweden)

    Fabrice Le Boeuf

    2017-09-01

    Full Text Available The reovirus fusion-associated small transmembrane (FAST proteins are the smallest known viral fusogens (∼100–150 amino acids and efficiently induce cell-cell fusion and syncytium formation in multiple cell types. Syncytium formation enhances cell-cell virus transmission and may also induce immunogenic cell death, a form of apoptosis that stimulates immune recognition of tumor cells. These properties suggest that FAST proteins might serve to enhance oncolytic virotherapy. The oncolytic activity of recombinant VSVΔM51 (an interferon-sensitive vesicular stomatitis virus [VSV] mutant encoding the p14 FAST protein (VSV-p14 was compared with a similar construct encoding GFP (VSV-GFP in cell culture and syngeneic BALB/c tumor models. Compared with VSV-GFP, VSV-p14 exhibited increased oncolytic activity against MCF-7 and 4T1 breast cancer spheroids in culture and reduced primary 4T1 breast tumor growth in vivo. VSV-p14 prolonged survival in both primary and metastatic 4T1 breast cancer models, and in a CT26 metastatic colon cancer model. As with VSV-GFP, VSV-p14 preferentially replicated in vivo in tumors and was cleared rapidly from other sites. Furthermore, VSV-p14 increased the numbers of activated splenic CD4, CD8, natural killer (NK, and natural killer T (NKT cells, and increased the number of activated CD4 and CD8 cells in tumors. FAST proteins may therefore provide a multi-pronged approach to improving oncolytic virotherapy via syncytium formation and enhanced immune stimulation.

  19. Vesicular stomatitis virus-based vaccines protect nonhuman primates against Bundibugyo ebolavirus.

    Directory of Open Access Journals (Sweden)

    Chad E Mire

    Full Text Available Ebola virus (EBOV causes severe and often fatal hemorrhagic fever in humans and nonhuman primates (NHPs. Currently, there are no licensed vaccines or therapeutics for human use. Recombinant vesicular stomatitis virus (rVSV-based vaccine vectors, which encode an EBOV glycoprotein in place of the VSV glycoprotein, have shown 100% efficacy against homologous Sudan ebolavirus (SEBOV or Zaire ebolavirus (ZEBOV challenge in NHPs. In addition, a single injection of a blend of three rVSV vectors completely protected NHPs against challenge with SEBOV, ZEBOV, the former Côte d'Ivoire ebolavirus, and Marburg virus. However, recent studies suggest that complete protection against the newly discovered Bundibugyo ebolavirus (BEBOV using several different heterologous filovirus vaccines is more difficult and presents a new challenge. As BEBOV caused nearly 50% mortality in a recent outbreak any filovirus vaccine advanced for human use must be able to protect against this new species. Here, we evaluated several different strategies against BEBOV using rVSV-based vaccines. Groups of cynomolgus macaques were vaccinated with a single injection of a homologous BEBOV vaccine, a single injection of a blended heterologous vaccine (SEBOV/ZEBOV, or a prime-boost using heterologous SEBOV and ZEBOV vectors. Animals were challenged with BEBOV 29-36 days after initial vaccination. Macaques vaccinated with the homologous BEBOV vaccine or the prime-boost showed no overt signs of illness and survived challenge. In contrast, animals vaccinated with the heterologous blended vaccine and unvaccinated control animals developed severe clinical symptoms consistent with BEBOV infection with 2 of 3 animals in each group succumbing. These data show that complete protection against BEBOV will likely require incorporation of BEBOV glycoprotein into the vaccine or employment of a prime-boost regimen. Fortunately, our results demonstrate that heterologous rVSV-based filovirus vaccine

  20. Infection of Melanoplus sanguinipes Grasshoppers following Ingestion of Rangeland Plant Species Harboring Vesicular Stomatitis Virus▿

    Science.gov (United States)

    Drolet, Barbara S.; Stuart, Melissa A.; Derner, Justin D.

    2009-01-01

    Knowledge of the many mechanisms of vesicular stomatitis virus (VSV) transmission is critical for understanding of the epidemiology of sporadic disease outbreaks in the western United States. Migratory grasshoppers [Melanoplus sanguinipes (Fabricius)] have been implicated as reservoirs and mechanical vectors of VSV. The grasshopper-cattle-grasshopper transmission cycle is based on the assumptions that (i) virus shed from clinically infected animals would contaminate pasture plants and remain infectious on plant surfaces and (ii) grasshoppers would become infected by eating the virus-contaminated plants. Our objectives were to determine the stability of VSV on common plant species of U.S. Northern Plains rangelands and to assess the potential of these plant species as a source of virus for grasshoppers. Fourteen plant species were exposed to VSV and assayed for infectious virus over time (0 to 24 h). The frequency of viable virus recovery at 24 h postexposure was as high as 73%. The two most common plant species in Northern Plains rangelands (western wheatgrass [Pascopyrum smithii] and needle and thread [Hesperostipa comata]) were fed to groups of grasshoppers. At 3 weeks postfeeding, the grasshopper infection rate was 44 to 50%. Exposure of VSV to a commonly used grasshopper pesticide resulted in complete viral inactivation. This is the first report demonstrating the stability of VSV on rangeland plant surfaces, and it suggests that a significant window of opportunity exists for grasshoppers to ingest VSV from contaminated plants. The use of grasshopper pesticides on pastures would decrease the incidence of a virus-amplifying mechanical vector and might also decontaminate pastures, thereby decreasing the inter- and intraherd spread of VSV. PMID:19286779

  1. Infection of Melanoplus sanguinipes grasshoppers following ingestion of rangeland plant species harboring vesicular stomatitis virus.

    Science.gov (United States)

    Drolet, Barbara S; Stuart, Melissa A; Derner, Justin D

    2009-05-01

    Knowledge of the many mechanisms of vesicular stomatitis virus (VSV) transmission is critical for understanding of the epidemiology of sporadic disease outbreaks in the western United States. Migratory grasshoppers [Melanoplus sanguinipes (Fabricius)] have been implicated as reservoirs and mechanical vectors of VSV. The grasshopper-cattle-grasshopper transmission cycle is based on the assumptions that (i) virus shed from clinically infected animals would contaminate pasture plants and remain infectious on plant surfaces and (ii) grasshoppers would become infected by eating the virus-contaminated plants. Our objectives were to determine the stability of VSV on common plant species of U.S. Northern Plains rangelands and to assess the potential of these plant species as a source of virus for grasshoppers. Fourteen plant species were exposed to VSV and assayed for infectious virus over time (0 to 24 h). The frequency of viable virus recovery at 24 h postexposure was as high as 73%. The two most common plant species in Northern Plains rangelands (western wheatgrass [Pascopyrum smithii] and needle and thread [Hesperostipa comata]) were fed to groups of grasshoppers. At 3 weeks postfeeding, the grasshopper infection rate was 44 to 50%. Exposure of VSV to a commonly used grasshopper pesticide resulted in complete viral inactivation. This is the first report demonstrating the stability of VSV on rangeland plant surfaces, and it suggests that a significant window of opportunity exists for grasshoppers to ingest VSV from contaminated plants. The use of grasshopper pesticides on pastures would decrease the incidence of a virus-amplifying mechanical vector and might also decontaminate pastures, thereby decreasing the inter- and intraherd spread of VSV.

  2. Serodiagnosis of Human Cysticercosis by Using Antigens from Vesicular Fluid of Taenia crassiceps Cysticerci

    Science.gov (United States)

    Bueno, Ednéia C.; Snege, Miriam; Vaz, Adelaide J.; Leser, Paulo G.

    2001-01-01

    Neurocysticercosis (NC), caused by the presence of Taenia solium metacestodes in tissues, is a severe parasitic infection of the central nervous system with universal distribution. To determine the efficiency of enzyme-linked immunosorbent assay (ELISA) and immunoblot with antigens of T. crassiceps vesicular fluid (Tcra) compared to standard techniques (indirect immunofluorescence test [IFT] and complement fixation test [CFT]) using T. solium cysticerci (Tso) for the serodiagnosis of NC, we studied serum samples from 24 patients with NC, 30 supposedly healthy individuals, 76 blood bank donors, 45 individuals with other non-NC parasitoses, and 97 samples from individuals screened for cysticercosis serology (SC). The sensitivity observed was 100% for ELISA-Tso and ELISA-Tcra, 91.7% for the IFT, and 87.5% for the CFT. The specificity was 90% for ELISA-Tso, 96.7% for ELISA-Tcra, 50% for IFT, and 63.3% for CFT. The efficiency was highest for ELISA-Tcra, followed by ELISA-Tso, IFT, and CFT. Of the 23 samples from SC group, which were reactive to ELISA-Tso and/or ELISA-Tcra, only 3 were positive to immunblot-Tcra (specific peptides of 14- and 18-kDa) and to glycoprotein peptides purified from Tcra antigen (gp-Tcra), showing the low predictive value of ELISA for screening. None of the samples from the remaining groups showed specific reactivity in immunoblot-Tcra. These results demonstrate that ELISA-Tcra can be used as a screening method for the serodiagnosis of NC and support the need for specific tests for confirmation of the results. The immunoblot can be used as a confirmatory test both with Tcra and gp-Tcra, with the latter having an advantage in terms of visualization of the results. PMID:11687454

  3. Vesicular stomatitis virus-based ebola vaccine is well-tolerated and protects immunocompromised nonhuman primates.

    Directory of Open Access Journals (Sweden)

    Thomas W Geisbert

    2008-11-01

    Full Text Available Ebola virus (EBOV is a significant human pathogen that presents a public health concern as an emerging/re-emerging virus and as a potential biological weapon. Substantial progress has been made over the last decade in developing candidate preventive vaccines that can protect nonhuman primates against EBOV. Among these prospects, a vaccine based on recombinant vesicular stomatitis virus (VSV is particularly robust, as it can also confer protection when administered as a postexposure treatment. A concern that has been raised regarding the replication-competent VSV vectors that express EBOV glycoproteins is how these vectors would be tolerated by individuals with altered or compromised immune systems such as patients infected with HIV. This is especially important as all EBOV outbreaks to date have occurred in areas of Central and Western Africa with high HIV incidence rates in the population. In order to address this concern, we evaluated the safety of the recombinant VSV vector expressing the Zaire ebolavirus glycoprotein (VSVDeltaG/ZEBOVGP in six rhesus macaques infected with simian-human immunodeficiency virus (SHIV. All six animals showed no evidence of illness associated with the VSVDeltaG/ZEBOVGP vaccine, suggesting that this vaccine may be safe in immunocompromised populations. While one goal of the study was to evaluate the safety of the candidate vaccine platform, it was also of interest to determine if altered immune status would affect vaccine efficacy. The vaccine protected 4 of 6 SHIV-infected macaques from death following ZEBOV challenge. Evaluation of CD4+ T cells in all animals showed that the animals that succumbed to lethal ZEBOV challenge had the lowest CD4+ counts, suggesting that CD4+ T cells may play a role in mediating protection against ZEBOV.

  4. Glycoprotein cytoplasmic domain sequences required for rescue of a vesicular stomatitis virus glycoprotein mutant

    International Nuclear Information System (INIS)

    Whitt, M.A.; Chong, L.; Rose, J.K.

    1989-01-01

    The authors have used transient expression of the wild-type vesicular stomatitis virus (VSV) glycoprotein (G protein) from cloned cDNA to rescue a temperature-sensitive G protein mutant of VSV in cells at the nonpermissive temperature. Using cDNAs encoding G proteins with deletions in the normal 29-amino-acid cytoplasmic domain, they determined that the presence of either the membrane-proximal 9 amino acids or the membrane-distal 12 amino acids was sufficient for rescue of the temperature-sensitive mutant. G proteins with cytoplasmic domains derived from other cellular or viral G proteins did not rescue the mutant, nor did G proteins with one or three amino acids of the normal cytoplasmic domain. Rescue correlated directly with the ability of the G proteins to be incorporated into virus particles. This was shown by analysis of radiolabeled particles separated on sucrose gradients as well as by electron microscopy of rescued virus after immunogold labeling. Quantitation of surface expression showed that all of the mutated G proteins were expressed less efficiently on the cell surface than was wild-type G protein. However, they were able to correct for differences in rescue efficiency resulting from differences in the level of surface expression by reducing wild-type G protein expression to levels equivalent to those observed for the mutated G proteins. The results provide evidence that at least a portion of the cytoplasmic domain is required for efficient assembly of the VSV G protein into virions during virus budding

  5. The value of Tzanck smear test in diagnosis of erosive, vesicular, bullous, and pustular skin lesions.

    Science.gov (United States)

    Durdu, Murat; Baba, Mete; Seçkin, Deniz

    2008-12-01

    Tzanck smear is generally used for the diagnosis of the pemphigus group of autoimmune bullous diseases and mucocutaneous herpesvirus infections. There are only a few studies in the literature investigating its diagnostic value. We aimed to investigate Tzanck smear findings and to determine the diagnostic value of this test in moist (erosive, vesicular, bullous, and pustular) skin lesions. We also aimed to develop an algorithmic approach for the diagnosis of these types of skin lesions according to the Tzanck smear findings. Samples were stained with May-Grünwald-Giemsa and evaluated by the same dermatologist. In some patients, methylene blue and Gram staining or direct immunofluorescence examinations were additionally performed. In all of the study cases, after the evaluation of clinical and laboratory findings (including, when appropriate, potassium hydroxide examination; viral serology; bacterial and fungal cultures; histopathology; direct and indirect immunofluorescence; patch testing), the definite diagnosis was established. We also determined the sensitivity and the specificity of certain Tzanck smear findings. Tzanck smear was performed in a total of 400 patients with moist skin lesions. The sensitivities of multinucleated giant cells and acantholytic cells in herpetic infections, dyskeratotic acantholytic cells and cocci in bullous impetigo, pseudohyphae in candidiasis, acantholytic cells in pemphigus and more than 10 tadpole cells (magnification x100) in spongiotic dermatitis were 84.7%, 92%, 100%, 100%, and 81.5%, respectively. Because Tzanck smears were evaluated by the same dermatologist, no comment could be made regarding the interobserver reliability of this test and how the level of experience with this technique might affect the results. Also, the sensitivity and the specificity of Tzanck smear test findings for certain diseases could not be calculated because of an insufficient number of patients. The Tzanck smear test is an inexpensive, useful

  6. Echinococcus multilocularis vesicular fluid inhibits activation and proliferation of natural killer cells.

    Science.gov (United States)

    Bellanger, Anne-Pauline; Mougey, Valentine; Pallandre, Jean-Rene; Gbaguidi-Haore, Houssein; Godet, Yann; Millon, Laurence

    2017-08-25

    Alveolar echinococcosis is a severe chronic helminthic disease that mimics slow-growing liver cancer. The immune evasion strategy of Echinococcus multilocularis Leuckart, 1863 remains poorly understood. The aim of this study was to investigate in vitro the impact of E. multilocularis vesicular fluid (Em-VF) on peripheral blood mononuclear cells (PBMC) and on natural killer (NK) cells. PBMC and NK cells were exposed to Em-VF (1 µg/ml) during six days. The effect of Em-VF was assessed on CD69, viability and proliferation, and on and transforming growth factor β (TGF-β), interferon γ (IFN-γ), interleukin 17 (IL-17) and interleukin 10, using flow cytometry and ELISA, respectively. Exposure to Em-VF had no bearing on PBMC's viability, proliferation and expression of CD69. In contrast, higher levels of IL-17 at day three and of TGF-β at day six were observed in PBMC supernatant after exposure to Em-VF (p Wilcoxon signed-rank test). Exposure to Em-VF induced a significant decrease of CD69 expression of NK cells at day three and a significant decrease of proliferation of NK cells at day six (p Wilcoxon signed-rank test). In contrast, NK cells viability and levels of cytokines did not vary significantly over Em-VF stimulation. Exposure to Em-VF had a significant bearing on activation and proliferation of NK cells. NK cells may play an important role in the immune response of the host against E. multilocularis.

  7. Dromedary milk exosomes as mammary transcriptome nano-vehicle: Their isolation, vesicular and phospholipidomic characterizatio

    Directory of Open Access Journals (Sweden)

    Aya M. Yassin

    2016-09-01

    Full Text Available Exosomes are extracellular nanovesicles that play a role in cellular trafficking and communication. Camel milk exosomes might carry the potential of recovery of several illnesses that coins the dromedary milk. This study shows for the first time their isolation and fine characterization. The differential ultracentrifugation was used for their isolation. Their recovery from dromedary milk during different lactation periods was evaluated. The vesicular characterization and stability testing of the recovered exosome were examined by transmission electron microscopy (TEM. The proteome footprinting was resolved by gel electrophoresis prior to their specific protein biomarker analysis. The immunoblotting of their specific protein biomarker TSG101 unexpectedly revealed a truncated 35 KDa protein specific for dromedary milk exosome rather than the previously reported 43 KDa mammalian one. The reversed-phase HPLC screening of their phospholipid makeup was compared with that of cattle milk exosomes at different lactation periods. Since dromedary milk exosomes reflect their mammary transcriptome outcome, further assessment of their content of αs1casein, αs2casein β-casein κ-casein mRNAs parallel with a constitutive glyceraldehyde dehydrogenase (GAPD gene was performed using real-time PCR. The TEM scanning indicated that dromedary milk exosomes are freeze-stress unstable homogeneous with average size of 30 nm. There was no significant difference in expression level of different casein genes in mid lactation period in dromedary milk exosomes over late lactation period. The phospholipidomic survey proved that phosphatidylcholine is the major candidate of the examined phospholipids in dromedary milk exosomes. The obtained data give novel interpretation about the content of camel milk exosomes with possible insight for use as potentially-safe nano carrier.

  8. Distribution of vesicular-arbuscular mycorrhizal fungi in coal, lignite and calcite mine spoils of India

    Energy Technology Data Exchange (ETDEWEB)

    Ganesan, V.; Ragupathy, S.; Parthipan, B.; Rani, D.B.R.; Mahadevan, A.

    1991-12-31

    Vesicular-arbuscular mycorhizzal (VAM) status was assessed for coal, lignite and calcite mine spoils. The three study sites were: The Kothagudem coal field in the south central region where waste materials are piled 1 to 2 m high on the soil surface. Samples were collected from plants growing on the waste. Neyveli, on the southeastern coast, is a lignite coal mine where the spoil is piled 70 to 100 m high on the soil surface. Samples were collected from recently revegetated mine spoil and from 25 year old revegetated sites. The calcite mine at Thazhaiyuthu in the south where the spoil is piled up 2 to 3 m on the soil surface. Samples were collected from 4 to 7 year old reclaimed sites. The wastes generally supported different plant species. The level of VAM infection of plants was markedly different in each mine spoil, with the maximum infection in the coal and calcite spoils, and the least in the lignite spoil. There was more infection in the 25 year old lignite spoil than in the newly revegetated spoil. There were different VAM species in each spoil, and no one species was present in all of the samples. The authors conclude that one of the factors leading to the differences between spoils is the amount of topsoil contained in the spoil (least in the lignite spoils which are very deep). The other is age of the spoils. Unfortunately the authors concluded that the best approach is to enrich the spoils with VAM rather than salvaging and replacing topsoil

  9. Interaction of Vesicular-arbuscular Mycorrhizal Fungi and Phosphorus with Meloidogyne incognita on Tomato.

    Science.gov (United States)

    Cason, K M; Hussey, R S; Roncadori, R W

    1983-07-01

    The influence of two vesicular-arbuscular mycorrhizal fungi and phosphorus (P) nutrition on penetration, development, and reproduction by Meloidogyne incognita on Walter tomato was studied in the greenhouse. Inoculation with either Gigaspora margarita or Glomus mosseae 2 wk prior to nematode inoculation did not alter infection by M. incognita compared with nonmycorrhizal plants, regardless of soil P level (either 3 mug [low P] or 30 mug [high P] available P/g soil). At a given soil P level, nematode penetration and reproduction did not differ in mycorrhizal and nonmycorrhizal plants. However, plants grown in high P soil had greater root weights, increased nematode penetration and egg production per plant, and decreased colonization by mycorrhizal fungi, compared with plants grown in low P soil. The number of eggs per female nematode on mycorrhizal and nonmycorrhizal plants was not influenced by P treatment. Tomato plants with split root systems grown in double-compartment containers which had either low P soil in both sides or high P in one side and low P in the other, were inoculated at transplanting with G. margarita and 2 wk later one-half of the split root system of each plant was inoculated with M. incognita larvae. Although the mycoorhizal fungus increased the inorganic P content of the root to a level comparable to that in plants grown in high P soil, nematode penetration and reproduction were not altered. In a third series of experiments, the rate of nematode development was not influenced by either the presence of G. margarita or high soil P, compared with control plants grown in low P soil. These data indicate that supplemental P (30 mu/g soil) alters root-knot nematode infection of tomato more than G. mosseae and G. margarita.

  10. Current good manufacturing practice production of an oncolytic recombinant vesicular stomatitis viral vector for cancer treatment.

    Science.gov (United States)

    Ausubel, L J; Meseck, M; Derecho, I; Lopez, P; Knoblauch, C; McMahon, R; Anderson, J; Dunphy, N; Quezada, V; Khan, R; Huang, P; Dang, W; Luo, M; Hsu, D; Woo, S L C; Couture, L

    2011-04-01

    Vesicular stomatitis virus (VSV) is an oncolytic virus currently being investigated as a promising tool to treat cancer because of its ability to selectively replicate in cancer cells. To enhance the oncolytic property of the nonpathologic laboratory strain of VSV, we generated a recombinant vector [rVSV(MΔ51)-M3] expressing murine gammaherpesvirus M3, a secreted viral chemokine-binding protein that binds to a broad range of mammalian chemokines with high affinity. As previously reported, when rVSV(MΔ51)-M3 was used in an orthotopic model of hepatocellular carcinoma (HCC) in rats, it suppressed inflammatory cell migration to the virus-infected tumor site, which allowed for enhanced intratumoral virus replication leading to increased tumor necrosis and substantially prolonged survival. These encouraging results led to the development of this vector for clinical translation in patients with HCC. However, a scalable current Good Manufacturing Practice (cGMP)-compliant manufacturing process has not been described for this vector. To produce the quantities of high-titer virus required for clinical trials, a process that is amenable to GMP manufacturing and scale-up was developed. We describe here a large-scale (50-liter) vector production process capable of achieving crude titers on the order of 10(9) plaque-forming units (PFU)/ml under cGMP. This process was used to generate a master virus seed stock and a clinical lot of the clinical trial agent under cGMP with an infectious viral titer of approximately 2 × 10(10) PFU/ml (total yield, 1 × 10(13) PFU). The lot has passed all U.S. Food and Drug Administration-mandated release testing and will be used in a phase 1 clinical translational trial in patients with advanced HCC.

  11. Covalent Trapping of Methyllycaconitine at the α4-α4 Interface of the α4β2 Nicotinic Acetylcholine Receptor

    DEFF Research Database (Denmark)

    Absalom, Nathan L; Quek, Gracia; Lewis, Trevor M

    2013-01-01

    The α4β2 nicotinic acetylcholine receptors (nAChRs) are widely expressed in the brain and are implicated in a variety of physiological processes. There are two stoichiometries of the α4β2 nAChR, (α4)2(β2)3 and (α4)3(β2)2, with different sensitivities to acetylcholine (ACh), but their pharmacologi......The α4β2 nicotinic acetylcholine receptors (nAChRs) are widely expressed in the brain and are implicated in a variety of physiological processes. There are two stoichiometries of the α4β2 nAChR, (α4)2(β2)3 and (α4)3(β2)2, with different sensitivities to acetylcholine (ACh...

  12. Strains of Lentinula edodes suppress growth of phytopathogenic fungi and inhibit Alagoas serotype of vesicular stomatitis virus Linhagens de Lentinula edodes inibem fungos fitopatogênicos e o vírus da estomatite vesicular, sorotipo Alagoas

    Directory of Open Access Journals (Sweden)

    Selma H. Sasaki

    2001-03-01

    Full Text Available Four Lentinula edodes strains (Le10, 46, K2, Assai were assessed for their antagonistic effect on four filamentous fungus species of agricultural importance (Helminthosporium euphorbiae, Helminthosporium sp, Fusarium solani and Phomopsis sojae and on Alagoas serotype of Vesicular Stomatitis Virus (VSA. The L. edodes strains studied had variable effects on the filamentous fungi and on VSA. The K2 and Le10 strains were antagonistic on the fungi assessed and the 46 and K2 strains were efficient on the Vesicular Stomatitis Virus. The results widened the list of beneficial effects of L. edodes on the control and prevention of animal pathogenic virus and filamentous fungi.Quatro linhagens de Lentinula edodes (Le10, 46, K2, ASSAI foram avaliadas quanto ao seu efeito inibitório sobre quatro espécies de fungos filamentosos de importância agrícola (Helminthosporium euphorbiae, Helminthosporium sp., Fusarium solani, Phomopsis sojae e sobre o sorotipo Alagoas vírus da estomatite vesicular (VSA. Foi observado que as linhagens de L. edodes estudadas apresentaram variabilidade quanto ao seu efeito, tanto sobre os fungos filamentosos quanto sobre o vírus VSA. As linhagens K2 e Le10 apresentaram-se antagônicas sobre os fungos e as linhagens 46 e K2 foram eficientes na inibição do vírus VSA. Os resultados obtidos permitem ampliar a lista de efeitos benéficos de algumas linhagens de L. edodes no controle e prevenção de vírus patogênicos animais e de fungos filamentosos.

  13. Acetylcholine produces contraction mediated by cyclooxigenase pathway in arterial vessels in the marine fish (Isacia conceptionis

    Directory of Open Access Journals (Sweden)

    FA. Moraga

    Full Text Available Preliminary studies showed that dorsal artery contraction mediated by acetylcholine (ACh is blocked with indomethacin in intertidal fish (G. laevifrons. Our objective was to characterize the cholinergic pathway in several artery vessels of the I. conceptionis. Afferent and efferent branchial, dorsal and mesenteric arteries were dissected of 6 juvenile specimens, isometric tension studies were done using doses response curves (DRC for Ach (10–13 to 10–3 M, and cholinergic pathways were obtained by blocking with atropine or indomethacin. CRC to ACh showed a pattern of high sensitivity only in efferente branchial artery and low sensibility in all vessels. Furthermore, these contractions were blocked in the presence of atropine and indomethacin in all vessels. Our results corroborate previous results observed in intertidal species that contraction induced by acetylcholine is mediated by receptors that activate a cyclooxygenase contraction pathway.

  14. Study of the Peripheral Nerve Fibers Myelin Structure Changes during Activation of Schwann Cell Acetylcholine Receptors.

    Directory of Open Access Journals (Sweden)

    Ekaterina E Verdiyan

    Full Text Available In the present paper we consider a new type of mechanism by which neurotransmitter acetylcholine (ACh regulates the properties of peripheral nerve fibers myelin. Our data show the importance of the relationship between the changes in the number of Schwann cell (SC acetylcholine receptors (AChRs and the axon excitation (different intervals between action potentials (APs. Using Raman spectroscopy, an effect of activation of SC AChRs on the myelin membrane fluidity was investigated. It was found, that ACh stimulates an increase in lipid ordering degree of the myelin lipids, thus providing evidence for specific role of the "axon-SC" interactions at the axon excitation. It was proposed, that during the axon excitation, the SC membrane K+- depolarization and the Ca2+-influx led to phospholipase activation or exocytosis of intracellular membrane vesicles and myelin structure reorganization.

  15. Changes in acetylcholine content, release and muscarinic receptors in rat hippocampus under cold stress

    International Nuclear Information System (INIS)

    Fatranska, M.; Budai, D.; Gulya, K; Kvetnansky, R.

    1989-01-01

    The aim was to study the mechanism of the previously established decrease in acetylcholine (ACh) concentration in the rat hippocampus under cold stress. Male rats were exposed for 14 days to cold (5 degree C) or kept (controls) at room temperature (24 degree C). Acetylcholine content, release and muscarinic receptor binding were investigated in the hippocampus. Cold exposure resulted in a decrease of ACh concentration in the dorsal hippocampus. Moreover, the potassium-evoked release of ACh from hippocampal slices was increased and an increase of maximal binding capacity of [ 3 H](-) quinuclidinyl benzilate in the dorsal hippocampus of cold exposed animals was also observed. Thus the decrease of hippocampal ACh concentration under cold exposure is probably due to its increased release. On balance then, our results demonstrate that cold stress in the rat induces significant activation of the hippocampal cholinergic system

  16. Functional Characterization of CCHamide and Muscarinic Acetylcholine Receptor Signalling in Drosophila melanogaster

    DEFF Research Database (Denmark)

    Ren, Guilin Robin

    G-protein coupled receptors (GPCRs) constitute a large and ancient superfamily of membraneproteins responsible for the transduction of extracellular signals to the inside of the cells. In thisPh.D. thesis, Drosophila melanogaster (Dm) was used as a model organism to investigate a numberof topics...... is a newly discovered insect peptide hormone. The function of this novel peptide hasnot been well characterised. In this Ph.D. thesis, I identified CCHamide-2 peptides in endocrinecells of the gut and neurones of the brain of larvae and endocrine cells of the gut of adultDrosophila. Behavioural assays...... little is known about muscarinic acetylcholine receptorsignalling in insects. In this study, I found that two types of mAChRs occur in D. melanogaster, onecoupling to Gq (A-type) and the other to Gi (B-type). Both A- and B-type Dm-mAChRs can beactivated by acetylcholine (ACh), but the classical...

  17. Fluoxetine Alleviates Behavioral Depression while Decreasing Acetylcholine Release in the Nucleus Accumbens Shell

    OpenAIRE

    Chau, David T; Rada, Pedro V; Kim, Kay; Kosloff, Rebecca A; Hoebel, Bartley G

    2011-01-01

    Selective serotonin reuptake inhibitors, such as fluoxetine, have demonstrated the ability to alleviate behavioral depression in the forced swim test; however, the sites and mechanisms of their actions remain to be further elucidated. Previous studies have suggested that behavioral depression in the swim test is mediated in part by acetylcholine (ACh) stimulating the cholinergic M1 receptors in the nucleus accumbens (NAc) shell. The current study tested whether acute, local, and chronic, subc...

  18. Classical and atypical agonists activate M1 muscarinic acetylcholine receptors through common mechanisms

    Czech Academy of Sciences Publication Activity Database

    Randáková, Alena; Dolejší, Eva; Rudajev, Vladimír; Zimčík, Pavel; Doležal, Vladimír; El-Fakahany, E. E.; Jakubík, Jan

    2015-01-01

    Roč. 97, Jul 2015 (2015), s. 27-39 ISSN 1043-6618 R&D Projects: GA ČR(CZ) GA305/09/0681; GA ČR(CZ) GBP304/12/G069; GA MŠk(CZ) EE2.3.30.0025 Institutional support: RVO:67985823 Keywords : muscarinic acetylcholine receptors * atypical agonists * xanomeline * activation mechanism Subject RIV: ED - Physiology Impact factor: 4.816, year: 2015

  19. Nicotine at clinically relevant concentrations affects atrial inward rectifier potassium current sensitive to acetylcholine.

    Science.gov (United States)

    Bébarová, Markéta; Matejovič, Peter; Švecová, Olga; Kula, Roman; Šimurdová, Milena; Šimurda, Jiří

    2017-05-01

    Nicotine abuse is associated with variety of diseases including arrhythmias, most often atrial fibrillation (AF). Altered inward rectifier potassium currents including acetylcholine-sensitive current I K(Ach) are known to be related to AF pathogenesis. Since relevant data are missing, we aimed to investigate I K(Ach) changes at clinically relevant concentrations of nicotine. Experiments were performed by the whole cell patch clamp technique at 23 ± 1 °C on isolated rat atrial myocytes. Nicotine was applied at following concentrations: 4, 40 and 400 nM; ethanol at 20 mM (∼0.09%). Nicotine at 40 and 400 nM significantly activated constitutively active component of I K(Ach) with the maximum effect at 40 nM (an increase by ∼100%); similar effect was observed at -110 and -50 mV. Changes at 4 nM nicotine were negligible on average. Coapplication of 40 nM nicotine and 20 mM ethanol (which is also known to activate this current) did not show cumulative effect. In the case of acetylcholine-induced component of I K(Ach) , a dual effect of nicotine and its correlation with the current magnitude in control were apparent: the current was increased by nicotine in the cells showing small current in control and vice versa. The effect of 40 and 400 nM nicotine on acetylcholine-induced component of I K(Ach) was significantly different at -110 and -50 mV. We conclude that nicotine at clinically relevant concentrations significantly increased constitutively active component of I K(Ach) and showed a dual effect on its acetylcholine-induced component, similarly as ethanol. Synchronous application of nicotine and ethanol did not cause additive effect.

  20. Brain nicotinic acetylcholine receptors are involved in stress-induced potentiation of nicotine reward in rats.

    Science.gov (United States)

    Javadi, Parastoo; Rezayof, Ameneh; Sardari, Maryam; Ghasemzadeh, Zahra

    2017-07-01

    The aim of the present study was to examine the possible role of nicotinic acetylcholine receptors of the dorsal hippocampus (CA1 regions), the medial prefrontal cortex or the basolateral amygdala in the effect of acute or sub-chronic stress on nicotine-induced conditioned place preference. Our results indicated that subcutaneous administration of nicotine (0.2 mg/kg) induced significant conditioned place preference. Exposure to acute or sub-chronic elevated platform stress potentiated the response of an ineffective dose of nicotine. Pre-conditioning intra-CA1 (0.5-4 µg/rat) or intra-medial prefrontal cortex (0.2-0.3 µg/rat) microinjection of mecamylamine (a non-selective nicotinic acetylcholine receptor antagonist) reversed acute stress-induced potentiation of nicotine reward as measured in the conditioned place preference paradigm. By contrast, pre-conditioning intra-basolateral amygdala microinjection of mecamylamine (4 µg/rat) potentiated the effects of acute stress on nicotine reward. Our findings also showed that intra-CA1 or intra-medial prefrontal cortex, but not intra-basolateral amygdala, microinjection of mecamylamine (4 µg/rat) prevented the effect of sub-chronic stress on nicotine reward. These findings suggest that exposure to elevated platform stress potentiates the rewarding effect of nicotine which may be associated with the involvement of nicotinic acetylcholine receptors. It seems that there is a different contribution of the basolateral amygdala, the medial prefrontal cortex or the CA1 nicotinic acetylcholine receptors in stress-induced potentiation of nicotine-induced conditioned place preference.

  1. Autoradiographic visualization of extrajunctional acetylcholine receptors in whole human biceps brachii muscle

    International Nuclear Information System (INIS)

    Askmark, H.; Gillberg, P.-G.; Aquilonius, S.-M.

    1985-01-01

    The morphological distribution of acetylcholine receptors in the whole biceps brachii muscle from a patient with amyotrophic lateral sclerosis (ALS) and from a control patient was studied by in vitro autoradiography with 3 H-alpha-bungarotoxin alphaBtx). In ALS, 3 H-alpha-Btx binding occurred over the entire muscle, while in the control case the binding was restricted to the motor end-plate region. (author)

  2. Nerve terminal contributes to acetylcholine receptor organization at the dystrophic neuromuscular junction of mdx mice.

    Science.gov (United States)

    Marques, Maria Julia; Taniguti, Ana Paula Tiemi; Minatel, Elaine; Neto, Humberto Santo

    2007-02-01

    Changes in the distribution of acetylcholine receptors have been reported to occur at the neuromuscular junction of mdx mice and may be a consequence of muscle fiber regeneration rather than the absence of dystrophin. In the present study, we examined whether the nerve terminal determines the fate of acetylcholine receptor distribution in the dystrophic muscle fibers of mdx mice. The left sternomastoid muscle of young (1-month-old) and adult (6-month-old) mdx mice was injected with 60 microl lidocaine hydrochloride to induce muscle degeneration-regeneration. Some mice had their sternomastoid muscle denervated at the time of lidocaine injection. After 10 days of muscle denervation, nerve terminals and acetylcholine receptors were labeled with 4-Di-2-ASP and rhodamine-alpha-bungarotoxin, respectively, for confocal microscopy. In young mdx mice, 75% (n = 137 endplates) of the receptors were distributed in islands. The same was observed in 100% (n = 114 endplates) of the adult junctions. In denervated-regenerated fibers of young mice, the receptors were distributed as branches in 89% of the endplates (n = 90). In denervated-regenerated fibers of adult mice, the receptors were distributed in islands in 100% of the endplates (n = 100). These findings show that nerve-dependent mechanisms are also involved in the changes in receptor distribution in young dystrophic muscles. In older dystrophic muscles, other factors may play a role in receptor distribution.

  3. The anthelmintic levamisole is an allosteric modulator of human neuronal nicotinic acetylcholine receptors.

    Science.gov (United States)

    Levandoski, Mark M; Piket, Barbara; Chang, Jane

    2003-06-13

    L-[-]-2,3,5,6-Tetrahydro-6-phenylimidazo[2,1b]-thiazole hydrochloride (levamisole) is an anthelmintic that targets the nicotinic acetylcholine receptors of parasitic nematodes. We report here the effects of levamisole on human neuronal alpha 3 beta 2 and alpha 3 beta 4 nicotinic receptors, heterologously expressed in Xenopus oocytes and studied with the voltage clamp method. Applied alone, levamisole was a very weak partial agonist for the two subunit combinations. When co-applied with acetylcholine, micromolar concentrations of levamisole potentiated responses, while millimolar concentrations inhibited them; these effects were complex functions of both acetylcholine and levamisole concentrations. The differences in the levamisole effects on the two receptor combinations suggest that the effects are mediated by the beta subunit. Several combinations of agonist and anthelmintic gave the dual potentiation/inhibition behavior, suggesting that the modulatory effects are general. Levamisole inhibition showed macroscopic characteristics of open channel block. Several results led us to conclude that levamisole potentiation occurs through noncompetitive binding to the receptor. We propose pseudo-site binding for noncompetitive potentiation by levamisole.

  4. Monkey Adrenal Chromaffin Cells Express α6β4* Nicotinic Acetylcholine Receptors

    Science.gov (United States)

    Scadden, Mick´l; Carmona-Hidalgo, Beatriz; McIntosh, J. Michael; Albillos, Almudena

    2014-01-01

    Nicotinic acetylcholine receptors (nAChRs) that contain α6 and β4 subunits have been demonstrated functionally in human adrenal chromaffin cells, rat dorsal root ganglion neurons, and on noradrenergic terminals in the hippocampus of adolescent mice. In human adrenal chromaffin cells, α6β4* nAChRs (the asterisk denotes the possible presence of additional subunits) are the predominant subtype whereas in rodents, the predominant nAChR is the α3β4* subtype. Here we present molecular and pharmacological evidence that chromaffin cells from monkey (Macaca mulatta) also express α6β4* receptors. PCR was used to show the presence of transcripts for α6 and β4 subunits and pharmacological characterization was performed using patch-clamp electrophysiology in combination with α-conotoxins that target the α6β4* subtype. Acetylcholine-evoked currents were sensitive to inhibition by BuIA[T5A,P6O] and MII[H9A,L15A]; α-conotoxins that inhibit α6-containing nAChRs. Two additional agonists were used to probe for the expression of α7 and β2-containing nAChRs. Cells with currents evoked by acetylcholine were relatively unresponsive to the α7-selctive agonist choline but responded to the agonist 5-I-A-85380. These studies provide further insights into the properties of natively expressed α6β4* nAChRs. PMID:24727685

  5. Dopamine modulates acetylcholine release via octopamine and CREB signaling in Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Satoshi Suo

    Full Text Available Animals change their behavior and metabolism in response to external stimuli. cAMP response element binding protein (CREB is a signal-activated transcription factor that enables the coupling of extracellular signals and gene expression to induce adaptive changes. Biogenic amine neurotransmitters regulate CREB and such regulation is important for long-term changes in various nervous system functions, including learning and drug addiction. In Caenorhabditis elegans, the amine neurotransmitter octopamine activates a CREB homolog, CRH-1, in cholinergic SIA neurons, whereas dopamine suppresses CREB activation by inhibiting octopamine signaling in response to food stimuli. However, the physiological role of this activation is unknown. In this study, the effect of dopamine, octopamine, and CREB on acetylcholine signaling was analyzed using the acetylcholinesterase inhibitor aldicarb. Mutants with decreased dopamine signaling exhibited reduced acetylcholine signaling, and octopamine and CREB functioned downstream of dopamine in this regulation. This study demonstrates that the regulation of CREB by amine neurotransmitters modulates acetylcholine release from the neurons of C. elegans.

  6. Development of a novel real-time RT-PCR assay to detect Seneca Valley virus-1 associated with emerging cases of vesicular disease in pigs.

    Science.gov (United States)

    Fowler, Veronica L; Ransburgh, Russell H; Poulsen, Elizabeth G; Wadsworth, Jemma; King, Donald P; Mioulet, Valerie; Knowles, Nick J; Williamson, Susanna; Liu, Xuming; Anderson, Gary A; Fang, Ying; Bai, Jianfa

    2017-01-01

    Seneca Valley virus 1 (SVV-1) can cause vesicular disease that is clinically indistinguishable from foot-and-mouth disease, vesicular stomatitis and swine vesicular disease. SVV-1-associated disease has been identified in pigs in several countries, namely USA, Canada, Brazil and China. Diagnostic tests are required to reliably detect this emerging virus, and this report describes the development and evaluation of a novel real-time (r) reverse-transcription (RT) PCR assay (rRT-PCR), targeting the viral polymerase gene (3D) of SVV-1. This new assay detected all historical and contemporary SVV-1 isolates examined (n=8), while no cross-reactivity was observed with nucleic acid templates prepared from other vesicular disease viruses or common swine pathogens. The analytical sensitivity of the rRT-PCR was 0.79 TCID 50 /ml and the limit of detection was equivalent using two different rRT-PCR master-mixes. The performance of the test was further evaluated using pig nasal (n=25) and rectal swab samples (n=25), where concordant results compared to virus sequencing were generated for 43/50 samples. The availability of this assay, will enable laboratories to rapidly detect SVV-1 in cases of vesicular disease in pigs, negated for notifiable diseases, and could enable existing knowledge gaps to be investigated surrounding the natural epidemiology of SVV-1. Copyright © 2016 Elsevier B.V. All rights reserved.

  7. Plasma biomarkers of decreased vesicular storage distinguish Parkinson disease with orthostatic hypotension from the parkinsonian form of multiple system atrophy.

    Science.gov (United States)

    Goldstein, David S; Kopin, Irwin J; Sharabi, Yehonatan; Holmes, Courtney

    2015-02-01

    Parkinson disease with orthostatic hypotension (PD + OH) and the parkinsonian form of multiple system atrophy (MSA-P) can be difficult to distinguish clinically. Recent studies indicate that PD entails a vesicular storage defect in catecholaminergic neurons. Although cardiac sympathetic neuroimaging by (18)F-dopamine positron emission tomography can identify decreased vesicular storage, this testing is not generally available. We assessed whether plasma biomarkers of a vesicular storage defect can separate PD + OH from MSA-P. We conceptualized that after F-dopamine injection, augmented production of F-dihydroxyphenylacetic acid (F-DOPAC) indicates decreased vesicular storage, and we therefore predicted that arterial plasma F-DOPAC would be elevated in PD + OH but not in MSA-P. We measured arterial plasma F-DOPAC after (18)F-dopamine administration (infused i.v. over 3 min) in patients with PD + OH (N = 12) or MSA-P (N = 21) and in healthy control subjects (N = 26). Peak F-DOPAC:dihydroxyphenylglycol (DHPG) was also calculated to adjust for effects of denervation on F-DOPAC production. Plasma F-DOPAC accumulated rapidly after initiation of (18)F-dopamine infusion. Peak F-DOPAC (5-10 min) in PD + OH averaged three times that in MSA-P (P 300 nCi-kg/cc-mCi, in contrast with 7 of 12 PD + OH patients (χ(2) = 16.6, P < 0.0001). DHPG was lower in PD + OH (3.83 ± 0.36 nmol/L) than in MSA-P (5.20 ± 0.29 nmol/L, P = 0.007). All MSA-P patients had peak F-DOPAC:DHPG < 60, in contrast with 9 of 12 PD + OH patients (χ(2) = 17.5, P < 0.0001). Adjustment of peak F-DOPAC for DHPG increased test sensitivity from 58 to 81% at similar high specificity. After F-dopamine injection, plasma F-DOPAC and F-DOPAC:DHPG distinguish PD + OH from MSA-P.

  8. Resistance Responses of Potato to Vesicular-Arbuscular Mycorrhizal Fungi under Varying Abiotic Phosphorus Levels.

    Science.gov (United States)

    McArthur, D A; Knowles, N R

    1992-09-01

    In mycorrhizal symbioses, susceptibility of a host plant to infection by fungi is influenced by environmental factors, especially the availability of soil phosphorus. This study describes morphological and biochemical details of interactions between a vesicular-arbuscular mycorrhizal (VAM) fungus and potato (Solanum tuberosum L. cv Russet Burbank) plants, with a particular focus on the physiological basis for P-induced resistance of roots to infection. Root infection by the VAM fungus Glomus fasciculatum ([Thaxt. sensu Gerdemann] Gerdemann and Trappe) was extensive for plants grown with low abiotic P supply, and plant biomass accumulation was enhanced by the symbiosis. The capacity of excised roots from P-deficient plants to produce ethylene in the presence or absence of exogenous 1-amino cyclopropane-1-carboxylic acid (ACC) was markedly reduced by VAM infection. This apparent inhibition of ACC oxidase (ACC(ox)) activity was localized to areas containing infected roots, as demonstrated in split-root studies. Furthermore, leachate from VAM roots contained a potent water-soluble inhibitor of ethylene generation from exogenous ACC by nonmycorrhizal (NM) roots. The leachate from VAM-infected roots had a higher concentration of phenolics, relative to that from NM roots. Moreover, the rates of ethylene formation and phenolic concentration in leachates from VAM roots were inversely correlated, suggesting that this inhibitor may be of a phenolic nature. The specific activity of extracellular peroxidase recovered in root leachates was not stimulated by VAM infection, although activity on a fresh weight basis was significantly enhanced, reflecting the fact that VAM roots had higher protein content than NM roots. Polyphenol oxidase activity of roots did not differ between NM and VAM roots. These results characterize the low resistance response of P-deficient plants to VAM infection. When plants were grown with higher abiotic P supply, the relative benefit of the VAM symbiosis

  9. Influence of Vesicular-Arbuscular Mycorrhizal Fungi on the Response of Potato to Phosphorus Deficiency.

    Science.gov (United States)

    McArthur, DAJ.; Knowles, N. R.

    1993-01-01

    Morphological and biochemical interactions between a vesicular-arbuscular mycorrhizal (VAM) fungus (Glomus fasciculatum [Thaxt. sensu Gerdemann] Gerdemann and Trappe) and potato (Solanum tuberosum L.) plants during the development of P deficiency were characterized. Nonmycorrhizal (NM) plants grown for 63 d with low abiotic P supply (0.5 mM) produced 34, 52, and 73% less root, shoot, and tuber dry matter, respectively, than plants grown with high P (2.5 mM). The total leaf area and the leaf area:plant dry weight ratio of low-P plants were substantially lower than those of high-P plants. Moreover, a lower shoot:root dry weight ratio and tuber:plant dry weight ratio in low-P plants than in high-P plants characterized a major effect of P deficiency stress on dry matter partitioning. In addition to a slower rate of growth, low-P plants accumulated nonreducing sugars and nitrate. Furthermore, root respiration and leaf nitrate reductase activity were lower in low-P plants than in high-P plants. Low abiotic P supply also induced physiological changes that contributed to the greater efficiency of P acquisition by low-P plants than by high-P plants. For example, allocation of dry matter and P to root growth was less restricted by P deficiency stress than to shoot and tuber growth. Also, the specific activities of root acid phosphatases and vanadate-sensitive microsomal ATPases were enhanced in P-deficient plants. The establishment of a VAM symbiosis by low-P plants was essential for efficient P acquisition, and a greater root infection level for P-stressed plants indicated increased compatibility to the VAM fungus. By 63 d after planting, low-P VAM plants had recovered 42% more of the available soil P than low-P NM plants. However, the VAM fungus only partially alleviated P deficiency stress and did not completely compensate for inadequate abiotic P supply. Although the specific activities of acid phosphatases and microsomal ATPases were only marginally influenced by VAM

  10. Pancreatic and snake venom presynaptically active phospholipases A2 inhibit nicotinic acetylcholine receptors.

    Science.gov (United States)

    Vulfius, Catherine A; Kasheverov, Igor E; Kryukova, Elena V; Spirova, Ekaterina N; Shelukhina, Irina V; Starkov, Vladislav G; Andreeva, Tatyana V; Faure, Grazyna; Zouridakis, Marios; Tsetlin, Victor I; Utkin, Yuri N

    2017-01-01

    Phospholipases A2 (PLA2s) are enzymes found throughout the animal kingdom. They hydrolyze phospholipids in the sn-2 position producing lysophospholipids and unsaturated fatty acids, agents that can damage membranes. PLA2s from snake venoms have numerous toxic effects, not all of which can be explained by phospholipid hydrolysis, and each enzyme has a specific effect. We have earlier demonstrated the capability of several snake venom PLA2s with different enzymatic, cytotoxic, anticoagulant and antiproliferative properties, to decrease acetylcholine-induced currents in Lymnaea stagnalis neurons, and to compete with α-bungarotoxin for binding to nicotinic acetylcholine receptors (nAChRs) and acetylcholine binding protein. Since nAChRs are implicated in postsynaptic and presynaptic activities, in this work we probe those PLA2s known to have strong presynaptic effects, namely β-bungarotoxin from Bungarus multicinctus and crotoxin from Crotalus durissus terrificus. We also wished to explore whether mammalian PLA2s interact with nAChRs, and have examined non-toxic PLA2 from porcine pancreas. It was found that porcine pancreatic PLA2 and presynaptic β-bungarotoxin blocked currents mediated by nAChRs in Lymnaea neurons with IC50s of 2.5 and 4.8 μM, respectively. Crotoxin competed with radioactive α-bungarotoxin for binding to Torpedo and human α7 nAChRs and to the acetylcholine binding protein. Pancreatic PLA2 interacted similarly with these targets; moreover, it inhibited radioactive α-bungarotoxin binding to the water-soluble extracellular domain of human α9 nAChR, and blocked acetylcholine induced currents in human α9α10 nAChRs heterologously expressed in Xenopus oocytes. These and our earlier results show that all snake PLA2s, including presynaptically active crotoxin and β-bungarotoxin, as well as mammalian pancreatic PLA2, interact with nAChRs. The data obtained suggest that this interaction may be a general property of all PLA2s, which should be proved by

  11. Two types of muscarinic acetylcholine receptors in Drosophila and other arthropods.

    Science.gov (United States)

    Collin, Caitlin; Hauser, Frank; Gonzalez de Valdivia, Ernesto; de Valdivia, Ernesto Gonzalez; Li, Shizhong; Reisenberger, Julia; Carlsen, Eva M M; Khan, Zaid; Hansen, Niels O; Puhm, Florian; Søndergaard, Leif; Niemiec, Justyna; Heninger, Magdalena; Ren, Guilin R; Grimmelikhuijzen, Cornelis J P

    2013-09-01

    Muscarinic acetylcholine receptors (mAChRs) play a central role in the mammalian nervous system. These receptors are G protein-coupled receptors (GPCRs), which are activated by the agonists acetylcholine and muscarine, and blocked by a variety of antagonists. Mammals have five mAChRs (m1-m5). In this study, we cloned two structurally related GPCRs from the fruit fly Drosophila melanogaster, which, after expression in Chinese hamster ovary cells, proved to be muscarinic acetylcholine receptors. One mAChR (the A-type; encoded by gene CG4356) is activated by acetylcholine (EC50, 5 × 10(-8) M) and muscarine (EC50, 6 × 10(-8) M) and blocked by the classical mAChR antagonists atropine, scopolamine, and 3-quinuclidinyl-benzilate (QNB), while the other (the B-type; encoded by gene CG7918) is also activated by acetylcholine, but has a 1,000-fold lower sensitivity to muscarine, and is not blocked by the antagonists. A- and B-type mAChRs were also cloned and functionally characterized from the red flour beetle Tribolium castaneum. Recently, Haga et al. (Nature 2012, 482: 547-551) published the crystal structure of the human m2 mAChR, revealing 14 amino acid residues forming the binding pocket for QNB. These residues are identical between the human m2 and the D. melanogaster and T. castaneum A-type mAChRs, while many of them are different between the human m2 and the B-type receptors. Using bioinformatics, one orthologue of the A-type and one of the B-type mAChRs could also be found in all other arthropods with a sequenced genome. Protostomes, such as arthropods, and deuterostomes, such as mammals and other vertebrates, belong to two evolutionarily distinct lineages of animal evolution that split about 700 million years ago. We found that animals that originated before this split, such as cnidarians (Hydra), had two A-type mAChRs. From these data we propose a model for the evolution of mAChRs.

  12. Pancreatic and snake venom presynaptically active phospholipases A2 inhibit nicotinic acetylcholine receptors.

    Directory of Open Access Journals (Sweden)

    Catherine A Vulfius

    Full Text Available Phospholipases A2 (PLA2s are enzymes found throughout the animal kingdom. They hydrolyze phospholipids in the sn-2 position producing lysophospholipids and unsaturated fatty acids, agents that can damage membranes. PLA2s from snake venoms have numerous toxic effects, not all of which can be explained by phospholipid hydrolysis, and each enzyme has a specific effect. We have earlier demonstrated the capability of several snake venom PLA2s with different enzymatic, cytotoxic, anticoagulant and antiproliferative properties, to decrease acetylcholine-induced currents in Lymnaea stagnalis neurons, and to compete with α-bungarotoxin for binding to nicotinic acetylcholine receptors (nAChRs and acetylcholine binding protein. Since nAChRs are implicated in postsynaptic and presynaptic activities, in this work we probe those PLA2s known to have strong presynaptic effects, namely β-bungarotoxin from Bungarus multicinctus and crotoxin from Crotalus durissus terrificus. We also wished to explore whether mammalian PLA2s interact with nAChRs, and have examined non-toxic PLA2 from porcine pancreas. It was found that porcine pancreatic PLA2 and presynaptic β-bungarotoxin blocked currents mediated by nAChRs in Lymnaea neurons with IC50s of 2.5 and 4.8 μM, respectively. Crotoxin competed with radioactive α-bungarotoxin for binding to Torpedo and human α7 nAChRs and to the acetylcholine binding protein. Pancreatic PLA2 interacted similarly with these targets; moreover, it inhibited radioactive α-bungarotoxin binding to the water-soluble extracellular domain of human α9 nAChR, and blocked acetylcholine induced currents in human α9α10 nAChRs heterologously expressed in Xenopus oocytes. These and our earlier results show that all snake PLA2s, including presynaptically active crotoxin and β-bungarotoxin, as well as mammalian pancreatic PLA2, interact with nAChRs. The data obtained suggest that this interaction may be a general property of all PLA2s, which

  13. Stimuli that induce a cholinergic neuronal phenotype of NG108-15 cells upregulate ChAT and VAChT mRNAs but fail to increase VAChT protein

    Czech Academy of Sciences Publication Activity Database

    Doležal, Vladimír; Castell, X.; Tomasi, M.; Diebler, M. F.

    2001-01-01

    Roč. 54, č. 4 (2001), s. 363-373 ISSN 0361-9230 R&D Projects: GA ČR GA305/98/0469 Institutional research plan: CEZ:AV0Z5011922 Keywords : choline acetyltransferase * vesicular acetylcholine transporter * NG108-15 Subject RIV: FH - Neurology Impact factor: 1.783, year: 2001

  14. The grain size dependency of vesicular particle shapes strongly affects the drag of particles. First results from microtomography investigations of Campi Flegrei fallout deposits

    Science.gov (United States)

    Mele, Daniela; Dioguardi, Fabio

    2018-03-01

    Acknowledging the grain size dependency of shape is important in volcanology, in particular when dealing with tephra produced and emplaced during and after explosive volcanic eruptions. A systematic measurement of the tridimensional shape of vesicular pyroclasts of Campi Flegrei fallout deposits (Agnano-Monte Spina, Astroni 6 and Averno 2 eruptions) varying in size from 8.00 to 0.016 mm has been carried out by means of X-Ray Microtomography. Data show that particle shape changes with size, especially for juvenile vesicular clasts, since it is dependent on the distribution and size of vesicles that contour the external clast outline. Two drag laws that include sphericity in the formula were used for estimating the dependency of settling velocity on shape. Results demonstrate that it is not appropriate to assume a size-independent shape for vesicular particles, in contrast with the approach commonly employed when simulating the ash dispersion in the atmosphere.

  15. Choline transporter mutations in severe congenital myasthenic syndrome disrupt transporter localization.

    Science.gov (United States)

    Wang, Haicui; Salter, Claire G; Refai, Osama; Hardy, Holly; Barwick, Katy E S; Akpulat, Ugur; Kvarnung, Malin; Chioza, Barry A; Harlalka, Gaurav; Taylan, Fulya; Sejersen, Thomas; Wright, Jane; Zimmerman, Holly H; Karakaya, Mert; Stüve, Burkhardt; Weis, Joachim; Schara, Ulrike; Russell, Mark A; Abdul-Rahman, Omar A; Chilton, John; Blakely, Randy D; Baple, Emma L; Cirak, Sebahattin; Crosby, Andrew H

    2017-11-01

    The presynaptic, high-affinity choline transporter is a critical determinant of signalling by the neurotransmitter acetylcholine at both central and peripheral cholinergic synapses, including the neuromuscular junction. Here we describe an autosomal recessive presynaptic congenital myasthenic syndrome presenting with a broad clinical phenotype due to homozygous choline transporter missense mutations. The clinical phenotype ranges from the classical presentation of a congenital myasthenic syndrome in one patient (p.Pro210Leu), to severe neurodevelopmental delay with brain atrophy (p.Ser94Arg) and extend the clinical outcomes to a more severe spectrum with infantile lethality (p.Val112Glu). Cells transfected with mutant transporter construct revealed a virtually complete loss of transport activity that was paralleled by a reduction in transporter cell surface expression. Consistent with these findings, studies to determine the impact of gene mutations on the trafficking of the Caenorhabditis elegans choline transporter orthologue revealed deficits in transporter export to axons and nerve terminals. These findings contrast with our previous findings in autosomal dominant distal hereditary motor neuropathy of a dominant-negative frameshift mutation at the C-terminus of choline transporter that was associated with significantly reduced, but not completely abrogated choline transporter function. Together our findings define divergent neuropathological outcomes arising from different classes of choline transporter mutation with distinct disease processes and modes of inheritance. These findings underscore the essential role played by the choline transporter in sustaining acetylcholine neurotransmission at both central and neuromuscular synapses, with important implications for treatment and drug selection. © The Author (2017). Published by Oxford University Press on behalf of the Guarantors of Brain. All rights reserved. For Permissions, please email: journals.permissions@oup.com.

  16. Depolarization by K+ and glutamate activates different neurotransmitter release mechanisms in GABAergic neurons: vesicular versus non-vesicular release of GABA

    DEFF Research Database (Denmark)

    Belhage, B; Hansen, Gert Helge; Schousboe, A

    1993-01-01

    differences in the mode of action of the two depolarizing stimuli were reflected in the properties of the increase in [Ca++]i elicited by 55 mM K+ and 100 microM glutamate, respectively. The K(+)-induced increase in [Ca++]i was reduced by both verapamil and Ca(++)-free media whereas the corresponding...... neurotransmitter glutamate (100 microM). Both depolarizing stimuli exerted prompt increases in the release of preloaded [3H]GABA as well as in [Ca++]i. However, the basic properties of transmitter release and the increase in [Ca++]i under a variety of conditions were different during stimulation with K...... was also reduced by organic (verapamil) and inorganic (Co++) Ca++ channel blockers but was insensitive to the GABA transport inhibitor SKF 89976A. In contrast, the second phase was less sensitive to nocodazole and Ca++ channel antagonists but could be inhibited by SKF 89976A. The glutamate-induced [3H...

  17. Didelphis marsupialis como un reservorio potencial u hospedero amplificador del virus de la estomatitis vesicular, serotipo new jersey en Antioquia

    Directory of Open Access Journals (Sweden)

    John Arboleda

    2004-02-01

    Full Text Available

    La Estomatitis Vesicular (EV es una enfermedad viral, aguda
    y autolimitante que afecta principalmente bovinos, equinos y
    porcinos. Es producida por el virus de estomatitis vesicular (VEV, serotipos New Jersey (VEV-NJ e Indiana (VEV-IN, que son los as importantes epidemiológicamente (1. Los estudios serológicos demuestran que VEV-NJ y VEV-IN infectan en forma natural una gran variedad de animales silvestres, que están posiblemente implicados en la  coepizootiología de la EV, como hospederos portadores, mplificadores o reservorios (2.

    La zarigüeya (Didelphis marsupialis es un buen candidato
    para cumplir esta función, debido a que es la especie silvestre
    mayormente capturada en zonas enzoóticas; presenta altos
    porcentajes de infección natural (3, resiste la antropización y
    además, su comportamiento le permite interactuar con
    diferentes poblaciones de vectores u otros reservorios en los
    bosques y servir como fuente de infección para las especies
    domésticas susceptibles.

     

     

  18. Eficacia de la Prueba del Cuy en el Diagnóstico de Patologia Vesicular en Pacientes del Consultorio Particular, Arequipa. 2007 - 2008

    OpenAIRE

    Zeballos Benavides María Candelaria

    2010-01-01

    En la investigación se trata de encontrar los hallazgos en la vesícula de la persona con la Prueba del Cuy y compararlos con los hallazgos de la Prueba Ecográfica para determinar su eficacia. El objetivo del presente estudio de investigación fue evaluar la Eficacia de la Prueba del Cuy (cobayo), en el diagnóstico de Patología Vesicular. Se escogió un grupo de estudio que fue constituido por 21 personas con sintomatología aparentemente vesicular, deseosos de encontrar resp...

  19. Prior nicotine self-administration attenuates subsequent dopaminergic deficits of methamphetamine in rats: role of nicotinic acetylcholine receptors.

    Science.gov (United States)

    Baladi, Michelle G; Nielsen, Shannon M; McIntosh, J Michael; Hanson, Glen R; Fleckenstein, Annette E

    2016-08-01

    Preclinical studies have demonstrated that oral nicotine exposure attenuates long-term dopaminergic damage induced by toxins, including repeated, high doses of methamphetamine. It is suggested that alterations in nicotinic acetylcholine receptor (nAChR) expression, including α4β2* and α6β2* subtypes, likely contribute to this protection. The current study extended these findings by investigating whether nicotine self-administration in male, Sprague-Dawley rats (a) attenuates short-term dopaminergic damage induced by methamphetamine and (b) causes alterations in levels of α4β2* and α6β2* nAChR subtypes. The findings indicate that nicotine self-administration (0.032 mg/kg/infusion for 14 days) per se did not alter α4β2* and α6β2* nAChR expression or dopamine transporter (DAT) expression and function. Interestingly, prior nicotine self-administration attenuated methamphetamine-induced decreases in DAT function when assessed 24 h, but not 1 h, after methamphetamine treatment (4×7.5 mg/kg/injection). The ability of nicotine to attenuate the effects of methamphetamine on DAT function corresponded with increases in α4β2*, but not α6β2*, nAChR binding density. Understanding the role of nAChRs in methamphetamine-induced damage has the potential to elucidate mechanisms underlying the etiology of disorders involving dopaminergic dysfunction, as well as to highlight potential new therapeutic strategies for prevention or reduction of dopaminergic neurodegeneration.

  20. Acetylcholine Attenuates Hypoxia/ Reoxygenation-Induced Mitochondrial and Cytosolic ROS Formation in H9c2 Cells via M2 Acetylcholine Receptor

    Directory of Open Access Journals (Sweden)

    Yi Miao

    2013-02-01

    Full Text Available Background: The anti-infammatory and cardioprotective effect of acetylcholine (ACh has been reported; nevertheless, whether and how ACh exhibits an antioxidant property against ischemia/reperfusion (I/R-induced oxidative stress remains obscure. Methods: In the present study, H9c2 rat cardiomyocytes were exposed to hypoxia/reoxygenation (H/R to mimic I/R injury. We estimated intracellular different sources of reactive oxygen species (ROS by measuring mitochondrial ROS (mtROS, mitochondrial DNA (mtDNA copy number, xanthine oxidase (XO and NADPH oxidase (NOX activity and expression of rac 1. Cell injury was determined by lactate dehydrogenase (LDH release and cleaved caspase-3 expression. The siRNA transfection was performed to knockdown of M2 acetylcholine receptor (M2 AChR expression. Results: 12-h hypoxia followed by 2-h reoxygenation resulted in an abrupt burst of ROS in H9c2 cells. Administration of ACh reduced the levels of ROS in a concentration-dependent manner. Compared to the H/R group, ACh decreased mtROS, recovered mtDNA copy number, diminished XO and NOX activity, rac 1 expression as well as cell injury. Co- treatment with atropine rather than hexamethonium abolished the antioxidant and cardioprotective effect of ACh. Moreover, knockdown of M2 AChR by siRNA showed the similar trends as atropine co-treatment group. Conclusions: ACh inhibits mitochondria-, XO- and NOX-derived ROS production thus protecting H9c2 cells against H/R-induced oxidative stress, and these benefcial effects are mainly mediated by M2 AChR. Our findings suggested that increasing ACh release could be a potential therapeutic strategy for treatment and prevention of I/R injury.

  1. Local application of SCH 39166 reversibly and dose-dependently decreases acetylcholine release in the rat striatum.

    Science.gov (United States)

    Acquas, E; Di Chiara, G

    1999-11-03

    The effect of local application by reverse dialysis of the dopamine D(1) receptor antagonist (-)-trans-6,7,7a,8,9, 13b-exahydro-3-chloro-2-hydroxy-N-methyl-5H-benzo-[d]-nap hto-[2, 1b]-azepine hydrochloride (SCH 39166) on acetylcholine release was studied in awake, freely moving rats implanted with concentric microdialysis probes in the dorsal striatum. In these experiments, the reversible acetylcholine esterase inhibitor, neostigmine, was added to the perfusion solution at two different concentrations, 0.01 and 0.1 microM. SCH 39166 (1, 5 and 10 microM), in the presence of 0.01 microM neostigmine, reversibly decreased striatal acetylcholine release (1 microM SCH 39166 by 8+/-4%; 5 microM SCH 39166 by 24+/-5%; 10 microM SCH 39166 by 27+/-7%, from basal). Similarly, SCH 39166, applied in the presence of a higher neostigmine concentration (0.1 microM), decreased striatal acetylcholine release by 14+/-4% at 1 microM, by 28+/-8% at 5 microM and by 30+/-5% at 10 microM, in a dose-dependent and time-dependent manner. These results are consistent with the existence of a facilitatory tone of dopamine on striatal acetylcholine transmission mediated by dopamine D(1) receptors located on striatal cholinergic interneurons.

  2. Investigation of the presence and antinociceptive function of muscarinic acetylcholine receptors in the African naked mole-rat (Heterocephalus glaber).

    Science.gov (United States)

    Jørgensen, Kristine B; Krogh-Jensen, Karen; Pickering, Darryl S; Kanui, Titus I; Abelson, Klas S P

    2016-01-01

    The present study investigated the cholinergic system in the African naked mole-rat (Heterocephalus glaber) with focus on the muscarinic acetylcholine receptor subtypes M1 and M4. The protein sequences for the subtypes m 1-5 of the naked mole-rat were compared to that of the house mouse (Mus musculus) using basic local alignment search tool (BLAST). The presence and function of M1 and M4 was investigated in vivo, using the formalin test with the muscarinic receptor agonists xanomeline and VU0152100. Spinal cord tissue from the naked mole-rat was used for receptor saturation binding studies with [(3)H]-N-methylscopolamine. The BLAST test revealed 95 % protein sequence homology showing the naked mole-rat to have the genetic potential to express all five muscarinic acetylcholine receptor subtypes. A significant reduction in pain behavior was demonstrated after administration of 8.4 mg/kg in the formalin test. Administration of 50 mg/kg VU0152100 resulted in a non-significant tendency towards antinociception. The antinociceptive effects were reversed by the muscarinic acetylcholine receptor antagonist atropine. Binding studies indicated presence of muscarinic acetylcholine receptors with a radioligand affinity comparable to that reported in mice. In conclusion, muscarinic acetylcholine receptor subtypes are present in the naked mole-rat and contribute to antinociception in the naked mole-rat.

  3. A neuronal acetylcholine receptor regulates the balance of muscle excitation and inhibition in Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Maelle Jospin

    2009-12-01

    Full Text Available In the nematode Caenorhabditis elegans, cholinergic motor neurons stimulate muscle contraction as well as activate GABAergic motor neurons that inhibit contraction of the contralateral muscles. Here, we describe the composition of an ionotropic acetylcholine receptor that is required to maintain excitation of the cholinergic motor neurons. We identified a gain-of-function mutation that leads to spontaneous muscle convulsions. The mutation is in the pore domain of the ACR-2 acetylcholine receptor subunit and is identical to a hyperactivating mutation in the muscle receptor of patients with myasthenia gravis. Screens for suppressors of the convulsion phenotype led to the identification of other receptor subunits. Cell-specific rescue experiments indicate that these subunits function in the cholinergic motor neurons. Expression of these subunits in Xenopus oocytes demonstrates that the functional receptor is comprised of three alpha-subunits, UNC-38, UNC-63 and ACR-12, and two non-alpha-subunits, ACR-2 and ACR-3. Although this receptor exhibits a partially overlapping subunit composition with the C. elegans muscle acetylcholine receptor, it shows distinct pharmacology. Recordings from intact animals demonstrate that loss-of-function mutations in acr-2 reduce the excitability of the cholinergic motor neurons. By contrast, the acr-2(gf mutation leads to a hyperactivation of cholinergic motor neurons and an inactivation of downstream GABAergic motor neurons in a calcium dependent manner. Presumably, this imbalance between excitatory and inhibitory input into muscles leads to convulsions. These data indicate that the ACR-2 receptor is important for the coordinated excitation and inhibition of body muscles underlying sinusoidal movement.

  4. Prostate stem cell antigen interacts with nicotinic acetylcholine receptors and is affected in Alzheimer's disease

    DEFF Research Database (Denmark)

    Jensen, Majbrit Myrup; Mikkelsen, Jens D.; Arvaniti, Maria

    2015-01-01

    Alzheimer's disease (AD) is a neurodegenerative disorder involving impaired cholinergic neurotransmission and dysregulation of nicotinic acetylcholine receptors (nAChRs). Ly-6/neurotoxin (Lynx) proteins have been shown to modulate cognition and neural plasticity by binding to nAChR subtypes...... are present in the human brain. We further showed that PSCA forms stable complexes with the α4 nAChR subunit and decreases nicotine-induced extracellular-signal regulated kinase phosphorylation in PC12 cells. In addition, we analyzed protein levels of PSCA and Lypd6 in postmortem tissue of medial frontal...

  5. Lynx1 and Aβ1-42 bind competitively to multiple nicotinic acetylcholine receptor subtypes

    DEFF Research Database (Denmark)

    Thomsen, Morten S; Arvaniti, Maria; Jensen, Majbrit M

    2016-01-01

    Lynx1 regulates synaptic plasticity in the brain by regulating nicotinic acetylcholine receptors (nAChRs). It is not known to which extent Lynx1 can bind to endogenous nAChR subunits in the brain or how this interaction is affected by Alzheimer's disease pathology. We apply affinity purification....... Incubation with Ws-Lynx1 decreases nicotine-mediated extracellular signal-regulated kinase phosphorylation in PC12 cells and striatal neurons, indicating that binding of Ws-Lynx1 is sufficient to inhibit signaling downstream of nAChRs. The effect of nicotine in PC12 cells is independent of α7 or α4β2 n...

  6. Regional distribution of muscarinic acetylcholine receptors in the telencephalon of the pigeon (Columba livia f. domestica)

    International Nuclear Information System (INIS)

    Waechtler, K.

    1985-01-01

    The distribution of muscarinic acetylcholine receptors was studied autoradiographically in croystat sections of the pigeon telencephalon using 3 H-quinuclidinylbenzylate as a ligand. Highest receptor density was observed in the hyperstriatum ventrale, palaeostriatum augmentatum, septum, and parts of the archistriatum. In sites of known sensory input of neostriatum (field L) and ectostriatum low receptor binding was observed. Acetylcholinesterase distribution is in good agreement with the receptor picture only in the basal telencephalon. In the pallium differences in the pattern of these two components can be seen. (author)

  7. Changes in acetylcholine release from the chick retina are not associated with myopia development

    International Nuclear Information System (INIS)

    Vessey, K.A.; Cotriall, C.L.; McBrien, N.A.

    2002-01-01

    Full text: The effectiveness of muscarinic receptor antagonists in inhibiting myopia progression in animal models and humans implicates cholinergic signalling in ocular growth regulation. Therefore to determine if changes in the release of acetylcholine from the retina are involved in myopia development, the efflux of acetylcholine from the in vitro retina of normal and myopic chick eyes was investigated. Chicks were monocularly deprived (MD) of pattern vision with translucent occluders for 2 or 7 days and refractive error of MD groups and age matched normals was monitored using retinoscopy (n=6 each group). 3 H-choline-Cl (1 Ci in 7μL) was injected into the vitreous of each eye under 2.5% halothane anaesthesia. After 1hr, the eyes were enucleated, under terminal anaesthesia (sodium pentobarbital, 120 mg/kg, im). Retinas were flat-mounted on acetate filter discs and superfused with oxygenated physiological saline solution (PSS) for 30min at 0.4mL/min. Five baseline fractions were collected (B1-B5), then three stimulated fractions were collected in the presence of PSS containing 50mM KCl (K1-K3) at 2min intervals. 3 H-acetylcholine ( 3 H-ACh) in each fraction was quantified by liquid scintillation counting. Significant amounts of myopia were induced in MD eyes after 2 (-5.1±0.8D) and 7 days (-18.8±2.4D) relative to control eyes (paired t-test p 3 H-ACh release was 146±15% above basal levels (K2/B1%) from retinas of normal animals. After 2 days MD, there was no significant difference between KCl-evoked release of 3 H-ACh from deprived eyes (147 39%) compared to control eyes (198±61%, paired t-test, p=0.27) or the eyes of normal animals (ANOVA, p>0.5). Similar results were obtained following 7 days MD. The results demonstrate that evoked acetylcholine release from the chick retina of myopic eyes is unaltered relative to control or normal eyes using an in vitro approach. Copyright (2002) Australian Neuroscience Society

  8. Temperature effect on proximal to distal gradient of quantal release of acetylcholine at frog endplate

    Czech Academy of Sciences Publication Activity Database

    Samigullin, D.; Bukharaeva, E.; Nikolsky, E.; Vyskočil, František

    2003-01-01

    Roč. 28, 3-4 (2003), s. 507-514 ISSN 0364-3190 R&D Projects: GA AV ČR IAA7011902; GA ČR GA305/02/1333; GA ČR GA202/02/1213 Grant - others:RFBR(RU) 02/04/48901 Institutional research plan: CEZ:AV0Z5011922; CEZ:MSM 113100003 Keywords : neuromuscular junction * acetylcholine release * temperature Subject RIV: ED - Physiology Impact factor: 1.511, year: 2003

  9. Characterization of the positive and negative inotropic effects of acetylcholine in the human myocardium

    OpenAIRE

    Du, Xiaoyi; Schoemaker, Regien; Bos, Egbert; Saxena, Pramod Ranjan

    1995-01-01

    textabstractIn the human isolated myocardium, acetylcholine (10−9 to 10−3 M) elicited a biphasic inotropic effect (a decrease in the lower and an increase in the higher concentration range) in atrial and a positive inotropic effect in ventricular trabeculae. However, under conditions of raised contractility achieved by exposure to noradrenaline (10−5 M), only negative inotropic effects were observed in both atria and ventricles. Atropine (10−6 M), but not propranolol (10−6 M), antagonized bot...

  10. [[sup 3]H]imidacloprid: synthesis of a candidate radioligand for the nicotinic acetylcholine receptor

    Energy Technology Data Exchange (ETDEWEB)

    Latli, B.; Casida, J.E. (California Univ., Berkeley, CA (United States). Dept. of Entomological Sciences)

    1992-08-01

    Imidacloprid is an exceptionally potent insecticide known from physiological studies to act at the nicotinic acetylcholine receptor. To prepare [[sup 3]H]imidacloprid as a candidate radioligand, 6-chloronicotinoyl chloride was reduced with NaB[sup 2]H[sub 4] (in model studies) or NaB[sup 3]H[sub 4] in absolute ethanol to 2-chloro-5-pyridinylmethanol which was transformed to 2-chloro-5-chloromethylpyridine on refluxing with thionyl chloride. Coupling with 4,5-dihydro-N-nitro-1H-imidazol-2-amine then gave [[sup 2]H[sub 2

  11. INFLUENCE OF ANTIBIOTICS ON THE MECHANICAL RESPONSES OF GUINEA-PIG ILEUM TO ACETYLCHOLINE AND HISTAMINE

    Directory of Open Access Journals (Sweden)

    Petroianu Andy

    1998-01-01

    Full Text Available The side effects of antibiotics have been extensively described during the last decades, however, their role on digestive motility must be better investigated. Following a line of research, the influence of penicillin, chloranfenicol tetracycline and gentamicine on longitudinal smooth muscle responses to acetylcholine and histamine were studied on guinea-pig ileum. There were no differences between the responses before and after the addition of each antibiotic. Further investigations must be performed in order to find a possible influence of antibiotics on digestive motility.

  12. Drugs interfering with Muscarinic Acetylcholine Receptors and Their effects on Place Navigation

    Czech Academy of Sciences Publication Activity Database

    Svoboda, Jan; Popelíková, Anna; Stuchlík, Aleš

    2017-01-01

    Roč. 8, Nov 9 (2017), č. článku 215. ISSN 1664-0640 R&D Projects: GA ČR(CZ) GA17-04047S; GA ČR NV17-30833A; GA MŠk(CZ) LM2015062 Grant - others:AV ČR(CZ) PAN-17-07 Program:Bilaterální spolupráce Institutional support: RVO:67985823 Keywords : scopolamine * biperiden * acetylcholine * receptor * behavior * learning Subject RIV: FH - Neurology OBOR OECD: Neurosciences (including psychophysiology Impact factor: 3.532, year: 2016

  13. Muscarinic Acetylcholine Receptor M3 Mutation Causes Urinary Bladder Disease and a Prune-Belly-like Syndrome.

    Science.gov (United States)

    Weber, Stefanie; Thiele, Holger; Mir, Sevgi; Toliat, Mohammad Reza; Sozeri, Betül; Reutter, Heiko; Draaken, Markus; Ludwig, Michael; Altmüller, Janine; Frommolt, Peter; Stuart, Helen M; Ranjzad, Parisa; Hanley, Neil A; Jennings, Rachel; Newman, William G; Wilcox, Duncan T; Thiel, Uwe; Schlingmann, Karl Peter; Beetz, Rolf; Hoyer, Peter F; Konrad, Martin; Schaefer, Franz; Nürnberg, Peter; Woolf, Adrian S

    2011-11-11

    Urinary bladder malformations associated with bladder outlet obstruction are a frequent cause of progressive renal failure in children. We here describe a muscarinic acetylcholine receptor M3 (CHRM3) (1q41-q44) homozygous frameshift mutation in familial congenital bladder malformation associated with a prune-belly-like syndrome, defining an isolated gene defect underlying this sometimes devastating disease. CHRM3 encodes the M3 muscarinic acetylcholine receptor, which we show is present in developing renal epithelia and bladder muscle. These observations may imply that M3 has a role beyond its known contribution to detrusor contractions. This Mendelian disease caused by a muscarinic acetylcholine receptor mutation strikingly phenocopies Chrm3 null mutant mice. Copyright © 2011 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

  14. Autonomous control of phosphatidylinositol turnover by histamine and acetylcholine receptors in the NIE-115 neuron-like cell line

    International Nuclear Information System (INIS)

    Large, T.H.; Lambert, M.P.; Cohen, N.M.; Klein, W.L.

    1986-01-01

    Histamine was found to stimulate the turnover of phosphatidylinositol (PI) in cultures of neuron-like NE-115 cells. Turnover was measured by increased production of ( 3 H)inositol phosphates (breakdown) and by accelerated incorporation of 32 P into PI (resynthesis). Data were consistent with hydrolysis of polyphosphoinositides being the initial event in receptor-stimulated PI turnover. This response to histamine desensitized within 10 min. Receptor systems for histamine and acetylcholine were tested for possible interactions: PI turnover in response to dual stimulation was approximately equal to the sum of the individual responses while prior desensitization of the acetylcholine receptor system had no effect on subsequent stimulation of the histamine receptor system. These results are consistent with the hypothesis that components of acetylcholine and histamine receptor systems responsible for PI turnover are autonomously organised and regulated. (author)

  15. An extract of lionfish (Pterois volitans) spine tissue contains acetylcholine and a toxin that affects neuromuscular transmission.

    Science.gov (United States)

    Cohen, A S; Olek, A J

    1989-01-01

    A soluble toxic extract derived from spine tissue of the lionfish (Pterois volitans) decreased heart rate and force of contraction in isolated clam and frog hearts. These actions were due to the presence of micromolar concentrations of acetylcholine in the extract. Toxicity was retained after hydrolysis of acetylcholine by exogenous acetylcholinesterase, but heart function was no longer affected. Toxin treated in this way induced muscle fibrillation in an isolated nerve-muscle preparation, followed by blockade of neuromuscular transmission. Bursts of transient depolarizations were recorded at the muscle endplate shortly after toxin addition that correlated in time with the duration of toxin-induced muscle fibrillation. These effects are thought to be due to the increased release and then depletion of acetylcholine from the nerve terminal.

  16. Analogues of neuroactive polyamine wasp toxins that lack inner basic sites exhibit enhanced antagonism toward a muscle-type mammalian nicotinic acetylcholine receptor

    DEFF Research Database (Denmark)

    Stromgaard, K; Brierley, M J; Andersen, K

    1999-01-01

    noncompetitively antagonized the nicotinic acetylcholine receptor (nAChR) in a concentration-, time-, and voltage-dependent manner. The amplitudes of acetylcholine-induced currents were compared at their peaks and at the end of a 1 s application in the presence or absence of the analogues. Most of the analogues...

  17. Human Secreted Ly-6/uPAR Related Protein-1 (SLURP-1) Is a Selective Allosteric Antagonist of α7 Nicotinic Acetylcholine Receptor

    DEFF Research Database (Denmark)

    Lyukmanova, Ekaterina N; Shulepko, Mikhail A; Kudryavtsev, Denis

    2016-01-01

    of nicotinic acetylcholine receptors (nAChRs) and muscarinic acetylcholine receptors, respectively, and anti-α7-nAChRs antibodies revealed α7 type nAChRs as an rSLURP-1 target in keratinocytes. Using affinity purification from human cortical extracts, we confirmed that rSLURP-1 binds selectively to the α7-n...

  18. The A- and B-type muscarinic acetylcholine receptors from Drosophila melanogaster couple to different second messenger pathways

    DEFF Research Database (Denmark)

    Ren, Guilin Robin; Folke, Jonas; Hauser, Frank

    2015-01-01

    Muscarinic acetylcholine receptors (mAChRs) are G protein-coupled receptors (GPCRs) that are activated by the agonists acetylcholine and muscarine and blocked by several antagonists, among them atropine. In mammals five mAChRs (m1-m5) exist of which m1, m3, and m5 are coupled to members of the Gq...

  19. Effects of BMS-902483, an α7 nicotinic acetylcholine receptor partial agonist, on cognition and sensory gating in relation to receptor occupancy in rodents.

    Science.gov (United States)

    Pieschl, Rick L; Miller, Regina; Jones, Kelli M; Post-Munson, Debra J; Chen, Ping; Newberry, Kimberly; Benitex, Yulia; Molski, Thaddeus; Morgan, Daniel; McDonald, Ivar M; Macor, John E; Olson, Richard E; Asaka, Yukiko; Digavalli, Siva; Easton, Amy; Herrington, James; Westphal, Ryan S; Lodge, Nicholas J; Zaczek, Robert; Bristow, Linda J; Li, Yu-Wen

    2017-07-15

    The α7 nicotinic acetylcholine receptor is thought to play an important role in human cognition. Here we describe the in vivo effects of BMS-902483, a selective potent α7 nicotinic acetylcholine receptor partial agonist, in relationship to α7 nicotinic acetylcholine receptor occupancy. BMS-902483 has low nanomolar affinity for rat and human α7 nicotinic acetylcholine receptors and elicits currents in cells expressing human or rat α7 nicotinic acetylcholine receptors that are about 60% of the maximal acetylcholine response. BMS-902483 improved 24h novel object recognition memory in mice with a minimal effective dose (MED) of 0.1mg/kg and reversed MK-801-induced deficits in a rat attentional set-shifting model of executive function with an MED of 3mg/kg. Enhancement of novel object recognition was blocked by the silent α7 nicotinic acetylcholine receptor agonist, NS6740, demonstrating that activity of BMS-902483 was mediated by α7 nicotinic acetylcholine receptors. BMS-902483 also reversed ketamine-induced deficits in auditory gating in rats, and enhanced ex vivo hippocampal long-term potentiation examined 24h after dosing in mice. Results from an ex vivo brain homogenate binding assay showed that α7 receptor occupancy ranged from 64% (novel object recognition) to ~90% (set shift and gating) at the MED for behavioral and sensory processing effects of BMS-902483. Copyright © 2017 Elsevier B.V. All rights reserved.

  20. Interaction of alpha-conotoxin ImII and its analogs with nicotinic receptors and acetylcholine-binding proteins: additional binding sites on Torpedo receptor

    NARCIS (Netherlands)

    Kasheverov, I.E.; Zhmak, M.N.; Fish, A.; Rucktooa, P.; Khruschov, A.Y.; Osipov, A.V.; Ziganshin, R.H.; D'Hoedt, D.; Bertrand, D.; Sixma, T.K.; Smit, A.B.; Tsetlin, V.I.

    2009-01-01

    α-Conotoxins interact with nicotinic acetylcholine receptors (nAChRs) and acetylcholine-binding proteins (AChBPs) at the sites for agonists/competitive antagonists. α-Conotoxins blocking muscle-type or α7 nAChRs compete with α-bungarotoxin. However, α-conotoxin ImII, a close homolog of the α7

  1. Topological dispositions of lysine α380 and lysine γ486 in the acetylcholine receptor from Torpedo californica

    International Nuclear Information System (INIS)

    Dwyer, B.P.

    1991-01-01

    The locations have been determined, with respect to the plasma membrane, of lysine α380 and lysine γ486 in the α subunit and the γ subunit, respectively, of the nicotinic acetylcholine receptor from Torpedo californica. Immunoadsorbents were constructed that recognize the carboxy terminus of the peptide GVKYIAE released by proteolytic digestion from positions 378-384 in the amino acid sequence of the α subunit of the acetylcholine receptor and the carboxy terminus of the peptide KYVP released by proteolytic digestion from positions 486-489 in the amino acid sequence of the γ subunit. They were used to isolate these peptides from proteolytic digests of polypeptides from the acetylcholine receptor. Sealed vesicles containing the native acetylcholine receptor were labeled with pyridoxal phosphate and sodium [ 3 H]-borohydride. The effect of saponin on the incorporation of pyridoxamine phosphate into lysine α380 and lysine γ486 from the acetylcholine receptor in these vesicles was assessed with the immunoadsorbents. The conclusions that follow from these results are that lysine α380 is on the inside surface of a vesicle and lysine γ486 is on the outside surface. Because a majority (85%) of the total binding sites for α-bungarotoxin bind the toxin in the absence of saponin, the majority of the vesicles are right side out with the inside of the vesicle corresponding to the cytoplasmic surface and the outside of the vesicle corresponding to the extracytoplasmic, synaptic surface. Because lysine α380 and lysine γ486 lie on opposite sides of the membrane, a membrane-spanning segment must be located between the two positions occupied by these two amino acids in the common sequence of a polypeptide of the acetylcholine receptor

  2. Critical Evaluation of Acetylcholine Determination in Rat Brain Microdialysates using Ion-Pair Liquid Chromatography with Amperometric Detection

    Directory of Open Access Journals (Sweden)

    Yvette Michotte

    2008-08-01

    Full Text Available Liquid chromatography with amperometric detection remains the most widely used method for acetylcholine quantification in microdialysis samples. Separation of acetylcholine from choline and other matrix components on a microbore chromatographic column (1 mm internal diameter, conversion of acetylcholine in an immobilized enzyme reactor and detection of the produced hydrogen peroxide on a horseradish peroxidase redox polymer coated glassy carbon electrode, achieves sufficient sensitivity for acetylcholine quantification in rat brain microdialysates. However, a thourough validation within the concentration range required for this application has not been carried out before. Furthermore, a rapid degradation of the chromatographic columns and enzyme systems have been reported. In the present study an ion-pair liquid chromatography assay with amperometric detection was validated and its long-term stability evaluated. Working at pH 6.5 dramatically increased chromatographic stability without a loss in sensitivity compared to higher pH values. The lower limit of quantification of the method was 0.3 nM. At this concentration the repeatability was 15.7%, the inter-day precision 8.7% and the accuracy 103.6%. The chromatographic column was stable over 4 months, the immobilized enzyme reactor up to 2-3 months and the enzyme coating of the amperometric detector up to 1-2 months. The concentration of acetylcholine in 30 μl microdialysates obtained under basal conditions from the hippocampus of freely moving rats was 0.40 ± 0.12 nM (mean ± SD, n = 30. The present method is therefore suitable for acetylcholine determination in rat brain microdialysates.

  3. Cellular Localization and Trafficking of the Human ABCG1 Transporter

    Science.gov (United States)

    Neufeld, Edward B.; O’Brien, Katherine; Walts, Avram D.; Stonik, John A.; Demosky, Steven J.; Malide, Daniela; Combs, Christian A.; Remaley, Alan T.

    2014-01-01

    We have developed a suitable heterologous cell expression system to study the localization, trafficking, and site(s) of function of the human ABCG1 transporter. Increased plasma membrane (PM) and late endosomal (LE) cholesterol generated by ABCG1 was removed by lipoproteins and liposomes, but not apoA-I. Delivery of ABCG1 to the PM and LE was required for ABCG1-mediated cellular cholesterol efflux. ABCG1 LEs frequently contacted the PM, providing a collisional mechanism for transfer of ABCG1-mobilized cholesterol, similar to ABCG1-mediated PM cholesterol efflux to lipoproteins. ABCG1-mobilized LE cholesterol also trafficked to the PM by a non-vesicular pathway. Transfer of ABCG1-mobilized cholesterol from the cytoplasmic face of LEs to the PM and concomitant removal of cholesterol from the outer leaflet of the PM bilayer by extracellular acceptors suggests that ABCG1 mobilizes cholesterol on both sides of the lipid bilayer for removal by acceptors. ABCG1 increased uptake of HDL into LEs, consistent with a potential ABCG1-mediated cholesterol efflux pathway involving HDL resecretion. Thus, ABCG1 at the PM mobilizes PM cholesterol and ABCG1 in LE/LYS generates mobile pools of cholesterol that can traffic by both vesicular and non-vesicular pathways to the PM where it can also be transferred to extracellular acceptors with a lipid surface. PMID:25405320

  4. Use of 1H-NMR to determine the distribution of lecithin between the micellar and vesicular phases in model bile

    NARCIS (Netherlands)

    Groen, A. K.; Goldhoorn, B. G.; Egbers, P. H.; Chamuleau, R. A.; Tytgat, G. N.; Bovée, W. M.

    1990-01-01

    Biliary cholesterol/phospholipid vesicles play an important role in the pathogenesis of gallstone disease. A prerequisite for the study of the lipid composition and stability of these vesicles is a reliable method to quantify the amount of vesicular lipid. In the present report we show that NMR can

  5. Effects of mutations in the VP2/VP4 cleavage site of Swine vesicular disease virus on RNA encapsidation and viral infectivity

    NARCIS (Netherlands)

    Rebel, J.M.J.; Leendertse, C.H.; Dekker, A.; Moormann, R.J.M.

    2003-01-01

    We studied VP0 cleavage of Swine vesicular disease virus (SVDV), a member of the Picornaviridae using a full-length cDNA copy of the Dutch SVDV isolate. The influences of mutations, introduced at the cleavage site of SVDV, on VP0 cleavage, RNA encapsidation and viral infection were studied. Double

  6. Vesicular trafficking of semaphorin 3A is activity-dependent and differs between axons and dendrites

    NARCIS (Netherlands)

    de Wit, Joris; Toonen, Ruud F; Verhaagen, J.; Verhage, Matthijs

    Secreted semaphorins act as guidance cues in the developing nervous system and may have additional functions in mature neurons. How semaphorins are transported and secreted by neurons is poorly understood. We find that endogenous semaphorin 3A (Sema3A) displays a punctate distribution in axons and

  7. A synthetic combinatorial strategy for developing a-conotoxin analogs as potent a7 nicotinic acetylcholine receptor antagonists

    DEFF Research Database (Denmark)

    Armishaw, Christopher J; Singh, Narender; Medina-Franco, Jose L

    2010-01-01

    alpha-Conotoxins are peptide neurotoxins isolated from venomous cone snails that display exquisite selectivity for different subtypes of nicotinic acetylcholine receptors (nAChR). They are valuable research tools that have profound implications in the discovery of new drugs for a myriad of neurop......alpha-Conotoxins are peptide neurotoxins isolated from venomous cone snails that display exquisite selectivity for different subtypes of nicotinic acetylcholine receptors (nAChR). They are valuable research tools that have profound implications in the discovery of new drugs for a myriad...

  8. Differential Regulation of Receptor Activation and Agonist Selectivity by Highly Conserved Tryptophans in the Nicotinic Acetylcholine Receptor Binding Site

    OpenAIRE

    Williams, Dustin K.; Stokes, Clare; Horenstein, Nicole A.; Papke, Roger L.

    2009-01-01

    We have shown previously that a highly conserved Tyr in the nicotinic acetylcholine receptor (nAChR) ligand-binding domain (LBD) (α7 Tyr188 or α4 Tyr195) differentially regulates the activity of acetylcholine (ACh) and the α7-selective agonist 3-(4-hydroxy,2-methoxybenzylidene)anabaseine (4OH-GTS-21) in α4β2 and α7 nAChR. In this study, we mutated two highly conserved LBD Trp residues in human α7 and α4β2 and expressed the receptors in Xenopus laevis oocytes. α7 Re...

  9. Nicotinic acetylcholine receptor polymorphism, smoking behavior, and tobacco-related cancer and lung and cardiovascular diseases: a cohort study

    DEFF Research Database (Denmark)

    Kaur-Knudsen, Diljit; Bojesen, Stig E; Tybjærg-Hansen, Anne

    2011-01-01

    We examined the associations between the nicotinic acetylcholine receptor polymorphism (rs1051730) on chromosome 15q25 marking the gene cluster CHRNA3-CHRNB4-CHRNA5, smoking behavior, and tobacco-related cancer and lung and cardiovascular diseases in the general population.......We examined the associations between the nicotinic acetylcholine receptor polymorphism (rs1051730) on chromosome 15q25 marking the gene cluster CHRNA3-CHRNB4-CHRNA5, smoking behavior, and tobacco-related cancer and lung and cardiovascular diseases in the general population....

  10. Quantitative multiplex assay for simultaneous detection and identification of Indiana and New Jersey serotypes of vesicular stomatitis virus

    DEFF Research Database (Denmark)

    Rasmussen, Thomas Bruun; Uttenthal, Åse; Fernandez, Jovita

    2005-01-01

    In order to establish a rapid and reliable system for the detection of vesicular stomatitis virus (VSV), we developed a quantitative reverse transcription-PCR assay for the detection, quantification, and differentiation of the major serotypes, VSV Indiana and VSV New Jersey, using a closed......-tube multiplex format. The detection system is based on the recently invented primer-probe energy transfer (PriProET) system. A region of the gene encoding the RNA-dependent RNA polymerase was amplified by using VSV-specific primers in the presence of two serotype-specific fluorescent probes. By incorporating...... probes. The limits of detection ware found to be less than 10 50% tissue culture infective doses/ml for both serotypes. The diagnostic value of the new method was tested with clinical materials from experimentally infected pigs, and it is concluded that the method is a powerful tool for the rapid...

  11. Reduction of bacterial growth by a vesicular-arbuscular mycorrhizal fungus in the rhizosphere of cucumber (Cucumis sativus L.)

    DEFF Research Database (Denmark)

    Christensen, H.; Jakobsen, I.

    1993-01-01

    Cucumber was grown in a partially sterilized sand-soil mixture with the vesicular-arbuscular mycorrhizal (VAM) fungus Glomus fasciculatum or left uninoculated. Fresh soil extract was places in polyvinyl chloride tubes without propagules of mycorrhizal fungi. Root tips and root segments...... and top of tubes, and of cocci with a diameter of 0.55-0.78 mum in the bulk soil in the center of tubes, were significantly reduced by VAM fungi. The extremely high bacterial biomass (1-7 mg C g-1 dry weight soil) was significant reduced by mycorrhizal colonization on root segments and in bulk soil...... biomass, and changed the spatial pattern of bacterial growth compared to non-mycorrhizal cucumbers. The [H-3]-thymidine incorporation was significantly higher on root tips in the top of tubes, and on root segments and bulk soil in the center of tubes on non-mycorrhizal plants compared to mycorrhizal...

  12. Calixarene-mediated liquid membrane transport of choline conjugates 3: The effect of handle variation on neurotransmitter transport.

    Science.gov (United States)

    Collins, James L; Fujii, Ayu; Roshandel, Sahar; To, Cuong-Alexander; Schramm, Michael P

    2017-07-01

    Upper rim phosphonic acid functionalized calix[4]arene affects selective transport of multiple molecular payloads through a liquid membrane. The secret is in the attachment of a receptor-complementary handle to the payload. We find that the trimethylammonium ethylene group present in choline is one of several general handles for the transport of drug and drug-like species. Herein we compare the effect of handle variation against the transport of serotonin and dopamine. We find that several ionizable amine termini handles are sufficient for transport and identify two ideal candidates. Their performance is significantly enhanced in HEPES buffered solutions. This inquiry completes a series of 3 studies aimed at optimization of this strategy. In completion a new approach towards synthetic receptor mediated selective small molecule transport has emerged; future work in vesicular and cellular systems will follow. Copyright © 2017 Elsevier Ltd. All rights reserved.

  13. Procaine rapidly inactivates acetylcholine receptors from Torpedo and competes with agonist for inhibition sites

    International Nuclear Information System (INIS)

    Forman, S.A.; Miller, K.W.

    1989-01-01

    The relationship between the high-affinity procaine channel inhibition site and the agonist self-inhibition site on acetylcholine receptors (AChRs) from Torpedo electroplaque was investigated by using rapid 86 Rb + quenched-flux assays at 4 degree C in native AChR-rich vesicles on which 50-60% of ACh activation sites were blocked with α-bungarotoxin (α-BTX). In the presence of channel-activating acetylcholine (ACh) concentrations alone, AChR undergoes one phase of inactivation in under a second. Addition of procaine produces two-phase inactivation similar to that seen with self-inhibiting ACh concentrations rapid inactivation complete in 30-75 ms is followed by fast desensitization at the same k d observed without procaine. The dependence of k r on [procaine] is consistent with a bimolecular association between procaine and its AChR site. Inhibition of AChR function by mixtures of procaine plus self-inhibiting concentrations of ACh or suberyldicholine was studied by reducing the level of α-BTX block in vesicles. The data support a mechanism where procaine binds preferentially to the open-channel AChR state, since no procaine-induced inactivation is observed without agonist and k r 's dependence on [ACh] in channel-activating range closely parallels that of 86 Rb + flux response to ACh

  14. cap alpha. -bungarotoxin binding properties of a central nervous system nicotinic acetylcholine receptor

    Energy Technology Data Exchange (ETDEWEB)

    Lukasiewicz, R J; Bennett, E L

    1978-01-01

    High-affinity, specific binding of radiolabeled ..cap alpha..-bungarotoxin to particulate fractions derived from rat brain shows saturability (B/sub max/ approx. = 37fmol/mg, K/sub D//sup app/ = 1.7 nM) and insensitivity to ionic strength, and is essentially irreversible (K/sub on/ = 5 x 10/sup 6/ min/sup -1/ x mol/sup -1/; K(displacement) = 1.9 x 10/sup -4/ min/sup -1/, tau/sub 1/2/ = 62 h). Subcellular distribution of specific sites is consistent with their location on synaptic junctional complex and post-synaptic membranes. These membrane-bound binding sites exhibit unique sensitivity to cholinergic ligands; pretreatment of membranes with cholinergic agonists (but not antagonists) induces transformation of ..cap alpha..-bungarotoxin binding sites to a high affinity form toward agonist. The effect is most marked for the natural agonist, acetylcholine. These results strongly support the notion that the entity under study is an authentic nicotinic acetylcholine receptor.

  15. Acetylcholine-induced inhibition of presynaptic calcium signals and transmitter release in the frog neuromuscular junction

    Directory of Open Access Journals (Sweden)

    Eduard Khaziev

    2016-12-01

    Full Text Available Acetylcholine (ACh, released from axonal terminals of motor neurones in neuromuscular junctions regulates the efficacy of neurotransmission through activation of presynaptic nicotinic and muscarinic autoreceptors. Receptor-mediated presynaptic regulation could reflect either direct action on exocytotic machinery or modulation of Ca2+ entry and resulting intra-terminal Ca2+ dynamics. We have measured free intra-terminal cytosolic Ca2+ ([Ca2+]i using Oregon-Green 488 microfluorimetry, in parallel with voltage-clamp recordings of spontaneous (mEPC and evoked (EPC postsynaptic currents in post-junctional skeletal muscle fibre. Activation of presynaptic muscarinic and nicotinic receptors with exogenous acetylcholine and its non-hydrolized analogue carbachol reduced amplitude of the intra-terminal [Ca2+]i transients and decreased quantal content (calculated by dividing the area under EPC curve by the area under mEPC curve. Pharmacological analysis revealed the role of muscarinic receptors of M2 subtype as well as d-tubocurarine-sensitive nicotinic receptor in presynaptic modulation of [Ca2+]i transients. Modulation of synaptic transmission efficacy by ACh receptors was completely eliminated by pharmacological inhibition of N-type Ca2+ channels. We conclude that ACh receptor-mediated reduction of Ca2+ entry into the nerve terminal through N-type Ca2+ channels represents one of possible mechanism of presynaptic modulation in frog neuromuscular junction.

  16. The Anti-Acetylcholine Receptor Antibody Test in Suspected Ocular Myasthenia Gravis

    Directory of Open Access Journals (Sweden)

    Jung Jin Lee

    2014-01-01

    Full Text Available Aim. To estimate the clinical significance of anti-acetylcholine receptor antibody (anti-AChR-Ab levels in suspected ocular myasthenia gravis. Methods. In total, 144 patients complaining of fluctuating diplopia and ptosis were evaluated for serum levels of anti-acetylcholine receptor antibody and their medical charts were retrospectively reviewed. Subjects were classified into three groups: variable diplopia only, ptosis only, and both variable diplopia and ptosis. We investigated serum anti-AChR-Ab titer levels and performed thyroid autoantibody tests. Results. Patients’ chief complaints were diplopia (N=103, ptosis (N=12, and their concurrence (N=29. Abnormal anti-AChR-Ab was observed in 21 of 144 patients (14.1%. Between the three groups, mean age, number of seropositive patients, and mean anti-AChR-Ab level were not significantly different (P=0.224, 0.073, and 0.062, resp.. Overall, 27.5% of patients had abnormal thyroid autoantibodies. Conclusion. The sensitivity of anti-AChR-Ab was 14.1% in suspected ocular myasthenia gravis and seropositivity in myasthenia gravis patients showed a high correlation with the presence of thyroid autoantibodies.

  17. Conformational changes in acetylcholine binding protein investigated by temperature accelerated molecular dynamics.

    Directory of Open Access Journals (Sweden)

    Zeynab Mohammad Hosseini Naveh

    Full Text Available Despite the large number of studies available on nicotinic acetylcholine receptors, a complete account of the mechanistic aspects of their gating transition in response to ligand binding still remains elusive. As a first step toward dissecting the transition mechanism by accelerated sampling techniques, we study the ligand-induced conformational changes of the acetylcholine binding protein (AChBP, a widely accepted model for the full receptor extracellular domain. Using unbiased Molecular Dynamics (MD and Temperature Accelerated Molecular Dynamics (TAMD simulations we investigate the AChBP transition between the apo and the agonist-bound state. In long standard MD simulations, both conformations of the native protein are stable, while the agonist-bound structure evolves toward the apo one if the orientation of few key sidechains in the orthosteric cavity is modified. Conversely, TAMD simulations initiated from the native conformations are able to produce the spontaneous transition. With respect to the modified conformations, TAMD accelerates the transition by at least a factor 10. The analysis of some specific residue-residue interactions points out that the transition mechanism is based on the disruption/formation of few key hydrogen bonds. Finally, while early events of ligand dissociation are observed already in standard MD, TAMD accelerates the ligand detachment and, at the highest TAMD effective temperature, it is able to produce a complete dissociation path in one AChBP subunit.

  18. Longitudinal study of Senecavirus a shedding in sows and piglets on a single United States farm during an outbreak of vesicular disease.

    Science.gov (United States)

    Tousignant, Steven J P; Bruner, Laura; Schwartz, Jake; Vannucci, Fabio; Rossow, Stephanie; Marthaler, Douglas G

    2017-08-31

    The study highlights the shedding pattern of Senecavirus A (SVA) during an outbreak of vesicular disease in a sow farm from the South-central Minnesota, USA. In this study, 34 individual, mixed parity sows with clinical signs of vesicular lesions and 30 individual piglets from 15 individual litters from sows with vesicular lesions were conveniently selected for individual, longitudinal sampling. Serum, tonsil, rectal, and vesicular swabs were collected on day1 post outbreak, and then again at 1, 2, 3, 4, 6, and 9 weeks post outbreak. Samples were tested at the University of Minnesota Veterinary Diagnostic Laboratory for SVA via Real Time Polymerase Chain Reaction (RT-PCR) RESULTS: In sows, vesicular lesions had the highest concentration of SVA, but had the shortest duration of detection lasting only 2 weeks. Viremia was detected for 1 week post outbreak, and quickly declined thereafter. SVA was detected at approximately the same frequency for both tonsil and rectal swabs with the highest percentage of SVA positive samples detected in the first 6 weeks post outbreak. In suckling piglets, viremia quickly declined 1 week post outbreak and was prevalent in low levels during the first week after weaning (4 weeks post outbreak) and was also detected in piglets that were co-mingled from a SVA negative sow farm. Similar to sows, SVA detection on rectal and tonsil swabs in piglets lasted approximately 6 weeks post outbreak. The study illustrates the variation of SVA shedding patterns in different sample types over a 9 week period in sows and piglets, and suggests the potential for viral spread between piglets at weaning.

  19. Mapping organelle motion reveals a vesicular conveyor belt spatially replenishing secretory vesicles in stimulated chromaffin cells.

    Science.gov (United States)

    Maucort, Guillaume; Kasula, Ravikiran; Papadopulos, Andreas; Nieminen, Timo A; Rubinsztein-Dunlop, Halina; Meunier, Frederic A

    2014-01-01

    How neurosecretory cells spatially adjust their secretory vesicle pools to replenish those that have fused and released their hormonal content is currently unknown. Here we designed a novel set of image analyses to map the probability of tracked organelles undergoing a specific type of movement (free, caged or directed). We then applied our analysis to time-lapse z-stack confocal imaging of secretory vesicles from bovine Chromaffin cells to map the global changes in vesicle motion and directionality occurring upon secretagogue stimulation. We report a defined region abutting the cortical actin network that actively transports secretory vesicles and is dissipated by actin and microtubule depolymerizing drugs. The directionality of this "conveyor belt" towards the cell surface is activated by stimulation. Actin and microtubule networks therefore cooperatively probe the microenvironment to transport secretory vesicles to the periphery, providing a mechanism whereby cells globally adjust their vesicle pools in response to secretagogue stimulation.

  20. Reserpine-induced Reduction in Norepinephrine Transporter Function Requires Catecholamine Storage Vesicles

    OpenAIRE

    Mandela, Prashant; Chandley, Michelle; Xu, Yao-Yu; Zhu, Meng-Yang; Ordway, Gregory A.

    2010-01-01

    Treatment of rats with reserpine, an inhibitor of the vesicular monoamine transporter (VMAT), depletes norepinephrine (NE) and regulates NE transporter (NET) expression. The present study examined the molecular mechanisms involved in regulation of the NET by reserpine using cultured cells. Exposure of rat PC12 cells to reserpine for a period as short as 5 min decreased [3H]NE uptake capacity, an effect characterized by a robust decrease in the Vmax of the transport of [3H]NE. As expected, res...

  1. Structure-Function Relationship of Transporters in the Glutamate–Glutamine Cycle of the Central Nervous System

    Directory of Open Access Journals (Sweden)

    Mariko Kato Hayashi

    2018-04-01

    Full Text Available Many kinds of transporters contribute to glutamatergic excitatory synaptic transmission. Glutamate is loaded into synaptic vesicles by vesicular glutamate transporters to be released from presynaptic terminals. After synaptic vesicle release, glutamate is taken up by neurons or astrocytes to terminate the signal and to prepare for the next signal. Glutamate transporters on the plasma membrane are responsible for transporting glutamate from extracellular fluid to cytoplasm. Glutamate taken up by astrocyte is converted to glutamine by glutamine synthetase and transported back to neurons through glutamine transporters on the plasma membranes of the astrocytes and then on neurons. Glutamine is converted back to glutamate by glutaminase in the neuronal cytoplasm and then loaded into synaptic vesicles again. Here, the structures of glutamate transporters and glutamine transporters, their conformational changes, and how they use electrochemical gradients of various ions for substrate transport are summarized. Pharmacological regulations of these transporters are also discussed.

  2. Molecular determinants of subtype-selective efficacies of cytisine and the novel compound NS3861 at heteromeric nicotinic acetylcholine receptors

    DEFF Research Database (Denmark)

    Harpsøe, Kasper; Hald, Helle; Timmermann, Daniel B

    2013-01-01

    Deciphering which specific agonist-receptor interactions affect efficacy levels is of high importance, because this will ultimately aid in designing selective drugs. The novel compound NS3861 and cytisine are agonists of nicotinic acetylcholine receptors (nAChRs) and both bind with high affinity...

  3. Hippocampal α7 nicotinic acetylcholine receptor levels in patients with schizophrenia, bipolar disorder, or major depressive disorder

    DEFF Research Database (Denmark)

    Thomsen, Morten Skøtt; Weyn, Annelies; Mikkelsen, Jens D

    2011-01-01

    The α7 nicotinic acetylcholine receptor (nAChR) is involved in cognitive function and synaptic plasticity. Consequently, changes in α7 nAChR function have been implicated in a variety of mental disorders, especially schizophrenia. However, there is little knowledge regarding the levels of the α7 n...

  4. Carbamoylcholine analogs as nicotinic acetylcholine receptor agonists--structural modifications of 3-(dimethylamino)butyl dimethylcarbamate (DMABC)

    DEFF Research Database (Denmark)

    Hansen, Camilla Petrycer; Jensen, Anders Asbjørn; Balle, Thomas

    2009-01-01

    Compounds based on the 3-(dimethylamino)butyl dimethylcarbamate (DMABC) scaffold were synthesized and pharmacologically characterized at the alpha(4)beta(2), alpha(3)beta(4,) alpha(4)beta(4) and alpha(7) neuronal nicotinic acetylcholine receptors (nAChRs). The carbamate functionality and a small...

  5. The Actions of Piperidine Alkaloids at Fetal Muscle-Type and Autonomic-Type Nicotinic Acetylcholine Receptors

    Science.gov (United States)

    Piperidine alkaloids are found in many species of plants including Conium maculatum, Nicotiana spp., and Lupinus spp. A pharmacodynamic comparison was made of the alkaloids ammodendrine, anabasine, anabaseine, and coniine in; SH-SY5Y cells which express autonomic-type nicotinic acetylcholine recept...

  6. NMR Structure and Action on Nicotinic Acetylcholine Receptors of Water-soluble Domain of Human LYNX1

    Czech Academy of Sciences Publication Activity Database

    Lyukmanova, E. N.; Shenkarev, Z. O.; Shulepko, M. A.; Mineev, K. S.; D´Hoedt, D.; Kasheverov, I. E.; Filkin, S. Yu.; Krivolapova, A. P.; Janíčková, Helena; Doležal, Vladimír; Dolgikh, D. A.; Arseniev, A. S.; Bertrand, D.; Tsetlin, V.I.; Kirpichnikov, M. P.

    2011-01-01

    Roč. 286, č. 12 (2011), s. 10618-10627 ISSN 0021-9258 R&D Projects: GA ČR(CZ) GA305/09/0681 Institutional research plan: CEZ:AV0Z50110509 Keywords : NMR structure * nicotinic acetylcholine receptor * water-soluble domain Subject RIV: FH - Neurology Impact factor: 4.773, year: 2011

  7. Utrophin abundance is reduced at neuromuscular junctions of patients with both inherited and acquired acetylcholine receptor deficiencies

    NARCIS (Netherlands)

    Slater, CR; Young, C; Wood, SJ; Bewick, GS; Anderson, LVB; Baxter, P; Fawcett, PRW; Roberts, M; Jacobson, L; Kuks, J; Vincent, A; NewsomDavis, J

    Congenital myasthenic syndromes are a heterogenous group of conditions in which muscle weakness resulting from impaired neuromuscular transmission is often present from infancy. One form of congenital myasthenic syndrome is due to a reduction of the number of acetylcholine receptors (AChRs) at the

  8. An allosteric enhancer of M4muscarinic acetylcholine receptor function inhibits behavioral and neurochemical effects of cocaine

    DEFF Research Database (Denmark)

    Dencker, Ditte; Weikop, Pia; Sørensen, Gunnar

    2012-01-01

    The mesostriatal dopamine system plays a key role in mediating the reinforcing effects of psychostimulant drugs like cocaine. The muscarinic M4 acetylcholine receptor subtype is centrally involved in the regulation of dopamine release in striatal areas. Consequently, striatal M4 receptors could...

  9. Determination of anti-acetylcholine receptor antibodies in myasthenic patients by use of time-resolved fluorescence

    Czech Academy of Sciences Publication Activity Database

    Říčný, Jan; Šimková, L.; Vincent, A.

    2002-01-01

    Roč. 48, č. 3 (2002), s. 549-554 ISSN 0009-9147 R&D Projects: GA MZd NF4646 Institutional research plan: CEZ:AV0Z5011922 Keywords : nicotinic acetylcholine receptor * time-resolved fluorescence method * myasthenia gravis Subject RIV: FR - Pharmacology ; Medidal Chemistry Impact factor: 4.788, year: 2002

  10. Intravenously administered oxotremorine and atropine, in doses known to affect pain threshold, affect the intraspinal release of acetylcholine in rats

    DEFF Research Database (Denmark)

    Abelson, Klas S P; Höglund, A Urban

    2002-01-01

    muscarinic agonists and antagonists modify nociceptive threshold by affecting intraspinal release of acetylcholine (ACh). Catheters were inserted into the femoral vein in rats maintained on isoflurane anaesthesia for administration of oxotremorine (10-300 microg/kg) and atropine (0.1, 10, 5000 microg...

  11. Activation and desensitization of peripheral muscle and neuronal nicotinic acetylcholine receptors by selected, naturally-occurring pyridine alkaloids

    Science.gov (United States)

    Teratogenic alkaloids can cause developmental defects due to inhibition of fetal movement that results from desensitization of fetal muscletype nicotinic acetylcholine receptors (nAChRs). We investigated the ability of two known teratogens, the piperidinyl-pyridine anabasine and its 1,2-dehydropiper...

  12. Allosteric modulation by persistent binding of xanomeline of the interaction of competitive ligands with the M1 muscarinic acetylcholine receptor

    Czech Academy of Sciences Publication Activity Database

    Jakubík, Jan; Tuček, Stanislav; El-Fakahany, E. E.

    2002-01-01

    Roč. 301, č. 3 (2002), s. 1033-1041 ISSN 0022-3565 R&D Projects: GA ČR GP305/01/D119 Grant - others:NIH(US) NS25743 Institutional research plan: CEZ:AV0Z5011922 Keywords : xanomeline * M(1) muscarinic acetylcholine receptor Subject RIV: FH - Neurology Impact factor: 3.991, year: 2002

  13. Cholinesterase inhibition and acetylcholine accumulation following intracerebral administration of paraoxon in rats

    International Nuclear Information System (INIS)

    Ray, A.; Liu, J.; Karanth, S.; Gao, Y.; Brimijoin, S.; Pope, C.

    2009-01-01

    We evaluated the inhibition of striatal cholinesterase activity following intracerebral administration of paraoxon assaying activity either in tissue homogenates ex vivo or by substrate hydrolysis in situ. Artificial cerebrospinal fluid (aCSF) or paraoxon in aCSF was infused unilaterally (0.5 μl/min for 2 h) and ipsilateral and contralateral striata were harvested for ChE assay ex vivo. High paraoxon concentrations were needed to inhibit ipsilateral striatal cholinesterase activity (no inhibition at < 0.1 mM; 27% at 0.1 mM; 79% at 1 mM paraoxon). With 3 mM paraoxon infusion, substantial ChE inhibition was also noted in contralateral striatum. ChE histochemistry generally confirmed these concentration- and side-dependent effects. Microdialysates collected for up to 4 h after paraoxon infusion inhibited ChE activity when added to striatal homogenate, suggesting prolonged efflux of paraoxon. Since paraoxon efflux could complicate acetylcholine analysis, we evaluated the effects of paraoxon (0, 0.03, 0.1, 1, 10 or 100 μM, 1.5 μl/min for 45 min) administered by reverse dialysis through a microdialysis probe. ChE activity was then monitored in situ by perfusing the colorimetric substrate acetylthiocholine through the same probe and measuring product (thiocholine) in dialysates. Concentration-dependent inhibition was noted but reached a plateau of about 70% at 1 μM and higher concentrations. Striatal acetylcholine was below the detection limit at all times with 0.1 μM paraoxon but was transiently elevated (0.5-1.5 h) with 10 μM paraoxon. In vivo paraoxon (0.4 mg/kg, sc) in adult rats elicited about 90% striatal ChE inhibition measured ex vivo, but only about 10% inhibition measured in situ. Histochemical analyses revealed intense AChE and glial fibrillary acidic protein staining near the cannula track, suggesting proliferation of inflammatory cells/glia. The findings suggest that ex vivo and in situ cholinesterase assays can provide very different views into enzyme

  14. Acetylcholine-induced vasodilation in the uterine vascular bed of pregnant rats with adriamycin-induced nephrosis.

    Science.gov (United States)

    Yousif, Mariam H; Adeagbo, Ayotunde S; Kadavil, Elizabeth A; Chandrasekhar, Bindu; Oriowo, Mabayoje A

    2002-01-01

    This project was designed to study endothelium-dependent vasodilation in the uterine vascular bed during experimentally induced preeclampsia in rats. Uterine vascular beds were isolated from non-pregnant and pregnant rats with or without treatment with adriamycin (ADR) and perfused with physiological solution. Thereafter, vasodilator responses to acetylcholine were recorded. RECORDS: Pregnant ADR-treated rats displayed symptoms of preeclampsia including hypertension and proteinuria. Blood pressure was 110.0 +/- 4.7 mm Hg (n = 5) in control pregnant rats and 136.0 +/- 5.3 mm Hg (n = 5) in ADR-treated pregnant rats, and urinary protein concentrations were 0.35 mg/ml (n = 5) and 13.2 +/- 3.6 mg/ml (n = 9), respectively. Both blood pressure and proteinuria values were significantly (p acetylcholine-induced dose-dependent vasodilator responses in the vascular beds were not significantly different between the pregnant and nonpregnant rats. Although acetylcholine-induced vasodilation was significantly reduced by N omega-nitro-L-arginine methyl ester hydrochloride (L-NAME) in both groups, the residual response to acetylcholine was not affected by indomethacin, suggesting that prostanoids were not involved in this response. The L-NAME-resistant component, endothelium-derived hyperpolarizing factor (EDHF), was greater in ADR-treated uterine beds than in those of the controls, indicating a significant contribution from EDHF in these vessels. In the presence of an elevated external potassium ion concentration, acetylcholine produced similar vasodilator responses, indicating that the release of nitric oxide was not impaired. These results indicate that endothelium-dependent vasodilation was not impaired in this model of preeclampsia.

  15. Expression of nicotinic acetylcholine receptors on human B-lymphoma cells

    Directory of Open Access Journals (Sweden)

    Skok M. V.

    2009-12-01

    Full Text Available Aim. To find a correlation between the level of nicotinic acetylcholine receptor (nAChR expression and B lymphocyte differentiation or activation state. Methods. Expression of nAChRs in the REH, Ramos and Daudi cell lines was studied by flow cytometry using nAChR subunit-specific antibodies; cell proliferation was studied by MTT test. Results. It is shown that the level of 42/4 and 7 nAChRs expression increased along with B lymphocyte differentiation (Ramos > REH and activation (Daudi > > Ramos and depended on the antigen-specific receptor expression. The nAChR stimulation/blockade did not influence the intensity of cell proliferation.

  16. Steroids induce acetylcholine receptors on cultured human muscle: Implications for myasthenia gravis

    International Nuclear Information System (INIS)

    Kaplan, I.; Blakely, B.T.; Pavlath, G.K.; Travis, M.; Blau, H.M.

    1990-01-01

    Antibodies to the acetylcholine receptor (AChR), which are diagnostic of the human autoimmune disease myasthenia gravis, block AChR function and increase the rate of AChR degradation leading to impaired neuromuscular transmission. Steroids are frequently used to alleviate symptoms of muscle fatigue and weakness in patients with myasthenia gravis because of their well-documented immunosuppressive effects. The authors show here that the steroid dexamethasone significantly increases total surface AChRs on cultured human muscle exposed to myasthenia gravis sera. The results suggest that the clinical improvement observed in myasthenic patients treated with steroids is due not only to an effect on the immune system but also a direct effect on muscle. They propose that the identification and development of pharmacologic agents that augment receptors and other proteins that are reduced by human genetic or autoimmune disease will have broad therapeutic applications

  17. Differential expression of muscarinic acetylcholine receptor subtypes in Jurkat cells and their signaling.

    Science.gov (United States)

    Alea, Mileidys Perez; Borroto-Escuela, Dasiel O; Romero-Fernandez, Wilber; Fuxe, Kjell; Garriga, Pere

    2011-08-15

    Muscarinic acetylcholine receptors expression and signaling in the human Jurkat T cell line were investigated. Semiquantitative real-time PCR and radioligand binding studies, using a wide set of antagonist compounds, showed the co-existence of M(3), M(4), and M(5) subtypes. Stimulation of these subpopulations caused a concentration and time- dependent activation of second messengers and ERK signaling pathways, with a major contribution of the M(3) subtype in a G(q/11)-mediated response. In addition, we found that T-cell stimulation leads to increased expression of M(3) and M(5) both at transcriptional and protein levels in a PLC/PKCθ dependent manner. Our data clarifies the functional role of AChR subtypes in Jurkat cells and pave the way to future studies on the potential cross-talk among these subpopulations and their regulation of T lymphocytes immune function. Copyright © 2011 Elsevier B.V. All rights reserved.

  18. The Nicotinic Acetylcholine Receptor as a Target for Antidepressant Drug Development

    Directory of Open Access Journals (Sweden)

    Noah S. Philip

    2012-01-01

    Full Text Available An important new area of antidepressant drug development involves targeting the nicotinic acetylcholine receptor (nAChR. This receptor, which is distributed widely in regions of the brain associated with depression, is also implicated in other important processes that are relevant to depression, such as stress and inflammation. The two classes of drugs that target nAChRs can be broadly divided into mecamylamine- and cytisine-based compounds. These drugs probably exert their effects via antagonism at α4β2 nAChRs, and strong preclinical data support the antidepressant efficacy of both classes when used in conjunction with other primary antidepressants (e.g., monoamine reuptake inhibitors. Although clinical data remain limited, preliminary results in this area constitute a compelling argument for further evaluation of the nAChR as a target for future antidepressant drug development.

  19. Heteromeric α7β2 Nicotinic Acetylcholine Receptors in the Brain

    DEFF Research Database (Denmark)

    Wu, Jie; Liu, Qiang; Tang, Pei

    2016-01-01

    The α7 nicotinic acetylcholine receptor (α7 nAChR) is highly expressed in the brain, where it maintains various neuronal functions including (but not limited to) learning and memory. In addition, the protein expression levels of α7 nAChRs are altered in various brain disorders. The classic rule...... governing α7 nAChR assembly in the mammalian brain was that it was assembled from five α7 subunits to form a homomeric receptor pentamer. However, emerging evidence demonstrates the presence of heteromeric α7 nAChRs in heterologously expressed systems and naturally in brain neurons, where α7 subunits are co...... nAChR, which have provided new insights into the understanding of a novel target of cholinergic signaling....

  20. Molecular basis of the functional heterogeneity of the muscarinic acetylcholine receptor

    International Nuclear Information System (INIS)

    Numa, S.; Fukuda, K.; Kubo, T.; Maeda, A.; Akiba, I.; Bujo, H.; Nakai, J.; Mishina, M.; Higashida, H.

    1988-01-01

    The muscarinic acetylcholine receptor (mAChR) mediates a variety of cellular responses, including inhibition of adenylate cyclase, breakdown of phosphoinositides, and modulation of potassium channels, through the action of guanine-nucleotide-binding regulatory proteins (G proteins). The question then arises as to whether multiple mAChR species exist that are responsible for the various biochemical and physiological effects. In fact, pharmacologically distinguishable forms of the mAChR occur in different tissues and have been provisionally classified into M 1 (I), M 2 cardiac (II), and M 2 glandular (III) subtypes on the basis of their difference in apparent affinity for antagonists. Here, the authors have made attempts to understand the molecular basis of the functional heterogeneity of the mAChR, using recombinant DNA technology

  1. [3H]imidacloprid: synthesis of a candidate radioligand for the nicotinic acetylcholine receptor

    International Nuclear Information System (INIS)

    Latli, B.; Casida, J.E.

    1992-01-01

    Imidacloprid is an exceptionally potent insecticide known from physiological studies to act at the nicotinic acetylcholine receptor. To prepare [ 3 H]imidacloprid as a candidate radioligand, 6-chloronicotinoyl chloride was reduced with NaB 2 H 4 (in model studies) or NaB 3 H 4 in absolute ethanol to 2-chloro-5-pyridinylmethanol which was transformed to 2-chloro-5-chloromethylpyridine on refluxing with thionyl chloride. Coupling with 4,5-dihydro-N-nitro-1H-imidazol-2-amine then gave [ 2 H 2 ]imidacloprid incorporating about 95% of the deuterium or [ 3 H 2 ]imidacloprid (25 Ci/mmol) in 80% radiochemical yield. In studies not detailed here [ 3 H] imidacloprid was found to undergo high affinity, specific and saturable binding to a site in insect brain. (author)

  2. Design of ligands for the nicotinic acetylcholine receptors: the quest for selectivity.

    Science.gov (United States)

    Bunnelle, William H; Dart, Michael J; Schrimpf, Michael R

    2004-01-01

    In the last decade, nicotinic acetylcholine receptors (nAChRs) have emerged as important targets for drug discovery. The therapeutic potential of nicotinic agonists depends substantially on the ability to selectively activate certain receptor subtypes that mediate beneficial effects. The design of such compounds has proceeded in spite of a general shortage of data pertaining to subtype selectivity. Medicinal chemistry efforts have been guided principally by binding affinities to the alpha4beta2 and/or alpha7 subtypes, even though these are not predictive of agonist activity at either subtype. Nevertheless, a diverse family of nAChR ligands has been developed, and several analogs with promising therapeutic potential have now advanced to human clinical trials. This paper provides an overview of the structure-affinity relationships that continue to drive development of new nAChR ligands.

  3. Reactivity of Acetylcholine Esterase in inner Ear Maculae of Fish after Development at Hypergravity

    Science.gov (United States)

    Feucht, I.; Hilbig, R.; Anken, R.

    It has been shown earlier that the growth of inner ear otoliths of larval fish is (among other environmental factors) guided by the gravity vector. This guidance most probably is effected by the efferent vestibular system in the brainstem, because a transection of the nervus vestibularis has been shown to effect a cessation of the supply of calcium to the otoliths. The efferent innervation of fish inner ear maculae uses the synaptic transmitter acetylcholine (ACh). Therefore, we were - in order to further assess the role of the efferent system for otolith growth - prompted to determine ACh esterase-reactivity in the sensory epithelium of the utricle and the saccule (as well as in a non-gravity relevant brain region for control) in larval cichlid fish (Oreochromis mossambicus), which had been maintained at hypergravity during their development. The respective data will be communicated at the meeting. Acknowledgement: This work was financially supported by the German Aerospace Center (DLR) (FKZ: 50 WB 9997).

  4. Steroids induce acetylcholine receptors on cultured human muscle: Implications for myasthenia gravis

    Energy Technology Data Exchange (ETDEWEB)

    Kaplan, I.; Blakely, B.T.; Pavlath, G.K.; Travis, M.; Blau, H.M. (Stanford Univ. School of Medicine, CA (USA))

    1990-10-01

    Antibodies to the acetylcholine receptor (AChR), which are diagnostic of the human autoimmune disease myasthenia gravis, block AChR function and increase the rate of AChR degradation leading to impaired neuromuscular transmission. Steroids are frequently used to alleviate symptoms of muscle fatigue and weakness in patients with myasthenia gravis because of their well-documented immunosuppressive effects. The authors show here that the steroid dexamethasone significantly increases total surface AChRs on cultured human muscle exposed to myasthenia gravis sera. The results suggest that the clinical improvement observed in myasthenic patients treated with steroids is due not only to an effect on the immune system but also a direct effect on muscle. They propose that the identification and development of pharmacologic agents that augment receptors and other proteins that are reduced by human genetic or autoimmune disease will have broad therapeutic applications.

  5. Acetylcholine causes rooting in leaf explants of in vitro raised tomato (Lycopersicon esculentum Miller) seedlings.

    Science.gov (United States)

    Bamel, Kiran; Gupta, Shrish Chandra; Gupta, Rajendra

    2007-05-30

    The animal neurotransmitter acetylcholine (ACh) induces rooting and promotes secondary root formation in leaf explants of tomato (Lycopersicon esculentum Miller var. Pusa Ruby), cultured in vitro on Murashige and Skoog's medium. The roots originate from the midrib of leaf explants and resemble taproot. ACh at 10(-5) M was found to be the optimum over a wide range of effective concentrations between 10(-7) and 10(-3) M. The breakdown products, choline and acetate were ineffective even at 10(-3) M concentration. ACh appears to have a natural role in tomato rhizogenesis because exogenous application of neostigmine, an inhibitor of ACh hydrolysis, could mimic the effect of ACh. Neostigmine, if applied in combination with ACh, potentiated the ACh effect.

  6. Escherichia coli Protein Expression System for Acetylcholine Binding Proteins (AChBPs.

    Directory of Open Access Journals (Sweden)

    Nikita Abraham

    Full Text Available Nicotinic acetylcholine receptors (nAChR are ligand gated ion channels, identified as therapeutic targets for a range of human diseases. Drug design for nAChR related disorders is increasingly using structure-based approaches. Many of these structural insights for therapeutic lead development have been obtained from co-crystal structures of nAChR agonists and antagonists with the acetylcholine binding protein (AChBP. AChBP is a water soluble, structural and functional homolog of the extracellular, ligand-binding domain of nAChRs. Currently, AChBPs are recombinantly expressed in eukaryotic expression systems for structural and biophysical studies. Here, we report the establishment of an Escherichia coli (E. coli expression system that significantly reduces the cost and time of production compared to the existing expression systems. E. coli can efficiently express unglycosylated AChBP for crystallography and makes the expression of isotopically labelled forms feasible for NMR. We used a pHUE vector containing an N-terminal His-tagged ubiquitin fusion protein to facilitate AChBP expression in the soluble fractions, and thus avoid the need to recover protein from inclusion bodies. The purified protein yield obtained from the E. coli expression system is comparable to that obtained from existing AChBP expression systems. E. coli expressed AChBP bound nAChR agonists and antagonists with affinities matching those previously reported. Thus, the E. coli expression system significantly simplifies the expression and purification of functional AChBP for structural and biophysical studies.

  7. Characterization of prejunctional serotonin receptors modulating [3H]acetylcholine release in the human detrusor.

    Science.gov (United States)

    D'Agostino, Gianluigi; Condino, Anna M; Gallinari, Paola; Franceschetti, Gian P; Tonini, Marcello

    2006-01-01

    Bladder overactivity (OAB) is a chronic and debilitating lower urinary tract (LUT) disorder that affects millions of individuals worldwide. LUT symptoms associated with OAB, such as urgency and urinary incontinence, cause a hygienic and social concern to patients, but their current pharmacological treatment is largely inadequate due to the lack of uroselectivity. Although OAB etiology remains multifactorial and poorly understood, increasing evidence indicates that serotonin [5-hydroxytryptamine (5-HT)] is an endogenous substance involved in the control of micturition at central and peripheral sites. In this study, we demonstrated the presence of three distinct 5-HT receptors localized at parasympathetic nerve terminals of the human bladder by measuring electrically evoked tritiated acetylcholine release in isolated detrusor strips. These prejunctional receptors, involved in both positive and negative feedback mechanisms regulating cholinergic transmission, have been characterized by means of three highly selective 5-HT antagonists for 5-HT(4), 5-HT(7), and 5-HT(1A) receptors, namely GR113808A ([1-[2-[(-methylsulphonyl) amino] ethyl]4-piperinidyl]methyl1-methyl-1H-indole-3-carboxylate succinate), SB269970 [(R)-3-(2-(2-(4-methylpiperidin-1-yl)ethyl)pyrrolidine-1-sulfonyl)phenol hydrochloride], and WAY100635 [N-(2-(4-(2-methoxyphenyl)-1-piperazinyl)ethyl)-N-(2-pyridyl)-cyclohexane-carboxamide trichloride]. Under these conditions, we confirmed the facilitatory role of 5-HT(4) heteroreceptors on acetylcholine release and revealed for the first time the occurrence of 5-HT(7) and 5-HT(1A) heteroreceptors with a facilitatory and an inhibitory action, respectively. Our findings strengthen the novel concept for the use of recently patented selective 5-HT agonists and antagonists for the control of OAB dysfunctions associated with inflammatory conditions, although their therapeutic efficacy needs to be explored in the clinical setting.

  8. Acetylcholine muscarinic receptors and response to anti-cholinesterase therapy in patients with Alzheimer's disease

    International Nuclear Information System (INIS)

    Brown, Derek; Chisholm, Jennifer A.; Patterson, Jim; Wyper, David; Owens, Jonathan; Pimlott, Sally

    2003-01-01

    An acetylcholine deficit remains the most consistent neurotransmitter abnormality found in Alzheimer's disease and various therapeutic agents have been targeted at this. In this study we investigated the action of Donepezil, a cholinesterase inhibitor that has few side-effects. In particular we set out to investigate whether muscarinic acetylcholine receptor (mAChR) availability influences the response to this therapy. We used the novel single-photon emission tomography (SPET) tracer (R,R)[ 123 I]I-quinuclidinyl benzilate (R,R[ 123 I]I-QNB), which has high affinity for the M1 subtype of mAChR. Regional cerebral perfusion was also assessed using technetium-99m hexamethylpropylene amine oxime. We investigated 20 patients on Donepezil treatment and ten age-matched controls. The results showed a reduction in (R,R)[ 123 I]I-QNB binding in the caudal anterior cingulate in patients compared with controls and relatively high binding in the putamen and rostral anterior cingulate, suggesting a relative sparing of mAChR in these regions. The main finding of the study was that mAChR availability as assessed by (R,R)[ 123 I]I-QNB binding did not distinguish responders from non-responders. Interestingly, we found that the extent of cognitive improvement showed no positive correlation with (R,R)[ 123 I]I-QNB binding in any brain region but was inversely related to binding in the insular cortex. This suggests that, within the advised cognitive performance band for use of Donepezil, response is greater in those patients with evidence of a more marked cholinergic deficit. A larger study should investigate this. (orig.)

  9. Carbon monoxide-induced delayed amnesia, delayed neuronal death and change in acetylcholine concentration in mice

    International Nuclear Information System (INIS)

    Nabeshima, T.; Katoh, A.; Ishimaru, H.; Yoneda, Y.; Ogita, K.; Murase, K.; Ohtsuka, H.; Inari, K.; Fukuta, T.; Kameyama, T.

    1991-01-01

    We investigated the interrelationship of delayed amnesia, delayed neuronal death and changes in acetylcholine concentration induced by carbon monoxide (CO)-exposure in mice. In the test for retention of the passive avoidance task, amnesia was observed 5 and 7 days after CO-exposure when the mice were exposed to CO 1 day after training; in the case when the mice were exposed to CO 5 and 7 days before training, amnesia was also observed in a retention test given 1 day after training. The number of pyramidal cells in the hippocampal CA1 subfield was lower than that of the control 3, 5 and 7 days after CO-exposure. But the neurodegeneration in the parietal cortex, area 1, was not observed until 7 days after CO-exposure. The findings indicated that the amnesia and the neuronal death were produced after a delay when the mice were exposed to CO. In addition, the delayed amnesia was closely related to the delayed neuronal death in the hippocampal CA1 subfield. Moreover, [3H]glutamate and [3H]glycine binding sites did not change after CO-exposure but, 7 days after CO-exposure, the concentration of acetylcholine and the binding of [3H]quinuclidinyl benzilate in the frontal cortex and the striatum were found to have significantly changed, but those in the hippocampus did not show significant change. Therefore, we suggest that delayed amnesia induced by CO-exposure may result from delayed neuronal death in the hippocampal CA1 subfield and dysfunction in the acetylcholinergic neurons, in the frontal cortex, the striatum and/or the hippocampus

  10. Carbon monoxide-induced delayed amnesia, delayed neuronal death and change in acetylcholine concentration in mice

    Energy Technology Data Exchange (ETDEWEB)

    Nabeshima, T.; Katoh, A.; Ishimaru, H.; Yoneda, Y.; Ogita, K.; Murase, K.; Ohtsuka, H.; Inari, K.; Fukuta, T.; Kameyama, T. (Meijo Univ., Nagoya (Japan))

    1991-01-01

    We investigated the interrelationship of delayed amnesia, delayed neuronal death and changes in acetylcholine concentration induced by carbon monoxide (CO)-exposure in mice. In the test for retention of the passive avoidance task, amnesia was observed 5 and 7 days after CO-exposure when the mice were exposed to CO 1 day after training; in the case when the mice were exposed to CO 5 and 7 days before training, amnesia was also observed in a retention test given 1 day after training. The number of pyramidal cells in the hippocampal CA1 subfield was lower than that of the control 3, 5 and 7 days after CO-exposure. But the neurodegeneration in the parietal cortex, area 1, was not observed until 7 days after CO-exposure. The findings indicated that the amnesia and the neuronal death were produced after a delay when the mice were exposed to CO. In addition, the delayed amnesia was closely related to the delayed neuronal death in the hippocampal CA1 subfield. Moreover, (3H)glutamate and (3H)glycine binding sites did not change after CO-exposure but, 7 days after CO-exposure, the concentration of acetylcholine and the binding of (3H)quinuclidinyl benzilate in the frontal cortex and the striatum were found to have significantly changed, but those in the hippocampus did not show significant change. Therefore, we suggest that delayed amnesia induced by CO-exposure may result from delayed neuronal death in the hippocampal CA1 subfield and dysfunction in the acetylcholinergic neurons, in the frontal cortex, the striatum and/or the hippocampus.

  11. Effect of ethanol at clinically relevant concentrations on atrial inward rectifier potassium current sensitive to acetylcholine.

    Science.gov (United States)

    Bébarová, Markéta; Matejovič, Peter; Pásek, Michal; Hořáková, Zuzana; Hošek, Jan; Šimurdová, Milena; Šimurda, Jiří

    2016-10-01

    Alcohol intoxication tends to induce arrhythmias, most often the atrial fibrillation. To elucidate arrhythmogenic mechanisms related to alcohol consumption, the effect of ethanol on main components of the ionic membrane current is investigated step by step. Considering limited knowledge, we aimed to examine the effect of clinically relevant concentrations of ethanol (0.8-80 mM) on acetylcholine-sensitive inward rectifier potassium current I K(Ach). Experiments were performed by the whole-cell patch clamp technique at 23 ± 1 °C on isolated rat and guinea-pig atrial myocytes, and on expressed human Kir3.1/3.4 channels. Ethanol induced changes of I K(Ach) in the whole range of concentrations applied; the effect was not voltage dependent. The constitutively active component of I K(Ach) was significantly increased by ethanol with the maximum effect (an increase by ∼100 %) between 8 and 20 mM. The changes were comparable in rat and guinea-pig atrial myocytes and also in expressed human Kir3.1/3.4 channels (i.e., structural correlate of I K(Ach)). In the case of the acetylcholine-induced component of I K(Ach), a dual ethanol effect was apparent with a striking heterogeneity of changes in individual cells. The effect correlated with the current magnitude in control: the current was increased by eth-anol in the cells showing small current in control and vice versa. The average effect peaked at 20 mM ethanol (an increase of the current by ∼20 %). Observed changes of action potential duration agreed well with the voltage clamp data. Ethanol significantly affected both components of I K(Ach) even in concentrations corresponding to light alcohol consumption.

  12. Adenosine receptors and muscarinic receptors cooperate in acetylcholine release modulation in the neuromuscular synapse.

    Science.gov (United States)

    Santafe, M M; Priego, M; Obis, T; Garcia, N; Tomàs, M; Lanuza, M A; Tomàs, J

    2015-07-01

    Adenosine receptors (ARs) are present in the motor terminals at the mouse neuromuscular junction. ARs and the presynaptic muscarinic acetylcholine receptors (mAChRs) share the functional control of the neuromuscular junction. We analysed their mutual interaction in transmitter release modulation. In electrophysiological experiments with unaltered synaptic transmission (muscles paralysed by blocking the voltage-dependent sodium channel of the muscle cells with μ-conotoxin GIIIB), we found that: (i) a collaborative action between different AR subtypes reduced synaptic depression at a moderate activity level (40 Hz); (ii) at high activity levels (100 Hz), endogenous adenosine production in the synaptic cleft was sufficient to reduce depression through A1 -type receptors (A1 Rs) and A2 A-type receptors (A2 A Rs); (iii) when the non-metabolizable 2-chloroadenosine (CADO) agonist was used, both the quantal content and depression were reduced; (iv) the protective effect of CADO on depression was mediated by A1 Rs, whereas A2 A Rs seemed to modulate A1 Rs; (v) ARs and mAChRs absolutely depended upon each other for the modulation of evoked and spontaneous acetylcholine release in basal conditions and in experimental conditions with CADO stimulation; (vi) the purinergic and muscarinic mechanisms cooperated in the control of depression by sharing a common pathway although the purinergic control was more powerful than the muscarinic control; and (vii) the imbalance of the ARs created by using subtype-selective and non-selective inhibitory and stimulatory agents uncoupled protein kinase C from evoked transmitter release. In summary, ARs (A1 Rs, A2 A Rs) and mAChRs (M1 , M2 ) cooperated in the control of activity-dependent synaptic depression and may share a common protein kinase C pathway. © 2015 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

  13. Activation of muscarinic acetylcholine receptors elicits pigment granule dispersion in retinal pigment epithelium isolated from bluegill.

    Science.gov (United States)

    González, Alfredo; Crittenden, Elizabeth L; García, Dana M

    2004-07-13

    In fish, melanin pigment granules in the retinal pigment epithelium disperse into apical projections as part of the suite of responses the eye makes to bright light conditions. This pigment granule dispersion serves to reduce photobleaching and occurs in response to neurochemicals secreted by the retina. Previous work has shown that acetylcholine may be involved in inducing light-adaptive pigment dispersion. Acetylcholine receptors are of two main types, nicotinic and muscarinic. Muscarinic receptors are in the G-protein coupled receptor superfamily, and five different muscarinic receptors have been molecularly cloned in human. These receptors are coupled to adenylyl cyclase, calcium mobilization and ion channel activation. To determine the receptor pathway involved in eliciting pigment granule migration, we isolated retinal pigment epithelium from bluegill and subjected it to a battery of cholinergic agents. The general cholinergic agonist carbachol induces pigment granule dispersion in isolated retinal pigment epithelium. Carbachol-induced pigment granule dispersion is blocked by the muscarinic antagonist atropine, by the M1 antagonist pirenzepine, and by the M3 antagonist 4-DAMP. Pigment granule dispersion was also induced by the M1 agonist 4-[N-(4-chlorophenyl) carbamoyloxy]-4-pent-2-ammonium iodide. In contrast the M2 antagonist AF-DX 116 and the M4 antagonist tropicamide failed to block carbachol-induced dispersion, and the M2 agonist arecaidine but-2-ynyl ester tosylate failed to elicit dispersion. Our results suggest that carbachol-mediated pigment granule dispersion occurs through the activation of Modd muscarinic receptors, which in other systems couple to phosphoinositide hydrolysis and elevation of intracellular calcium. This conclusion must be corroborated by molecular studies, but suggests Ca2+-dependent pathways may be involved in light-adaptive pigment dispersion.

  14. Activation of muscarinic acetylcholine receptors elicits pigment granule dispersion in retinal pigment epithelium isolated from bluegill

    Directory of Open Access Journals (Sweden)

    Crittenden Elizabeth L

    2004-07-01

    Full Text Available Abstract Background In fish, melanin pigment granules in the retinal pigment epithelium disperse into apical projections as part of the suite of responses the eye makes to bright light conditions. This pigment granule dispersion serves to reduce photobleaching and occurs in response to neurochemicals secreted by the retina. Previous work has shown that acetylcholine may be involved in inducing light-adaptive pigment dispersion. Acetylcholine receptors are of two main types, nicotinic and muscarinic. Muscarinic receptors are in the G-protein coupled receptor superfamily, and five different muscarinic receptors have been molecularly cloned in human. These receptors are coupled to adenylyl cyclase, calcium mobilization and ion channel activation. To determine the receptor pathway involved in eliciting pigment granule migration, we isolated retinal pigment epithelium from bluegill and subjected it to a battery of cholinergic agents. Results The general cholinergic agonist carbachol induces pigment granule dispersion in isolated retinal pigment epithelium. Carbachol-induced pigment granule dispersion is blocked by the muscarinic antagonist atropine, by the M1 antagonist pirenzepine, and by the M3 antagonist 4-DAMP. Pigment granule dispersion was also induced by the M1 agonist 4-[N-(4-chlorophenyl carbamoyloxy]-4-pent-2-ammonium iodide. In contrast the M2 antagonist AF-DX 116 and the M4 antagonist tropicamide failed to block carbachol-induced dispersion, and the M2 agonist arecaidine but-2-ynyl ester tosylate failed to elicit dispersion. Conclusions Our results suggest that carbachol-mediated pigment granule dispersion occurs through the activation of Modd muscarinic receptors, which in other systems couple to phosphoinositide hydrolysis and elevation of intracellular calcium. This conclusion must be corroborated by molecular studies, but suggests Ca2+-dependent pathways may be involved in light-adaptive pigment dispersion.

  15. Neuronal nicotinic acetylcholine receptors: Common molecular substrates of nicotine and alcohol dependence

    Directory of Open Access Journals (Sweden)

    Linzy M. Hendrickson

    2013-04-01

    Full Text Available Alcohol and nicotine are often co-abused. As many as 80-95% of alcoholics are also smokers, suggesting that ethanol and nicotine, the primary addictive component of tobacco smoke, may functionally interact in the central nervous system and/or share a common mechanism of action. While nicotine initiates dependence by binding to and activating neuronal nicotinic acetylcholine receptors (nAChRs, ligand-gated cation channels normally activated by endogenous acetylcholine (ACh, ethanol is much less specific with the ability to modulate multiple gene products including those encoding voltage-gated ion channels, and excitatory/inhibitory neurotransmitter receptors. However, emerging data indicate that ethanol interacts with nAChRs, both directly and indirectly, in the mesocorticolimbic dopaminergic (DAergic reward circuitry to affect brain reward systems. Like nicotine, ethanol activates DAergic neurons of the ventral tegmental area (VTA which project to the nucleus accumbens (NAc. Blockade of VTA nAChRs reduces ethanol-mediated activation of DAergic neurons, NAc DA release, consumption, and operant responding for ethanol in rodents. Thus, ethanol may increase ACh release into the VTA driving activation of DAergic neurons through nAChRs. In addition, ethanol potentiates distinct nAChR subtype responses to ACh and nicotine in vitro and in DAergic neurons. The smoking cessation therapeutic and nAChR partial agonist, varenicline, reduces alcohol consumption in heavy drinking smokers and rodent models of alcohol consumption. Finally, single nucleotide polymorphisms in nAChR subunit genes are associated with alcohol dependence phenotypes and smoking behaviors in human populations. Together, results from preclinical, clinical, and genetic studies indicate that nAChRs may have an inherent role in the abusive properties of ethanol, as well as in nicotine and alcohol co-dependence.

  16. Modulation of release of [3H]acetylcholine in the major pelvic ganglion of the rat.

    Science.gov (United States)

    Somogyi, G T; de Groat, W C

    1993-06-01

    Cholinergic modulation of [3H]acetylcholine release evoked by electrical stimulation was studied in the rat major pelvic ganglion, which was prelabeled with [3H]choline. Acetylcholine (ACh) release was independent of the frequency of stimulation; 0.3 Hz produced the same volley output as 10 Hz. Tetrodotoxin (1 microM) or omission of Ca2+ from the medium abolished ACh release. The M1 receptor agonist (4-hydroxy-2-butynyl)-1-trimethylammonium m-chlorocarbanilate chloride (McN-A 343, 50 microM) increased release (by 136%), whereas the M2 muscarinic agonist oxotremorine (1 microM) decreased ACh release (by 22%). The muscarinic antagonists, atropine (1 microM) or pirenzepine (M1 selective, 1 microM), did not change ACh release. However, pirenzepine (1 microM) blocked the facilitatory effect of McN-A 343, and atropine (1 microM) blocked the inhibitory effect of oxotremorine. The cholinesterase inhibitor physostigmine (1-5 microM), the nicotinic agonist 1,1-dimethyl-4-phenylpiperazinium (DMPP, 10 microM), and the nicotinic antagonist D-tubocurarine (50 microM) did not change ACh release. 4-Aminopyridine, a K+ channel blocker, significantly increased the release (by 146%). Seven days after decentralization of the major pelvic ganglion, the evoked release of ACh was abolished. It is concluded that release of ACh occurs from the preganglionic nerve terminals rather than from the cholinergic cell bodies and is not modulated by actions of endogenous ACh on either muscarinic or nicotinic autoreceptors. These data confirm and extend previous electrophysiological findings indicating that synapses in the major pelvic ganglion have primarily a relay function.

  17. Modes of Action, Resistance and Toxicity of Insecticides Targeting Nicotinic Acetylcholine Receptors.

    Science.gov (United States)

    Ihara, Makoto; Buckingham, Steven D; Matsuda, Kazuhiko; Sattelle, David B

    2017-01-01

    Nicotinic acetylcholine receptors (nAChRs) of insects play a key role in fast excitatory neurotransmission. Several classes of insecticides target insect nAChRs, which are composed of subunit members of a family of multiple subunit encoding genes. Alternative splicing and RNA A-to-I editing can add further to receptor diversity. Native and recombinant receptors have been explored as sites of insecticide action using radioligands, electrophysiology and site-directed mutagenesis. We have reviewed the properties of native and recombinant insect nAChRs, the challenges of functional recombinant insect nAChR expression, nAChR interactions with ligands acting at orthosteric and allosteric sites and in particular their interactions with insecticides. Actions on insect nAChRs of cartap, neonicotinoids, spinosyns, sulfoxamines, butenolides and mesoionic insecticides are reviewed and current knowledge of their modes of action are addressed. Mutations that add to our understanding of insecticide action and those leading to resistance are discussed. Co-crystallisation of neonicotinoids with the acetylcholine binding protein (AChBP), a surrogate for the nAChR ligand binding domain, has proved instructive. Toxicity issues relating to insecticides targeting nAChRs are also considered. An overview of insecticide classes targeting insect nAChRs has enhanced our understanding of these important receptors and their insecticide binding sites. However, the subunit composition of native nAChRs remains poorly understood and functional expression still presents difficulties. These topics together with improved understanding of the precise sites of insecticide actions on insect nAChRs will be the subject of future research. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  18. Super agonist actions of clothianidin and related compounds on the SAD beta 2 nicotinic acetylcholine receptor expressed in Xenopus laevis oocytes.

    Science.gov (United States)

    Ihara, Makoto; Matsuda, Kazuhiko; Shimomura, Masaru; Sattelle, David B; Komai, Koichiro

    2004-03-01

    To compare the actions of clothianidin, a neonicotinoid acting on insect nicotinic acetylcholine receptors, and related compounds with that of imidacloprid, the compounds were tested on the Drosophila SAD-chicken beta2 nicotinic acetylcholine receptor expressed in Xenopus laevis oocytes using two-electrode voltage-clamp electrophysiology. The maximum response of the SAD beta 2 nicotinic receptor to clothianidin was larger than that observed for acetylcholine. Ring breakage of the imidazolidine ring of imidacloprid resulting in the generation of a guanidine group was critical for this super agonist action.

  19. Discovery of isoxazole analogues of sazetidine-A as selective α4β2-nicotinic acetylcholine receptor partial agonists for the treatment of depression.

    Science.gov (United States)

    Liu, Jianhua; Yu, Li-Fang; Eaton, J Brek; Caldarone, Barbara; Cavino, Katie; Ruiz, Christina; Terry, Matthew; Fedolak, Allison; Wang, Daguang; Ghavami, Afshin; Lowe, David A; Brunner, Dani; Lukas, Ronald J; Kozikowski, Alan P

    2011-10-27

    Depression, a common neurological condition, is one of the leading causes of disability and suicide worldwide. Standard treatment, targeting monoamine transporters selective for the neurotransmitters serotonin and noradrenaline, is not able to help many patients that are poor responders. This study advances the development of sazetidine-A analogues that interact with α4β2 nicotinic acetylcholine receptors (nAChRs) as partial agonists and that possess favorable antidepressant profiles. The resulting compounds that are highly selective for the α4β2 subtype of nAChR over α3β4-nAChRs are partial agonists at the α4β2 subtype and have excellent antidepressant behavioral profiles as measured by the mouse forced swim test. Preliminary absorption, distribution, metabolism, excretion, and toxicity (ADMET) studies for one promising ligand revealed an excellent plasma protein binding (PPB) profile, low CYP450-related metabolism, and low cardiovascular toxicity, suggesting it is a promising lead as well as a drug candidate to be advanced through the drug discovery pipeline.

  20. A review of reagents for fluorescence microscopy of cellular compartments and structures, part I: BacMam labeling and reagents for vesicular structures.

    Science.gov (United States)

    Dolman, Nick J; Kilgore, Jason A; Davidson, Michael W

    2013-07-01

    Fluorescent labeling of vesicular structures in cultured cells, particularly for live cells, can be challenging for a number of reasons. The first challenge is to identify a reagent that will be specific enough where some structures have a number of potential reagents and others very few options. The emergence of BacMam constructs has allowed more easy-to-use choices. Presented here is a discussion of BacMam constructs as well as a review of commercially-available reagents for labeling vesicular structures in cells, including endosomes, peroxisomes, lysosomes, and autophagosomes, complete with a featured reagent for each structure, recommended protocol, troubleshooting guide, and example image. © 2013 by John Wiley & Sons, Inc.

  1. Integrated femtosecond stimulated Raman scattering and two-photon fluorescence imaging of subcellular lipid and vesicular structures

    Science.gov (United States)

    Li, Xuesong; Lam, Wen Jiun; Cao, Zhe; Hao, Yan; Sun, Qiqi; He, Sicong; Mak, Ho Yi; Qu, Jianan Y.

    2015-11-01

    The primary goal of this study is to demonstrate that stimulated Raman scattering (SRS) as a new imaging modality can be integrated into a femtosecond (fs) nonlinear optical (NLO) microscope system. The fs sources of high pulse peak power are routinely used in multimodal nonlinear microscopy to enable efficient excitation of multiple NLO signals. However, with fs excitations, the SRS imaging of subcellular lipid and vesicular structures encounters significant interference from proteins due to poor spectral resolution and a lack of chemical specificity, respectively. We developed a unique NLO microscope of fs excitation that enables rapid acquisition of SRS and multiple two-photon excited fluorescence (TPEF) signals. In the in vivo imaging of transgenic C. elegans animals, we discovered that by cross-filtering false positive lipid signals based on the TPEF signals from tryptophan-bearing endogenous proteins and lysosome-related organelles, the imaging system produced highly accurate assignment of SRS signals to lipid. Furthermore, we demonstrated that the multimodal NLO microscope system could sequentially image lipid structure/content and organelles, such as mitochondria, lysosomes, and the endoplasmic reticulum, which are intricately linked to lipid metabolism.

  2. The Role of Teak Leaves (Tectona grandis), Rhizobium, and Vesicular-Arbuscular Mycorrhizae on Improving Soil Structure and Soil Nutrition

    Science.gov (United States)

    Yuliani; Rahayu, Y. S.

    2018-01-01

    Calcium is the largest mineral in calcareous soils. High levels of calcium carbonate lead to phosphate deposition. Nutrient deficiencies in calcareous soil (mainly Phosphate and Nitrogen) resulted only certain crops with a wide range of tolerances that can grow. Meanwhile, dynamics nutrient in calcareous soils also depend on the topography and decomposition of the litter in the growing vegetation. The purpose of this study was to describe the pattern of nutrient enhancement and soil-texture structures on calcareous soils after littering the teak leaves, Rhizobium and Vesicular Arbuscular Mycorrhiza. The research parameters were the concentration of N, P, K; C/N ratio, humid acid content, and soil structure, which measured at days 30, 60, and 85 of soil decomposition process. The results showed that at days 30, the texture and structure of the soil tend to be stable (porosity 31.2, DMR 1.93, moisture content 0.36, sandy clay) while at days 85 has been very stable (porosity 49.8; Water content 0.28, sandy clay). While C and N organic, N and K concentration at days 30 showed low value (C organic 1.03, N 0.12, K 0.49, C / N ratio 9). This condition is almost unchanged at days 85. While the P value shows very high value (60.53) at days 30 although after 60 days the P content showed a decrease.

  3. Dynamics of melanoma tumor therapy with vesicular stomatitis virus: explaining the variability in outcomes using mathematical modeling.

    Science.gov (United States)

    Rommelfanger, D M; Offord, C P; Dev, J; Bajzer, Z; Vile, R G; Dingli, D

    2012-05-01

    Tumor selective, replication competent viruses are being tested for cancer gene therapy. This approach introduces a new therapeutic paradigm due to potential replication of the therapeutic agent and induction of a tumor-specific immune response. However, the experimental outcomes are quite variable, even when studies utilize highly inbred strains of mice and the same cell line and virus. Recognizing that virotherapy is an exercise in population dynamics, we utilize mathematical modeling to understand the variable outcomes observed when B16ova malignant melanoma tumors are treated with vesicular stomatitis virus in syngeneic, fully immunocompetent mice. We show how variability in the initial tumor size and the actual amount of virus delivered to the tumor have critical roles on the outcome of therapy. Virotherapy works best when tumors are small, and a robust innate immune response can lead to superior tumor control. Strategies that reduce tumor burden without suppressing the immune response and methods that maximize the amount of virus delivered to the tumor should optimize tumor control in this model system.

  4. Activation of Nrf2 Signaling Augments Vesicular Stomatitis Virus Oncolysis via Autophagy-Driven Suppression of Antiviral Immunity.

    Science.gov (United States)

    Olagnier, David; Lababidi, Rassin R; Hadj, Samar Bel; Sze, Alexandre; Liu, Yiliu; Naidu, Sharadha Dayalan; Ferrari, Matteo; Jiang, Yuan; Chiang, Cindy; Beljanski, Vladimir; Goulet, Marie-Line; Knatko, Elena V; Dinkova-Kostova, Albena T; Hiscott, John; Lin, Rongtuan

    2017-08-02

    Oncolytic viruses (OVs) offer a promising therapeutic approach to treat multiple types of cancer. In this study, we show that the manipulation of the antioxidant network via transcription factor Nrf2 augments vesicular stomatitis virus Δ51 (VSVΔ51) replication and sensitizes cancer cells to viral oncolysis. Activation of Nrf2 signaling by the antioxidant compound sulforaphane (SFN) leads to enhanced VSVΔ51 spread in OV-resistant cancer cells and improves the therapeutic outcome in different murine syngeneic and xenograft tumor models. Chemoresistant A549 lung cancer cells that display constitutive dominant hyperactivation of Nrf2 signaling are particularly vulnerable to VSVΔ51 oncolysis. Mechanistically, enhanced Nrf2 signaling stimulated viral replication in cancer cells and disrupted the type I IFN response via increased autophagy. This study reveals a previously unappreciated role for Nrf2 in the regulation of autophagy and the innate antiviral response that complements the therapeutic potential of VSV-directed oncolysis against multiple types of OV-resistant or chemoresistant cancer. Copyright © 2017 The Author(s). Published by Elsevier Inc. All rights reserved.

  5. Oncolytic Vesicular Stomatitis Virus as a Viro-Immunotherapy: Defeating Cancer with a “Hammer” and “Anvil”

    Directory of Open Access Journals (Sweden)

    Michael Karl Melzer

    2017-02-01

    Full Text Available Oncolytic viruses have gained much attention in recent years, due, not only to their ability to selectively replicate in and lyse tumor cells, but to their potential to stimulate antitumor immune responses directed against the tumor. Vesicular stomatitis virus (VSV, a negative-strand RNA virus, is under intense development as an oncolytic virus due to a variety of favorable properties, including its rapid replication kinetics, inherent tumor specificity, and its potential to elicit a broad range of immunomodulatory responses to break immune tolerance in the tumor microenvironment. Based on this powerful platform, a multitude of strategies have been applied to further improve the immune-stimulating potential of VSV and synergize these responses with the direct oncolytic effect. These strategies include: 1. modification of endogenous virus genes to stimulate interferon induction; 2. virus-mediated expression of cytokines or immune-stimulatory molecules to enhance anti-tumor immune responses; 3. vaccination approaches to stimulate adaptive immune responses against a tumor antigen; 4. combination with adoptive immune cell therapy for potentially synergistic therapeutic responses. A summary of these approaches will be presented in this review.

  6. Immunogenicity and efficacy of immunodeficiency virus-like particles pseudotyped with the G protein of vesicular stomatitis virus

    International Nuclear Information System (INIS)

    Kuate, Seraphin; Stahl-Hennig, Christiane; Stoiber, Heribert; Nchinda, Godwin; Floto, Anja; Franz, Monika; Sauermann, Ulrike; Bredl, Simon; Deml, Ludwig; Ignatius, Ralf; Norley, Steve; Racz, Paul; Tenner-Racz, Klara; Steinman, Ralph M.; Wagner, Ralf; Uberla, Klaus

    2006-01-01

    Vaccination with exogenous antigens such as recombinant viral proteins, immunodeficiency virus-derived whole inactivated virus particles, or virus-like particles (VLP) has generally failed to provide sufficient protection in animal models for AIDS. Pseudotyping VLPs with the vesicular stomatitis virus G protein (VSV-G), which is known to mediate entry into dendritic cells, might allow more efficient stimulation of immune responses. Therefore, we pseudotyped noninfectious immunodeficiency virus-like particles with VSV-G and carried out a preliminary screen of their immunogenicity and vaccination efficacy. Incorporation of VSV-G into HIV-1 VLPs led to hundred-fold higher antibody titers to HIV-1 Gag and enhancement of T cell responses in mice. Repeated vaccination of rhesus monkeys for 65 weeks with VSV-G pseudotyped simian immunodeficiency virus (SIV)-like particles (VLP[G]) provided initial evidence for efficient suppression of viral load after mucosal challenge with the SIVmac239 virus. Challenge of monkeys after a 28 week vaccination regimen with VLP[G] led to a reduction in peak viremia, but persistent suppression of viral load was not achieved. Due to limitations in the number of animals available for this study, improved efficacy of VSV-G pseudotyped VLPs in nonhuman primates could not be demonstrated. However, mouse experiments revealed that pseudotyping of VLPs with fusion-competent VSV-G clearly improves their immunogenicity. Additional strategies, particularly adjuvants, should be considered to provide greater protection against a challenge with pathogenic immunodeficiency virus

  7. Long-Term Single-Dose Efficacy of a Vesicular Stomatitis Virus-Based Andes Virus Vaccine in Syrian Hamsters

    Directory of Open Access Journals (Sweden)

    Joseph Prescott

    2014-01-01

    Full Text Available Andes virus (ANDV is highly pathogenic in humans and is the primary etiologic agent of hantavirus cardiopulmonary syndrome (HCPS in South America. Case-fatality rates are as high as 50% and there are no approved vaccines or specific therapies for infection. Our laboratory has recently developed a replication-competent recombinant vesicular stomatitis virus (VSV-based vaccine that expressed the glycoproteins of Andes virus in place of the native VSV glycoprotein (G. This vaccine is highly efficacious in the Syrian hamster model of HCPS when given 28 days before challenge with ANDV, or when given around the time of challenge (peri-exposure, and even protects when administered post-exposure. Herein, we sought to test the durability of the immune response to a single dose of this vaccine in Syrian hamsters. This vaccine was efficacious in hamsters challenged intranasally with ANDV 6 months after vaccination (p = 0.025, but animals were not significantly protected following 1 year of vaccination (p = 0.090. The decrease in protection correlated with a reduction of measurable neutralizing antibody responses, and suggests that a more robust vaccination schedule might be required to provide long-term immunity.

  8. Immunogenicity in African Green Monkeys of M Protein Mutant Vesicular Stomatitis Virus Vectors and Contribution of Vector-Encoded Flagellin

    Directory of Open Access Journals (Sweden)

    Marlena M. Westcott

    2018-03-01

    Full Text Available Recombinant vesicular stomatitis virus (VSV is a promising platform for vaccine development. M51R VSV, an attenuated, M protein mutant strain, is an effective inducer of Type I interferon and dendritic cell (DC maturation, which are desirable properties to exploit for vaccine design. We have previously evaluated M51R VSV (M51R and M51R VSV that produces flagellin (M51R-F as vaccine vectors using murine models, and found that flagellin enhanced DC activation and VSV-specific antibody production after low-dose vaccination. In this report, the immunogenicity of M51R vectors and the adjuvant effect of virus-produced flagellin were evaluated in nonhuman primates following high-dose (108 pfu and low-dose (105 pfu vaccination. A single intramuscular vaccination of African green monkeys with M51R or M51R-F induced VSV-specific, dose-dependent humoral immune responses. Flagellin induced a significant increase in antibody production (IgM, IgG and neutralizing antibody at the low vaccination dose. A VSV-specific cellular response was detected at 6 weeks post-vaccination, but was neither dose-dependent nor enhanced by flagellin; similar numbers of VSV-specific, IFNγ-producing cells were detected in lymph node and spleen of all animals. These results indicate that virus-directed, intracellular flagellin production may improve VSV-based vaccines encoding heterologous antigens by lowering the dose required to achieve humoral immunity.

  9. The effect of vesicular-arbuscular mycorrhiza isolated from Syrian soil on alfalfa growth and nitrogen fixation in saline soil

    International Nuclear Information System (INIS)

    El Atrash, F

    2001-01-01

    The influence of vesicular - arbuscular Mycorrhiza fungi (VAM) on symbiotic fixation of N 2 n alfalfa plants has been observed. Beneficial effects of study the effect of VAM or phosphorous fertilization on alfalfa (Medicago sativa L,) yields, umber of nodules and N 2 fixation by N 15 isotope dilution at different salinity levels. This experiment was realized in green house conditions, using soil of 2.3 dsm -1 conductivity mixed with sand (5: 2V) for alfalfa plants growing at various levels of phosphorus, or infected by Mycorrhiza fungi. Different conductivities (13.18, 22.2, 28.8, 43.5 dsm -1 ) were applied on these treatment by increasing concentrations of Nacl, CaCl 2 and MgCl 2 and MgCl 2 by salinity soil irrigation. Ten days after planting, soil was enriched with 2 ppm of (NH 4 15 ) 2 SO 4 . Plant were grown under greenhouse condition for ten weeks. Our results confirmed that increased salinity reduced nitrogen - fixation and the number of nodules. The negative effect with increasing salinity was less in Mycorrhiza plants than in plants fertilized with various levels of phosphorus, and only the higher levels of salinity reduced significantly, the percentage of Mycorrhiza colonization, However, at all levels of salinity, VAM stimulated plant growth and nutrient uptake. (author)

  10. The Effect of Vesicular Arbuscular Mycorrhizal (VAM on Yield and Yield Components of Three Sorghum (Sorghum bicolor Cultivars

    Directory of Open Access Journals (Sweden)

    A. Mehraban

    2012-10-01

    Full Text Available To evaluate the influence of vesicular arbuscular mycorrhizal (VAM on yield and yield components of three sorghum cultivars, a factorial experiment based randomized complete block design with four replications was carried out in 2007, at the Agricultural Research Center of Zahak, Iran. The treatments were different mycorrhiza species in three levels: without mycorrhiza (M1, Glomus etanicatum (M2 and G. mosseae(M3 and three cultivars of sorghum: local cultivars (C1, KGS25 (C2 and KGS29 (C3. The results showed that all of the traits measured were increased by inoculation of cultivars with mycorrhiza. The highest plant height (165.1 cm, stem diameter (1.61 cm, flag leaf length (27.22 cm, flag leaf width (3.67 cm and ear width (5.00 cm was obtained by inoculation of seed with Glumus etanicatum, and highest ear length (19.21 cm, ear number (2.51, seed number per ear (10252.11, 1000-seed weight (17.56 g and grain yield (1967.32 kg/ha by using Glumus mossea. The highest leaf width and length belonged to local cultivar, and the highest seed yield to KGS 29 cultivar. However, differences of other traits among sorghum cultivars were not significant. Based on the experimental results it can be concluded that highest grain yield may be obtained by inoculating seeds of KGS 29 with Glumus mossea.

  11. Existing of Vesicular Arbuscular Mycorrhizal on The Corn Field Subjected by Long-term Organic and Inorganic Fertilizers

    Directory of Open Access Journals (Sweden)

    Sri Yusnaini

    2009-09-01

    Full Text Available The existing of vesicular arbuscular mycorrhizal fungi was determined in continuously cropping systems which had applied by organic and/or inorganic fertilizers for a long term (4 years application of corn and upland rice rotation. The experiment was established at Taman Bogo, Probolinggo sub-district, East Lampung district. The experiment were: control (without fertilizer, 20 Mg ha-1 chicken manure (CK, 20 Mg ha-1 green manure Glyricidium sp. (GM, 100 % inorganic fertilizers (IF (urea 300 kg ha-1, SP 36 200 kg ha-1, and KCl 100 kg ha-1, 50% CK + 50% IF, 50% GM + 50% IF, 75% CK + 25% IF, and 75% GM + 25% IF. Soil samples were taken at the first corn growing season (2001 and the eight corn growing season (2004. VAM fungi spores were examined by wet sieving methods. The results showed that application of chicken manure or its combination with inorganic fertilizers had higher VAM fungi spore. The dominant species of VAM fungi at all treatment was Glomus constrictum.

  12. Establishment of vesicular-arbuscular mycorrhizal fungi and other microorganisms on a beach replenishment site in Florida.

    Science.gov (United States)

    Sylvia, D M; Will, M E

    1988-02-01

    Beach replenishment is a widely used method of controlling coastal erosion. To reduce erosional losses from wind, beach grasses are often planted on the replenishment sands. However, there is little information on the microbial populations in this material that may affect plant establishment and growth. The objectives of this research were to document changes in the populations of vesicular-arbuscular mycorrhizal (VAM) fungi and other soil microorganisms in replenishment materials and to determine whether roots of transplanted beach grasses become colonized by beneficial microbes. The study was conducted over a 2-year period on a replenishment project in northeastern Florida. Three sampling locations were established at 1-km intervals along the beach. Each location consisted of three plots: an established dune, replenishment sand planted with Uniola paniculata and Panicum sp., and replenishment sand left unplanted. Fungal and bacterial populations increased rapidly in the rhizosphere of beach grasses in the planted plots. However, no bacteria were recovered that could fix significant amounts of N(2). The VAM fungi established slowly on the transplanted grasses. Even after two growing seasons, levels of root colonization and sporulation were significantly below those found in the established dune. There was a shift in the dominant VAM fungi found in the planted zone with respect to those in the established dunes. The most abundant species recovered from the established dunes were Glomus deserticola, followed by Acaulospora scrobiculata and Scutellospora weresubiae. The VAM fungi that colonized the planted zone most rapidly were Glomus globiferum, followed by G. deserticola and Glomus aggregatum.

  13. Matrix protein of vesicular stomatitis virus: a potent inhibitor of vascular endothelial growth factor and malignant ascites formation.

    Science.gov (United States)

    Zhou, Y; Wen, F; Zhang, P; Tang, R; Li, Q

    2013-03-01

    Malignant ascites is common in various types of cancers and is difficult to manage. Vascular endothelial growth factor (VEGF) has a pivotal role in malignant ascites. The matrix protein of vesicular stomatitis virus (VSVMP) has been shown to inhibit host gene expression and induce the apoptosis of cancer cells. The present study was designed to determine whether VSVMP suppresses the formation of ascites in ascites-producing peritoneal carcinomatosis. BALB/c female mice, 6-8 weeks old, bearing peritoneal tumors of H22 or MethA cells received an intraperitoneal administration of 50 μg VSVMP/250 μg liposome complexes, 50 μg empty plasmid/250 μg liposome complexes or 0.9% NaCl solution, respectively, every 2 days for 3 weeks. Administration of VSVMP resulted in a significant inhibition in ascites formation, improvement in health condition and prolonged survival of the treated mice. Decreased peritoneum osmolarity and reduced tumor vascularity coincided with dramatic reductions in the VEGF level in ascites fluid and plasma. Examination of floating tumor cells collected from the peritoneal wash revealed an apparently increased number of apoptotic cells and profound downregulation of VEGF mRNA in the VSVMP-treated mice. Our data indicate for the first time that in BALB/c mice bearing H22 or MethA cell peritoneal tumors, VSVMP may inhibit VEGF production and suppress angiogenesis, consequently abolishing ascites formation.

  14. A case of vesicular cutaneous lupus erythematosus in a Border collie successfully treated with topical tacrolimus and nicotinamide-tetracycline.

    Science.gov (United States)

    Lehner, Georg M; Linek, Monika

    2013-12-01

    Canine vesicular cutaneous lupus erythematosus (VCLE) is an autoimmune skin disease of the Shetland sheepdog and rough collie, which manifests as an erosive dermatitis of sparsely haired skin of the ventrum and concave pinnae. Reported treatment consists of immunosuppression with glucocorticoids alone or in combination with azathioprine, but successful treatment is unpredictable. To report on the treatment of VCLE in a Border collie dog with topical 0.1% tacrolimus and nicotinamide in combination with tetracycline. An 8-year-old male neutered Border collie was presented with multiple coalescing erosions on the ventral abdomen, groin and axillae and ulceration on the oral commissures. Clinical presentation, routine diagnostics, histology and immunohistochemistry were consistent with VCLE. Remission was achieved with topical 0.1% tacrolimus and combination therapy of nicotinamide and tetracycline. This dog responded well to treatment with topical 0.1% tacrolimus, nicotinamide-tetracycline and sun avoidance. Complete remission was achieved after 2.5 months, and the dog was lesion free during a 1 year follow-up period. © 2013 ESVD and ACVD.

  15. From the Cover: 7,8-Dihydroxyflavone Rescues Lead-Induced Impairment of Vesicular Release: A Novel Therapeutic Approach for Lead Intoxicated Children.

    Science.gov (United States)

    Zhang, Xiao-Lei; McGlothan, Jennifer L; Miry, Omid; Stansfield, Kirstie H; Loth, Meredith K; Stanton, Patric K; Guilarte, Tomás R

    2018-01-01

    Childhood lead (Pb2+) intoxication is a public health problem of global proportion. Lead exposure during development produces multiple effects on the central nervous system including impaired synapse formation, altered synaptic plasticity, and learning deficits. In primary hippocampal neurons in culture and hippocampal slices, Pb2+ exposure inhibits vesicular release and reduces the number of fast-releasing sites, an effect associated with Pb2+ inhibition of NMDA receptor-mediated trans-synaptic Brain-Derived Neurotrophic Factor (BDNF) signaling. The objective of this study was to determine if activation of TrkB, the cognate receptor for BDNF, would rescue Pb2+-induced impairments of vesicular release. Rats were chronically exposed to Pb2+ prenatally and postnatally until 50 days of age. This chronic Pb2+ exposure paradigm enhanced paired-pulse facilitation of synaptic potentials in Schaffer collateral-CA1 synapses in the hippocampus, a phenomenon indicative of reduced vesicular release probability. Decreased vesicular release probability was confirmed by both mean-variance analysis and direct 2-photon imaging of vesicular release from hippocampal slices of rats exposed to Pb2+in vivo. We also found a Pb2+-induced impairment of calcium influx in Schaffer collateral-CA1 synaptic terminals. Intraperitoneal injections of Pb2+ rats with the TrkB receptor agonist 7,8-dihydroxyflavone (5 mg/kg) for 14-15 days starting at postnatal day 35, reversed all Pb2+-induced impairments of presynaptic transmitter release at Schaffer collateral-CA1 synapses. This study demonstrates for the first time that in vivo pharmacological activation of TrkB receptors by small molecules such as 7,8-dihydroxyflavone can reverse long-term effects of chronic Pb2+ exposure on presynaptic terminals, pointing to TrkB receptor activation as a promising therapeutic intervention in Pb2+-intoxicated children. © The Author 2017. Published by Oxford University Press on behalf of the Society of

  16. Ultrasound effects on brain-targeting mannosylated liposomes: in vitro and blood-brain barrier transport investigations.

    Science.gov (United States)

    Zidan, Ahmed S; Aldawsari, Hibah

    2015-01-01

    Delivering drugs to intracerebral regions can be accomplished by improving the capacity of transport through blood-brain barrier. Using sertraline as model drug for brain targeting, the current study aimed at modifying its liposomal vesicles with mannopyranoside. Box-Behnken design was employed to statistically optimize the ultrasound parameters, namely ultrasound amplitude, time, and temperature, for maximum mannosylation capacity, sertraline entrapment, and surface charge while minimizing vesicular size. Moreover, in vitro blood-brain barrier transport model was established to assess the transendothelial capacity of the optimized mannosylated vesicles. Results showed a dependence of vesicular size, mannosylation capacity, and sertraline entrapment on cavitation and bubble implosion events that were related to ultrasound power amplitude, temperature. However, short ultrasound duration was required to achieve >90% mannosylation with nanosized vesicles (ultrasound parameters of 65°C, 27%, and 59 seconds for ultrasound temperature, amplitude, and time were elucidated to produce 81.1%, 46.6 nm, and 77.6% sertraline entrapment, vesicular size, and mannosylation capacity, respectively. Moreover, the transendothelial ability was significantly increased by 2.5-fold by mannosylation through binding with glucose transporters. Hence, mannosylated liposomes processed by ultrasound could be a promising approach for manufacturing and scale-up of brain-targeting liposomes.

  17. Neuroprotection of rat retinal ganglion cells mediated through alpha7 nicotinic acetylcholine receptors.

    Science.gov (United States)

    Iwamoto, K; Mata, D; Linn, D M; Linn, C L

    2013-05-01

    Glutamate-induced excitotoxicity is thought to play an important role in several neurodegenerative diseases in the central nervous system (CNS). In this study, neuroprotection against glutamate-induced excitotoxicity was analyzed using acetylcholine (ACh), nicotine and the α7 specific nicotinic acetylcholine receptor (α7 nAChR) agonist, N-[(3R)-1-azabicyclo[2.2.2]oct-3-yl]-4-chlorobenzamide hydrochloride (PNU-282987), in cultured adult rat retinal neurons. Adult Long Evans rat retinas were dissociated and retinal ganglion cells (RGCs) were isolated from all other retinal tissue using a two-step panning technique. Once isolated, RGCs were cultured under various pharmacological conditions to demonstrate excitotoxicity and neuroprotection against excitotoxicity. After 3 days, RGCs were immunostained with antibodies against the glycoprotein, Thy 1.1, counted and cell survival was assessed relative to control untreated conditions. 500 μM glutamate induced excitotoxicity in large and small RGCs in an adult rat dissociated culture. After 3 days in culture with glutamate, the cell survival of large RGCs decreased by an average of 48.16% while the cell survival of small RGCs decreased by an average of 42.03%. Using specific glutamate receptor agonists and antagonists, we provide evidence that the excitotoxic response was mediated through α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/kainic acid (KA) and N-methyl-d-aspartate (NMDA) glutamate receptors through an apoptotic mechanism. However, the excitotoxic effect of glutamate on all RGCs was eliminated if cells were cultured for an hour with 10 μM ACh, 100 μM nicotine or 100 nM of the α7 nAChR agonist, PNU-282987, before the glutamate insult. Inhibition studies using 10nM methyllycaconitine (MLA) or α-bungarotoxin (α-Bgt) supported the hypothesis that neuroprotection against glutamate-induced excitotoxicity on rat RGCs was mediated through α7 nAChRs. In immunocytochemical studies, double

  18. Simultaneous improvement in production of microalgal biodiesel and high-value alpha-linolenic acid by a single regulator acetylcholine

    OpenAIRE

    Parsaeimehr, Ali; Sun, Zhilan; Dou, Xiao; Chen, Yi-Feng

    2015-01-01

    Background Photoautotrophic microalgae are a promising avenue for sustained biodiesel production, but are compromised by low yields of biomass and lipids at present. We are developing a chemical approach to improve microalgal accumulation of feedstock lipids as well as high-value alpha-linolenic acid which in turn might provide a driving force for biodiesel production. Results We demonstrate the effectiveness of the small bioactive molecule ?acetylcholine? on accumulation of biomass, total li...

  19. Lipophilicity of acetylcholine and related ions examined by ion transfer voltammetry at a polarized room-temperature ionic liquid membrane

    Czech Academy of Sciences Publication Activity Database

    Langmaier, Jan; Záliš, Stanislav; Samec, Zdeněk

    2018-01-01

    Roč. 815, APR 2018 (2018), s. 183-188 ISSN 1572-6657 R&D Projects: GA ČR(CZ) GA15-03139S; GA ČR(CZ) GA17-05387S Institutional support: RVO:61388955 Keywords : acetylcholine * choline * ion transfer voltammetry Subject RIV: CG - Electrochemistry OBOR OECD: Electrochemistry (dry cells, batteries, fuel cells, corrosion metals, electrolysis) Impact factor: 3.012, year: 2016

  20. R+-methanandamide inhibits tracheal response to endogenously released acetylcholine via capsazepine-sensitive receptors.

    Science.gov (United States)

    Nieri, Paola; Martinotti, Enrica; Testai, Lara; Adinolfi, Barbara; Calderone, Vincenzo; Breschi, Maria Cristina

    2003-01-10

    The effects of cannabinoid drugs on the cholinergic response evoked by electrical field stimulation (0.2 ms pulse width, 20 V amplitude, 10 Hz, 7.5 s train duration) in guinea-pig tracheal preparations were investigated. The stable analogue of the endocannabinoid anandamide, R(+)-methanandamide (10(-7)-10(-4) M), produced a dose-dependent inhibition (up to 27+/-5% of control) of electrical field stimulation-mediated atropine-sensitive response. This effect was not blocked by the selective cannabinoid CB(1) receptor antagonist N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3 carboxamide hydrochloride (SR 141716A; 10(-6) M), and was not reproduced with the cannabinoid CB(1)/CB(2) receptor agonist R(+)-[2,3-dihydro-5-methyl-[(morpholinyl)methyl]pyrrolo [1,2,3-de]-1,4-benzoxazin-6-yl]-(1-naphthalenyl)methanone mesylate) (WIN 55,212-2; 10(-8)-10(-5) M) or the cannabinoid CB(2) receptor selective agonist 1-propyl-2-methyl-3-(1-naphthoyl)indole (JWH-015; 10(-8)-10(-5) M); it was, on the contrary, antagonized by the vanilloid antagonist 2-[2-(4-chlorophenyl)ethyl-amino-thiocarbonyl]-7,8-dihydroxy-2,3,4,5-tetrahydro-1H-2 benzazepine (capsazepine; 10(-6) M). At the postjunctional level, neither R(+)-methanandamide nor WIN 55,212-2 nor JWH-015 did affect tracheal contractions induced by exogenous acetylcholine (10(-6) M). An inhibitory vanilloid receptor-mediated effect on the cholinergic response evoked by electrical stimulation was confirmed with the vanilloid agonist capsaicin, at doses (3-6 x 10(-8) M) which poorly influenced the basal smooth muscle tone of trachea. In conclusion, our data indicate that in guinea-pig trachea (a) neither CB(1) nor CB(2) cannabinoid receptor-mediated modulation of acetylcholine release occurs; (b) vanilloid VR1-like receptors appear involved in R(+)-methanandamide inhibitory activity on the cholinergic response to electrical field stimulation.