Sample records for t-2 toxin ht-2

  1. Metabolism of HT-2 Toxin and T-2 Toxin in Oats

    PubMed Central

    Meng-Reiterer, Jacqueline; Bueschl, Christoph; Rechthaler, Justyna; Berthiller, Franz; Lemmens, Marc; Schuhmacher, Rainer

    2016-01-01

    The Fusarium mycotoxins HT-2 toxin (HT2) and T-2 toxin (T2) are frequent contaminants in oats. These toxins, but also their plant metabolites, may contribute to toxicological effects. This work describes the use of 13C-assisted liquid chromatography–high-resolution mass spectrometry for the first comprehensive study on the biotransformation of HT2 and T2 in oats. Using this approach, 16 HT2 and 17 T2 metabolites were annotated including novel glycosylated and hydroxylated forms of the toxins, hydrolysis products, and conjugates with acetic acid, putative malic acid, malonic acid, and ferulic acid. Further targeted quantitative analysis was performed to study toxin metabolism over time, as well as toxin and conjugate mobility within non-treated plant tissues. As a result, HT2-3-O-β-d-glucoside was identified as the major detoxification product of both parent toxins, which was rapidly formed (to an extent of 74% in HT2-treated and 48% in T2-treated oats within one day after treatment) and further metabolised. Mobility of the parent toxins appeared to be negligible, while HT2-3-O-β-d-glucoside was partly transported (up to approximately 4%) through panicle side branches and stem. Our findings demonstrate that the presented combination of untargeted and targeted analysis is well suited for the comprehensive elucidation of mycotoxin metabolism in plants. PMID:27929394

  2. Occurrence of Fusarium langsethiae and T-2 and HT-2 Toxins in Italian Malting Barley

    PubMed Central

    Morcia, Caterina; Tumino, Giorgio; Ghizzoni, Roberta; Badeck, Franz W.; Lattanzio, Veronica M.T.; Pascale, Michelangelo; Terzi, Valeria

    2016-01-01

    T-2 and HT-2 toxins are two of the most toxic members of type-A trichothecenes, produced by a number of Fusarium species. The occurrence of these mycotoxins was studied in barley samples during a survey carried out in the 2011–2014 growing seasons in climatically different regions in Italy. The percentage of samples found positive ranges from 22% to 53%, with values included between 26 and 787 μg/kg. The percentage of samples with a T-2 and HT-2 content above the EU indicative levels for barley of 200 μg/kg ranges from 2% to 19.6% in the 2011–2014 period. The fungal species responsible for the production of these toxins in 100% of positive samples has been identified as Fusarium langsethiae, a well-known producer of T-2 and HT-2 toxins. A positive correlation between the amount of F. langsethiae DNA and of the sum of T-2 and HT-2 toxins was found. This is the first report on the occurrence of F. langsethiae—and of its toxic metabolites T-2 and HT-2—in malting barley grown in Italy. PMID:27556490

  3. Occurrence of Fusarium langsethiae and T-2 and HT-2 Toxins in Italian Malting Barley.

    PubMed

    Morcia, Caterina; Tumino, Giorgio; Ghizzoni, Roberta; Badeck, Franz W; Lattanzio, Veronica M T; Pascale, Michelangelo; Terzi, Valeria

    2016-08-20

    T-2 and HT-2 toxins are two of the most toxic members of type-A trichothecenes, produced by a number of Fusarium species. The occurrence of these mycotoxins was studied in barley samples during a survey carried out in the 2011-2014 growing seasons in climatically different regions in Italy. The percentage of samples found positive ranges from 22% to 53%, with values included between 26 and 787 μg/kg. The percentage of samples with a T-2 and HT-2 content above the EU indicative levels for barley of 200 μg/kg ranges from 2% to 19.6% in the 2011-2014 period. The fungal species responsible for the production of these toxins in 100% of positive samples has been identified as Fusarium langsethiae, a well-known producer of T-2 and HT-2 toxins. A positive correlation between the amount of F. langsethiae DNA and of the sum of T-2 and HT-2 toxins was found. This is the first report on the occurrence of F. langsethiae-and of its toxic metabolites T-2 and HT-2-in malting barley grown in Italy.

  4. Comparison of T-2 Toxin and HT-2 Toxin Distributed in the Skeletal System with That in Other Tissues of Rats by Acute Toxicity Test.

    PubMed

    Yu, Fang Fang; Lin, Xia Lu; Yang, Lei; Liu, Huan; Wang, Xi; Fang, Hua; Lammi, ZMikko J; Guo, Xiong

    2017-11-01

    Twelve healthy rats were divided into the T-2 toxin group receiving gavage of 1 mg/kg T-2 toxin and the control group receiving gavage of normal saline. Total relative concentrations of T-2 toxin and HT-2 toxin in the skeletal system (thighbone, knee joints, and costal cartilage) were significantly higher than those in the heart, liver, and kidneys (P < 0.05). The relative concentrations of T-2 toxin and HT-2 toxin in the skeletal system (thighbone and costal cartilage) were also significantly higher than those in the heart, liver, and kidneys. The rats administered T-2 toxin showed rapid metabolism compared with that in rats administered HT-2 toxin, and the metabolic conversion rates in the different tissues were 68.20%-90.70%. Copyright © 2017 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.

  5. Comparison of Anorectic Potencies of the Trichothecenes T-2 Toxin, HT-2 Toxin and Satratoxin G to the Ipecac Alkaloid Emetine.

    PubMed

    Wu, Wenda; Zhou, Hui-Ren; Pan, Xiao; Pestka, James J

    Trichothecene mycotoxins, potent translational inhibitors that are associated with human food poisonings and damp-building illnesses, are of considerable concern to animal and human health. Food refusal is a hallmark of exposure of experimental animals to deoxynivalenol (DON) and other Type B trichothecenes but less is known about the anorectic effects of foodborne Type A trichothecenes (e.g., T-2 toxin, HT-2 toxin), airborne Type D trichothecenes (e.g. satratoxin G [SG]) or functionally analogous metabolites that impair protein synthesis. Here, we utilized a well-described mouse model of food intake to compare the anorectic potencies of T-2 toxin, HT-2 toxin, and SG to that of emetine, a medicinal alkaloid derived from ipecac that inhibits translation. Intraperitoneal (IP) administration with T-2 toxin, HT-2 toxin, emetine and SG evoked anorectic responses that occurred within 0.5 h that lasted up to 96, 96, 3 and 96 h, respectively, with lowest observed adverse effect levels (LOAELs) being 0.1, 0.1, 2.5 and 0.25 mg/kg BW, respectively. When delivered via natural routes of exposure, T-2 toxin, HT-2 toxin, emetine (oral) and SG (intranasal) induced anorectic responses that lasted up to 48, 48, 3 and 6 h, respectively with LOAELs being 0.1, 0.1, 0.25, and 0.5 mg/kg BW, respectively. All four compounds were generally much more potent than DON which was previously observed to have LOAELs of 1 and 2.5 mg/kg BW after IP and oral dosing, respectively. Taken together, these anorectic potency data will be valuable in discerning the relative risks from trichothecenes and other translational inhibitors of natural origin.

  6. Role of Peptide YY3-36 and Glucose-Dependent Insulinotropic Polypeptide in Anorexia Induction by Trichothecences T-2 Toxin, HT-2 Toxin, Diacetoxyscirpenol, and Neosolaniol.

    PubMed

    Zhang, Jie; Jia, Hui; Wang, Qingqing; Zhang, Yajie; Wu, Wenda; Zhang, Haibin

    2017-09-01

    Trichothecences, secondary metabolites produced by Fusarium, are serious health risks to humans and animals worldwide. Although type A trichothecence-induced food refusal has been observed, the mechanism underlying the anorexia caused by these compounds is not fully understood. In this study, we hypothesized that anorexia induced by type A trichothecenes, including T-2 toxin (T-2), HT-2 toxin (HT-2), diacetoxyscirpenol (DAS), and neosolaniol (NEO), in mice corresponds to the changes in the gut satiety hormones peptide YY3-36 (PYY3-36) and glucose-dependent insulinotropic polypeptide (GIP) in plasma. A well-characterized mouse food refusal model was used in this assay. Oral exposure to or intraperitoneal (ip) injection of 1 mg/kg bw T-2, HT-2, DAS, or NEO resulted in dramatically decreased food intake, and PYY3-36 and GIP concentrations were elevated accordingly. Specifically, the PYY3-36 and GIP concentrations peaked at 2 h following oral exposure to these 4 toxins individually, although the durations were not identical. After ip administration of T-2 or HT-2, PYY3-36 significantly increased within 6 h. However, no significant difference was found in the DAS and NEO groups. The GIP levels peaked within 2, 2, 0.5, and 0.5 h, respectively, and remained increased up to 6, 6, 2, and 6 h, respectively, following T-2, HT-2, DAS, or NEO ip exposure. The increase in GIP was greater than that of PYY3-36 after exposure to the 4 toxins using 2 administration routes. Together, these findings suggest that PYY3-36 and GIP play a role in T-2-, HT-2-, DAS-, and NEO-induced anorexia. © The Author 2017. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

  7. Gut satiety hormones cholecystokinin and glucagon-like Peptide-17-36 amide mediate anorexia induction by trichothecenes T-2 toxin, HT-2 toxin, diacetoxyscirpenol and neosolaniol.

    PubMed

    Zhang, Jie; Liu, Shengli; Zhang, Hua; Li, Yuanyuan; Wu, Wenda; Zhang, Haibin

    2017-11-15

    The food-borne trichothecene mycotoxins have been documented to cause human and animal food poisoning. Anorexia is a hallmark of the trichothecene mycotoxins-induced adverse effects. Type B trichothecenes have been previously demonstrated to elicit robust anorectic responses, and this response has been directly linked to secretion of the gut satiety hormones cholecystokinin (CCK) and glucagon-like peptide-1 7-36 amide (GLP-1). However, less is known about the anorectic effects and underlying mechanisms of the type A trichothecenes, including T-2 toxin (T-2), HT-2 toxin (HT-2), diacetoxyscirpenol (DAS), neosolaniol (NEO). The purpose of this study was to relate type A trichothecenes T-2, HT-2, DAS and NEO-induced anorectic response to changes plasma concentrations of CCK and GLP-1. Following both oral gavage and intraperitoneal (IP) administration of 1mg/kg bw T-2, HT-2, DAS and NEO evoked robust anorectic response and secretion of CCK and GLP-1. Elevations of plasma CCK markedly corresponded to anorexia induction by T-2, HT-2, DAS and NEO. Following oral exposure, plasma CCK was peaked at 6h, 6h, 2h, 2h and lasted up to 24h, 24h, > 6h, > 6h for T-2, HT-2, DAS and NEO, respectively. IP exposed to four toxins all induced elevation of CCK with peak point and duration at 6h and >24h, respectively. In contrast to CCK, GLP-1 was moderately elevated by these toxins. Following both oral and IP exposure, T-2 and HT-2 evoked plasma GLP-1 elevation with peak point and duration at 2h and 6h, respectively. Plasma GLP-1 was peaked at 2h and still increased at 6h for IP and oral administration with DAS and NEO, respectively. In conclusion, CCK plays a contributory role in anorexia induction but GLP-1 might play a lesser role in this response. Copyright © 2017 Elsevier Inc. All rights reserved.

  8. Determination of T-2 and HT-2 toxins from maize by direct analysis in real time mass spectrometry

    USDA-ARS?s Scientific Manuscript database

    Direct analysis in real time (DART) ionization coupled to mass spectrometry (MS) was used for the rapid quantitative analysis of T-2 toxin, and the related HT-2 toxin, extracted from corn. Sample preparation procedures and instrument parameters were optimized to obtain sensitive and accurate determi...

  9. Tracing the metabolism of HT-2 toxin and T-2 toxin in barley by isotope-assisted untargeted screening and quantitative LC-HRMS analysis

    USDA-ARS?s Scientific Manuscript database

    An extensive study of the metabolism of the type-A trichothecene mycotoxins HT-2 toxin and T-2 toxin in barley using liquid chromatography coupled to high resolution mass spectrometry (LC-HRMS) is reported. A recently developed untargeted approach based on stable isotopic labelling, LC-Orbitrap-MS a...

  10. Interactions between cyclodextrins and fluorescent T-2 and HT-2 toxin derivatives: a physico-chemical study

    USDA-ARS?s Scientific Manuscript database

    T-2 and HT-2 toxins are mycotoxins produced by several Fusarium species that are commonly found in various cereal grains, including oats, barley, wheat and maize. Intake estimates indicate that the presence of these mycotoxins in the diet can be of concern for public health.In this work, the inclusi...

  11. T-2 Toxin/HT-2 Toxin and Ochratoxin A ELISAs Development and In-House Validation in Food in Accordance with Commission Regulation (EU) No 519/2014

    PubMed Central

    Oplatowska-Stachowiak, Michalina; Sajic, Nermin; Haasnoot, Willem; Campbell, Katrina; Elliott, Christopher T.; Salden, Martin

    2017-01-01

    T-2 toxin/HT-2 toxin (T-2/HT-2) and ochratoxin A (OTA) are mycotoxins that can contaminate a variety of agricultural commodities. To protect consumers’ health, indicative limits for T-2/HT-2 and maximum limits for OTA have been set by the European Commission, requiring food business operators and controlling agencies to conduct routine checks for the presence of these harmful contaminants. Screening methods are increasingly used for monitoring purposes. Due to the demand for new and improved screening tools, two individual detection methods, T-2/HT-2 and OTA enzyme-linked immunosorbent assays (ELISAs), were developed in this study. The T-2/HT-2 ELISA was based on a T-2 monoclonal antibody with an IC50 (50% inhibitory concentration) of 0.28 ng/mL and 125% cross-reactivity with HT-2. As regards the OTA ELISA, a new sensitive monoclonal antibody specific to OTA with an IC50 of 0.13 ng/mL was produced. Both developed ELISA tests were then validated in agricultural commodities in accordance with the new performance criteria guidelines for the validation of screening methods for mycotoxins included in Commission Regulation (EU) No 519/2014. The T-2/HT-2 ELISA was demonstrated to be suitable for the detection of T-2/HT-2 in cereals and baby food at and above the screening target concentration (STC) of 12.5 μg/kg and 7.5 μg/kg, respectively. The OTA ELISA was shown to be applicable for the detection of OTA in cereals, coffee, cocoa and wine at and above the STC of 2 μg/kg, 2.5 μg/kg, 2.5 μg/kg and 0.4 ng/mL, respectively. The accuracy of both ELISAs was further confirmed by analysing proficiency test and reference samples. The developed methods can be used for sensitive and high-throughput screening for the presence of T-2/HT-2 and OTA in agricultural commodities. PMID:29189752

  12. T-2 Toxin/HT-2 Toxin and Ochratoxin A ELISAs Development and In-House Validation in Food in Accordance with the Commission Regulation (EU) No 519/2014.

    PubMed

    Oplatowska-Stachowiak, Michalina; Kleintjens, Tim; Sajic, Nermin; Haasnoot, Willem; Campbell, Katrina; Elliott, Christopher T; Salden, Martin

    2017-11-30

    T-2 toxin/HT-2 toxin (T-2/HT-2) and ochratoxin A (OTA) are mycotoxins that can contaminate a variety of agricultural commodities. To protect consumers' health, indicative limits for T-2/HT-2 and maximum limits for OTA have been set by the European Commission, requiring food business operators and controlling agencies to conduct routine checks for the presence of these harmful contaminants. Screening methods are increasingly used for monitoring purposes. Due to the demand for new and improved screening tools, two individual detection methods, T-2/HT-2 and OTA enzyme-linked immunosorbent assays (ELISAs), were developed in this study. The T-2/HT-2 ELISA was based on a T-2 monoclonal antibody with an IC 50 (50% inhibitory concentration) of 0.28 ng/mL and 125% cross-reactivity with HT-2. As regards the OTA ELISA, a new sensitive monoclonal antibody specific to OTA with an IC 50 of 0.13 ng/mL was produced. Both developed ELISA tests were then validated in agricultural commodities in accordance with the new performance criteria guidelines for the validation of screening methods for mycotoxins included in Commission Regulation (EU) No 519/2014. The T-2/HT-2 ELISA was demonstrated to be suitable for the detection of T-2/HT-2 in cereals and baby food at and above the screening target concentration (STC) of 12.5 μg/kg and 7.5 μg/kg, respectively. The OTA ELISA was shown to be applicable for the detection of OTA in cereals, coffee, cocoa and wine at and above the STC of 2 μg/kg, 2.5 μg/kg, 2.5 μg/kg and 0.4 ng/mL, respectively. The accuracy of both ELISAs was further confirmed by analysing proficiency test and reference samples. The developed methods can be used for sensitive and high-throughput screening for the presence of T-2/HT-2 and OTA in agricultural commodities.

  13. Study of the natural occurrence of T-2 and HT-2 toxins and their glucosyl derivatives from field barley to malt by high-resolution Orbitrap mass spectrometry

    USDA-ARS?s Scientific Manuscript database

    This paper reports a new method for the determination of T-2 and HT-2 toxins and their glucosylated derivatives in cereals, and some survey data aimed at obtaining more comprehensive information on the co-occurrence of T-2 and HT-2 toxins and their glucosylated derivatives in naturally contaminated ...

  14. Survey of T-2/HT-2 toxins in unprocessed cereals, food and feed coming from Croatia and Bosnia & Herzegovina.

    PubMed

    Pleadin, Jelka; Vasilj, Višnja; Kudumija, Nina; Petrović, Danijela; Vilušić, Milica; Škrivanko, Mario

    2017-06-01

    The aim of this study was to investigate into the level of T-2/HT-2 toxins in different unprocessed cereals (n=201), as well as in marketed cereal-based products (n=58), feed components (n=191) and feedstuffs (n=91) coming from Croatia and Bosnia & Herzegovina. The number of positive samples of unprocessed cereals for food production (>LOD) ranged from 30.4% in barley to 68.8% in oat whereas for feed components ranged from 26.9% in wheat to 86.1% in oat. The maximal values found in unprocessed oat and oat-based feed components were 304.2μg/kg and 521.0μg/kg, respectively. As for final products, the highest T-2/HT-2 concentrations were determined in oat flakes (89.4μg/kg) and calf feed (129.3μg/kg). Despite of the increased T-2/HT-2 concentrations found in some of the samples, the obtained values were unanimously lower than the indicative levels given as recommendations above which further investigations should be necessary performed. Copyright © 2016 Elsevier Ltd. All rights reserved.

  15. Effects of Phenolic Acids on the Growth and Production of T-2 and HT-2 Toxins by Fusarium langsethiae and F. sporotrichioides.

    PubMed

    Ferruz, Elena; Atanasova-Pénichon, Vessela; Bonnin-Verdal, Marie-Noëlle; Marchegay, Gisèle; Pinson-Gadais, Laëtitia; Ducos, Christine; Lorán, Susana; Ariño, Agustín; Barreau, Christian; Richard-Forget, Florence

    2016-04-04

    The effect of natural phenolic acids was tested on the growth and production of T-2 and HT-2 toxins by Fusarium langsethiae and F. sporotrichioides, on Mycotoxin Synthetic medium. Plates treated with 0.5 mM of each phenolic acid (caffeic, chlorogenic, ferulic and p-coumaric) and controls without phenolic acid were incubated for 14 days at 25 °C. Fungal biomass of F. langsethiae and F. sporotrichioides was not reduced by the phenolic acids. However, biosynthesis of T-2 toxin by F. langsethiae was significantly reduced by chlorogenic (23.1%) and ferulic (26.5%) acids. Production of T-2 by F. sporotrichioides also decreased with ferulic acid by 23% (p < 0.05). In contrast, p-coumaric acid significantly stimulated the production of T-2 and HT-2 toxins for both strains. A kinetic study of F. langsethiae with 1 mM ferulic acid showed a significant decrease in fungal biomass, whereas T-2 production increased after 10 days of incubation. The study of gene expression in ferulic supplemented cultures of F. langsethiae revealed a significant inhibition for Tri5, Tri6 and Tri12 genes, while for Tri16 the decrease in gene expression was not statistically significant. Overall, results indicated that phenolic acids had a variable effect on fungal growth and mycotoxin production, depending on the strain and the concentration and type of phenolic acid assayed.

  16. Determination of T-2 toxin, HT-2 toxin, and three other type A trichothecenes in layer feed by high-performance liquid chromatography-tandem mass spectrometry (LC-MS/MS)--comparison of two sample preparation methods.

    PubMed

    Bernhardt, Katrin; Valenta, Hana; Kersten, Susanne; Humpf, Hans-Ulrich; Dänicke, Sven

    2016-05-01

    A sensitive method for the simultaneous determination of T-2 toxin, HT-2 toxin, neosolaniol, T-2 triol, and T-2 tetraol in layer feed using high-performance liquid chromatography coupled to triple quadrupole mass spectrometry in the positive ionization mode (LC-ESI-MS/MS) is described. Two fast and easy clean-up methods-with BondElut Mycotoxin and MycoSep 227 columns, respectively-were tested. The separation of the toxins was conducted on a Pursuit XRs Ultra 2.8 HPLC column using 0.13 mM ammonium acetate as eluent A and methanol as eluent B. Detection of the mycotoxins was carried out in the multiple reaction monitoring (MRM) mode using ammonium adducts as precursor ions. Quantification of all analytes was performed with d3-T-2 toxin as an internal standard. The clean-up method with MycoSep 227 columns gave slightly better results for layer feed compared to the method using BondElut Mycotoxin columns (MycoSep 227: recovery between 50 and 63%, BondElut Mycotoxin: recovery between 32 and 67%) and was therefore chosen as the final method. The limits of detection ranged between 0.9 and 7.5 ng/g depending on the mycotoxin. The method was developed for the analysis of layer feed used at carry-over experiments with T-2 toxin in laying hens. For carry-over experiments, it is necessary that the method includes not only T-2 toxin but also the potential metabolites in animal tissues HT-2 toxin, neosolaniol, T-2 triol, and T-2 tetraol which could naturally occur in cereals used as feed stuff as well.

  17. Development of an LC-MS/MS Determination Method for T-2 Toxin and Its Glucoside and Acetyl Derivatives for Estimating the Contamination of Total T-2 Toxins in Staple Flours.

    PubMed

    Nakagawa, Hiroyuki; Matsuo, Yosuke; McCormick, Susan; Lim, Chee Wei

    2018-05-01

    A determination method previously validated for trichothecenes and zearalenone by means of liquid chromatography-tandem mass spectrometry (LC-MS/MS) was adapted for the quantification of T-2 toxin (T-2) as well as its glucoside and acetyl derivatives, T-2-3-glucoside (T-2-3G) and 3-acetyl-T-2 (3A-T-2). HT-2 toxin (HT-2) and its acetyl derivative 3-acetyl-HT-2 (3A-HT-2) were also included as the target chemicals. Staple flours (56 samples collected from the Singapore market) were examined for contamination from T-2 and/or HT-2 and their derivatives. Among them, 16 flours were found to be contaminated with T-2 and/or HT-2, whereas none was contaminated with T-2-3G and 3A-HT-2, except for trace 3A-T-2 detected in 2 rye samples. Rye flour samples were frequently contaminated with both T-2 and HT-2. Some of the reference materials (RMs) were further analyzed, and T-2-3G and 3A-T-2 were quantitatively detected in corn and wheat RMs. The ratio of T-2-3G to T-2 in the RMs seemed to be much lower than the ratio of deoxynivalenol-3-glucoside to deoxynivalenol usually reported in former studies. To the best of our knowledge, the natural contamination of 3A-T-2 in staple flour is reported here for the first time.

  18. Study of natural occurrence of T-2 and HT-2 toxins and their glucosyl derivatives from field barley to malt by high resolution orbitrap mass spectrometry

    USDA-ARS?s Scientific Manuscript database

    A recent European Union Commission Recommendation (2013/165/EU), asked for collection of more data on the occurrence of T-2 and HT-2 toxins in cereals and cereal products and emphasized that if the method of analysis enables it, it would be appropriate to collect data of the occurrence of masked myc...

  19. A non-linear model for temperature-dependent sporulation and T-2 and HT-2 production of Fusarium langsethiae and Fusarium sporotrichioides.

    PubMed

    Nazari, Leyla; Manstretta, Valentina; Rossi, Vittorio

    2016-04-01

    This research has produced new quantitative data on the sporulation and T-2+HT-2 toxin production that could be further integrated to develop a comprehensive disease or toxin prediction model for Fusarium langsethiae and Fusarium sporotrichioides. Experiments were conducted to determine the effect of temperature or incubation time on sporulation and the effect of temperature on T-2+HT-2 toxin production of strains of the two species. F. sporotrichioides demonstrated a preference for higher temperatures than F. langsethiae during sporulation; the optimum temperature was 24.5 ± 0.7 °C for F. langsethiae and 32.3 ± 2.1 °C for F. sporotrichioides, according to the Beta equation fitted to the data. The dynamics of sporulation over different incubation times were fitted by a Gompertz function. The maximum spore production was estimated to be after 18 and 8 d incubation at optimum temperatures for F. langsethiae and F. sporotrichioides, respectively. F. sporotrichioides produced more T-2+HT-2 than F. langsethiae. The best fit of the effect of temperature on T-2+HT-2 production in wheat grains was obtained with a Beta equation showing an optimum temperature of 14.7 ± 0.8 °C for F. langsethiae and 12.1 ± 0.2 °C for F. sporotrichioides. The optimum temperature for mycotoxin production was lower than for sporulation. Copyright © 2016 The British Mycological Society. Published by Elsevier Ltd. All rights reserved.

  20. Observation of T-2 and HT-2 glucosides from Fusarium sporotrichioides by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS)

    USDA-ARS?s Scientific Manuscript database

    Cultures of Fusarium sporotrichioides were extracted and subjected to evaluation by high performance liquid chromatography – tandem mass spectrometry (LC-MS/MS). Along with the expected T-2 and HT-2 toxins, compounds 162 m/z higher than the toxins were observed. Fragmentation behavior of the larger ...

  1. Aphid Infestation Increases Fusarium langsethiae and T-2 and HT-2 Mycotoxins in Wheat

    PubMed Central

    Drakulic, Jassy; Ajigboye, Olubukola; Swarup, Ranjan; Bruce, Toby

    2016-01-01

    ABSTRACT Fusarium langsethiae is a fungal pathogen of cereal crops that is an increasing problem in northern Europe, but much of its epidemiology is poorly understood. The species produces the mycotoxins T-2 and HT-2, which are highly toxic. It was hypothesized that grain aphids, Sitobion avenae, may transmit F. langsethiae inoculum between wheat plants, and a series of transmission experiments and volatile chemical analyses was performed to test this. Manual translocation of aphids from inoculated to uninfected hosts resulted in pathogen DNA accumulation in hosts. However, the free movement of wingless aphids from infected to healthy plants did not. The addition of winged aphids reared on F. langsethiae-inoculated wheat seedlings to wheat plants also did not achieve successful pathogen transfer. While our data suggested that aphid transmission of the pathogen was not very efficient, we observed an increase in disease when aphids were present. After seedling inoculation, an increase in pathogen DNA accumulation in seedling leaves was observed upon treatment with aphids. Furthermore, the presence of aphids on wheat plants with F. langsethiae-inoculated ears not only led to a rise in the amount of F. langsethiae DNA in infected grain but also to an increase in the concentrations of T-2 and HT-2 toxins, with more than 3-fold higher toxin levels than diseased plants without aphids. This work highlights that aphids increase the susceptibility of wheat host plants to F. langsethiae and that aphid infestation is a risk factor for accumulating increased levels of T-2 and HT-2 in wheat products. IMPORTANCE Fusarium langsethiae is shown here to cause increased contamination levels of grain with toxins produced by fungus when aphids share the host plant. This effect has also recently been demonstrated with Fusarium graminearum, yet the two fungal species show stark differences in their effect on aphid populations. In both cases, aphids improve the ability of the pathogens to

  2. The roles of carboxylesterase and CYP isozymes on the in vitro metabolism of T-2 toxin.

    PubMed

    Lin, Ni-Ni; Chen, Jia; Xu, Bin; Wei, Xia; Guo, Lei; Xie, Jian-Wei

    2015-01-01

    T-2 toxin poses a great threat to human health because it has the highest toxicity of the currently known trichothecene mycotoxins. To understand the in vivo toxicity and transformation mechanism of T-2 toxin, we investigated the role of one kind of principal phase I drug-metabolizing enzymes (cytochrome P450 [CYP450] enzymes) on the metabolism of T-2 toxin, which are crucial to the metabolism of endogenous substances and xenobiotics. We also investigated carboxylesterase, which also plays an important role in the metabolism of toxic substances. A chemical inhibition method and a recombinant method were employed to investigate the metabolism of the T-2 toxin by the CYP450 enzymes, and a chemical inhibition method was used to study carboxylesterase metabolism. Samples incubated with human liver microsomes were analyzed by high performance liquid chromatography-triple quadrupole mass spectrometry (HPLC- QqQ MS) after a simple pretreatment. In the presence of a carboxylesterase inhibitor, only 20 % T-2 toxin was metabolized. When CYP enzyme inhibitors and a carboxylesterase inhibitor were both present, only 3 % of the T-2 toxin was metabolized. The contributions of the CYP450 enzyme family to T-2 toxin metabolism followed the descending order CYP3A4, CYP2E1, CYP1A2, CYP2B6 or CYP2D6 or CYP2C19. Carboxylesterase and CYP450 enzymes are of great importance in T-2 toxin metabolism, in which carboxylesterase is predominant and CYP450 has a subordinate role. CYP3A4 is the principal member of the CYP450 enzyme family responsible for T-2 toxin metabolism. The primary metabolite produced by carboxylesterase is HT-2, and the main metabolite produced by CYP 3A4 is 3'-OH T-2. The different metabolites show different toxicities. Our results will provide useful data concerning the toxic mechanism, the safety evaluation, and the health risk assessment of T-2 toxin.

  3. Use of T-2 toxin-immobilized amine-activated beads as an efficient affinity purification matrix for the isolation of specific IgY.

    PubMed

    Edupuganti, Soujanya Ratna; Edupuganti, Om Prakash; O'Kennedy, Richard; Defrancq, Eric; Boullanger, Stéphanie

    2013-04-01

    An affinity purification method that isolates T-2 toxin-specific IgY utilizing a T-2-toxin-immobilized column was developed. The T-2 toxin was covalently coupled via a carbonyldiimidazole-activated hydroxyl functional group to amine-activated sepharose beads. The affinity-purified IgY was characterized by gel electrophoresis, fast protein liquid chromatography, enzyme-linked immunosorbant assay, surface plasmon resonance and mass spectrometry. A competitive inhibition ELISA (CI-ELISA) was performed using affinity-purified IgY with a T-2 toxin detection sensitivity of 30 ng/mL, which falls within the maximum permissible limit of 100 ng/mL. The cross reactivity of IgY towards deoxynivalenol, zearalenone, fumonisin B1 and HT-2 was significantly reduced after affinity purification. A surface plasmon resonance (SPR)-based inhibition assay was also applied for quantitative determination of T-2 toxin in spiked wheat samples. The results obtained indicate the feasibility of utilizing this IgY-based assay for the detection of T-2 toxin in food samples.

  4. An imaging surface plasmon resonance biosensor assay for the detection of T-2 toxin and masked T-2 toxin-3-glucoside in wheat

    USDA-ARS?s Scientific Manuscript database

    A sensitive, rapid, and reproducible imaging surface plasmon resonance (iSPR) biosensor assay was developed to detect T-2 toxin and T-2 toxin-3-glucoside (T2-G) in wheat. In this competitive assay, an amplification strategy was used after conjugating a secondary antibody (Ab2) with gold nanoparticle...

  5. Comparison of penetration and metabolism of [3H]diacetoxyscirpenol, [3H]verrucarin A and [3H]T-2 toxin in skin.

    PubMed

    Kemppainen, B W; Riley, R T; Biles-Thurlow, S

    1987-05-01

    The purpose of this research was to determine the rate of cutaneous penetration and metabolism of [3H]diacetoxyscirpenol (DAS) and [3H]verrucarin A (VCA) and compare these values to previously determined values for [3H]T-2 toxin (T-2), to compare the cutaneous penetration and metabolism of DAS in human and guinea-pig skin, and to compare the effects of dose and of two vehicles, methanol and dimethylsulphoxide (DMSO), on penetration rates. DAS or VCA was applied to the epidermal surface of excised skin, and the receptor fluid bathing the dermal surface was sampled periodically for 48 hr. Whether the applied dose (581 ng/cm2) was dissolved in methanol or DMSO, the rate of penetration through human skin was lower for VCA than for DAS or T-2, the rates for the two latter compounds being similar at this dose. Metabolism of DAS occurred during penetration through excised human skin and did not occur in the receptor fluid as a result of enzymes leaching out of the skin. VCA appeared to be metabolized by human skin, but this conclusion is tentative because of the relative instability of this compound. DAS penetrated significantly (P less than 0.05) faster through excised guinea-pig skin than through human skin. Metabolism of DAS was greater in human skin than in guinea-pig skin. When compared with methanol, DMSO increased the penetration of DAS and VCA by factors of between 7 and 52. At the low dose (79 ng/cm2) DAS penetrated human and guinea-pig skin significantly (P less than 0.05) faster than T-2 using either vehicle.

  6. Anomericity of T-2 toxin-glucosides; masked mycotoxins in cereal crops

    USDA-ARS?s Scientific Manuscript database

    T-2 toxin is a trichothecene mycotoxin produced when the fungus Fusarium infects small grains, especially oats. Ingestion of T-2 toxin contaminated grain can cause diarrhea, hemorrhaging, and feed refusal. Cereal crops infected with mycotoxin-producing fungi form toxin glycosides, sometimes called m...

  7. Electrophysiological response of chicken's jejunal epithelium to increasing levels of T-2 toxin.

    PubMed

    Yunus, Agha Waqar; Kröger, Susan; Tichy, Alexander; Zentek, Jürgen; Böhm, Josef

    2013-02-01

    The present investigations were conducted to test the effects of T-2 toxin on electrophysiological variables of jejunal epithelium of chicken. Jejunal segments of broilers were monitored in Ussing chambers in the presence of T-2 toxin at the levels of 0 (negative control), 0 (methanol/vehicle control), 0.1, 1, 5, and 10 μg/ml of buffer. T-2 toxin did not affect basal values of short circuit current (I(sc)), transmural potential difference, or tissue conductivity in the jejunal epithelium. T-2 toxin also did not statistically affect glucose-induced electrophysiological variables during the first 3 min of glucose induction. Compared to the vehicle control, the ouabain-sensitive I(sc) was negatively affected (P = 0.008) only under 5 μg of T-2 toxin/ml. Increasing levels of T-2 toxin negatively affected the ouabain-sensitive I(sc) in a cubic (P = 0.007) fashion. These data indicate that acute exposure to moderate levels of T-2 toxin may progressively impair the cation gradient across the jejunal epithelium.

  8. The inhibition of cholera toxin-induced 5-HT release by the 5-HT3 receptor antagonist, granisetron, in the rat

    PubMed Central

    Turvill, J L; Connor, P; Farthing, M J G

    2000-01-01

    The secretagogue 5-hydroxytryptamine (5-HT) is implicated in the pathophysiology of cholera. 5-HT released from enterochromaffin cells after cholera toxin exposure is thought to activate non-neuronally (5-HT2 dependent) and neuronally (5-HT3 dependent) mediated water and electrolyte secretion. CT-secretion can be reduced by preventing the release of 5-HT. Enterochromaffin cells possess numerous receptors that, under basal conditions, modulate 5-HT release. These include basolateral 5-HT3 receptors, the activation of which is known to enhance 5-HT release. Until now, 5-HT3 receptor antagonists (e.g. granisetron) have been thought to inhibit cholera toxin-induced fluid secretion by blockading 5-HT3 receptors on secretory enteric neurones. Instead we postulated that they act by inhibiting cholera toxin-induced enterochromaffin cell degranulation. Isolated intestinal segments in anaesthetized male Wistar rats, pre-treated with granisetron 75 μg kg−1, lidoocaine 6 mg kg−1 or saline, were instilled with a supramaximal dose of cholera toxin or saline. Net fluid movement was determined by small intestinal perfusion or gravimetry and small intestinal and luminal fluid 5-HT levels were determined by HPLC with fluorimetric detection. Intraluminal 5-HT release was proportional to the reduction in tissue 5-HT levels and to the onset of water and electrolyte secretion, suggesting that luminal 5-HT levels reflect enterochromaffin cell activity. Both lidocaine and granisetron inhibited fluid secretion. However, granisetron alone, and proportionately, reduced 5-HT release. The simultaneous inhibition of 5-HT release and fluid secretion by granisetron suggests that 5-HT release from enterochromaffin cells is potentiated by endogenous 5-HT3 receptors. The accentuated 5-HT release promotes cholera toxin-induced fluid secretion. PMID:10882387

  9. Influence of Selenium on the Production of T-2 Toxin by Fusarium poae.

    PubMed

    Cheng, Bolun; Zhang, Yan; Tong, Bei; Yin, Hong

    2017-07-01

    The objective of this study was to investigate the effects of selenium on the production of T-2 toxin by a Fusarium poae strain cultured in a synthetic medium containing different concentrations of selenium. The T-2 toxin contents in fermentative products were evaluated by a high performance liquid chromatography (HPLC). The results showed that the production of T-2 toxin was correlated with the concentration of selenium added to the medium. In all three treatments, the addition of 1 mg/L selenium to the medium resulted in a lower toxin yield than the control (0 mg/L); the yield of the toxin began to increase when selenium concentration was 10 mg/L, while it decreased again at 20 mg/L. In summary, T-2 toxin yield in the fermentative product was affected by the addition of selenium to the medium, and a selenium concentration of 20 mg/L produced the maximum inhibitory effect of T-2 toxin yield in the fermentative product of F. poae.

  10. Imaging surface plasmon resonance detection for T-2 toxin in wheat

    USDA-ARS?s Scientific Manuscript database

    T-2 toxin is a trichothecene (Type A) mycotoxin harmful to humans and animals. It is produced by mainly Fusarium species, particularly F. sporotrichoides. Fusarium species often contaminate grains such as corn, wheat, barley, oats, rice and rye. T-2 toxin contamination in grains has been observed mo...

  11. Glucosylation and Other Biotransformations of T-2 Toxin by Yeasts of the Trichomonascus Clade

    PubMed Central

    Price, Neil P. J.; Kurtzman, Cletus P.

    2012-01-01

    Trichothecenes are sesquiterpenoid toxins produced by Fusarium species. Since these mycotoxins are very stable, there is interest in microbial transformations that can remove toxins from contaminated grain or cereal products. Twenty-three yeast species assigned to the Trichomonascus clade (Saccharomycotina, Ascomycota), including four Trichomonascus species and 19 anamorphic species presently classified in Blastobotrys, were tested for their ability to convert the trichothecene T-2 toxin to less-toxic products. These species gave three types of biotransformations: acetylation to 3-acetyl T-2 toxin, glycosylation to T-2 toxin 3-glucoside, and removal of the isovaleryl group to form neosolaniol. Some species gave more than one type of biotransformation. Three Blastobotrys species converted T-2 toxin into T-2 toxin 3-glucoside, a compound that has been identified as a masked mycotoxin in Fusarium-infected grain. This is the first report of a microbial whole-cell method for producing trichothecene glycosides, and the potential large-scale availability of T-2 toxin 3-glucoside will facilitate toxicity testing and development of methods for detection of this compound in agricultural and other products. PMID:23042183

  12. Acute Respiratory Tract Toxicity of the Trichothecene Mycotoxin, T-2 Toxin.

    DTIC Science & Technology

    1987-03-31

    to 10-week-old, castrated , e male, cross-bred, specific pathogen-free pigs were exposed to an aerosol of T-2 toxin for 45 to 61 minutes with 8 mg of... castrated , crossbred, specific pathogen-free swine to aerosols of T-2 toxin for 42 to 65 minutes with 9 mg of the toxin nebulized per kilogram of body... cattle , Ph.D. Thesis, University of Illinois, Urbana, IL, 1983. 28. Lorenzana, R.M., Beasley, V.R., Buck, W.B., Ghent, A.W., Lundeen, G.R., and

  13. Stability of fumonisin B1, deoxynivalenol, zearalenone, and T-2 toxin during processing of traditional Nigerian beer and spices.

    PubMed

    Chilaka, Cynthia Adaku; De Boevre, Marthe; Atanda, Olusegun Oladimeji; De Saeger, Sarah

    2018-05-03

    The stability of the Fusarium mycotoxins fumonisin B 1 , deoxynivalenol, T-2 toxin, and zearalenone during processing of Nigerian traditional spices (dawadawa, okpehe, and ogiri) and beer (burukutu) using artificially contaminated raw materials was investigated. Results revealed the reduction of these toxins in all the final products. Boiling played a significant role (p < 0.05) in Fusarium mycotoxin reduction in the traditional spices. The highest percentage reduction of deoxynivalenol (76%) and zearalenone (74%) was observed during okpehe processing (boiled for 12 h). Dehulling and fermentation further demonstrated a positive influence on the reduction of these toxins with a total reduction ranging from 85 to 98% for dawadawa, 86 to 100% for okpehe, and 57 to 81% for ogiri. This trend was also observed during the production of traditional beer (burukutu), with malting and brewing playing a major impact in observed reduction. In addition, other metabolites including deoxynivalenol-3-glucoside, 15-acetyl-deoxynivalenol, α-zearalenol, and β-zearalenol which were initially not present in the raw sorghum were detected in the final beer product at the following concentrations 26 ± 11, 16 ± 7.7, 22 ± 18, and 31 ± 16 μg/kg, respectively. HT-2 toxin was also detected at a concentration of 36 ± 13 μg/kg along the processing chain (milled malted fraction) of the traditional beer. For the traditional spices, HT-2 toxin was detected (12 μg/kg) in ogiri. Although there was a reduction of mycotoxins during processing, appreciable concentrations of these toxins were still detected in the final products. Thus, the use of good quality raw materials significantly reduces mycotoxin contamination in final products.

  14. Conduction-band valley spin splitting in single-layer H-T l2O

    NASA Astrophysics Data System (ADS)

    Ma, Yandong; Kou, Liangzhi; Du, Aijun; Huang, Baibiao; Dai, Ying; Heine, Thomas

    2018-02-01

    Despite numerous studies, coupled spin and valley physics is currently limited to two-dimensional (2D) transition-metal dichalcogenides (TMDCs). Here, we predict an exceptional 2D valleytronic material associated with the spin-valley coupling phenomena beyond 2D TMDCs—single-layer (SL) H-T l2O . It displays large valley spin splitting (VSS), significantly larger than that of 2D TMDCs, and a finite band gap, which are both critically attractive for the integration of valleytronics and spintronics. More importantly, in sharp contrast to all the experimentally confirmed 2D valleytronic materials, where the strong valence-band VSS (0.15-0.46 eV) supports the spin-valley coupling, the VSS in SL H-T l2O is pronounced in its conduction band (0.61 eV), but negligibly small in its valence band (21 meV), thus opening a way for manipulating the coupled spin and valley physics. Moreover, SL H-T l2O possesses extremely high carrier mobility, as large as 9.8 ×103c m2V-1s-1 .

  15. Down-regulation of ATF2 in the inhibition of T-2-toxin-induced chondrocyte apoptosis by selenium chondroitin sulfate nanoparticles

    NASA Astrophysics Data System (ADS)

    Han, Jing; Guo, Xiong

    2013-12-01

    Selenium chondroitin sulfate nanoparticles (SeCS) with a size range of 30-200 nm were obtained in our previous study. Meanwhile, the up-regulated expression of ATF2 mRNA and protein levels could be observed in the cartilage from Kashin-Beck disease (KBD) patients. In this paper, we investigated the inhibition effect of SeCS on T-2-toxin-induced apoptosis of chondrocyte from KBD patients. Here, we found that when the chondrocytes were treated with T-2 toxin, the chondrocyte apoptosis performed in a concentration-dependent manner. The apoptosis of chondrocyte induced by T-2 toxin involved the increased levels of ATF2, JNK and p38 mRNAs and related protein expression. SeCS could partly block the T-2-toxin-induced chondrocyte apoptosis by decreasing the expression of ATF2, JNK and p38 mRNAs and p-JNK, p-38, ATF2 and p-ATF2 proteins. JNK and p38 pathways involved in the apoptosis of chondrocyte induced by T-2 toxin, and SeCS was efficient in the inhibition of chondrocyte apoptosis by T-2 toxin. These results suggested that SeCS had a potential for further prevention and treatment for KBD as well as other selenium deficiency disease.

  16. On the Toxicity and Metabolism of the Trichothecene Mycotoxin T-2 Toxin,

    DTIC Science & Technology

    1988-06-20

    on r~atiado d nfto..ayl --The present study deals witL1 toxicolegical effects anid :btoi of the trichothecene mycotoxin T-2 toxi.. T-2 toxin was shoeen...AND METABOLISM OF THE TRICHOTHECENE MYCOTOXIN T-2 TOXIN SUMMARY The present study deals with toxicological effects and metabolism of the trichothecene...directly related to this effect (McLaughlin et al., 1977). 15 1.4 DNA synthesis It is well established that trichothecene mycotoxins block DNA synthesis in

  17. Short-term effects of T-2 toxin or deoxynivalenol on lipid peroxidation and the glutathione system in common carp.

    PubMed

    Pelyhe, Csilla; Kövesi, Benjámin; Zándoki, Erika; Kovács, Balázs; Szabó-Fodor, Judit; Mézes, Miklós; Balogh, Krisztián

    2016-12-01

    The purpose of this study was to investigate the short-term effects of a single oral dose of T-2 and HT-2 toxin at 0.15, 0.33 and 1.82 mg kg -1 body weight, or deoxynivalenol (DON) and 15-acetyl-DON at 0.13, 0.31 and 1.75 mg kg -1 body weight in common carp. Conjugated dienes and trienes (the early markers of lipid peroxidation) were elevated in all DON-treated groups at the 16th hour, while thiobarbituric acid reactive substances (TBARS; termination marker) were increased at the highest dose of DON at the 16th and 24th hours. T-2 toxin did not cause changes in these parameters. Glutathione content and glutathione peroxidase activity showed higher levels at the 16th hour as the effect of both mycotoxins. The expression of glutathione peroxidase (GPx4) genes (gpx4a and gpx4b) revealed a dual response. Downregulation was observed at the 8th hour, followed by an induction at the 16th hour, at the lowest dose of both mycotoxins. Higher doses revealed long-drawn emergence and an elevation was observed only at the 24th hour. However, at the lowest and highest doses of DON or T-2 toxin the changes in gene expression were delayed, which may be related to the low oxidative stress response, as suggested by the expression profiles of the nrf2, keap1, gpx4a and gpx4b genes.

  18. Selenium deficiency aggravates T-2 toxin-induced injury of primary neonatal rat cardiomyocytes through ER stress.

    PubMed

    Xu, Jing; Pan, Shengchi; Gan, Fang; Hao, Shu; Liu, Dandan; Xu, Haibin; Huang, Kehe

    2018-04-01

    Keshan disease is a potentially fatal cardiomyopathy in humans. Selenium deficiency, T-2 toxin, and myocarditis virus are thought to be the major factors contributing to Keshan disease. But the relationship among these three factors is poorly described. This study aims to explore whether selenium deficiency aggravates T-2 toxin-induced cardiomyocyte injury and its underlying mechanism. Cardiomyocytes were isolated from neonatal rat and cultured at the physiological (2.0 μM) or lower concentrations of selenium with different concentrations of T-2 toxin. Our results showed that selenium deficiencies aggravated T-2 toxin-induced cardiomyocyte injury in a concentration-dependent manner as demonstrated by MTT bioassay, LDH activity, reactive oxygen species levels and caspase 3 protein expressions. T-2 toxin treatment significantly increased mRNA expressions for stress proteins GRP78 and CHOP in cardiomyocytes compared with the control. Selenium deficiencies further promoted GRP78, CHOP and p-eIF2α expressions. Knockdown of CHOP by the specific small interfering RNA eliminated the effect of selenium deficiencies on T-2 toxin-induced injury. It could be concluded that selenium deficiency aggravates T-2 toxin-induced cardiomyocyte injury through initiating more aggressive endoplasmic reticulum stress. Copyright © 2018 Elsevier B.V. All rights reserved.

  19. Immunopathological effect of the mycotoxins cyclopiazonic acid and T-2 toxin on broiler chicken.

    PubMed

    Kamalavenkatesh, P; Vairamuthu, S; Balachandran, C; Manohar, B Murali; raj, G Dhinakar

    2005-02-01

    Forty, newly hatched, unsexed broiler chicks were fed diets containing 10 ppm cyclopiazonic acid (CPA) and 1 ppm T-2 toxin (T2) either individually or in combination for 28 days to study the immunopathological effects. Lymphoid organs revealed lymphocytolysis and lymphoid depletion in all toxin fed birds. Thymic and splenic CD+4 and CD+8 lymphocytes decreased significantly (p<0.01) in toxin fed birds when compared to the control. Thymic CD+8 lymphocytes of T2 and CPA-T2 showed significant (p<0.01) decrease from that of CPA and control groups. Splenic CD+4 and CD+8 lymphocytes showed significant (p<0.01) decrease in CPA and CPA-T2 fed groups when compared to the control. The T2 group did not differ significantly from that of control. The stimulation index (SI) of splenocytes to concavalin A revealed significant (p<0.01) decrease in all toxin fed birds. Significant (p<0.01) decrease were observed for the haemagglutination inhibition (HI) titres to Newcastle disease virus vaccine F strain (NDV) of birds fed CPA, T2 and in combination. Significant (p<0.01) interaction was found for lymphocyte subsets, SI and HI titres to NDV. The study indicated the immunosuppressive effect of these toxins either alone or in combination in broiler chicks.

  20. Functional expression of 5-HT{sub 2A} receptor in osteoblastic MC3T3-E1 cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Hirai, Takao; Kaneshige, Kota; Kurosaki, Teruko

    2010-05-28

    In the previous study, we reported the gene expression for proteins related to the function of 5-hydroxytryptamine (5-HT, serotonin) and elucidated the expression patterns of 5-HT{sub 2} receptor subtypes in mouse osteoblasts. In the present study, we evaluated the possible involvement of 5-HT receptor subtypes and its inactivation system in MC3T3-E1 cells, an osteoblast cell line. DOI, a 5-HT{sub 2A} and 5-HT{sub 2C} receptor selective agonist, as well as 5-HT concentration-dependently increased proliferative activities of MC3T3-E1 cells in their premature period. This effect of 5-HT on cell proliferation were inhibited by ketanserin, a 5-HT{sub 2A} receptor specific antagonist. Moreover, bothmore » DOI-induced cell proliferation and phosphorylation of ERK1 and 2 proteins were inhibited by PD98059 and U0126, selective inhibitors of MEK in a concentration-dependent manner. Furthermore, treatment with fluoxetine, a 5-HT specific re-uptake inhibitor which inactivate the function of extracellular 5-HT, significantly increased the proliferative activities of MC3T3-E1 cells in a concentration-dependent manner. Our data indicate that 5-HT fill the role for proliferation of osteoblast cells in their premature period. Notably, 5-HT{sub 2A} receptor may be functionally expressed to regulate mechanisms underlying osteoblast cell proliferation, at least in part, through activation of ERK/MAPK pathways in MC3T3-E1 cells.« less

  1. Regulatory mechanism of CCN2 production by serotonin (5-HT) via 5-HT2A and 5-HT2B receptors in chondrocytes.

    PubMed

    Hori, Ayaka; Nishida, Takashi; Takashiba, Shogo; Kubota, Satoshi; Takigawa, Masaharu

    2017-01-01

    Serotonin (5-hydroxytryptamine: 5-HT) is recognized as a neurotransmitter in the central nerve system and as a regulator of systemic blood pressure in the peripheral tissues. Recently, it was reported that 5-HT2 receptors (5-HT2Rs) were expressed in cartilage tissues lacking both vessels and neurons, suggesting possible novel functions of 5-HT during cartilage development and regeneration. Our previous data indicated that CCN family protein 2/connective tissue growth factor (CCN2/CTGF) plays a central role in cartilage development and regeneration. Therefore, the aim of this study was to investigate the effect of 5-HT on the production of CCN2 in chondrocytes. Firstly, we showed that the mRNAs of 5-HT2R subtypes 5-HT2AR and 5-HT2BR, were expressed in a human chondrocytic cell line, HCS-2/8; however, 5-HT2CR mRNA was not detected. In addition, exogenously added 5-HT did not affect the 5-HT2AR and 5-HT2BR expressions. Next, we demonstrated that CCN2 production was increased by treatment with a 5-HT2AR agonist and the combination of 5-HT and 5-HT2BR antagonist. In contrast, treatment with a 5-HT2BR agonist and the combination of 5-HT and 5-HT2AR antagonist decreased CCN2 production. Furthermore, we showed that phosphorylation of Akt and p38 MAPK were increased by treatment with 5-HT2AR agonist, and that phosphorylation of PKCε, PKCζ, ERK1/2 and JNK were increased by treatment with 5-HT2BR agonist. Finally, we found that 5-HT2AR was localized in the growth plate, whereas 5-HT2BR was localized in the articular cartilage. These findings suggest that 5-HT promotes CCN2 production through the 5-HT2AR in growth plates, and that it represses CCN2 production through the 5-HT2BR in articular cartilage for harmonized development of long bones.

  2. Effects of T-2 toxin on turkey herpesvirus–induced vaccinal immunity against Marek’s disease

    USDA-ARS?s Scientific Manuscript database

    T-2 toxin, a very potent immunotoxic Type A trichothecene, is a secondary metabolite produced primarily by Fusarium spp., which grows on cereal grains and can lead to contaminated livestock feed. Repeated exposure to T-2 toxin has been shown to cause immunosuppression and decrease the resistance of ...

  3. Adverse effects of T-2 toxin on chicken lymphocytes blastogenesis and its protection with Vitamin E.

    PubMed

    Jaradat, Ziad W; Viià, Borja; Marquardt, Ronal R

    2006-08-15

    T-2 toxin, a trichothecene mycotoxin that is produced by fusarium species, is prevalent mainly in cereal crops and poultry feed. One of the major effects of this toxin is immunomodulation. The effect of T-2 toxin on chicken lymphocyte proliferation in the presence of mitogens and the subsequent protection with Vitamin E in both fat and water soluble forms was studied using an MTT colorimetric assay. T-2 toxin was administered in concentrations ranging from 0 to 10ng/mL of lymphocytes in the presence of either concanavalin A (ConA) or phytohemagglutinine (PHA-M) at optimum concentration of 333ng/mL and a dilution of 1:160 for ConA and PHA-M, respectively. Lymphocyte proliferation in response to ConA and PHA-M mitogens was depressed at T-2 doses of 1ng/mL or higher (p<0.05). The proliferation was completely abolished at 10ng/mL when the toxin was added at 0 time, while it was decreased by 80% when the toxin was added to the lymphocytes after 24h. The addition of Vitamin E in the fat soluble form (alpha-tocopheryl acetate) did not exert any protection effect against the toxin when it was added at either 25 or 100microg. However, when the water soluble form (Trolox) was added at a concentration of (200microg) (equivalent to 100microM of alpha-tocopherol), it provided considerable protection (p<0.05) against T-2 toxin inhibition of lymphocyte proliferation. The difference in the effect between the two forms of Vitamin E might be related to their relative solubility in the culture media which in turn may affect their availability for protection.

  4. Modulation of sweet taste by umami compounds via sweet taste receptor subunit hT1R2.

    PubMed

    Shim, Jaewon; Son, Hee Jin; Kim, Yiseul; Kim, Ki Hwa; Kim, Jung Tae; Moon, Hana; Kim, Min Jung; Misaka, Takumi; Rhyu, Mee-Ra

    2015-01-01

    Although the five basic taste qualities-sweet, sour, bitter, salty and umami-can be recognized by the respective gustatory system, interactions between these taste qualities are often experienced when food is consumed. Specifically, the umami taste has been investigated in terms of whether it enhances or reduces the other taste modalities. These studies, however, are based on individual perception and not on a molecular level. In this study we investigated umami-sweet taste interactions using umami compounds including monosodium glutamate (MSG), 5'-mononucleotides and glutamyl-dipeptides, glutamate-glutamate (Glu-Glu) and glutamate-aspartic acid (Glu-Asp), in human sweet taste receptor hT1R2/hT1R3-expressing cells. The sensitivity of sucrose to hT1R2/hT1R3 was significantly attenuated by MSG and umami active peptides but not by umami active nucleotides. Inhibition of sweet receptor activation by MSG and glutamyl peptides is obvious when sweet receptors are activated by sweeteners that target the extracellular domain (ECD) of T1R2, such as sucrose and acesulfame K, but not by cyclamate, which interact with the T1R3 transmembrane domain (TMD). Application of umami compounds with lactisole, inhibitory drugs that target T1R3, exerted a more severe inhibitory effect. The inhibition was also observed with F778A sweet receptor mutant, which have the defect in function of T1R3 TMD. These results suggest that umami peptides affect sweet taste receptors and this interaction prevents sweet receptor agonists from binding to the T1R2 ECD in an allosteric manner, not to the T1R3. This is the first report to define the interaction between umami and sweet taste receptors.

  5. Damaging effects of Clostridium perfringens delta toxin on blood platelets and their relevance to ganglioside GM2.

    PubMed

    Jolivet-Reynaud, C; Launay, J M; Alouf, J E

    1988-04-01

    The lytic effect of Clostridium perfringens delta toxin was investigated on goat, human, rabbit, and guinea pig platelets. In contrast to erythrocytes from the latter three species, which are insensitive to the toxin, the platelets were equally lysed by the same amount of toxin. These results suggest the presence of GM2 or GM2-like ganglioside(s) as a specific recognition site of the toxin on platelet plasmic membrane as previously established for sensitive erythrocytes. Plasmic membrane damage of human platelets was evidenced by the release of entrapped alpha-[14C]aminoisobutyric acid used as a cytoplasmic marker. The specific binding of hemolytically active 125I-delta toxin by human and rabbit platelets was practically identical, dose dependent, and inhibitable by GM2. Labeled toxin was also bound by various subcellular organelles separated from rabbit platelets except the 5-hydroxytryptamine (5-HT)-containing dense bodies, suggesting the absence or inaccessibility of GM2 on the surface of the latter organelles. This result correlates with the low amounts of 5-[3H]HT liberated after platelet challenge with delta toxin whereas this mediator was massively liberated upon lysis by the sulfhydryl-activated toxin alveolysin. The levels of M and P forms of phenol sulfotransferase (PST), involved in 5-HT catabolism, were determined in human platelet lysates after challenge with delta toxin, alveolysin, and other disruptive treatments. The low PST-M activities detected after lysis by delta toxin suggest that this isoenzyme is very likely associated to dense bodies in contrast to PST-P which is cytoplasmic. Platelet lysis by the toxin allows easy separation of these organelles.

  6. The role of serotonin-2 (5-HT2) and dopamine receptors in the behavioral actions of the 5-HT2A/2C agonist, DOI, and putative 5-HT2C inverse agonist, SR46349B.

    PubMed

    Scarlota, Laura C; Harvey, John A; Aloyo, Vincent J

    2011-02-01

    Atypical antipsychotic efficacy is often attributed to actions at serotonin-2 (5-HT(2)) and dopamine receptors, indicating a potential benefit of understanding the interplay between these systems. Currently, it is known that 5-HT(2) receptors modulate dopamine release, although the role of specific dopamine receptors in 5-HT(2)-mediated behavior is not well understood. We examined the role of 5-HT(2A), 5-HT(2C), and dopamine (D1 and D2) receptors in the behavioral response to a 5-HT(2A/2C) agonist (DOI) and 5-HT(2A/2C) antagonist (SR46349B). Effects were assessed by measuring rabbit head bobs (previously characterized as 5-HT(2A) receptor-mediated) and body shakes (5-HT(2C)-mediated). As expected, DOI produced head bobs and body shakes, and these DOI-elicited behaviors were attenuated by the SR46349B pretreatment. Unexpectedly, SR46349B also induced head bobs when administered alone. However, SR46349B-elicited head bobs are distinguishable from those produced by DOI since the 5-HT(2A) antagonist, ketanserin, only attenuated DOI-elicited head bobs. Conversely, 5-HT(2C) ligands (SB242084 and SB206553) inhibited SR46349B but not DOI-induced head bobs. Furthermore, when administered alone, SB206553 (a 5-HT(2C) inverse agonist) produced head bobs, indicating the behavior can be either 5-HT(2A) or 5-HT(2C) mediated. Next, it was revealed that D1 and D2 receptors play a role in DOI-elicited head bobs, but only D1 receptors are required for SR46349B-elicited head bobs. 5-HT(2A) receptor agonism and 5-HT(2C) inverse agonism produce the same behavior, likely due to similar downstream actions at D1 receptors. Consequently, 5-HT(2C) agonism or D1 agonism may be effective therapies for disorders, such as schizophrenia, currently being treated with 5-HT(2A) antagonists.

  7. Agonist properties of N,N-dimethyltryptamine at serotonin 5-HT2A and 5-HT2C receptors.

    PubMed

    Smith, R L; Canton, H; Barrett, R J; Sanders-Bush, E

    1998-11-01

    Extensive behavioral and biochemical evidence suggests an agonist role at the 5-HT2A receptor, and perhaps the 5-HT2C receptor, in the mechanism of action of hallucinogenic drugs. However the published in vitro pharmacological properties of N,N-dimethyltryptamine (DMT), an hallucinogenic tryptamine analog, are not consistent with this hypothesis. We, therefore, undertook an extensive investigation into the properties of DMT at 5-HT2A and 5-HT2C receptors. In fibroblasts transfected with the 5-HT2A receptor or the 5-HT2C receptor, DMT activated the major intracellular signaling pathway (phosphoinositide hydrolysis) to an extent comparable to that produced by serotonin. Because drug efficacy changes with receptor density and cellular microenvironment, we also examined the properties of DMT in native preparations using a behavioral and biochemical approach. Rats were trained to discriminate an antagonist ketanserin from an agonist 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) in a two-lever choice paradigm. Pharmacological studies showed that responding on the DOI and ketanserin lever reflected agonist and antagonist activity at 5-HT2A receptors, and hence, was a suitable model for evaluating the in vivo functional properties of DMT. Like other 5-HT2A receptor agonists, DMT substituted fully for DOI. Intact choroid plexus was used to evaluate the agonist properties at endogenous 5-HT2C receptors; DMT was a partial agonist at 5-HT2C receptors in this native preparation. Thus, we conclude that DMT behaves as an agonist at both 5-HT2A and 5-HT2A receptors. One difference was evident in that the 5-HT2C, but not the 5-HT2A, receptor showed a profound desensitization to DMT over time. This difference is interesting in light of the recent report that the hallucinogenic activity of DMT does not tolerate in humans and suggests the 5-HT2C receptor plays a less prominent role in the action of DMT.

  8. H-T Magnetic Phase Diagram of a Frustrated Triangular Lattice Antiferromagnet CuFeO 2

    NASA Astrophysics Data System (ADS)

    Mitsuda, Setsuo; Mase, Motoshi; Uno, Takahiro; Kitazawa, Hideaki; Katori, Hiroko

    2000-01-01

    By magnetization and specific heat measurements in an applied magnetic field up to 12 T, we obtained the magnetic field (H) versus temperature (T) phase diagram of a frustrated triangular lattice antiferromagnet (TLA), CuFeO2, where a partially disordered phase typical to Ising TLA exists as a thermally induced state for the 4-sublattice ground state as well as for the first-field-induced 5-sublattice-like state. The experimentally obtained H-T magnetic phase diagram is compared with that from Monte-Carlo simulation of a 2D Ising TLA model with competing exchange interactions up to 3rd neighbors.

  9. Histopathology of chondronecrosis development in knee articular cartilage in a rat model of Kashin-Beck disease using T-2 toxin and selenium deficiency conditions.

    PubMed

    Guan, Fang; Li, Siyuan; Wang, Zhi-Lun; Yang, Haojie; Xue, Senghai; Wang, Wei; Song, Daiqing; Zhou, Xiaorong; Zhou, Wang; Chen, Jing-Hong; Caterson, Bruce; Hughes, Clare

    2013-01-01

    The objective of this study is to observe pathogenic lesions of joint cartilages in rats fed with T-2 toxin under a selenium deficiency nutrition status in order to determine possible etiological factors causing Kashin-Beck disease (KBD). Sprague-Dawley rats were fed selenium-deficient or control diets for 4 weeks prior to their being exposed to T-2 toxin. Six dietary groups were formed and studied 4 weeks later, i.e., controls, selenium-deficient, low T-2 toxin, high T-2 toxin, selenium-deficient diet plus low T-2 toxin, and selenium-deficient diet plus high T-2 toxin. Selenium deficiencies were confirmed by the determination of glutathione peroxidase activity and selenium levels in serum. The morphology and pathology (chondronecrosis) of knee joint cartilage of experimental rats were observed using light microscopy and the expression of proteoglycans was determined by histochemical staining. Chondronecrosis in deep zone of articular cartilage of knee joints was seen in both the low and high T-2 toxin plus selenium-deficient diet groups, these chondronecrotic lesions being very similar to chondronecrosis observed in human KBD. However, the chondronecrosis observed in the rat epiphyseal growth plates of animals treated with T-2 toxin alone or T-2 toxin plus selenium-deficient diets were not similar to that found in human KBD. Our results indicate that the rat can be used as a suitable animal model for studying etiological factors contributing to the pathogenesis (chondronecrosis) observed in human KBD. However, those changes seen in epiphyseal growth plate differ from those seen in human KBD probably because of the absence of growth plate closure in the rat.

  10. The critical role of p16/Rb pathway in the inhibition of GH3 cell cycle induced by T-2 toxin.

    PubMed

    Fatima, Zainab; Guo, Pu; Huang, Deyu; Lu, Qirong; Wu, Qinghua; Dai, Menghong; Cheng, Guyue; Peng, Dapeng; Tao, Yanfei; Ayub, Muhammad; Ul Qamar, Muhammad Tahir; Ali, Muhammad Waqar; Wang, Xu; Yuan, Zonghui

    2018-05-01

    T-2 toxin is a worldwide trichothecenetoxin and can cause various toxicities.T-2 toxin is involved in G1 phase arrest in several cell lines but molecular mechanism is still not clear. In present study, we used rat pituitary GH3 cells to investigate the mechanism involved in cell cycle arrest against T-2 toxin (40 nM) for 12, 24, 36 and 48 h as compared to control cells. GH3 cells showed a considerable increase in reactive oxygen species (ROS) as well as loss in mitochondrial membrane potential (△Ym) upon exposure to the T-2 toxin. Flow cytometry showed a significant time-dependent increase in percentage of apoptotic cells and gel electrophoresis showed the hallmark of apoptosis oligonucleosomal DNA fragmentation. Additionally, T-2 toxin-induced oxidative stress and DNA damage with a time-dependent significant increased expression of p53 favors the apoptotic process by the activation of caspase-3 in T-2 toxin treated cells. Cell cycle analysis by flow cytometry revealed a time-dependent increase ofG1 cell population along with the significant time-dependent up-regulation of mRNA and protein expression of p16 and p21 and significant down-regulation of cyclin D1, CDK4, and p-RB levels further verify the G1 phase arrest in GH3 cells. Morphology of GH3 cells by TEM clearly showed the damage and dysfunction to mitochondria and the cell nucleus. These findings for the first time demonstrate that T-2 toxin induces G1 phase cell cycle arrest by the involvement of p16/Rb pathway, along with ROS mediated oxidative stress and DNA damage with p53 and caspase cascade interaction, resulting in apoptosis in GH3 cells. Copyright © 2018 Elsevier B.V. All rights reserved.

  11. Glucosylation and other biotransformations of T-2 toxin by yeasts of the Trichomonascus clade

    USDA-ARS?s Scientific Manuscript database

    Twenty-five yeasts assigned to the Trichomonascus clade (Saccharomycotina, Ascomycota), including three Trichomonascus species and 22 anamorphic species presently classified in Blastobotrys, were tested for their ability to convert T-2 toxin, a Fusarium trichothecene mycotoxin, to less toxic product...

  12. Role of 2 common variants of 5HT2A gene in medication overuse headache.

    PubMed

    Terrazzino, Salvatore; Sances, Grazia; Balsamo, Francesca; Viana, Michele; Monaco, Francesco; Bellomo, Giorgio; Martignoni, Emilia; Tassorelli, Cristina; Nappi, Giuseppe; Canonico, Pier Luigi; Genazzani, Armando A

    2010-11-01

    The aim of the present study was to evaluate a possible involvement of 2 polymorphisms of the serotonin 5HT2A receptor gene (A-1438G and C516T) as risk factors for medication overuse headache (MOH) and whether the presence of these polymorphic variants might determine differences within MOH patients in monthly drug consumption. Despite a growing scientific interest in the mechanisms underlying the pathophysiology of MOH, few studies have focused on the role of genetics in the development of the disease, as well as on the genetic determinants of the inter-individual variability in the number of drug doses taken per month. Our study was performed by polymerase chain reaction (PCR) and PCR-restriction fragment length polymorphism on genomic DNA extracted from peripheral blood of 227 MOH patients and 312 control subjects. Genotype-specific risks were estimated as odds ratios with associated 95% confidence intervals by unconditional logistic regression and adjusted for age and gender. A stepwise multiple linear regression analysis was employed to identify significant predictors of the number of drug doses taken per month. No significant association was found between 5HT2A A and 1438G and C516T gene polymorphisms and MOH risk. In contrast, a higher consumption of monthly drug doses was observed among 516T 5HT2A carriers (median 50, range 13-120) compared to 516CC patients (median 30, range 12-128) (Mann-Whitney U-test, P = .018). In the stepwise multiple regression analysis, C516T 5HT2A polymorphism (P = .018) and class of overused drug (P = .047) emerged as significant, independent predictors of the monthly drug consumption in MOH patients. Although our results do not support a major role of the A-1438G and C516T polymorphic variants of the 5HT2A gene in the susceptibility of MOH, our findings support an influence of the C516T polymorphism on the number of symptomatic drug doses taken and, possibly, on the drug-seeking behavior in these patients. © 2010 American

  13. Association study of T102C 5-HT2A polymorphism in schizophrenic patients: diagnosis, psychopathology, and suicidal behavior

    PubMed Central

    Correa, Humberto; De Marco, Luiz; Boson, Wolfanga; Nicolato, Rodrigo; Teixeira, Antó L.; Campo, Valdir R.; Romano-Silva, Marco A.

    2007-01-01

    The objective of this study was to examine the association between the serotonin (5-HT)2A gene polymorphism (102T/C) and suicidal behavior in schizophrenic inpatients. We studied 129 subjects who met the diagnostic criteria for schizophrenia according to a structured clinicai interview (MINI-PLUS), Patients underwent a semistructured interview to assess suicide attempt history and its characteristics, in addition, at least one close relative of the patient was interviewed to assess prohand and family suicidal behavior. Healthy controls were students and hospital staff members free of psychiatric and medical illness. Genotypes were determined after polymerase chain reaction amplification of the region of 5-HT2A/T102C containing the polymorphic site and digestion with the restriction enzyme Hpall, We found no association between suicidal attempt history and suicide attempt characteristics and genotypic or aileie frequencies. Suicidal behavior was also not associated with demographic or psychopathological characteristics. These results suggest that the S-HT2A gene polymorphism (102T/C) is not involved in genetic susceptibility to suicidal behavior, but further studies in a larger sample are needed. PMID:17506229

  14. Hallucinogenic 5-HT2AR agonists LSD and DOI enhance dopamine D2R protomer recognition and signaling of D2-5-HT2A heteroreceptor complexes.

    PubMed

    Borroto-Escuela, Dasiel O; Romero-Fernandez, Wilber; Narvaez, Manuel; Oflijan, Julia; Agnati, Luigi F; Fuxe, Kjell

    2014-01-03

    Dopamine D2LR-serotonin 5-HT2AR heteromers were demonstrated in HEK293 cells after cotransfection of the two receptors and shown to have bidirectional receptor-receptor interactions. In the current study the existence of D2L-5-HT2A heteroreceptor complexes was demonstrated also in discrete regions of the ventral and dorsal striatum with in situ proximity ligation assays (PLA). The hallucinogenic 5-HT2AR agonists LSD and DOI but not the standard 5-HT2AR agonist TCB2 and 5-HT significantly increased the density of D2like antagonist (3)H-raclopride binding sites and significantly reduced the pKiH values of the high affinity D2R agonist binding sites in (3)H-raclopride/DA competition experiments. Similar results were obtained in HEK293 cells and in ventral striatum. The effects of the hallucinogenic 5-HT2AR agonists on D2R density and affinity were blocked by the 5-HT2A antagonist ketanserin. In a forskolin-induced CRE-luciferase reporter gene assay using cotransfected but not D2R singly transfected HEK293 cells DOI and LSD but not TCB2 significantly enhanced the D2LR agonist quinpirole induced inhibition of CRE-luciferase activity. Haloperidol blocked the effects of both quinpirole alone and the enhancing actions of DOI and LSD while ketanserin only blocked the enhancing actions of DOI and LSD. The mechanism for the allosteric enhancement of the D2R protomer recognition and signalling observed is likely mediated by a biased agonist action of the hallucinogenic 5-HT2AR agonists at the orthosteric site of the 5-HT2AR protomer. This mechanism may contribute to the psychotic actions of LSD and DOI and the D2-5-HT2A heteroreceptor complex may thus be a target for the psychotic actions of hallunicogenic 5-HT2A agonists. Copyright © 2013 Elsevier Inc. All rights reserved.

  15. Multi-mycotoxin stable isotope dilution LC-MS/MS method for Fusarium toxins in beer.

    PubMed

    Habler, Katharina; Gotthardt, Marina; Schüler, Jan; Rychlik, Michael

    2017-03-01

    A stable isotope dilution LC-MS/MS multi-mycotoxin method was developed for 12 different Fusarium toxins including modified mycotoxins in beer (deoxynivalenol-3-glucoside, deoxynivalenol, 3-acetyldeoxynivalenol, 15-acetyl-deoxynivalenol, HT2-toxin, T2-toxin, enniatin B, B1, A1, A, beauvericin and zearalenone). As sample preparation and purification of beer a combined solid phase extraction for trichothecenes, enniatins, beauvericin and zearalenone was firstly developed. The validation of the new method gave satisfying results: intra-day and inter-day precision and recoveries were 1-5%, 2-8% and 72-117%, respectively. In total, 61 different organic and conventional beer samples from Germany and all over the world were analyzed by using the newly developed multi-mycotoxin method. In summary, deoxynivalenol, deoxynivalenol-3-glucoside, 3-acetyldeoxynivaleneol and enniatin B were quantified in rather low contents in the investigated beer samples. None of the other monitored Fusarium toxins like 15-acetyldeoxynivalenol, HT2- and T2-toxin, zearalenone, enniatin B1, A1, A or beauvericin were detectable. Copyright © 2016 Elsevier Ltd. All rights reserved.

  16. Induction of apoptosis by fungal culture materials containing cyclopiazonic acid and T-2 toxin in primary lymphoid organs of broiler chickens.

    PubMed

    Venkatesh, P Kamala; Vairamuthu, S; Balachandran, C; Manohar, B Murali; Raj, G Dhinakar

    2005-04-01

    Thirty-six, twenty-eight-day-old broiler chicks were randomly distributed into three groups of 12 birds each. Two groups were fed diets containing 10 ppm cyclopiazonic acid (CPA) and 1ppm T-2 toxin, respectively, to determine the mechanism of cell death in spleen and thymus at 6, 12, 24, and 36 h of post-treatment. The other group served as control. T-2 toxin treated group showed significant (P < 0.01) induction of apoptosis in thymus with peak induction at 24 h post-treatment where as, no significant differences were observed between the control and CPA groups. The CPA toxin treated group showed significant (P < 0.01) induction of apoptosis in spleen with peak induction at 24 h post-treatment. No significant differences were observed between the control and T-2 toxin group even though the latter showed a slight increase in the quantity of apoptotic cells at 36 h post-treatment in spleen. The semi-thin sections stained with toluidine blue from the spleen of CPA treated group exhibited crescent margination of chromatin against the nuclear envelope and shrinkage of lymphoid cells without any surrounding inflammation, the characteristics of apoptosis. The apoptotic thymocytes from T-2 fed birds appeared shrunken with condensed nucleus and showed crescent margination of chromatin against the nuclear envelope without any surrounding inflammation when compared with well-defined nuclei with dispersed chromatin in normal thymocytes. Ultrastructurally, splenocytes of the CPA treated group and thymocytes of the T-2 toxin treated birds showed apoptotic bodies characterized by crescent margination of the chromatin against the nuclear envelope. The study indicates that one route of the CPA and T-2 toxin induced cell death in lymphoid organs of broiler chicken is by apoptosis.

  17. Selection and Evaluation of Indexes Commonly Used to Determine Contamination with T-2 Toxin in Pacific White Shrimp Litopenaeus vannamei by the Grey Relational Method.

    PubMed

    Lu, Pengli; Wang, Yaling; Dai, Zhe; Sun, Lijun; Xu, Defeng; Liu, Ying; Ye, Riying; Gooneratne, Ravi; Bi, Siyuan

    2017-09-01

    The objectives of the present study were to evaluate the effects of different concentrations of the mycotoxin T-2 toxin in feed on muscle performance in the Pacific white shrimp Litopenaeus vannamei, evaluate indexes of physiological variables that indicate T-2 toxin contamination in the shrimp using the grey relational method, and determine the dose-response relationships between T-2 toxin and the indexes. Of the 6 physical, 7 biochemical, and 17 nutritional indexes examined, the values of the grey relational coefficients were highest for the hepatopancreas: body weight ratio (HBR), alanine aminotransferase (ALT) activity, and serine (SER) content (0.83, 0.68, and 0.82, respectively). Therefore, the HBR, ALT activity, and SER content were selected as appropriate indexes for contamination of Pacific white shrimp muscle with T-2 toxin. Based on their dose-response relationship curves, mean effective doses of 1.45, 1.69, and 1.33 mg of T-2 toxin/kg of feed were obtained for the HBR, ALT activity, and SER content, respectively. These results offer technical reference points for the evaluation and control of T-2 toxin in shrimp feed. Received April 28, 2016; accepted April 9, 2017.

  18. 5-HT2A receptor activation is necessary for CO2-induced arousal

    PubMed Central

    Smith, Haleigh R.; MacAskill, Amanda; Richerson, George B.

    2015-01-01

    Hypercapnia-induced arousal from sleep is an important protective mechanism pertinent to a number of diseases. Most notably among these are the sudden infant death syndrome, obstructive sleep apnea and sudden unexpected death in epilepsy. Serotonin (5-HT) plays a significant role in hypercapnia-induced arousal. The mechanism of 5-HT's role in this protective response is unknown. Here we sought to identify the specific 5-HT receptor subtype(s) involved in this response. Wild-type mice were pretreated with antagonists against 5-HT receptor subtypes, as well as antagonists against adrenergic, cholinergic, histaminergic, dopaminergic, and orexinergic receptors before challenge with inspired CO2 or hypoxia. Antagonists of 5-HT2A receptors dose-dependently blocked CO2-induced arousal. The 5-HT2C receptor antagonist, RS-102221, and the 5-HT1A receptor agonist, 8-OH-DPAT, attenuated but did not completely block CO2-induced arousal. Blockade of non-5-HT receptors did not affect CO2-induced arousal. None of these drugs had any effect on hypoxia-induced arousal. 5-HT2 receptor agonists were given to mice in which 5-HT neurons had been genetically eliminated during embryonic life (Lmx1bf/f/p) and which are known to lack CO2-induced arousal. Application of agonists to 5-HT2A, but not 5-HT2C, receptors, dose-dependently restored CO2-induced arousal in these mice. These data identify the 5-HT2A receptor as an important mediator of CO2-induced arousal and suggest that, while 5-HT neurons can be independently activated to drive CO2-induced arousal, in the absence of 5-HT neurons and endogenous 5-HT, 5-HT receptor activation can act in a permissive fashion to facilitate CO2-induced arousal via another as yet unidentified chemosensor system. PMID:25925320

  19. Differences in potency and efficacy of a series of phenylisopropylamine/phenylethylamine pairs at 5-HT(2A) and 5-HT(2C) receptors.

    PubMed

    Acuña-Castillo, Claudio; Villalobos, Claudio; Moya, Pablo R; Sáez, Patricio; Cassels, Bruce K; Huidobro-Toro, J Pablo

    2002-06-01

    The pharmacological profile of a series of (+/-)-2,5-dimethoxy-4-(X)-phenylisopropylamines (X=I, Br, NO(2), CH(3), or H) and corresponding phenylethylamines, was determined in Xenopus laevis oocytes injected with cRNA coding for rat 5-HT(2A) or 5-HT(2C) receptors. The efficacy and relative potency of these drugs were determined and compared to classical 5-HT(2) receptor agonists and antagonists. The rank order of agonist potency at the 5-HT(2A) receptor was: alpha-methyl-5-HT=5-HT>m-CPP>MK-212; at the 5-HT(2C) receptor the order was: 5-HT>alpha-methyl-5-HT>MK-212>m-CPP. All these compounds were full agonists at the 5-HT(2C) receptor, but alpha-methyl-5-HT and m-CPP showed lower efficacy at the 5-HT(2A) receptor. 4-(4-Fluorobenzoyl)-1-(4-phenylbutyl)piperidine (4F 4PP) was 200 times more potent as a 5-HT(2A) antagonist than at 5-HT(2C) receptors. Conversely, RS 102221 was 100 times more potent as a 5-HT(2C) antagonist, confirming their relative receptor selectivities. The phenylisopropylamines were partial agonists at the 5-HT(2A) receptor, with I(max) relative to 5-HT in the 22+/-7 to 58+/-15% range; the corresponding phenylethylamines had lower or undetectable efficacies. All these drugs had higher efficacies at 5-HT(2C) receptors; DOI was a full 5-HT(2C) agonist. 2C-I and the other phenylethylamines examined showed relative efficacies at the 5-HT(2C) receptor ranging from 44+/-10% to 76+/-16%. 2C-N was a 5-HT(2) receptor antagonist; the mechanism was competitive at the 5-HT(2A), but non-competitive at the 5-HT(2C) receptor. The antagonism was time-dependent at the 5-HT(2C) receptor but independent of pre-incubation time at the 5-HT(2A) receptor subtype. The alpha-methyl group determines the efficacy of these phenylalkylamines at the 5-HT(2A) and 5-HT(2C) receptors.

  20. A method of analysis for T-2 toxin and neosolaniol by UPLC-MS/MS in apple fruit inoculated with Trichothecium roseum.

    PubMed

    Tang, Yamei; Xue, Huali; Bi, Yang; Li, Yongcai; Wang, Yi; Zhao, Ying; Shen, Keping

    2015-01-01

    Trichothecenes are one of the most important groups of mycotoxins produced by Trichothecium roseum, which causes core rot of apple. A reliable and sensitive method was developed and successfully applied for the rapid detection of trichothecenes including T-2 toxin and neosolaniol in harvested apple using UPLC-MS/MS. After the extraction of the two mycotoxins from the apple matrix with methanol/water (80/20, v/v), the concentrated extracts were cleaned-up by PriboFast M270 columns and then analysed by UPLC-MS/MS. T-2 toxin and neosolaniol were effectively separated as unique peaks. The validity of this method was established by its linearity (R(2) ≥ 0.9995), precision (relative standard deviation ≤ 3.6%), accuracy, selectivity, limit of detection of 2-5 μg kg(-1), limit of quantification of 5-10 μg kg(-1) and average recovery of 73-96%. Levels of T-2 toxin were found in the range 7.1-128.4 µg kg(-1) in the core rot lesion of three cultivars apple (cvs. Red Delicious, Fuji and Ralls). T-2 was detected not only in the lesion, but also in the tissue without any disease symptoms. However, neosolaniol was only detected in the lesion on 'Red Delicious' apples. In addition, the concentration of T-2 toxin in the susceptible cultivar (cv. Fuji) was significantly higher than that in the resistant one (cv. Ralls). This method proved to be suitable at detecting T-2 and neosolaniol simultaneously in apples infected with T. roseum.

  1. Incubation of cocaine cue reactivity associates with neuroadaptations in the cortical serotonin (5-HT) 5-HT2C receptor (5-HT2CR) system.

    PubMed

    Swinford-Jackson, S E; Anastasio, N C; Fox, R G; Stutz, S J; Cunningham, K A

    2016-06-02

    Intensification of craving elicited by drug-associated cues during abstinence occurs over time in human cocaine users while elevation of cue reactivity ("incubation") is observed in rats exposed to extended forced abstinence from cocaine self-administration. Incubation in rodents has been linked to time-dependent neuronal plasticity in the medial prefrontal cortex (mPFC). We tested the hypothesis that incubation of cue reactivity during abstinence from cocaine self-administration is accompanied by lower potency and/or efficacy of the selective serotonin (5-HT) 5-HT2C​ receptor (5-HT2CR) agonist WAY163909 to suppress cue reactivity and a shift in the subcellular localization profile of the mPFC 5-HT2CR protein. We observed incubation of cue reactivity (measured as lever presses reinforced by the discrete cue complex) between Day 1 and Day 30 of forced abstinence from cocaine relative to sucrose self-administration. Pharmacological and biochemical analyses revealed that the potency of the selective 5-HT2CR agonist WAY163909 to suppress cue reactivity, the expression of synaptosomal 5-HT2CR protein in the mPFC, and the membrane to cytoplasmic expression of the 5-HT2CR in mPFC were lower on Day 30 vs. Day 1 of forced abstinence from cocaine self-administration. Incubation of cue reactivity assessed during forced abstinence from sucrose self-administration did not associate with 5-HT2CR protein expression in the mPFC. Collectively, these outcomes are the first indication that neuroadaptations in the 5-HT2CR system may contribute to incubation of cocaine cue reactivity. Copyright © 2016 IBRO. Published by Elsevier Ltd. All rights reserved.

  2. Effects of acute MDMA intoxication on mood and impulsivity: role of the 5-HT2 and 5-HT1 receptors.

    PubMed

    van Wel, Janelle H P; Kuypers, Kim P C; Theunissen, Eef L; Bosker, Wendy M; Bakker, Katja; Ramaekers, Johannes G

    2012-01-01

    MDMA induces positive mood and increases impulse control during intoxication, but only a few studies on the neuropharmacological mechanisms underlying these processes have been conducted. It was hypothesized that pretreatment with 5-HT(1) and 5-HT(2) receptor blockers would prevent MDMA effects on mood and impulsivity. Subjects (N = 17) participated in a double-blind, placebo controlled, within-subject design involving 6 experimental conditions consisting of pretreatment (T1) and treatment (T2). T1 preceded T2 by 30 minutes. T1-T2 combinations were: placebo-placebo, 20 mg pindolol-placebo, 50 mg ketanserin-placebo, placebo-75 mg MDMA, 20 mg pindolol-75 mg MDMA and 50 mg ketanserin-75 g MDMA. Subjects completed a Profile of Mood States (POMS) questionnaire and several impulsivity tasks (Stop signal task, Matching familiar figures task, Cue dependent reversal learning task) at 1.5 hrs post-treatment. MDMA alone increased both positive (vigor, arousal, friendliness, elation, positive mood) and negative affect (anxiety, confusion) as assessed by the POMS questionnaire. MDMA also increased stop reaction time in the Stop signal task and reaction time in the Matching familiar figures task. Pretreatment with ketanserin blocked MDMA effects on positive affect, but not negative affect. Ketanserin did not influence the effects of MDMA on impulsivity. Pindolol did not interact with MDMA on any of the measures. In conclusion, 5-HT(2) receptors mediate positive moods induced by MDMA but not negative moods or impulsivity. 5-HT(1) receptors do not appear to be involved in MDMA effects on mood and impulse control. Nederlands Trial Register NTR2352.

  3. A Novel Aminotetralin-Type Serotonin (5-HT) 2C Receptor-Specific Agonist and 5-HT2A Competitive Antagonist/5-HT2B Inverse Agonist with Preclinical Efficacy for Psychoses

    PubMed Central

    Morgan, Drake; Felsing, Daniel; Kondabolu, Krishnakanth; Rowland, Neil E.; Robertson, Kimberly L.; Sakhuja, Rajeev; Booth, Raymond G.

    2014-01-01

    Development of 5-HT2C agonists for treatment of neuropsychiatric disorders, including psychoses, substance abuse, and obesity, has been fraught with difficulties, because the vast majority of reported 5-HT2C selective agonists also activate 5-HT2A and/or 5-HT2B receptors, potentially causing hallucinations and/or cardiac valvulopathy. Herein is described a novel, potent, and efficacious human 5-HT2C receptor agonist, (−)-trans-(2S,4R)-4-(3′[meta]-bromophenyl)-N,N-dimethyl-1,2,3,4-tetrahydronaphthalen-2-amine (−)-MBP), that is a competitive antagonist and inverse agonist at human 5-HT2A and 5-HT2B receptors, respectively. (−)-MBP has efficacy comparable to the prototypical second-generation antipsychotic drug clozapine in three C57Bl/6 mouse models of drug-induced psychoses: the head-twitch response elicited by [2,5]-dimethoxy-4-iodoamphetamine; hyperlocomotion induced by MK-801 [(5R,10S)-(+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate (dizocilpine maleate)]; and hyperlocomotion induced by amphetamine. (−)-MBP, however, does not alter locomotion when administered alone, distinguishing it from clozapine, which suppresses locomotion. Finally, consumption of highly palatable food by mice was not increased by (−)-MBP at a dose that produced at least 50% maximal efficacy in the psychoses models. Compared with (−)-MBP, the enantiomer (+)-MBP was much less active across in vitro affinity and functional assays using mouse and human receptors and also translated in vivo with comparably lower potency and efficacy. Results indicate a 5-HT2C receptor-specific agonist, such as (−)-MBP, may be pharmacotherapeutic for psychoses, without liability for obesity, hallucinations, heart disease, sedation, or motoric disorders. PMID:24563531

  4. T-2 Toxin Alters the Levels of Collagen II and Its Regulatory Enzymes MMPs/TIMP-1 in a Low-Selenium Rat Model of Kashin-Beck Disease.

    PubMed

    Zhou, Xiaorong; Yang, Haojie; Guan, Fang; Xue, Senhai; Song, Daiqin; Chen, Jinghong; Wang, Zhilun

    2016-02-01

    The objectives of this study are to assess T-2 toxin's involvement in low selenium (Se)-induced Kashin-Beck disease (KBD) in rats and unveil the mechanisms underlying this disease. Two hundred thirty rats were randomly divided into two groups after weaning and fed normal or low-Se diets (n = 115), respectively, for a month. After low-Se model confirmation, rats in each group were subdivided into five: two subgroups (n = 20) were fed their current diets (normal or low-Se diets, respectively) for 30 and 90 days, respectively; two other subgroups (n = 25) received their current diets + low T-2 toxin (100 ng/g BW/day) for 30 and 90 days, respectively; and 25 rats were fed their current diets + high T-2 toxin (200 ng/g BW/day) for 30 days. Articular cartilage samples were extracted for hematoxylin and eosin (H&E) staining and immunohistochemistry. Western blot and reverse transcription-polymerase chain reaction (RT-PCR) were used to assess protein and mRNA levels, respectively, of collagen II, matrix metalloproteinase (MMP-1), MMP -3, MMP-13, and tissue inhibitor of metalloproteinase-1 (TIMP-1). Low Se and T-2 toxin synergistically affected animal fitness. Interestingly, low Se + T-2 toxin groups showed KBD characteristics. MMP-1, -3, and -13 mRNA and protein levels generally increased in low-Se groups, while collagen II and TIMP-1 levels showed a downward trend, compared with normal diet fed animals for the same treatment (P < 0.05). T-2 toxin's effect was dose but not time dependent. Low Se and T-2 toxin synergistically alter the expression levels of collagen II as well as its regulatory enzymes MMP-1, MMP-3, MMP-13, and TIMP-1, inducing cartilage damage. Therefore, T-2 toxin may cause KBD in low-Se conditions.

  5. Effects of Acute MDMA Intoxication on Mood and Impulsivity: Role of the 5-HT2 and 5-HT1 Receptors

    PubMed Central

    van Wel, Janelle H. P.; Kuypers, Kim P. C.; Theunissen, Eef L.; Bosker, Wendy M.; Bakker, Katja; Ramaekers, Johannes G.

    2012-01-01

    MDMA induces positive mood and increases impulse control during intoxication, but only a few studies on the neuropharmacological mechanisms underlying these processes have been conducted. It was hypothesized that pretreatment with 5-HT1 and 5-HT2 receptor blockers would prevent MDMA effects on mood and impulsivity. Subjects (N = 17) participated in a double-blind, placebo controlled, within-subject design involving 6 experimental conditions consisting of pretreatment (T1) and treatment (T2). T1 preceded T2 by 30 minutes. T1–T2 combinations were: placebo-placebo, 20 mg pindolol-placebo, 50 mg ketanserin-placebo, placebo-75 mg MDMA, 20 mg pindolol-75 mg MDMA and 50 mg ketanserin-75 g MDMA. Subjects completed a Profile of Mood States (POMS) questionnaire and several impulsivity tasks (Stop signal task, Matching familiar figures task, Cue dependent reversal learning task) at 1.5 hrs post-treatment. MDMA alone increased both positive (vigor, arousal, friendliness, elation, positive mood) and negative affect (anxiety, confusion) as assessed by the POMS questionnaire. MDMA also increased stop reaction time in the Stop signal task and reaction time in the Matching familiar figures task. Pretreatment with ketanserin blocked MDMA effects on positive affect, but not negative affect. Ketanserin did not influence the effects of MDMA on impulsivity. Pindolol did not interact with MDMA on any of the measures. In conclusion, 5-HT2 receptors mediate positive moods induced by MDMA but not negative moods or impulsivity. 5-HT1 receptors do not appear to be involved in MDMA effects on mood and impulse control. Trial Registration Nederlands Trial Register NTR2352 PMID:22808116

  6. Incubation of cocaine cue reactivity associates with neuroadaptations in the cortical serotonin (5-HT) 5-HT2C receptor (5-HT2CR) system

    PubMed Central

    Swinford-Jackson, Sarah E.; Anastasio, Noelle C.; Fox, Robert G.; Stutz, Sonja J.; Cunningham, Kathryn A.

    2016-01-01

    Intensification of craving elicited by drug-associated cues during abstinence occurs over time in human cocaine users while elevation of cue reactivity (“incubation”) is observed in rats exposed to extended forced abstinence from cocaine self-administration. Incubation in rodents has been linked to time-dependent neuronal plasticity in the medial prefrontal cortex (mPFC). We tested the hypothesis that incubation of cue reactivity during abstinence from cocaine self-administration is accompanied by lower potency and/or efficacy of the selective 5-HT2CR agonist WAY163909 to suppress cue reactivity and a shift in the subcellular localization profile of the mPFC 5-HT2CR protein. We observed incubation of cue reactivity (measured as lever presses reinforced by the discrete cue complex) between Day 1 and Day 30 of forced abstinence from cocaine relative to sucrose self-administration. Pharmacological and biochemical analyses revealed that the potency of the selective 5-HT2CR agonist WAY163909 to suppress cue reactivity, the expression of synaptosomal 5-HT2CR protein in the mPFC, and the membrane to cytoplasmic expression of the 5-HT2CR in mPFC were lower on Day 30 vs. Day 1 of forced abstinence from cocaine self-administration. Incubation of cue reactivity assessed during forced abstinence from sucrose self-administration did not associate with 5-HT2CR protein expression in the mPFC. Collectively, these outcomes are the first indication that neuroadaptations in the 5-HT2CR system may contribute to incubation of cocaine cue reactivity. PMID:26926963

  7. Immunological techniques for detection of fungal and dinoflagellate toxins. Annual report, 1 May 1987-30 April 1988

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Chu, F.S.

    1988-05-12

    1) A new direct ELISA, which involves the use of a generic type antibody for T-2 toxin and 3-acetyl-neosolaniol-hemisuccinate (3-Ac-NEOS-HS) conjugated to horseradish peroxidase for the analysis of T-2 toxin metabolites, was established. 2) Analysis of 5 metabolites, i.e., T-2 toxin, HT-2, T-2-4ol, 3'-OH-HT-2, NEOS and a mixture of these five toxins, added at toxin concentrations of 50 and 200 ng/mL of urine, revealed that 87% of the added toxins were recovered analytically at both concentrations in the ELISA, and 69% in RIA and the 200 ppb level. 3) A series of monkey urine samples obtained from a metabolic studymore » was subjected to the new ELISA protocol was as well as to RIA. 4) An extensive study to evaluate a fluorometric method, i.e., hit-and-run assay was made. 5) A direct ELISA for deoxynivalenol (DON) was established. 6) Further improvements for the ELISA for saxitoxin were made. 7) New approaches to produce antibody against nivalenol and saxitoxin were made, but the antibody titers were low. 8) Antibody against microcystin (MCS) was obtained from rabbits after immunizing the animals with MCS-BSA conjugate. 9) A chemical method for the preparation of different monoacetoxyscirpenol (MAS) was established. 10) Routine preparation of different immunochemical reagents continued. Phytotoxins Biological warfare agents; Dinoflagellates. (AW)« less

  8. Serum Metabonomics of Articular Cartilage Destruction Induced by T-2 Toxin in Wistar Rats.

    PubMed

    Zhu, Lei; Zhao, Zhi Jun; Ren, Xiao Bin; Li, Qiang; Ding, Hua; Sun, Zhou; Kao, Qing Jun; Wang, Li Hua

    2018-01-01

    The molecular pathogenesis of T-2 toxin-induced cartilage destruction has not been fully unraveled yet. The aim of this study was to detect changes in serum metabolites in a rat anomaly model with articular cartilage destruction. Thirty healthy male Wistar rats were fed a diet containing T-2 toxin (300 ng/kg chow) for 3 months. Histopathological changes in femorotibial cartilage were characterized in terms of chondrocyte degeneration/necrosis and superficial cartilage defect, and the endogenous metabolite profile of serum was determined by UPLC/Q-TOF MS. Treated rats showed extensive areas of chondrocyte necrosis and superficial cartilage defect in the articular cartilage. In addition, 8 metabolites were found to change significantly in these rats compared to the control group, including lysoPE (18:0/0:0), lysoPC(14:0), lysoPC[18:4 (6Z,9Z,12Z,15Z)], lysoPC[(16:1(9Z)], lysoPC(16:0), L-valine, hippuric acid, and asparaginyl-glycine. These 8 metabolites associated with cartilage injury are mainly involved in phospholipid and amino acid metabolic pathways. Copyright © 2018 The Editorial Board of Biomedical and Environmental Sciences. Published by China CDC. All rights reserved.

  9. The Adenylate Cyclase Toxins of Bacillus anthracis and Bordetella pertussis Promote Th2 Cell Development by Shaping T Cell Antigen Receptor Signaling

    PubMed Central

    Rossi Paccani, Silvia; Benagiano, Marisa; Capitani, Nagaja; Zornetta, Irene; Ladant, Daniel; Montecucco, Cesare; D'Elios, Mario M.; Baldari, Cosima T.

    2009-01-01

    The adjuvanticity of bacterial adenylate cyclase toxins has been ascribed to their capacity, largely mediated by cAMP, to modulate APC activation, resulting in the expression of Th2–driving cytokines. On the other hand, cAMP has been demonstrated to induce a Th2 bias when present during T cell priming, suggesting that bacterial cAMP elevating toxins may directly affect the Th1/Th2 balance. Here we have investigated the effects on human CD4+ T cell differentiation of two adenylate cyclase toxins, Bacillus anthracis edema toxin (ET) and Bordetella pertussis CyaA, which differ in structure, mode of cell entry, and subcellular localization. We show that low concentrations of ET and CyaA, but not of their genetically detoxified adenylate cyclase defective counterparts, potently promote Th2 cell differentiation by inducing expression of the master Th2 transcription factors, c-maf and GATA-3. We also present evidence that the Th2–polarizing concentrations of ET and CyaA selectively inhibit TCR–dependent activation of Akt1, which is required for Th1 cell differentiation, while enhancing the activation of two TCR–signaling mediators, Vav1 and p38, implicated in Th2 cell differentiation. This is at variance from the immunosuppressive toxin concentrations, which interfere with the earliest step in TCR signaling, activation of the tyrosine kinase Lck, resulting in impaired CD3ζ phosphorylation and inhibition of TCR coupling to ZAP-70 and Erk activation. These results demonstrate that, notwithstanding their differences in their intracellular localization, which result in focalized cAMP production, both toxins directly affect the Th1/Th2 balance by interfering with the same steps in TCR signaling, and suggest that their adjuvanticity is likely to result from their combined effects on APC and CD4+ T cells. Furthermore, our results strongly support the key role of cAMP in the adjuvanticity of these toxins. PMID:19266022

  10. Family-based association study of 5-HT(2A) receptor T102C polymorphism and suicidal behavior in Ashkenazi inpatient adolescents.

    PubMed

    Zalsman, Gil; Frisch, Amos; Baruch-Movshovits, Ruth; Sher, Leo; Michaelovsky, Elena; King, Robert A; Fischel, Tsvi; Hermesh, Haggai; Goldberg, Pablo; Gorlyn, Marianne; Misgav, Sagit; Apter, Alan; Tyano, Sam; Weizman, Abraham

    2005-01-01

    Suicidal behavior runs in families and is partially genetically determined. Since greater serotonin 5-HT(2A) receptor binding has been reported in postmortem brain and platelets of suicide victims, the 5-HT(2A) receptor gene polymorphism T102C became one of the candidate sites in the study of suicide and impulsive-aggressive traits. However, studies that examined the association of this polymorphism with suicidality have contradictory results. This study used a family-based method and one homogenous ethnic group to overcome ethnic stratification in order to test this association. Thirty families of inpatient adolescents from Jewish Ashkenazi origin, with a recent suicide attempt, were genotyped. All subjects were interviewed for clinical diagnosis, depressive and impulsive-aggressive traits and demographic data. Allele frequencies were assessed using the Haplotype Relative Risk method for trios. No difference was found in allelic distribution between transmitted and non-transmitted alleles. There was no significant association of genotype with any of the clinical traits These preliminary results suggest that the 5-HT(2A) T102C polymorphism is unlikely to be associated with suicidal behavior and related traits in adolescent suicide attempters.

  11. T-2 toxin contamination in grains and selenium concentration in drinking water and grains in Kaschin-Beck disease endemic areas of Qinghai Province.

    PubMed

    Sun, Li-Yan; Li, Qiang; Meng, Fan-Gang; Fu, Ying; Zhao, Zhi-Jun; Wang, Li-Hua

    2012-12-01

    It has been strongly suggested that two factors are involved in the development of Kaschin-Beck Disease (KBD), namely grains contamination with T-2 toxin and selenium deficiency. So our team undertook a survey about grains and drinking water in three rural KBD endemic villages and one non-KBD village in Qinghai Province. The level of T-2 toxin contamination in 364 grain samples was assayed using an ELISA kit. The selenium concentration in these grains and 15 drinking water samples from three KBD endemic villages were determined using the 2,3-diaminonaphthalene fluorometric assay. The results revealed that the level of T-2 toxin contamination in the samples from three KBD endemic villages was relatively high with an average level of 78.91 ng/g in wheat and 47.47 ng/g in flour. The T-2 toxin level in samples from the non-KBD village (12.23 ng/g) was significantly lower than that of local grains from the three KBD endemic villages. The average selenium content in wheat and flour from KBD areas was 0.0045 and 0.0067 μg/g, respectively. The selenium concentration in local grain samples was significantly lower than that in samples from the non-KBD village (0.0604 μg/g). In addition, the selenium concentration in drinking water from three KBD endemic villages was also low (0.156 μg/L). These results support a potential role of T-2 toxin contamination and selenium deficiency in KBD. Compared with non-KBD endemic areas, health hazards in grains and in the environment of KBD endemic areas were observed.

  12. A Versatile Family 3 Glycoside Hydrolase from Bifidobacterium adolescentis Hydrolyzes β-Glucosides of the Fusarium Mycotoxins Deoxynivalenol, Nivalenol, and HT-2 Toxin in Cereal Matrices

    PubMed Central

    Varga, Elisabeth; Malachova, Alexandra; Nguyen, Nhung Thi; Lorenz, Cindy; Haltrich, Dietmar; Berthiller, Franz; Adam, Gerhard

    2015-01-01

    Glycosylation plays a central role in plant defense against xenobiotics, including mycotoxins. Glucoconjugates of Fusarium toxins, such as deoxynivalenol-3-O-β-d-glucoside (DON-3G), often cooccur with their parental toxins in cereal-based food and feed. To date, only limited information exists on the occurrence of glucosylated mycotoxins and their toxicological relevance. Due to a lack of analytical standards and the requirement of high-end analytical instrumentation for their direct determination, hydrolytic cleavage of β-glucosides followed by analysis of the released parental toxins has been proposed as an indirect determination approach. This study compares the abilities of several fungal and recombinant bacterial β-glucosidases to hydrolyze the model analyte DON-3G. Furthermore, substrate specificities of two fungal and two bacterial (Lactobacillus brevis and Bifidobacterium adolescentis) glycoside hydrolase family 3 β-glucosidases were evaluated on a broader range of substrates. The purified recombinant enzyme from B. adolescentis (BaBgl) displayed high flexibility in substrate specificity and exerted the highest hydrolytic activity toward 3-O-β-d-glucosides of the trichothecenes deoxynivalenol (DON), nivalenol, and HT-2 toxin. A Km of 5.4 mM and a Vmax of 16 μmol min−1 mg−1 were determined with DON-3G. Due to low product inhibition (DON and glucose) and sufficient activity in several extracts of cereal matrices, this enzyme has the potential to be used for indirect analyses of trichothecene-β-glucosides in cereal samples. PMID:25979885

  13. The effect of chronic feeding of diacetoxyscirpenol and T-2 toxin on performance, health, small intestinal physiology and antibody production in turkey poults.

    PubMed

    Sklan, D; Shelly, M; Makovsky, B; Geyra, A; Klipper, E; Friedman, A

    2003-03-01

    1. The effects of feeding T-2 toxin or diacetoxyscirpenol (DAS) at levels up to 1 ppm for 32 d on performance, health, small intestinal physiology and immune response to enteral and parenteral immunisation were examined in young poults. 2. Slight improvement in growth was observed in some groups of poults fed T-2 or DAS mycotoxins for 32 d, with no change in feed efficiency. Feeding both T-2 and DAS resulted in oral lesions which had maximal severity after 7-15 d. 3. Mild intestinal changes were observed at 32 d but no pathological or histopathological lesions were found. Both mycotoxins altered small intestinal morphology, especially in the jejunum where villi were shorter and thinner. In addition, both DAS and T-2 mycotoxins enhanced the proportion of proliferating cells both in the crypts and along the villi. Migration rates were reduced in the jejunum of poults fed T-2 toxin but did not change in the duodenum or in poults fed DAS. 4. No significant effects of T-2 or DAS were observed on antibody production to antigens administered by enteral or parenteral routes. 5. This study indicates that tricothecene toxins at concentrations of up to 1 ppm for more than 30 d influenced small intestinal morphology but did not affect growth or antibody production.

  14. Haliotis tuberculata hemocyanin (HtH): analysis of oligomeric stability of HtH1 and HtH2, and comparison with keyhole limpet hemocyanin KLH1 and KLH2.

    PubMed

    Harris, J R; Scheffler, D; Gebauer, W; Lehnert, R; Markl, J

    2000-12-01

    The multimeric/higher oligomeric states of the two isoforms of Haliotis tuberculata hemocyanin (HtH1 and HtH2) have been assessed by transmission electron microscopy (TEM) of negatively stained specimens, for comparison with previously published structural data from keyhole limpet hemocyanin (KLH1 and KLH2) [see Harris, J.R., Gebauer, W., Guderian, F.U., Markl, J., 1997a. Keyhole limpet hemocyanin (KLH), I: Reassociation from Immucothel followed by separation of KLH1 and KLH2. Micron, 28, 31-41; Harris, J.R., Gebauer, W., Söhngen, S.M., Nermut, M.V., Markl, J., 1997b. Keyhole limpet hemocyanin (KLH). II: Characteristic reassociation properties of purified KLH1 and KLH2. Micron, 28, 43-56; Harris, J.R., Gebauer, W., Adrian, M., Markl, J., 1998. Keyhole limpet hemocyanin (KLH): Slow in vitro reassociation of KLH1 and KLH2 from Immucothel. Micron, 29, 329-339]. In purified samples of both HtH isoforms, the hollow cylindrical ca 8MDa didecamer predominates together with a small number of decamers, but tri- and longer multidecamers are detectable only in the HtH2. The stability of the two HtH isoforms under varying ionic conditions have been monitored, thereby enabling conditions for the production of stable decamers to be established. The ability of these decamers to reform multimers in the presence of 10 and 100mM concentrations of calcium and magnesium ions in Tris-HCl buffer (pH 7.4), and also of individual HtH1 and HtH2 subunits (produced by pH 9.6 dissociation in glycine-NaOH buffer), to reassociate in the presence of calcium and magnesium ions, has been assessed. For the HtH1 decamers, the predominant multimeric product is the didecamer at 10 and 100mM calcium and magnesium concentrations, whereas for the HtH2 decamers, large numbers of multidecamers are produced, with the reaction proceeding more completely at the higher calcium and magnesium concentration. With the HtH1 subunit, reassociation in the presence of 10 and 100mM calcium and magnesium ions yielded

  15. Monoamine receptor interaction profiles of 4-thio-substituted phenethylamines (2C-T drugs).

    PubMed

    Luethi, Dino; Trachsel, Daniel; Hoener, Marius C; Liechti, Matthias E

    2018-05-15

    4-Thio-substituted phenethylamines (2C-T drugs) are potent psychedelics with poorly defined pharmacological properties. Because of their psychedelic effects, 2C-T drugs are sometimes sold as new psychoactive substances (NPSs). The aim of the present study was to characterize the monoamine receptor and transporter interaction profiles of a series of 2C-T drugs. We determined the binding affinities of 2C-T drugs at monoamine receptors and transporters in human cells that were transfected with the respective receptors or transporters. We also investigated the functional activation of serotonergic 5-hydroxytryptamine 2A (5-HT 2A ) and 5-HT 2B receptors, activation of human trace amine-associated receptor 1 (TAAR 1 ), and inhibition of monoamine uptake transporters. 2C-T drugs had high affinity for 5-HT 2A and 5-HT 2C receptors (1-54 nM and 40-350 nM, respectively). With activation potencies of 1-53 nM and 44-370 nM, the drugs were potent 5-HT 2A receptor and 5-HT 2B receptor, respectively, partial agonists. An exception to this were the benzylthiophenethylamines, which did not potently activate the 5-HT 2B receptor (EC 50  > 3000 nM). Furthermore, the compounds bound to serotonergic 5-HT 1A and adrenergic receptors. The compounds had high affinity for the rat TAAR 1 (5-68 nM) and interacted with the mouse but not human TAAR 1 . The 2C-T drugs did not potently interact with monoamine transporters (K i  > 4000 nM). The receptor binding profile of 2C-T drugs predicts psychedelic effects that are mediated by potent 5-HT 2 receptor interactions. This article is part of the Special Issue entitled 'Designer Drugs and Legal Highs.' Copyright © 2017 Elsevier Ltd. All rights reserved.

  16. Design and Discovery of Functionally Selective Serotonin 2C (5-HT2C) Receptor Agonists.

    PubMed

    Cheng, Jianjun; McCorvy, John D; Giguere, Patrick M; Zhu, Hu; Kenakin, Terry; Roth, Bryan L; Kozikowski, Alan P

    2016-11-10

    On the basis of the structural similarity of our previous 5-HT 2C agonists with the melatonin receptor agonist tasimelteon and the putative biological cross-talk between serotonergic and melatonergic systems, a series of new (2,3-dihydro)benzofuran-based compounds were designed and synthesized. The compounds were evaluated for their selectivity toward 5-HT 2A , 5-HT 2B , and 5-HT 2C receptors in the calcium flux assay with the ultimate goal to generate selective 5-HT 2C agonists. Selected compounds were studied for their functional selectivity by comparing their transduction efficiency at the G protein signaling pathway versus β-arrestin recruitment. The most functionally selective compound (+)-7e produced weak β-arrestin recruitment and also demonstrated less receptor desensitization compared to serotonin in both calcium flux and phosphoinositide (PI) hydrolysis assays. We report for the first time that selective 5-HT 2C agonists possessing weak β-arrestin recruitment can produce distinct receptor desensitization properties.

  17. Effect of disruption of actin filaments by Clostridium botulinum C2 toxin on insulin secretion in HIT-T15 cells and pancreatic islets.

    PubMed Central

    Li, G; Rungger-Brändle, E; Just, I; Jonas, J C; Aktories, K; Wollheim, C B

    1994-01-01

    To examine their role in insulin secretion, actin filaments (AFs) were disrupted by Clostridium botulinum C2 toxin that ADP-ribosylates G-actin. Ribosylation also prevents polymerization of G-actin to F-actin and inhibits AF assembly by capping the fast-growing end of F-actin. Pretreatment of HIT-T15 cells with the toxin inhibited stimulated insulin secretion in a time- and dose-dependent manner. The toxin did not affect cellular insulin content or nonstimulated secretion. In static incubation, toxin treatment caused 45-50% inhibition of secretion induced by nutrients alone (10 mM glucose + 5 mM glutamine + 5 mM leucine) or combined with bombesin (phospholipase C-activator) and 20% reduction of that potentiated by forskolin (stimulator of adenylyl cyclase). In perifusion, the stimulated secretion during the first phase was marginally diminished, whereas the second phase was inhibited by approximately 80%. Pretreatment of HIT cells with wartmannin, a myosin light chain kinase inhibitor, caused a similar pattern of inhibition of the biphasic insulin release as C2 toxin. Nutrient metabolism and bombesin-evoked rise in cytosolic free Ca2+ were not affected by C2 toxin, indicating that nutrient recognition and the coupling between receptor activation and second messenger generation was not changed. In the toxin-treated cells, the AF web beneath the plasma membrane and the diffuse cytoplasmic F-actin fibers disappeared, as shown both by staining with an antibody against G- and F-actin and by staining F-actin with fluorescent phallacidin. C2 toxin dose-dependently reduced cellular F-actin content. Stimulation of insulin secretion was not associated with changes in F-actin content and organization. Treatment of cells with cytochalasin E and B, which shorten AFs, inhibited the stimulated insulin release by 30-50% although differing in their effects on F-actin content. In contrast to HIT-T15 cells, insulin secretion was potentiated in isolated rat islets after disruption of

  18. Acute Inhalation Toxicity of T-2 Mycotoxin in the Rat and Guinea Pig

    DTIC Science & Technology

    1990-01-01

    2/kg body weight for the guinea pig . These data show that inhaled T-2 toxin is approximately 20 times more toxic to the rat (0.05 mg T-2/kg body wt...inhaled vs 1.0 mg T-2/kg body wt ip) and at least twice as toxic to the guinea pig (0.4 mg T-2/kg body wt inhaled vs 1-2 mg T-2/kg body wt ip) than ip...administered T-2 toxin. Histopathologic examination of major organs in both the rat and guinea pig after respiratory exposure to T-2 toxin indicated

  19. Spectrodensitometric Quantitative Determination of Trichothecenes in Water: Application to T-2 Toxin and T-2 Tetraol

    DTIC Science & Technology

    1984-10-01

    3-hydroxy-8-( 3-methybutyryloxy)-1 2,13-epoxytrichothec- *. 9-ene] (la) is an important member of the 12,13-epoxytrichochecene class of mycotoxins ...produced in nat by a number of genera of fungi and recognized increasingly since the late .960’s as responsible for toxic effects in live- stock... poultry , and humans from consumption of contaminated food grains. 1 More recently, since 1981, the issue of possible use of these toxins as " chemical

  20. Chronic 5-HT2 receptor blockade unmasks the role of 5-HT1F receptors in the inhibition of rat cardioaccelerator sympathetic outflow.

    PubMed

    García-Pedraza, José Ángel; Hernández-Abreu, Oswaldo; García, Mónica; Morán, Asunción; Villalón, Carlos M

    2018-04-01

    Serotonin (5-hydroxytryptamine; 5-HT) inhibits the rat cardioaccelerator sympathetic outflow by 5-HT 1B/1D/5 receptors. Because chronic blockade of sympatho-excitatory 5-HT 2 receptors is beneficial in several cardiovascular pathologies, this study investigated whether sarpogrelate (a 5-HT 2 receptor antagonist) alters the pharmacological profile of the above sympatho-inhibition. Rats were pretreated for 2 weeks with sarpogrelate in drinking water (30 mg/kg per day; sarpogrelate-treated group) or equivalent volumes of drinking water (control group). Animals were pithed and prepared for spinal stimulation (C 7 -T 1 ) of the cardioaccelerator sympathetic outflow or for intravenous (i.v.) bolus injections of noradrenaline. Both procedures produced tachycardic responses remaining unaltered after saline. Continuous i.v. infusions of 5-HT induced a cardiac sympatho-inhibition that was mimicked by the 5-HT receptor agonists 5-carboxamidotryptamine (5-CT; 5-HT 1/5A ), CP 93,129 (5-HT 1B ), or PNU 142633 (5-HT 1D ), but not by indorenate (5-HT 1A ) in both groups; whereas LY344864 (5-HT 1F ) mimicked 5-HT only in sarpogrelate-treated rats. In sarpogrelate-treated animals, i.v. GR 127935 (310 μg/kg; 5-HT 1B/1D/1F receptor antagonist) attenuated 5-CT-induced sympatho-inhibition and abolished LY344864-induced sympatho-inhibition; while GR 127935 plus SB 699551 (1 mg/kg; 5-HT 5A receptor antagonist) abolished 5-CT-induced inhibition. These results confirm the cardiac sympatho-inhibitory role of 5-HT 1B , 5-HT 1D , and 5-HT 5A receptors in both groups; nevertheless, sarpogrelate treatment specifically unmasked a cardiac sympatho-inhibition mediated by 5-HT 1F receptors.

  1. Increased hypothalamic 5-HT2A receptor gene expression and effects of pharmacologic 5-HT2A receptor inactivation in obese A{sup y} mice

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Nonogaki, Katsunori; Nozue, Kana; Oka, Yoshitomo

    2006-12-29

    Serotonin (5-hydroxytryptamine; 5-HT) 2A receptors contribute to the effects of 5-HT on platelet aggregation and vascular smooth muscle cell proliferation, and are reportedly involved in decreases in plasma levels of adiponectin, an adipokine, in diabetic subjects. Here, we report that systemic administration of sarpogrelate, a 5-HT2A receptor antagonist, suppressed appetite and increased hypothalamic pro-opiomelanocortin and cocaine- and amphetamine-regulated transcript, corticotropin releasing hormone, 5-HT2C, and 5-HT1B receptor gene expression. A{sup y} mice, which have ectopic expression of the agouti protein, significantly increased hypothalamic 5-HT2A receptor gene expression in association with obesity compared with wild-type mice matched for age. Systemic administration ofmore » sarpogrelate suppressed overfeeding, body weight gain, and hyperglycemia in obese A{sup y} mice, whereas it did not increase plasma adiponectin levels. These results suggest that obesity increases hypothalamic 5-HT2A receptor gene expression, and pharmacologic inactivation of 5-HT2A receptors inhibits overfeeding and obesity in A{sup y} mice, but did not increase plasma adiponectin levels.« less

  2. 5-HT2A/5-HT2C Receptor Pharmacology and Intrinsic Clearance of N-Benzylphenethylamines Modified at the Primary Site of Metabolism.

    PubMed

    Leth-Petersen, Sebastian; Petersen, Ida N; Jensen, Anders A; Bundgaard, Christoffer; Bæk, Mathias; Kehler, Jan; Kristensen, Jesper L

    2016-11-16

    The toxic hallucinogen 25B-NBOMe is very rapidly degraded by human liver microsomes and has low oral bioavailability. Herein we report on the synthesis, microsomal stability, and 5-HT 2A /5-HT 2C receptor profile of novel analogues of 25B-NBOMe modified at the primary site of metabolism. Although microsomal stability could be increased while maintaining potent 5-HT 2 receptor agonist properties, all analogues had an intrinsic clearance above 1.3 L/kg/h predictive of high first-pass metabolism.

  3. 5-HT2C Agonists Modulate Schizophrenia-Like Behaviors in Mice.

    PubMed

    Pogorelov, Vladimir M; Rodriguiz, Ramona M; Cheng, Jianjun; Huang, Mei; Schmerberg, Claire M; Meltzer, Herbert Y; Roth, Bryan L; Kozikowski, Alan P; Wetsel, William C

    2017-10-01

    All FDA-approved antipsychotic drugs (APDs) target primarily dopamine D 2 or serotonin (5-HT 2A ) receptors, or both; however, these medications are not universally effective, they may produce undesirable side effects, and provide only partial amelioration of negative and cognitive symptoms. The heterogeneity of pharmacological responses in schizophrenic patients suggests that additional drug targets may be effective in improving aspects of this syndrome. Recent evidence suggests that 5-HT 2C receptors may be a promising target for schizophrenia since their activation reduces mesolimbic nigrostriatal dopamine release (which conveys antipsychotic action), they are expressed almost exclusively in CNS, and have weight-loss-promoting capabilities. A difficulty in developing 5-HT 2C agonists is that most ligands also possess 5-HT 2B and/or 5-HT 2A activities. We have developed selective 5-HT 2C ligands and herein describe their preclinical effectiveness for treating schizophrenia-like behaviors. JJ-3-45, JJ-3-42, and JJ-5-34 reduced amphetamine-stimulated hyperlocomotion, restored amphetamine-disrupted prepulse inhibition, improved social behavior, and novel object recognition memory in NMDA receptor hypofunctioning NR1-knockdown mice, and were essentially devoid of catalepsy. However, they decreased motivation in a breakpoint assay and did not promote reversal learning in MK-801-treated mice. Somewhat similar effects were observed with lorcaserin, a 5-HT 2C agonist with potent 5-HT 2B and 5-HT 2A agonist activities, which is approved for treating obesity. Microdialysis studies revealed that both JJ-3-42 and lorcaserin reduced dopamine efflux in the infralimbic cortex, while only JJ-3-42 decreased it in striatum. Collectively, these results provide additional evidence that 5-HT 2C receptors are suitable drug targets with fewer side effects, greater therapeutic selectivity, and enhanced efficacy for treating schizophrenia and related disorders than current APDs.

  4. Evaluation of two mycotoxin binders to reduce toxicity of broiler diets containing ochratoxin A and T-2 toxin contaminated grain.

    PubMed

    García, A R; Avila, E; Rosiles, R; Petrone, V M

    2003-01-01

    In order to assess ochratoxin A (OA) and T-2 toxin (T-2) binding ability of two commercial sorbents, both in vitro and in vivo trials with broilers were performed. Crude OA and T-2 extracts from contaminated grain were used to assess in vitro binding ability of two sorbents (Zeotek [Zk] and Mycofix [Mx]), by quantifying free mycotoxin through an enzyme-linked immunosorbent assay (ELISA) test. For in vivo trial, a 3 x 2 x 2 factorial arrangement was used for this experiment, being the factors: adsorbents (none, Zk, and Mx), OA (0 and 567 parts per billion [ppb]) and T-2 (0 and 927 ppb). OA and T-2 contaminated wheat and corn, respectively, were added to sorghum-soybean meal diets to meet 567 ppb of OA and 927 ppb of T-2. Mycotoxins were fed alone or combined in treatments. After 21 days, blood chemistry, gross, and histological evaluations were performed. Relative weights of liver, kidney, and bursa of Fabricius were obtained. Zk had the highest OA and T-2 in vitro binding ability (100% and 8.67%, respectively). Chickens fed OA with or without sorbents had a lower body weight and feed intake reduction. However, those birds fed T-2 were partly protected by a sorbent. Birds fed both toxins showed toxic additive effects, and no protection of any adsorbent was observed. A significant reduction in plasma proteins, albumin, and globulins was a characteristic observed in all birds fed diets with OA both with or without adsorbents. Uric acid level in blood was increased in all chickens fed OA-contaminated diets. Histological findings observed in birds fed OA-contaminated diets were necrosis of kidney tubular cells, swollen and necrotic hepatocytes, bile ducts hyperplasia, and increased diameter of proventriculus glands. In birds that received T-2 alone, only the liver, with the same kind of lesions, was affected. According to these results, it can be concluded that there is not a relation between in vitro and in vivo trials. OA toxic effects could not be counteracted by any

  5. The antidepressant effects of curcumin in the forced swimming test involve 5-HT1 and 5-HT2 receptors.

    PubMed

    Wang, Rui; Xu, Ying; Wu, Hong-Li; Li, Ying-Bo; Li, Yu-Hua; Guo, Jia-Bin; Li, Xue-Jun

    2008-01-06

    Curcuma longa is a main constituent of many traditional Chinese medicines, such as Xiaoyao-san, used to manage mental disorders effectively. Curcumin is a major active component of C. longa and its antidepressant-like effect has been previously demonstrated in the forced swimming test. The purpose of this study was to explore the possible contribution of serotonin (5-HT) receptors in the behavioral effects induced by curcumin in this animal model of depression. 5-HT was depleted by the tryptophan hydroxylase inhibitor p-chlorophenylalanine (PCPA, 100 mg/kg, i.p.) prior to the administration of curcumin, and the consequent results showed that PCPA blocked the anti-immobility effect of curcumin in forced swimming test, suggesting the involvement of the serotonergic system. Moreover, pre-treatment of pindolol (10 mg/kg, i.p., a beta-adrenoceptors blocker/5-HT(1A/1B) receptor antagonist), 4-(2'-methoxy-phenyl)-1-[2'-(n-2''-pyridinyl)-p-iodobenzamino-]ethyl-piperazine (p-MPPI, 1 mg/kg, s.c., a selective 5-HT(1A) receptor antagonist), or 1-(2-(1-pyrrolyl)-phenoxy)-3-isopropylamino-2-propanol (isamoltane, 2.5 mg/kg, i.p., a 5-HT(1B) receptor antagonist) was found to prevent the effect of curcumin (10 mg/kg) in forced swimming test. On the other hand, a sub-effective dose of curcumin (2.5 mg/kg, p.o.) produced a synergistic effect when given jointly with (+)-8-hydroxy-2-(di-n-propylamino)tetralin, (8-OH-DPAT, 1 mg/kg, i.p., a 5-HT(1A) receptor agonist), anpirtoline (0.25 mg/kg, i.p., a 5-HT(1B) receptor agonist) or ritanserin (4 mg/kg, i.p., a 5-HT(2A/2C) receptor antagonist), but not with ketanserin (5 mg/kg, i.p., a 5-HT(2A/2C) receptor antagonist with higher affinity to 5-HT(2A) receptor) or R(-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI, 1 mg/kg, i.p., a 5-HT(2A) receptor agonist). Taken together, these results indicate that the antidepressant-like effect of curcumin in the forced swimming test is related to serotonergic system and may be mediated by, at least

  6. Synthesis and structure-activity relationships of N-benzyl phenethylamines as 5-HT2A/2C agonists.

    PubMed

    Hansen, Martin; Phonekeo, Karina; Paine, James S; Leth-Petersen, Sebastian; Begtrup, Mikael; Bräuner-Osborne, Hans; Kristensen, Jesper L

    2014-03-19

    N-Benzyl substitution of 5-HT2A receptor agonists of the phenethylamine structural class of psychedelics (such as 4-bromo-2,5-dimethoxyphenethylamine, often referred to as 2C-B) confer a significant increase in binding affinity as well as functional activity of the receptor. We have prepared a series of 48 compounds with structural variations in both the phenethylamine and N-benzyl part of the molecule to determine the effects on receptor binding affinity and functional activity at 5-HT2A and 5-HT2C receptors. The compounds generally had high affinity for the 5-HT2A receptor with 8b having the highest affinity at 0.29 nM but with several other compounds also exhibiting subnanomolar binding affinities. The functional activity of the compounds was distributed over a wider range with 1b being the most potent at 0.074 nM. Most of the compounds exhibited low to moderate selectivity (1- to 40-fold) for the 5-HT2A receptor in the binding assays, although one compound 6b showed an impressive 100-fold selectivity for the 5-HT2A receptor. In the functional assay, selectivity was generally higher with 1b being more than 400-fold selective for the 5-HT2A receptor.

  7. The role of serotonin 5-HT2A receptors in memory and cognition

    PubMed Central

    Zhang, Gongliang; Stackman, Robert W.

    2015-01-01

    Serotonin 5-HT2A receptors (5-HT2ARs) are widely distributed in the central nervous system, especially in brain region essential for learning and cognition. In addition to endogenous 5-HT, several hallucinogens, antipsychotics, and antidepressants function by targeting 5-HT2ARs. Preclinical studies show that 5-HT2AR antagonists have antipsychotic and antidepressant properties, whereas agonist ligands possess cognition-enhancing and hallucinogenic properties. Abnormal 5-HT2AR activity is associated with a number of psychiatric disorders and conditions, including depression, schizophrenia, and drug addiction. In addition to its traditional activity as a G protein-coupled receptor (GPCR), recent studies have defined novel operations of 5-HT2ARs. Here we review progress in the (1) receptor anatomy and biology: distribution, signaling, polymerization and allosteric modulation; and (2) receptor functions: learning and memory, hallucination and spatial cognition, and mental disorders. Based on the recent progress in basic research on the 5-HT2AR, it appears that post-training 5-HT2AR activation enhances non-spatial memory consolidation, while pre-training 5-HT2AR activation facilitates fear extinction. Further, the potential influence that 5-HT2AR-elicited visual hallucinations may have on visual cue (i.e., landmark) guided spatial cognition is discussed. We conclude that the development of selective 5-HT2AR modulators to target distinct signaling pathways and neural circuits represents a new possibility for treating emotional, neuropsychiatric, and neurodegenerative disorders. PMID:26500553

  8. On the magnetism of the C14 Nb0.975Fe2.025 Laves phase compound: Determination of the H-T phase diagram

    NASA Astrophysics Data System (ADS)

    Bałanda, Maria; Dubiel, Stanisław M.

    2018-05-01

    A C14 Nb0.975Fe2.025 Laves phase compound was investigated aimed at determining the H-T magnetic phase diagram. Magnetization, M, and AC magnetic susceptibility measurement were performed. Concerning the former field-cooled and zero-field-cooled M-curves were recorded in the temperature range of 2-200 K and in applied magnetic field, H, up to 1000 Oe, isothermal M(H) curves at 2 K, 5 K, 50 K, 80 K and 110 K as well as hysteresis loops at several temperatures over the field range of ±10 kOe were measured. Regarding the AC susceptibility, χ, both real and imaginary components were registered as a function of increasing temperature in the interval of 2 K-150 K at the frequencies of the oscillating field, f, from 3 Hz up to 999 Hz. An influence of the external DC magnetic field on the temperature dependence of χ was investigated, too. The measurements clearly demonstrated that the magnetism of the studied sample is weak, itinerant and has a reentrant character. Based on the obtained results a magnetic phase diagram has been constructed in the H-T coordinates.

  9. Binding properties of Clostridium botulinum type C progenitor toxin to mucins.

    PubMed

    Nakamura, Toshio; Takada, Noriko; Tonozuka, Takashi; Sakano, Yoshiyuki; Oguma, Keiji; Nishikawa, Atsushi

    2007-04-01

    It has been reported that Clostridium botulinum type C 16S progenitor toxin (C16S toxin) first binds to the sialic acid on the cell surface of mucin before invading cells [A. Nishikawa, N. Uotsu, H. Arimitsu, J.C. Lee, Y. Miura, Y. Fujinaga, H. Nakada, T. Watanabe, T. Ohyama, Y. Sakano, K. Oguma, The receptor and transporter for internalization of Clostridium botulinum type C progenitor toxin into HT-29 cells, Biochem. Biophys. Res. Commun. 319 (2004) 327-333]. In this study we investigated the binding properties of the C16S toxin to glycoproteins. Although the toxin bound to membrane blotted mucin derived from the bovine submaxillary gland (BSM), which contains a lot of sialyl oligosaccharides, it did not bind to neuraminidase-treated BSM. The binding of the toxin to BSM was inhibited by N-acetylneuraminic acid, N-glycolylneuraminic acid, and sialyl oligosaccharides strongly, but was not inhibited by neutral oligosaccharides. Both sialyl alpha2-3 lactose and sialyl alpha2-6 lactose prevented binding similarly. On the other hand, the toxin also bound well to porcine gastric mucin. In this case, neutral oligosaccharides might play an important role as ligand, since galactose and lactose inhibited binding. These results suggest that the toxin is capable of recognizing a wide variety of oligosaccharide structures.

  10. Resistance to Bacillus thuringiensis Toxin Cry2Ab in Trichoplusia ni Is Conferred by a Novel Genetic Mechanism

    PubMed Central

    Song, Xiaozhao; Kain, Wendy; Cassidy, Douglas

    2015-01-01

    The resistance to the Bacillus thuringiensis (Bt) toxin Cry2Ab in a greenhouse-originated Trichoplusia ni strain resistant to both Bt toxins Cry1Ac and Cry2Ab was characterized. Biological assays determined that the Cry2Ab resistance in the T. ni strain was a monogenic recessive trait independent of Cry1Ac resistance, and there existed no significant cross-resistance between Cry1Ac and Cry2Ab in T. ni. From the dual-toxin-resistant T. ni strain, a strain resistant to Cry2Ab only was isolated, and the Cry2Ab resistance trait was introgressed into a susceptible laboratory strain to facilitate comparative analysis of the Cry2Ab resistance with the susceptible T. ni strain. Results from biochemical analysis showed no significant difference between the Cry2Ab-resistant and -susceptible T. ni larvae in midgut proteases, including caseinolytic proteolytic activity and zymogram profile and serine protease activities, in midgut aminopeptidase and alkaline phosphatase activity, and in midgut esterases and hemolymph plasma melanization activity. For analysis of genetic linkage of Cry2Ab resistance with potential Cry toxin receptor genes, molecular markers for the midgut cadherin, alkaline phosphatase (ALP), and aminopeptidase N (APN) genes were identified between the original greenhouse-derived dual-toxin-resistant and the susceptible laboratory T. ni strains. Genetic linkage analysis showed that the Cry2Ab resistance in T. ni was not genetically associated with the midgut genes coding for the cadherin, ALP, and 6 APNs (APN1 to APN6) nor associated with the ABC transporter gene ABCC2. Therefore, the Cry2Ab resistance in T. ni is conferred by a novel but unknown genetic mechanism. PMID:26025894

  11. The Role of 5-HT2A, 5-HT2C and mGlu2 Receptors in the Behavioral Effects of Tryptamine Hallucinogens N,N-Dimethyltryptamine and N,N-Diisopropyltryptamine in Rats and Mice

    PubMed Central

    Carbonaro, Theresa M.; Eshleman, Amy J.; Forster, Michael J.; Cheng, Kejun; Rice, Kenner C.; Gatch, Michael B.

    2014-01-01

    Rationale: Serotonin 5-HT2A and 5-HT2C receptors are thought to be the primary pharmacological mechanisms for serotonin-mediated hallucinogenic drugs, but recently there has been interest in metabotropic glutamate (mGluR2) receptors as contributors to the mechanism of hallucinogens. Objective: The present study assesses the role of these 5-HT and glutamate receptors as molecular targets for two tryptamine hallucinogens, N,N-dimethyltryptamine (DMT) and N,N-diisopropyltryptamine (DiPT). Methods: Drug discrimination, head twitch and radioligand binding assays were used. A 5-HT2AR inverse agonist (MDL100907), 5-HT2CR antagonist (SB242084) and mGluR2/3 agonist (LY379268) were tested for their ability to attenuate the discriminative stimulus effects of DMT and DiPT; an mGluR2/3 antagonist (LY341495) was tested for potentiation. MDL100907 was used to attenuate head twitches induced by DMT and DiPT. Radioligand binding studies and inosital-1-phosphate (IP-1) accumulation were performed at the 5-HT2CR for DiPT. Results: MDL100907 fully blocked the discriminative stimulus effects of DMT, but only partially blocked DiPT. SB242084 partially attenuated the discriminative stimulus effects of DiPT, but produced minimal attenuation of DMT’s effects. LY379268 produced potent, but only partial blockade of the discriminative stimulus effects of DMT. LY341495 facilitated DMT- and DiPT-like effects. Both compounds elicited head twitches (DiPT>DMT) which were blocked by MDL1000907. DiPT was a low potency full agonist at 5-HT2CR in vitro. Conclusions: The 5-HT2AR likely plays a major role in mediating the effects of both compounds. 5-HT2C and mGluR2 receptors likely modulate the discriminative stimulus effects of both compounds to some degree. PMID:24985890

  12. The role of 5-HT2A, 5-HT 2C and mGlu2 receptors in the behavioral effects of tryptamine hallucinogens N,N-dimethyltryptamine and N,N-diisopropyltryptamine in rats and mice.

    PubMed

    Carbonaro, Theresa M; Eshleman, Amy J; Forster, Michael J; Cheng, Kejun; Rice, Kenner C; Gatch, Michael B

    2015-01-01

    Serotonin 5-HT2A and 5-HT2C receptors are thought to be the primary pharmacological mechanisms for serotonin-mediated hallucinogenic drugs, but recently there has been interest in metabotropic glutamate (mGluR2) receptors as contributors to the mechanism of hallucinogens. The present study assesses the role of these 5-HT and glutamate receptors as molecular targets for two tryptamine hallucinogens, N,N-dimethyltryptamine (DMT) and N,N-diisopropyltryptamine (DiPT). Drug discrimination, head twitch, and radioligand binding assays were used. A 5-HT2AR inverse agonist (MDL100907), 5-HT2CR antagonist (SB242084), and mGluR2/3 agonist (LY379268) were tested for their ability to attenuate the discriminative stimulus effects of DMT and DiPT; an mGluR2/3 antagonist (LY341495) was tested for potentiation. MDL100907 was used to attenuate head twitches induced by DMT and DiPT. Radioligand binding studies and inosital-1-phosphate (IP-1) accumulation were performed at the 5-HT2CR for DiPT. MDL100907 fully blocked the discriminative stimulus effects of DMT, but only partially blocked DiPT. SB242084 partially attenuated the discriminative stimulus effects of DiPT, but produced minimal attenuation of DMT's effects. LY379268 produced potent, but only partial blockade of the discriminative stimulus effects of DMT. LY341495 facilitated DMT- and DiPT-like effects. Both compounds elicited head twitches (DiPT>DMT) which were blocked by MDL1000907. DiPT was a low-potency full agonist at 5-HT2CR in vitro. The 5-HT2AR likely plays a major role in mediating the effects of both compounds. 5-HT2C and mGluR2 receptors likely modulate the discriminative stimulus effects of both compounds to some degree.

  13. Role of the serotonin 5-HT(2A) receptor in learning.

    PubMed

    Harvey, John A

    2003-01-01

    This study reviews the role of the serotonin 5-HT2A receptor in learning as measured by the acquisition of the rabbit's classically conditioning nictitating membrane response, a component of the eyeblink response. Agonists at the 5-HT2A receptor including LSD (d-lysergic acid diethylamide) enhanced associative learning at doses that produce cognitive effects in humans. Some antagonists such as BOL (d-bromolysergic acid diethylamide), LY53,857, and ketanserin acted as neutral antagonists in that they had no effect on learning, whereas others (MDL11,939, ritanserin, and mianserin) acted as inverse agonists in that they retarded learning through an action at the 5-HT2A receptor. These results were placed in the context of what is known concerning the anatomical distribution and electrophysiological effects of 5-HT2A receptor activation in frontal cortex and hippocampus, as well as the role of cortical 5-HT2A receptors in schizophrenia. It was concluded that the 5-HT2A receptor demonstrates constitutive activity, and that variations in this activity can produce profound alterations in cognitive states.

  14. Assessment of Efficacy of Activated Charcoal for Treatment of Acute T-2 Toxin Poisoning,

    DTIC Science & Technology

    1986-11-14

    toxic secondary metabolite produced by a variety of Fusarium fungi-i. This trichothecene mycotoxin is highly toxic and has produced illness and death...Kern, and H. Richter, eds.), Verlag Paul Parly, BerLin, 1978, p. 58-93. 7 4. M. Saito and T. Tatsuna, "Toxins of Fusarium nivale," in Microbial Toxin...mechanisms of diarrhea induced by fusarenon-X, a trichothecene mycotoxin from Fusarium species, Toxicol. %ppl.Pharmacol. 57, 293-301 (1981). 7. Y. Ueno

  15. Synthesis and Structure–Activity Relationships of N-Benzyl Phenethylamines as 5-HT2A/2C Agonists

    PubMed Central

    2014-01-01

    N-Benzyl substitution of 5-HT2A receptor agonists of the phenethylamine structural class of psychedelics (such as 4-bromo-2,5-dimethoxyphenethylamine, often referred to as 2C-B) confer a significant increase in binding affinity as well as functional activity of the receptor. We have prepared a series of 48 compounds with structural variations in both the phenethylamine and N-benzyl part of the molecule to determine the effects on receptor binding affinity and functional activity at 5-HT2A and 5-HT2C receptors. The compounds generally had high affinity for the 5-HT2A receptor with 8b having the highest affinity at 0.29 nM but with several other compounds also exhibiting subnanomolar binding affinities. The functional activity of the compounds was distributed over a wider range with 1b being the most potent at 0.074 nM. Most of the compounds exhibited low to moderate selectivity (1- to 40-fold) for the 5-HT2A receptor in the binding assays, although one compound 6b showed an impressive 100-fold selectivity for the 5-HT2A receptor. In the functional assay, selectivity was generally higher with 1b being more than 400-fold selective for the 5-HT2A receptor. PMID:24397362

  16. Selective increases in serotonin 5-HT1B/1D and 5-HT2A/2C binding sites in adult rat basal ganglia following lesions of serotonergic neurons.

    PubMed

    Compan, V; Segu, L; Buhot, M C; Daszuta, A

    1998-05-18

    Quantitative autoradiography was used to examine possible adaptive changes in serotonin 5-HT1B/1D and 5-HT2A/2C receptor binding sites in adult rat basal ganglia, after partial or severe lesions of serotonergic neurons produced by intraraphe injections of variable amounts of 5,7-dihydroxytryptamine. In controls, the 5-HT1B/1D sites labeled with S-CM-G[125I]TNH2 were evenly distributed in the core and the shell of the nucleus accumbens. The density of 5-HT1B/1D sites was higher in the ventral than dorsal part of the striatum and no regional differences were detected along the rostrocaudal axis of the structure. The 5-HT2A/2C sites labeled with [125I]DOI were preferentially distributed in the mediodorsal striatum and higher densities were detected in the shell than core of the nucleus accumbens. Following 5,7-dihydroxytryptamine injections, there were no changes in binding of either receptor subtype after partial lesions entailing 80-90% 5-HT depletions. After severe 5-HT depletions (over 95%), large increases in 5-HT1B/1D binding were observed in the substantia nigra (78%), but no changes took place in the globus pallidus. Increases in 5-HT1B/1D binding were also detected in the shell of the nucleus accumbens (27%). Similar sized increases in 5-HT2A/2C binding (22%) were restricted to the medial striatum. The present results suggest a preferential association between 5-HT1B/1D receptors and the striatonigral neurons containing substance P, as indicated by the striatal distribution of these receptors and their selective increases in the substantia nigra after severe 5-HT deprivation. We recently proposed a similar relationship between the 5-HT4 receptors and the striatopallidal neurons containing met-enkephalin. Moreover, the increases in 5-HT1B/1D binding in the substantia nigra and in the shell of the nucleus accumbens reinforce the view of an implication of this receptor subtype in motor functions. In contrast, the prominent increases in 5-HT2A/2C binding after

  17. Polymorphisms of the 5-HT2A receptor gene and clinical response to olanzapine in paranoid schizophrenia.

    PubMed

    Olajossy-Hilkesberger, Luiza; Godlewska, Beata; Schosser-Haupt, Alexandra; Olajossy, Marcin; Wojcierowski, Jacek; Landowski, Jerzy; Marmurowska-Michałowska, Halina; Kasper, Siegfried

    2011-01-01

    5-HT2A receptor is strongly implicated in the mode of action of atypical antipsychotic drugs. The aim of the study was to investigate whether the 5-HT2A receptor gene's polymorphisms (His452Tyr and T102C) have an influence on the response to olanzapine in patients with schizophrenia. We studied 99 Caucasian schizophrenia patients treated with olanzapine. Psychopathology was measured before and after 6 weeks of treatment. Clinical improvement was quantified as change in Positive and Negative Syndrome Scale (PANSS) total scores and subscores as shown by percentage improvement below the baseline score. The clinical response to antipsychotic treatment was defined as 30% improvement from baseline in PANSS scores. The His/Tyr polymorphism was significantly associated with a percentage improvement in PANSS positive symptom subscore (better response in His/His homozygotes; p<0.05) after treatment with olanzapine. As for the T102C polymorphism, a better response in terms of PANSS positive subscore improvement was observed for C/C homozygotes (p<0.01). A significant association of 5-HT2A genotype distribution of the T102C polymorphism with a categorical measure of response, but only in terms of PANSS positive symptom subscores, was observed (p<0.01). Variations in the 5-HT2A receptor gene may influence individual and particularly positive symptom response to olanzapine. Copyright © 2011 S. Karger AG, Basel.

  18. Involvement of P-type Ca2+ channels in the K(+)- and d-fenfluramine-induced [3H]5-HT release from rat hippocampal synaptosomes.

    PubMed

    Frittoli, E; Gobbi, M; Mennini, T

    1994-06-01

    The Ca2(+)-dependent [3H]5-HT release induced by depolarization or by 0.5 microM d-fenfluramine in rat hippocampal synaptosomes, was significantly reduced (35-42%) by three different P-type Ca2+ channels blockers (omega-Agatoxin-IVA, 100 nM, funnel-web spider toxin, FTX, 0.05 microliters/ml, and its synthetic analogue, sFTX, 1 mM), indicating the major role of these channels in the Ca2+ influx preceding neurotransmitter release.

  19. Influence of nonylphenol and octylphenol exposure on 5-HT, 5-HT transporter, and 5-HT2A receptor.

    PubMed

    Liu, Chunhong; Lai, Yuting; Ouyang, Junyan; Yang, Tongwang; Guo, Youting; Yang, Jie; Huang, Shaowen

    2017-03-01

    Nonylphenol (NP) and octylphenol (OP) are priority environmental contaminants that have a potential role as endocrine disruptors. They can be biomagnified in the food chain and pose an estrogenic health risk to human health. A 28-day oral toxicity study was performed to observe the impact of single and combined exposure to NP and OP on 5-HT transporter (SERT) as well as 5-HT 2A receptor. Results showed that the 5-HT levels in rat plasma increased with exposure to middle-dose and high-dose NP, to high-dose OP, and to low, middle, and high doses of combined NP and OP (P < 0.05), while the 5-HT levels in rat platelets increased when exposed to NP/OP or combined NP and OP of middle or high dose (P < 0.05). The expression levels of SERT in rat platelets decreased when exposed to high-dose NP/OP or high dose of combined NP and OP (P < 0.05). Meanwhile, the expression levels of 5-HT 2A in rat platelets decreased when exposed to high-dose NP/OP as well as combined NP and OP (P < 0.05). These findings suggested that exposure to NP and OP could influence the metabolic network of 5-hydroxytryptamine via transportation and receptor binding pathways.

  20. Activation of Ventral Tegmental Area 5-HT2C Receptors Reduces Incentive Motivation.

    PubMed

    Valencia-Torres, Lourdes; Olarte-Sánchez, Cristian M; Lyons, David J; Georgescu, Teodora; Greenwald-Yarnell, Megan; Myers, Martin G; Bradshaw, Christopher M; Heisler, Lora K

    2017-06-01

    Obesity is primarily due to food intake in excess of the body's energetic requirements, intake that is not only associated with hunger but also the incentive value of food. The 5-hydroxytryptamine 2C receptor (5-HT 2C R) is a target for the treatment of human obesity. Mechanistically, 5-HT 2C Rs are positioned to influence both homeostatic feeding circuits within the hypothalamus and reward circuits within the ventral tegmental area (VTA). Here we investigated the role of 5-HT 2C Rs in incentive motivation using a mathematical model of progressive ratio (PR) responding in mice. We found that the 5-HT 2C R agonist lorcaserin significantly reduced both ad libitum chow intake and PR responding for chocolate pellets and increased c-fos expression in VTA 5-HT 2C R expressing γ-aminobutyric acid (GABA) neurons, but not 5-HT 2C R expressing dopamine (DA) neurons. We next adopted a chemogenetic approach using a 5-HT 2C R CRE line to clarify the function of subset of 5-HT 2C receptor expressing VTA neurons in the modulation of appetite and food-motivated behavior. Activation of VTA 5-HT 2C receptor expressing neurons significantly reduced ad libitum chow intake, operant responding for chocolate pellets, and the incentive value of food. In contrast, chemogenetic inhibition of VTA 5-HT 2C receptor expressing neurons had no effect on the feeding behavior. These results indicate that activation of the subpopulation of 5-HT 2C R neurons within the VTA is sufficient to significantly reduce homeostatic feeding and effort-based intake of palatable food, and that this subset has an inhibitory role in motivational processes. These findings are relevant to the treatment of obesity.

  1. Activation of Ventral Tegmental Area 5-HT2C Receptors Reduces Incentive Motivation

    PubMed Central

    Valencia-Torres, Lourdes; Olarte-Sánchez, Cristian M; Lyons, David J; Georgescu, Teodora; Greenwald-Yarnell, Megan; Myers, Martin G; Bradshaw, Christopher M; Heisler, Lora K

    2017-01-01

    Obesity is primarily due to food intake in excess of the body's energetic requirements, intake that is not only associated with hunger but also the incentive value of food. The 5-hydroxytryptamine 2C receptor (5-HT2CR) is a target for the treatment of human obesity. Mechanistically, 5-HT2CRs are positioned to influence both homeostatic feeding circuits within the hypothalamus and reward circuits within the ventral tegmental area (VTA). Here we investigated the role of 5-HT2CRs in incentive motivation using a mathematical model of progressive ratio (PR) responding in mice. We found that the 5-HT2CR agonist lorcaserin significantly reduced both ad libitum chow intake and PR responding for chocolate pellets and increased c-fos expression in VTA 5-HT2CR expressing γ-aminobutyric acid (GABA) neurons, but not 5-HT2CR expressing dopamine (DA) neurons. We next adopted a chemogenetic approach using a 5-HT2CRCRE line to clarify the function of subset of 5-HT2C receptor expressing VTA neurons in the modulation of appetite and food-motivated behavior. Activation of VTA 5-HT2C receptor expressing neurons significantly reduced ad libitum chow intake, operant responding for chocolate pellets, and the incentive value of food. In contrast, chemogenetic inhibition of VTA 5-HT2C receptor expressing neurons had no effect on the feeding behavior. These results indicate that activation of the subpopulation of 5-HT2CR neurons within the VTA is sufficient to significantly reduce homeostatic feeding and effort-based intake of palatable food, and that this subset has an inhibitory role in motivational processes. These findings are relevant to the treatment of obesity. PMID:27882999

  2. D2 dopaminergic and 5-HT1A serotonergic activity of 2-(1-naphthyl)ethyl- and 2-(2-naphthyl)ethyl amines.

    PubMed

    Šukalović, V; Roglić, G; Husinec, S; Kostić-Rajaćić, S; Andrić, D; Šoškić, Vukić

    2003-11-01

    Several tertiary 2-phenylethyl, 2-(1-naphthyl)ethyl and 2-(2-naphthyl)ethyl amines were synthesized and their binding affinities for dopamine D(1), D(2) and serotonin 5-HT(1A) receptors evaluated in radioligand binding assays. All compounds were inactive in D(1) dopamine radioligand binding assay. The 2-(1-naphthyl)ethyl analogues expressed a low but significant binding affinity for the D(2) and moderate one for the 5-HT(1A) receptor subtypes. Most of the remaining compounds expressed binding affinity at the 5-HT(1A) receptor subtype but were inactive in D(2) receptor binding assay. Based on these results and considering the chemical characteristics of the compounds synthesized and evaluated for dopaminergic and serotonergic activity throughout the present study it can be concluded that hydrophobic type of interaction (stacking or edge-to-face) plays a significant role in the formation of receptor-ligand complexes of 2-(1-naphthyl)ethyl amines. This structural motive can be applied to design and synthesize new, more potent dopaminergic/serotonergic ligands by slight chemical modifications.

  3. Purification, immunotoxic effects, and cellular uptake of trichothecene mycotoxins

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Witt, M.F.

    1989-01-01

    Studies were carried out to better understand how the trichothecenes alter immune function in animals and humans. Deoxynivalenol (DON) was purified for use in animal feeding studies. Dietary exposure to DON for 8 weeks altered the serum immunoglobulin profile in mice and decreased the splenic plaque-forming cell response to the antigen sheep red blood cells. The uptake of ({sup 3}H)T-2 toxin by a murine B-cell hybridoma was studied in order to learn more about the way in which trichothecenes interact with immune cells. A simple procedure was developed for the laboratory production and purification of gram quantities of crystalline DON.more » When Fusarium graminearum R6576 was grown on rice, concentrations of 600 to 700 ppm DON accumulated after 13 to 18 days of incubation. A DON derivative, 15-acetylDON, was also found at concentrations of 100 to 300 ppm after 7 to 10 days. DON was purified from crude culture extracts by water-saturated silica gel chromatography. Alpha-({sup 3}H)T-2 toxin of 99% chemical and radiochemical purity was prepared for use in uptake studies. Both the rate of uptake of ({sup 3}H)T-2 toxin by hybridomas and the time required for accumulation of ({sup 3}H)T-2 to reach equilibrium were proportional to the concentration of ({sup 3}H)T-2. ({sup 3}H)T-2 toxin accumulated by hybridomas was proportional to the concentration of ({sup 3}H)T-2 between 10{sup {minus}8} and 10{sup {minus}3} M. The rate of uptake of ({sup 3}H)jT-2 toxin by hybridomas was inhibited by the trichothecenes T-2 toxin, DON, verrucarin A, and roridin A, as well as the antibiotic anisomycin. The kinetics and concentration dependence of accumulation, along with the inhibition patterns, suggest that uptake of ({sup 3}H)T-2 toxin by hybridomas is mediated by binding of toxin to ribosomes.« less

  4. Enhancement of cortical extracellular 5-HT by 5-HT1A and 5-HT2C receptor blockade restores the antidepressant-like effect of citalopram in non-responder mice.

    PubMed

    Calcagno, Eleonora; Guzzetti, Sara; Canetta, Alessandro; Fracasso, Claudia; Caccia, Silvio; Cervo, Luigi; Invernizzi, Roberto W

    2009-07-01

    We recently found that the response of DBA/2 mice to SSRIs in the forced swim test (FST) was impaired and they also had a smaller basal and citalopram-stimulated increase in brain extracellular serotonin (5-HT) than 'responder' strains. We employed intracerebral microdialysis, FST and selective antagonists of 5-HT1A and 5-HT2C receptors to investigate whether enhancing the increase in extracellular 5-HT reinstated the anti-immobility effect of citalopram in the FST. WAY 100635 (0.3 mg/kg s.c.) or SB 242084 (1 mg/kg s.c.), respectively a selective 5-HT1A and 5-HT2C receptor antagonist, raised the effect of citalopram (5 mg/kg) on extracellular 5-HT in the medial prefrontal cortex of DBA/2N mice (citalopram alone 5.2+/-0.3 fmol/20 microl, WAY 100635+citalopram 9.9+/-2.1 fmol/20 microl, SB 242084+ citalopram 7.6+/-1.0 fmol/20 microl) to the level reached in 'responder' mice given citalopram alone. The 5-HT receptor antagonists had no effect on the citalopram-induced increase in extracellular 5-HT in the dorsal hippocampus. The combination of citalopram with WAY 100635 or SB 242084 significantly reduced immobility time in DBA/2N mice that otherwise did not respond to either drug singly. Brain levels of citalopram in mice given citalopram alone or with 5-HT antagonists did not significantly differ. The results confirm that impaired 5-HT transmission accounts for the lack of effect of citalopram in the FST and suggest that enhancing the effect of SSRIs on extracellular 5-HT, through selective blockade of 5-HT1A and 5-HT2C receptors, could be a useful strategy to restore the response in treatment-resistant depression.

  5. Spinal 5-HT2 and 5-HT3 receptors mediate low, but not high, frequency TENS-induced antihyperalgesia in rats

    PubMed Central

    Radhakrishnan, Rajan; King, Ellen W.; Dickman, Janelle K.; Herold, Carli A.; Johnston, Natalie F.; Spurgin, Megan L.; Sluka, Kathleen A.

    2009-01-01

    Transcutaneous electrical nerve stimulation (TENS) is a form of non-pharmacological treatment for pain. Involvement of descending inhibitory systems is implicated in TENS-induced analgesia. In the present study, the roles of spinal 5-HT and α2-adrenoceptors in TENS analgesia were investigated in rats. Hyperalgesia was induced by inflaming the knee joint with 3% kaolin—carrageenan mixture and assessed by measuring paw withdrawal latency (PWL) to heat before and 4 h after injection. The (1) α2-adrenergic antagonist yohimbine (30 μg), (2) 5-HT antagonist methysergide (5-HT1 and 5-HT2,30 μg), one of the 5-HT receptor subtype antagonists, (3) NAN-190 (5-HT1A, 15 μg), (4) ketanserin (5-HT2A, 30 μg), (5) MDL-72222 (5-HT3, 12 μg), or (6) vehicle was administered intrathecally prior to TENS treatment. Low (4 Hz) or high (100 Hz) frequency TENS at sensory intensity was then applied to the inflamed knee for 20 min and PWL was determined. Selectivity of the antagonists used was confirmed using respective agonists administered intrathecally. Yohimbine had no effect on the antihyperalgesia produced by low or high frequency TENS. Methysergide and MDL-72222 prevented the antihyperalgesia produced by low, but not high, frequency TENS. Ketanserin attenuated the antihyperalgesic effects of low frequency TENS whereas NAN-190 had no effect. The results from the present study show that spinal 5-HT receptors mediate low, but not high, frequency TENS-induced antihyperalgesia through activation of 5-HT2A and 5-HT3 receptors in rats. Furthermore, spinal noradrenergic receptors are not involved in either low or high frequency TENS antihyperalgesia. PMID:14499437

  6. Occurrence and distribution of 13 trichothecene toxins in naturally contaminated maize plants in Germany.

    PubMed

    Schollenberger, Margit; Müller, Hans-Martin; Ernst, Katrin; Sondermann, Sarah; Liebscher, Melanie; Schlecker, Claudia; Wischer, Gerald; Drochner, Winfried; Hartung, Karin; Piepho, Hans-Peter

    2012-10-01

    The objective of the present study was to monitor the occurrence and distribution of a spectrum of trichothecene toxins in different parts of maize plants. Therefore maize plants were sampled randomly from 13 fields in southwest Germany and the fractions kernels, cobs, husks, stalks, leaves and rudimentary ears were analyzed for eight A-type and five B-type trichothecenes. Each of the toxins was found in at least three of the total of 78 samples. The study revealed that both A-type and B-type trichothecenes may be present in all parts of the maize plant but may be unevenly distributed. For the contents of deoxynivalenol, 3- and 15-acetyldeoxynivalenol, nivalenol, scirpentriol, 15-monoacetoxyscirpenol, HT-2 and T-2 toxin significant differences (p < 0.05) were found between different parts of the maize plants whereas no significant differences were observed for fusarenon-X, 4,15-diacetoxyscirpenol, neosolaniol, T-2 triol and T-2 tetraol. Up to twelve toxins co-occurring in one sample were detected. As a group B-type trichothecenes dominated over A-type trichothecenes concerning incidences and levels. Contamination was strongest with rudimentary ears based on incidence and mean and maximum contents; mean contents with few exceptions tended towards a higher level than in other fractions with significant (p < 0.05) differences compared to leaves for seven toxins.

  7. Development of the 5-HT2CR-Tango System Combined with an EGFP Reporter Gene.

    PubMed

    Watanabe, Yoshihisa; Tsujimura, Atsushi; Aoki, Miku; Taguchi, Katsutoshi; Tanaka, Masaki

    2016-02-01

    The serotonin 2C receptor (5-HT2CR) is a G-protein-coupled receptor implicated in emotion, feeding, reward, and cognition. 5-HT2CRs are pharmacological targets for mental disorders and metabolic and reward system abnormalities, as alterations in 5-HT2CR expression, RNA editing, and SNPs are involved in these disturbances. To date, 5-HT2CR activity has mainly been measured by quantifying inositol phosphate production and intracellular Ca(2+) release, but these assays are not suitable for in vivo analysis. Here, we developed a 5-HT2CR-Tango assay system, a novel analysis tool of 5-HT2CR activity based on the G-protein-coupled receptor (GPCR)-arrestin interaction. With desensitization of activated 5-HT2CR by arrestin, this system converts the 5-HT2CR-arrestin interaction into EGFP reporter gene signal via the LexA transcriptional activation system. For validation of our system, we measured activity of two 5-HT2CR RNA-editing isoforms (INI and VGV) in HEK293 cells transfected with EGFP reporter gene. The INI isoform displayed both higher basal- and 5-HT-stimulated activities than the VGV isoform. Moreover, an inhibitory effect of 5-HT2CR antagonist SB242084 was also detected by 5-HT2CR-Tango system. This novel tool is useful for in vitro high-throughput targeted 5-HT2CR drug screening and can be applied to future in vivo brain function studies associated with 5-HT2CRs in transgenic animal models.

  8. 3,4-Dihydro-2H-benzoxazinones are 5-HT(1A) receptor antagonists with potent 5-HT reuptake inhibitory activity.

    PubMed

    Atkinson, Peter J; Bromidge, Steven M; Duxon, Mark S; Gaster, Laramie M; Hadley, Michael S; Hammond, Beverley; Johnson, Christopher N; Middlemiss, Derek N; North, Stephanie E; Price, Gary W; Rami, Harshad K; Riley, Graham J; Scott, Claire M; Shaw, Tracey E; Starr, Kathryn R; Stemp, Geoffrey; Thewlis, Kevin M; Thomas, David R; Thompson, Mervyn; Vong, Antonio K K; Watson, Jeannette M

    2005-02-01

    Starting from a high throughput screening hit, a series of 3,4-dihydro-2H-benzoxazinones has been identified with both high affinity for the 5-HT(1A) receptor and potent 5-HT reuptake inhibitory activity. The 5-(2-methyl)quinolinyloxy derivative combined high 5-HT(1A/1B/1D) receptor affinities with low intrinsic activity and potent inhibition of the 5-HT reuptake site (pK(i)8.2). This compound also had good oral bioavailability and brain penetration in the rat.

  9. Dysfunctional attitudes and 5-HT2 receptors during depression and self-harm.

    PubMed

    Meyer, Jeffrey H; McMain, Shelley; Kennedy, Sidney H; Korman, Lorne; Brown, Gregory M; DaSilva, Jean N; Wilson, Alan A; Blak, Thomas; Eynan-Harvey, Rahel; Goulding, Verdell S; Houle, Sylvain; Links, Paul

    2003-01-01

    Dysfunctional attitudes are negatively biased assumptions and beliefs regarding oneself, the world, and the future. In healthy subjects, increasing serotonin (5-HT) agonism with a single dose of d-fenfluramine lowered dysfunctional attitudes. To investigate whether the converse, a low level of 5-HT agonism, could account for the higher levels of dysfunctional attitudes observed in patients with major depression or with self-injurious behavior, cortex 5-HT(2) receptor binding potential and dysfunctional attitudes were measured in patients with major depressive disorder, patients with a history of self-injurious behavior, and healthy comparison subjects (5-HT(2) receptor density increases during 5-HT depletion). Twenty-nine healthy subjects were recruited to evaluate the effect of d-fenfluramine or of clonidine (control condition) on dysfunctional attitudes. Dysfunctional attitudes were assessed with the Dysfunctional Attitude Scale 1 hour before and 1 hour after drug administration. In a second experiment, dysfunctional attitudes and 5-HT(2) binding potential were measured in 22 patients with a major depressive episode secondary to major depressive disorder, 18 patients with a history of self-injurious behavior occurring outside of a depressive episode, and another 29 age-matched healthy subjects. Cortex 5-HT(2) binding potential was measured with [(18)F]setoperone positron emission tomography. In the first experiment, dysfunctional attitudes decreased after administration of d-fenfluramine. In the second experiment, in the depressed group, dysfunctional attitudes were positively associated with cortex 5-HT(2) binding potential, especially in Brodmann's area 9 (after adjustment for age). Depressed subjects with extremely dysfunctional attitudes had higher 5-HT(2) binding potential, compared to healthy subjects, particularly in Brodmann's area 9. Low levels of 5-HT agonism in the brain cortex may explain the severely pessimistic, dysfunctional attitudes associated

  10. 5-HT2CRs expressed by pro-opiomelanocortin neurons regulate insulin sensitivity in liver

    USDA-ARS?s Scientific Manuscript database

    Mice lacking 5-HT 2C receptors displayed hepatic insulin resistance, a phenotype normalized by re-expression of 5-HT2CRs only in pro-opiomelanocortin (POMC) neurons. 5-HT2CR deficiency also abolished the anti-diabetic effects of meta-chlorophenylpiperazine (a 5-HT2CR agonist); these effects were re...

  11. Therapeutic Potential of 5-HT2C Receptor Agonists for Addictive Disorders.

    PubMed

    Higgins, Guy A; Fletcher, Paul J

    2015-07-15

    The neurotransmitter 5-hydroxytryptamine (5-HT; serotonin) has long been associated with the control of a variety of motivated behaviors, including feeding. Much of the evidence linking 5-HT and feeding behavior was obtained from studies of the effects of the 5-HT releaser (dex)fenfluramine in laboratory animals and humans. Recently, the selective 5-HT2C receptor agonist lorcaserin received FDA approval for the treatment of obesity. This review examines evidence to support the use of selective 5-HT2C receptor agonists as treatments for conditions beyond obesity, including substance abuse (particularly nicotine, psychostimulant, and alcohol dependence), obsessive compulsive, and excessive gambling disorder. Following a brief survey of the early literature supporting a role for 5-HT in modulating food and drug reinforcement, we propose that intrinsic differences between SSRI and serotonin releasers may have underestimated the value of serotonin-based pharmacotherapeutics to treat clinical forms of addictive behavior beyond obesity. We then highlight the critical involvement of the 5-HT2C receptor in mediating the effect of (dex)fenfluramine on feeding and body weight gain and the evidence that 5-HT2C receptor agonists reduce measures of drug reward and impulsivity. A recent report of lorcaserin efficacy in a smoking cessation trial further strengthens the idea that 5-HT2C receptor agonists may have potential as a treatment for addiction. This review was prepared as a contribution to the proceedings of the 11th International Society for Serotonin Research Meeting held in Hermanus, South Africa, July 9-12, 2014.

  12. Molecular and pharmacological characterization of serotonin 5-HT2α and 5-HT7 receptors in the salivary glands of the blowfly Calliphora vicina.

    PubMed

    Röser, Claudia; Jordan, Nadine; Balfanz, Sabine; Baumann, Arnd; Walz, Bernd; Baumann, Otto; Blenau, Wolfgang

    2012-01-01

    Secretion in blowfly (Calliphora vicina) salivary glands is stimulated by the biogenic amine serotonin (5-hydroxytryptamine, 5-HT), which activates both inositol 1,4,5-trisphosphate (InsP(3))/Ca(2+) and cyclic adenosine 3',5'-monophosphate (cAMP) signalling pathways in the secretory cells. In order to characterize the signal-inducing 5-HT receptors, we cloned two cDNAs (Cv5-ht2α, Cv5-ht7) that share high similarity with mammalian 5-HT(2) and 5-HT(7) receptor genes, respectively. RT-PCR demonstrated that both receptors are expressed in the salivary glands and brain. Stimulation of Cv5-ht2α-transfected mammalian cells with 5-HT elevates cytosolic [Ca(2+)] in a dose-dependent manner (EC(50) = 24 nM). In Cv5-ht7-transfected cells, 5-HT produces a dose-dependent increase in [cAMP](i) (EC(50) = 4 nM). We studied the pharmacological profile for both receptors. Substances that appear to act as specific ligands of either Cv5-HT(2α) or Cv5-HT(7) in the heterologous expression system were also tested in intact blowfly salivary gland preparations. We observed that 5-methoxytryptamine (100 nM) activates only the Cv5-HT(2α) receptor, 5-carboxamidotryptamine (300 nM) activates only the Cv5-HT(7) receptor, and clozapine (1 µM) antagonizes the effects of 5-HT via Cv5-HT(7) in blowfly salivary glands, providing means for the selective activation of each of the two 5-HT receptor subtypes. This study represents the first comprehensive molecular and pharmacological characterization of two 5-HT receptors in the blowfly and permits the analysis of the physiological role of these receptors, even when co-expressed in cells, and of the modes of interaction between the Ca(2+)- and cAMP-signalling cascades.

  13. Ebselen has lithium-like effects on central 5-HT2A receptor function.

    PubMed

    Antoniadou, I; Kouskou, M; Arsiwala, T; Singh, N; Vasudevan, S R; Fowler, T; Cadirci, E; Churchill, G C; Sharp, T

    2018-02-27

    Lithium's antidepressant action may be mediated by inhibition of inositol monophosphatase (IMPase), a key enzyme in G q protein coupled receptor signalling. Recently, the antioxidant agent ebselen was identified as an IMPase inhibitor. Here we investigated both ebselen and lithium in models of the 5-HT 2A receptor, a G q protein coupled receptor implicated in lithium's actions. 5-HT 2A receptor function was modelled in mice by measuring the behavioural (head-twitches) and cortical immediate early gene (IEG; Arc, c-fos and Erg2 mRNA) responses to 5-HT 2A receptor agonist administration. Ebselen and lithium were administered either acutely or chronically prior to assessment of 5-HT 2A receptor function. Given the SSRI augmenting action of lithium and 5-HT 2A antagonists, ebselen was also tested for this action by co-administration with the SSRI citalopram in microdialysis (extracellular 5-HT) experiments. Acute and repeated administration of ebselen inhibited behavioural and IEG responses to the 5-HT 2A receptor agonist DOI. Repeated lithium also inhibited DOI-evoked behavioural and IEG responses. In comparison, a selective IMPase inhibitor (L-690,330) attenuated the behavioural response to DOI whereas glycogen synthase kinase inhibitor (AR-A014418) did not. Finally, ebselen increased regional brain 5-HT synthesis and enhanced the increase in extracellular 5-HT induced by citalopram. The current data demonstrate lithium-mimetic effects of ebselen in different experimental models of 5-HT 2A receptor function, likely mediated by IMPase inhibition. This evidence of lithium-like neuropharmacological effects of ebselen adds further support for the clinical testing of ebselen in mood disorder, including as an antidepressant augmenting agent. This article is protected by copyright. All rights reserved.

  14. Association of 5-HT2A receptor gene polymorphisms with gastrointestinal disorders in Egyptian children with autistic disorder.

    PubMed

    Abdelrahman, Hadeel M; Sherief, Laila M; Alghobashy, Ashgan A; Abdel Salam, Sanaa M; Hashim, Haitham M; Abdel Fattah, Nelly R; Mohamed, Randa H

    2014-11-12

    Gastrointestinal disturbances (GID) are frequently reported in children with autism spectrum disorders (ASD). Recently, mounting evidence suggests that there may be a genetic link for autism with gastrointestinal disturbances. We aimed to investigate whether there were any association between the -1438A/G, 102T/C and His452Tyr polymorphisms of the serotonin 2A receptor gene (5-HT2A) in Egyptian children with ASD and GID. Eighty children with autistic disorder and 100 healthy control children were examined. -1438A/G, 102T/C and His452Tyr polymorphisms of 5-HT2A were genotyped by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method. Significant increase of the G allele and the GG genotype of the -1438A/G polymorphism was observed in children with autism than control, but there were no significant differences in the frequencies either of the 102T/C genotype or His452Tyr genotype between the two groups. There was a significant increase of homozygote A allele of the -1438A/G and CC genotype of the 102T/C polymorphism in ASD children with GID. This study supports the possible involvement of the 5-HT2A receptor in the development of ASD and associated GID. Copyright © 2014 Elsevier Ltd. All rights reserved.

  15. Effects of Constant Flickering Light on Refractive Status, 5-HT and 5-HT2A Receptor in Guinea Pigs.

    PubMed

    Li, Bing; Luo, Xiumei; Li, Tao; Zheng, Changyue; Ji, Shunmei; Ma, Yuanyuan; Zhang, Shuangshuang; Zhou, Xiaodong

    2016-01-01

    To investigate the effects of constant flickering light on refractive development, the role of serotonin (i.e.5-hydroxytryptamine, 5-HT)and 5-HT2A receptor in myopia induced by flickering light in guinea pigs. Forty-five guinea pigs were randomly divided into three groups: control, form deprivation myopia (FDM) and flickering light induced myopia (FLM) groups(n = 15 for each group). The right eyes of the FDM group were covered with semitransparent hemispherical plastic shells serving as eye diffusers. Guinea pigs in FLM group were raised with illumination of a duty cycle of 50% at a flash frequency of 0.5Hz. The refractive status, axial length (AL), corneal radius of curvature(CRC) were measured by streak retinoscope, A-scan ultrasonography and keratometer, respectively. Ultramicroscopy images were taken by electron microscopy. The concentrations of 5-HTin the retina, vitreous body and retinal pigment epithelium (RPE) were assessed by high performance liquid chromatography, the retinal 5-HT2A receptor expression was evaluated by immunohistofluorescence and western blot. The refraction of FDM and FLM eyes became myopic from some time point (the 4th week and the 6th week, respectively) in the course of the experiment, which was indicated by significantly decreased refraction and longer AL when compared with the controls (p<0.05). The concentrations of 5-HT in the retina, vitreous body and RPE of FDM and FLM eyes were significantly increased in comparison with those of control eyes (both p<0.05). Similar to FDM eyes, the expression of retinal 5-HT2A receptor in FLM eyes was significantly up-regulated compared to that of control eyes (both p<0.05). Western blot analysis showed that retinal 5-HT2A receptor level elevated less in the FLM eyes than that in the FDM eyes. Moreover, the levels of norepinephrine and epinephrine in FDM and FLM groups generally decreased when compared with control groups (all p<0.05). Constant flickering light could cause progressive myopia in

  16. Influence of esterified-glucomannan on performance and organ morphology, serum biochemistry and haematology in broilers exposed to individual and combined mycotoxicosis (aflatoxin, ochratoxin and T-2 toxin).

    PubMed

    Raju, M V; Devegowda, G

    2000-12-01

    1. A study was conducted to evaluate the individual and combined effects of aflatoxin B1 (AF), ochratoxin A (OA) and T-2 toxin (T-2) on performance, organ morphology serum biochemistry and haematology of broiler chickens and the efficacy of esterified-glucomannan (E-GM), a cell wall derivative of Saccharomyces cerevisiae1026 in their counteraction. 2. Two dietary inclusion rates of AF (0 and 0.3 mg/kg), OA (0 and 2 mg/kg), T-2 (0 and 3 mg/kg) and E-GM (0 and 1 g/kg) were tested in a 2 x 2 x 2 x 2 factorial manner on a total of 960 broiler chickens from 1 to 35 d of age in an open sided deep litter pen house. 3. Body weight and food intake were depressed by all the mycotoxins, OA being the most toxic during early life. 4. Weights of kidney and adrenals were increased by AF and OA. Liver weight was increased by AF (17.8%), while OA increased gizzard weight (14.6%) and reduced bone ash content (8.1%). T-2 toxin showed no effect on these variables. 5. Serum cholesterol content was decreased and activity of serum gamma glutamyl transferase (GGT) was increased by AF and OA while serum protein content was decreased by AF. These effects were more pronounced at 21 d than at 35 d of age. Inconsistent responses were seen in the other variables: blood urea nitrogen (BUN) content, activities of serum alanine amino transferase and aspertate amino transferase. Blood haemoglobin content was depressed by AF and T-2, whereas blood coagulation time was prolonged by OA. 6. Significant interactions were observed between any 2 toxins for their additive effects on body weight, food intake, bone ash content and serum GGT activity at 21 d. Conversely, antagonistic interactions were observed among any 2 of the toxins for their effects on variables such as serum protein and serum cholesterol content. Simultaneous feeding of all 3 mycotoxins did not show increased toxicity above that seen with any 2. 7. Esterified-glucomannan increased body weight (2.26%) and food intake (1.6%), decreased

  17. Hydrogen peroxide (H/sub 2/O/sub 2/) stimulates the active transport of 5-hydroxytryptamine (5-HT) into platelets

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Bosin, T.R.

    1986-03-01

    Platelets function in a variety of physiological and pathological processes which may be altered by oxidant injury. One such process is the active transport 5-HT, which is an important mechanism in the control of circulating 5-HT levels. Exposure of mouse platelets (10/sup 8//ml) to H/sub 2/O/sub 2/ caused a time-dependent and dose-dependent increase in 5-HT (10/sup -7/M) uptake. The uptake 4 and 10 min following H/sub 2/O/sub 2/ (50 ..mu..M) was 228% and 145% of control values, respectively. Fluoxetine (10/sup -6/M) blocked all 5-HT uptake and catalase (1500 U/ml) blocked the H/sub 2/O/sub 2/-stimulated uptake. Enzymatically produced H/sub 2/O/sub 2/more » (glucose/glucose oxidase) and xanthine (X)/xanthine oxidase (XO) generated oxygen radicals produced quantitatively and qualitatively similar results. The stimulatory response of platelets to X/XO generated oxidants was unaffected by superoxide dismutase (250 U/ml) but, was inhibited using heat-denatured XO, allopurinol (0.5 mM) and catalase; fluoxetine inhibited all 5-HT uptake. Platelets exposed to X/XO in the presence of chelated (EDTA, 100 ..mu..M) or unchelated FeSO/sub 4/, FeNH/sub 4/(SO/sub 4/)/sub 2/ or CuCl (50 ..mu..M) did not have altered 5-HT uptake. These data indicate that brief exposure of platelets to physiological levels of H/sub 2/O/sub 2/ results in marked, reversible stimulation of active 5-HT uptake which may represent a homeostatic defense mechanism when H/sub 2/O/sub 2/ is elevated in the platelet microenvironment.« less

  18. Function and distribution of 5-HT2 receptors in the honeybee (Apis mellifera).

    PubMed

    Thamm, Markus; Rolke, Daniel; Jordan, Nadine; Balfanz, Sabine; Schiffer, Christian; Baumann, Arnd; Blenau, Wolfgang

    2013-01-01

    Serotonin plays a pivotal role in regulating and modulating physiological and behavioral processes in both vertebrates and invertebrates. In the honeybee (Apis mellifera), serotonin has been implicated in division of labor, visual processing, and learning processes. Here, we present the cloning, heterologous expression, and detailed functional and pharmacological characterization of two honeybee 5-HT2 receptors. Honeybee 5-HT2 receptor cDNAs were amplified from brain cDNA. Recombinant cell lines were established constitutively expressing receptor variants. Pharmacological properties of the receptors were investigated by Ca(2+) imaging experiments. Quantitative PCR was applied to explore the expression patterns of receptor mRNAs. The honeybee 5-HT2 receptor class consists of two subtypes, Am5-HT2α and Am5-HT2β. Each receptor gene also gives rise to alternatively spliced mRNAs that possibly code for truncated receptors. Only activation of the full-length receptors with serotonin caused an increase in the intracellular Ca(2+) concentration. The effect was mimicked by the agonists 5-methoxytryptamine and 8-OH-DPAT at low micromolar concentrations. Receptor activities were blocked by established 5-HT receptor antagonists such as clozapine, methiothepin, or mianserin. High transcript numbers were detected in exocrine glands suggesting that 5-HT2 receptors participate in secretory processes in the honeybee. This study marks the first molecular and pharmacological characterization of two 5-HT2 receptor subtypes in the same insect species. The results presented should facilitate further attempts to unravel central and peripheral effects of serotonin mediated by these receptors.

  19. Function and Distribution of 5-HT2 Receptors in the Honeybee (Apis mellifera)

    PubMed Central

    Thamm, Markus; Rolke, Daniel; Jordan, Nadine; Balfanz, Sabine; Schiffer, Christian; Baumann, Arnd; Blenau, Wolfgang

    2013-01-01

    Background Serotonin plays a pivotal role in regulating and modulating physiological and behavioral processes in both vertebrates and invertebrates. In the honeybee (Apis mellifera), serotonin has been implicated in division of labor, visual processing, and learning processes. Here, we present the cloning, heterologous expression, and detailed functional and pharmacological characterization of two honeybee 5-HT2 receptors. Methods Honeybee 5-HT2 receptor cDNAs were amplified from brain cDNA. Recombinant cell lines were established constitutively expressing receptor variants. Pharmacological properties of the receptors were investigated by Ca2+ imaging experiments. Quantitative PCR was applied to explore the expression patterns of receptor mRNAs. Results The honeybee 5-HT2 receptor class consists of two subtypes, Am5-HT2α and Am5-HT2β. Each receptor gene also gives rise to alternatively spliced mRNAs that possibly code for truncated receptors. Only activation of the full-length receptors with serotonin caused an increase in the intracellular Ca2+ concentration. The effect was mimicked by the agonists 5-methoxytryptamine and 8-OH-DPAT at low micromolar concentrations. Receptor activities were blocked by established 5-HT receptor antagonists such as clozapine, methiothepin, or mianserin. High transcript numbers were detected in exocrine glands suggesting that 5-HT2 receptors participate in secretory processes in the honeybee. Conclusions This study marks the first molecular and pharmacological characterization of two 5-HT2 receptor subtypes in the same insect species. The results presented should facilitate further attempts to unravel central and peripheral effects of serotonin mediated by these receptors. PMID:24324783

  20. Differences in 5-HT2A and mGlu2 Receptor Expression Levels and Repressive Epigenetic Modifications at the 5-HT2A Promoter Region in the Roman Low- (RLA-I) and High- (RHA-I) Avoidance Rat Strains.

    PubMed

    Fomsgaard, Luna; Moreno, Jose L; de la Fuente Revenga, Mario; Brudek, Tomasz; Adamsen, Dea; Rio-Alamos, Cristobal; Saunders, Justin; Klein, Anders Bue; Oliveras, Ignasi; Cañete, Toni; Blazquez, Gloria; Tobeña, Adolf; Fernandez-Teruel, Albert; Gonzalez-Maeso, Javier; Aznar, Susana

    2018-03-01

    The serotonin 2A (5-HT 2A ) and metabotropic glutamate 2 (mGlu2) receptors regulate each other and are associated with schizophrenia. The Roman high- (RHA-I) and the Roman low- (RLA-I) avoidance rat strains present well-differentiated behavioral profiles, with the RHA-I strain emerging as a putative genetic rat model of schizophrenia-related features. The RHA-I strain shows increased 5-HT 2A and decreased mGlu2 receptor binding levels in prefrontal cortex (PFC). Here, we looked for differences in gene expression and transcriptional regulation of these receptors. The striatum (STR) was included in the analysis. 5-HT 2A , 5-HT 1A , and mGlu2 mRNA and [ 3 H]ketanserin binding levels were measured in brain homogenates. As expected, 5-HT 2A binding was significantly increased in PFC in the RHA-I rats, while no difference in binding was observed in STR. Surprisingly, 5-HT 2A gene expression was unchanged in PFC but significantly decreased in STR. mGlu2 receptor gene expression was significantly decreased in both PFC and STR. No differences were observed for the 5-HT 1A receptor. Chromatin immunoprecipitation assay revealed increased trimethylation of histone 3 at lysine 27 (H3K27me3) at the promoter region of the HTR2A gene in the STR. We further looked at the Akt/GSK3 signaling pathway, a downstream point of convergence of the serotonin and glutamate system, and found increased phosphorylation levels of GSK3β at tyrosine 216 and increased β-catenin levels in the PFC of the RHA-I rats. These results reveal region-specific regulation of the 5-HT 2A receptor in the RHA-I rats probably due to absence of mGlu2 receptor that may result in differential regulation of downstream pathways.

  1. Molecular and Pharmacological Characterization of Serotonin 5-HT2α and 5-HT7 Receptors in the Salivary Glands of the Blowfly Calliphora vicina

    PubMed Central

    Röser, Claudia; Jordan, Nadine; Balfanz, Sabine; Baumann, Arnd; Walz, Bernd; Baumann, Otto; Blenau, Wolfgang

    2012-01-01

    Secretion in blowfly (Calliphora vicina) salivary glands is stimulated by the biogenic amine serotonin (5-hydroxytryptamine, 5-HT), which activates both inositol 1,4,5-trisphosphate (InsP3)/Ca2+ and cyclic adenosine 3′,5′-monophosphate (cAMP) signalling pathways in the secretory cells. In order to characterize the signal-inducing 5-HT receptors, we cloned two cDNAs (Cv5-ht2α, Cv5-ht7) that share high similarity with mammalian 5-HT2 and 5-HT7 receptor genes, respectively. RT-PCR demonstrated that both receptors are expressed in the salivary glands and brain. Stimulation of Cv5-ht2α-transfected mammalian cells with 5-HT elevates cytosolic [Ca2+] in a dose-dependent manner (EC50 = 24 nM). In Cv5-ht7-transfected cells, 5-HT produces a dose-dependent increase in [cAMP]i (EC50 = 4 nM). We studied the pharmacological profile for both receptors. Substances that appear to act as specific ligands of either Cv5-HT2α or Cv5-HT7 in the heterologous expression system were also tested in intact blowfly salivary gland preparations. We observed that 5-methoxytryptamine (100 nM) activates only the Cv5-HT2α receptor, 5-carboxamidotryptamine (300 nM) activates only the Cv5-HT7 receptor, and clozapine (1 µM) antagonizes the effects of 5-HT via Cv5-HT7 in blowfly salivary glands, providing means for the selective activation of each of the two 5-HT receptor subtypes. This study represents the first comprehensive molecular and pharmacological characterization of two 5-HT receptors in the blowfly and permits the analysis of the physiological role of these receptors, even when co-expressed in cells, and of the modes of interaction between the Ca2+- and cAMP-signalling cascades. PMID:23145175

  2. Increasing spinal 5-HT2A receptor responsiveness mediates anti-allodynic effect and potentiates fluoxetine efficacy in neuropathic rats. Evidence for GABA release.

    PubMed

    Dupuis, Amandine; Wattiez, Anne-Sophie; Pinguet, Jérémy; Richard, Damien; Libert, Frédéric; Chalus, Maryse; Aissouni, Youssef; Sion, Benoit; Ardid, Denis; Marin, Philippe; Eschalier, Alain; Courteix, Christine

    2017-04-01

    Antidepressants are one of the first line treatments for neuropathic pain but their use is limited by the incidence and severity of side effects of tricyclics and the weak effectiveness of selective serotonin reuptake inhibitors (SSRIs). Serotonin type 2A (5-HT 2A ) receptors interact with PDZ proteins that regulate their functionality and SSRI efficacy to alleviate pain. We investigated whether an interfering peptide (TAT-2ASCV) disrupting the interaction between 5-HT 2A receptors and associated PDZ proteins would improve the treatment of traumatic neuropathic allodynia. Tactile allodynia was assessed in spinal nerve ligation-induced neuropathic pain in rats using von Frey filaments after acute treatment with TAT-2ASCV and/or 5-HT 2A receptor agonist, alone or in combination with repeated treatment with fluoxetine. In vivo microdialysis was performed in order to examine the involvement of GABA in TAT-2ASCV/fluoxetine treatment-associated analgesia. TAT-2ASCV (100ng, single i.t. injection) improved SNL-induced tactile allodynia by increasing 5-HT 2A receptor responsiveness to endogenous 5-HT. Fluoxetine alone (10mg/kg, five i.p. injections) slightly increased tactile thresholds and its co-administration with TAT-2ASCV (100ng, single i.t. injection) further enhanced the anti-allodynic effect. This effect depends on the integrity of descending serotonergic bulbospinal pathways and spinal release of GABA. The anti-allodynic effect of fluoxetine can be enhanced by disrupting 5-HT 2A receptor-PDZ protein interactions. This enhancement depends on 5-HT 2A receptor activation, spinal GABA release and GABAA receptor activation. Copyright © 2016 Elsevier Ltd. All rights reserved.

  3. Effects of Constant Flickering Light on Refractive Status, 5-HT and 5-HT2A Receptor in Guinea Pigs

    PubMed Central

    Li, Tao; Zheng, Changyue; Ji, Shunmei; Ma, Yuanyuan; Zhang, Shuangshuang; Zhou, Xiaodong

    2016-01-01

    Purpose To investigate the effects of constant flickering light on refractive development, the role of serotonin (i.e.5-hydroxytryptamine, 5-HT)and 5-HT2A receptor in myopia induced by flickering light in guinea pigs. Methods Forty-five guinea pigs were randomly divided into three groups: control, form deprivation myopia (FDM) and flickering light induced myopia (FLM) groups(n = 15 for each group). The right eyes of the FDM group were covered with semitransparent hemispherical plastic shells serving as eye diffusers. Guinea pigs in FLM group were raised with illumination of a duty cycle of 50% at a flash frequency of 0.5Hz. The refractive status, axial length (AL), corneal radius of curvature(CRC) were measured by streak retinoscope, A-scan ultrasonography and keratometer, respectively. Ultramicroscopy images were taken by electron microscopy. The concentrations of 5-HTin the retina, vitreous body and retinal pigment epithelium (RPE) were assessed by high performance liquid chromatography, the retinal 5-HT2A receptor expression was evaluated by immunohistofluorescence and western blot. Results The refraction of FDM and FLM eyes became myopic from some time point (the 4th week and the 6th week, respectively) in the course of the experiment, which was indicated by significantly decreased refraction and longer AL when compared with the controls (p<0.05). The concentrations of 5-HT in the retina, vitreous body and RPE of FDM and FLM eyes were significantly increased in comparison with those of control eyes (both p<0.05). Similar to FDM eyes, the expression of retinal 5-HT2A receptor in FLM eyes was significantly up-regulated compared to that of control eyes (both p<0.05). Western blot analysis showed that retinal 5-HT2A receptor level elevated less in the FLM eyes than that in the FDM eyes. Moreover, the levels of norepinephrine and epinephrine in FDM and FLM groups generally decreased when compared with control groups (all p<0.05). Conclusions Constant flickering

  4. High-voltage electroencephalogram spindles in rats, aging and 5-HT2 antagonism.

    PubMed

    Moyanova, S; Kortenska, L; Kirov, R

    1998-03-09

    We examined the effects of serotonin-2 (5-hydroxytryptamine-2, 5-HT2) receptor antagonists on the so-called high-voltage spindles (HVS, electroencephalographic patterns, characterized by large amplitude rhythmic waves mainly in the alpha band), recorded from the frontal cortex of young, middle-aged and old freely-moving rats during waking immobility. The study was based on the assumption that the effects of 5-HT2 receptor antagonists on the HVS activity depend on the age of rats, because there is evidence for an age-related decrease in the 5-HT2 binding sites density. Four parameters of the electroencephalogram (EEG) were used to characterize the HVS activity: the square root-transformed EEG peak power in the alpha band, the frequency corresponding to this peak (both measured from the EEG power spectra using the fast Fourier transform), the HVS mean duration, and the HVS incidence (both measured from the EEG records). The EEG parameters were analyzed after i.p. administration of three 5-HT2 receptor antagonists: ketanserin, ritanserin and cyproheptadine. In young rats, the three drugs increased the alpha power, but did not change the alpha peak-corresponding frequency. Ketanserin and ritanserin did not change the HVS mean duration and HVS incidence, while cyproheptadine increased both these parameters in young rats. In middle-aged and old untreated rats, the HVS activity was significantly increased. The three 5-HT2 antagonists did not change the HVS activity in aged rats, which could be due to age-related suppression of the 5-HT2 receptor functions. Copyright 1998 Elsevier Science B.V.

  5. Rational Design, Pharmacomodulation, and Synthesis of Dual 5-Hydroxytryptamine 7 (5-HT7)/5-Hydroxytryptamine 2A (5-HT2A) Receptor Antagonists and Evaluation by [(18)F]-PET Imaging in a Primate Brain.

    PubMed

    Deau, Emmanuel; Robin, Elodie; Voinea, Raluca; Percina, Nathalie; Satała, Grzegorz; Finaru, Adriana-Luminita; Chartier, Agnès; Tamagnan, Gilles; Alagille, David; Bojarski, Andrzej J; Morisset-Lopez, Séverine; Suzenet, Franck; Guillaumet, Gérald

    2015-10-22

    We report the synthesis of 46 tertiary amine-bearing N-alkylated benzo[d]imidazol-2(3H)-ones, imidazo[4,5-b]pyridin-2(3H)-ones, imidazo[4,5-c]pyridin-2(3H)-ones, benzo[d]oxazol-2(3H)-ones, oxazolo[4,5-b]pyridin-2(3H)-ones and N,N'-dialkylated benzo[d]imidazol-2(3H)-ones. These compounds were evaluated against 5-HT7R, 5-HT2AR, 5-HT1AR, and 5-HT6R as potent dual 5-HT7/5-HT2A serotonin receptors ligands. A thorough study of the structure-activity relationship of the aromatic rings and their substituents, the alkyl chain length and the tertiary amine was conducted. 1-(4-(4-(4-Fluorobenzoyl)piperidin-1-yl)butyl)-1H-benzo[d]imidazol-2(3H)-one (79) and 1-(6-(4-(4-fluorobenzoyl)piperidin-1-yl)hexyl)-1H-benzo[d]imidazol-2(3H)-one (81) were identified as full antagonist ligands on cyclic adenosine monophosphate (cAMP, KB = 4.9 and 5.9 nM, respectively) and inositol monophosphate (IP1, KB = 0.6 and 16 nM, respectively) signaling pathways of 5-HT7R and 5-HT2AR. Both antagonists crossed the blood-brain barrier as evaluated with [(18)F] radiolabeled compounds [(18)F]79 and [(18)F]81 in a primate's central nervous system using positron emission tomography. Both radioligands showed standard uptake values ranging from 0.8 to 1.1, a good plasmatic stability, and a distribution consistent with 5-HT7R and 5-HT2AR in the CNS.

  6. Clostridial binary toxins: iota and C2 family portraits.

    PubMed

    Stiles, Bradley G; Wigelsworth, Darran J; Popoff, Michel R; Barth, Holger

    2011-01-01

    There are many pathogenic Clostridium species with diverse virulence factors that include protein toxins. Some of these bacteria, such as C. botulinum, C. difficile, C. perfringens, and C. spiroforme, cause enteric problems in animals as well as humans. These often fatal diseases can partly be attributed to binary protein toxins that follow a classic AB paradigm. Within a targeted cell, all clostridial binary toxins destroy filamentous actin via mono-ADP-ribosylation of globular actin by the A component. However, much less is known about B component binding to cell-surface receptors. These toxins share sequence homology amongst themselves and with those produced by another Gram-positive, spore-forming bacterium also commonly associated with soil and disease: Bacillus anthracis. This review focuses upon the iota and C2 families of clostridial binary toxins and includes: (1) basics of the bacterial source; (2) toxin biochemistry; (3) sophisticated cellular uptake machinery; and (4) host-cell responses following toxin-mediated disruption of the cytoskeleton. In summary, these protein toxins aid diverse enteric species within the genus Clostridium.

  7. Clostridial Binary Toxins: Iota and C2 Family Portraits

    PubMed Central

    Stiles, Bradley G.; Wigelsworth, Darran J.; Popoff, Michel R.; Barth, Holger

    2011-01-01

    There are many pathogenic Clostridium species with diverse virulence factors that include protein toxins. Some of these bacteria, such as C. botulinum, C. difficile, C. perfringens, and C. spiroforme, cause enteric problems in animals as well as humans. These often fatal diseases can partly be attributed to binary protein toxins that follow a classic AB paradigm. Within a targeted cell, all clostridial binary toxins destroy filamentous actin via mono-ADP-ribosylation of globular actin by the A component. However, much less is known about B component binding to cell-surface receptors. These toxins share sequence homology amongst themselves and with those produced by another Gram-positive, spore-forming bacterium also commonly associated with soil and disease: Bacillus anthracis. This review focuses upon the iota and C2 families of clostridial binary toxins and includes: (1) basics of the bacterial source; (2) toxin biochemistry; (3) sophisticated cellular uptake machinery; and (4) host–cell responses following toxin-mediated disruption of the cytoskeleton. In summary, these protein toxins aid diverse enteric species within the genus Clostridium. PMID:22919577

  8. Anatomical relationships between serotonin 5-HT2A and dopamine D2 receptors in living human brain.

    PubMed

    Ishii, Tatsuya; Kimura, Yasuyuki; Ichise, Masanori; Takahata, Keisuke; Kitamura, Soichiro; Moriguchi, Sho; Kubota, Manabu; Zhang, Ming-Rong; Yamada, Makiko; Higuchi, Makoto; Okubo, Yoshinori; Suhara, Tetsuya

    2017-01-01

    Seven healthy volunteers underwent PET scans with [18F]altanserin and [11C]FLB 457 for 5-HT2A and D2 receptors, respectively. As a measure of receptor density, a binding potential (BP) was calculated from PET data for 76 cerebral cortical regions. A correlation matrix was calculated between the binding potentials of [18F]altanserin and [11C]FLB 457 for those regions. The regional relationships were investigated using a bicluster analysis of the correlation matrix with an iterative signature algorithm. We identified two clusters of regions. The first cluster identified a distinct profile of correlation coefficients between 5-HT2A and D2 receptors, with the former in regions related to sensorimotor integration (supplementary motor area, superior parietal gyrus, and paracentral lobule) and the latter in most cortical regions. The second cluster identified another distinct profile of correlation coefficients between 5-HT2A receptors in the bilateral hippocampi and D2 receptors in most cortical regions. The observation of two distinct clusters in the correlation matrix suggests regional interactions between 5-HT2A and D2 receptors in sensorimotor integration and hippocampal function. A bicluster analysis of the correlation matrix of these neuroreceptors may be beneficial in understanding molecular networks in the human brain.

  9. Purification of the Clostridium spiroforme binary toxin and activity of the toxin on HEp-2 cells.

    PubMed

    Popoff, M R; Milward, F W; Bancillon, B; Boquet, P

    1989-08-01

    The two components Sa (Mr, 44,000) and Sb (Mr, 92,000) of Clostridium spiroforme toxin were identified and characterized. Serological data permitted the identification of two groups of actin ADP-ribosylating clostridial toxins. The first consists of only C. botulinum C2. The second group includes spiroforme toxin, iota toxin of C. perfringens E, and an enzyme called CDT found in one strain of C. difficile, antibodies against which cross-react with all of the members of both groups. C. spiroforme toxin acted on cells by disrupting microfilaments by ADP-ribosylation of G actin. Toxicity was not blocked by 10 or 20 mM ammonium chloride and was only moderately inhibited by 30 mM NH4Cl. Inhibition of coated-pit formation in HEp-2 cells by potassium depletion strongly protected against the effect of C. spiroforme toxin. Toxicity was not blocked by incubation of HEp-2 cells and spiroforme toxin at 15 degrees C. These results suggest that this new binary toxin enters cells via the coated-pit-coated-vesicle pathway and might reach the cytoplasm at the same time as or before transfer to early endosomes.

  10. Purification of the Clostridium spiroforme binary toxin and activity of the toxin on HEp-2 cells.

    PubMed Central

    Popoff, M R; Milward, F W; Bancillon, B; Boquet, P

    1989-01-01

    The two components Sa (Mr, 44,000) and Sb (Mr, 92,000) of Clostridium spiroforme toxin were identified and characterized. Serological data permitted the identification of two groups of actin ADP-ribosylating clostridial toxins. The first consists of only C. botulinum C2. The second group includes spiroforme toxin, iota toxin of C. perfringens E, and an enzyme called CDT found in one strain of C. difficile, antibodies against which cross-react with all of the members of both groups. C. spiroforme toxin acted on cells by disrupting microfilaments by ADP-ribosylation of G actin. Toxicity was not blocked by 10 or 20 mM ammonium chloride and was only moderately inhibited by 30 mM NH4Cl. Inhibition of coated-pit formation in HEp-2 cells by potassium depletion strongly protected against the effect of C. spiroforme toxin. Toxicity was not blocked by incubation of HEp-2 cells and spiroforme toxin at 15 degrees C. These results suggest that this new binary toxin enters cells via the coated-pit-coated-vesicle pathway and might reach the cytoplasm at the same time as or before transfer to early endosomes. Images PMID:2545625

  11. Clostridium botulinum C2 toxin--new insights into the cellular up-take of the actin-ADP-ribosylating toxin.

    PubMed

    Aktories, Klaus; Barth, Holger

    2004-04-01

    Clostridium botulinum C2 toxin is a member of the family of binary actin-ADP-ribosylating toxins. It consists of the enzyme component C2I, and the separated binding/translocation component C2II. Proteolytically activated C2II forms heptamers and binds to a carbohydrate cell surface receptor. After attachment of C2I, the toxin complex is endocytosed to reach early endosomes. At low pH of endosomes, C2II-heptamers insert into the membrane, form pores and deliver C2I into the cytosol. Here, C2I ADP-ribosylates actin at Arg177 to block actin polymerization and to induce depolymerization of actin filaments. The mini-review describes main properties of C2 toxin and discusses new findings on the involvement of chaperones in the up-take process of the toxin.

  12. High Specific Activity Tritium-Labeled N-(2-methoxybenzyl)-2,5-dimethoxy-4-iodophenethylamine (INBMeO): A High Affinity 5-HT2A Receptor-Selective Agonist Radioligand

    PubMed Central

    Nichols, David E.; Frescas, Stewart P.; Chemel, Benjamin R.; Rehder, Kenneth S.; Zhong, Desong; Lewin, Anita H.

    2009-01-01

    The title compound ([3H]INBMeO) was prepared by an O,O-dimethylation reaction of a t-BOC protected diphenolic precursor using no carrier added tritiated iodomethane in DMF with K2CO3. Removal of the t-BOC protecting group and purification by HPLC afforded an overall yield of 43%, with a radiochemical purity of 99% and specific activity of 164 Ci/mmol. The new radioligand was suitable for labeling human 5-HT2A receptors in two heterologous cell lines and had about 20-fold higher affinity than [3H]ketanserin. PMID:18468904

  13. Effects of T-2 Toxin on Pacific White Shrimp Litopenaeus vannamei: Growth, and Antioxidant Defenses and Capacity and Histopathology in the Hepatopancreas.

    PubMed

    Deng, Yijia; Wang, Yaling; Zhang, Xiaodi; Sun, Lijun; Wu, Chaojin; Shi, Qi; Wang, Rundong; Sun, Xiaodong; Bi, Siyuan; Gooneratne, Ravi

    2017-03-01

    Modified-masked T-2 toxin (mT-2) formed during metabolism in edible aquatic animals may go undetected by traditional analytical methods, thereby underestimating T-2 toxicity. The effects of T-2 on growth and antioxidant capacity and histopathological changes in the hepatopancreas were studied in Pacific white shrimp Litopenaeus vannamei exposed for 20 d to 0, 0.5, 1.2, 2.4, 4.8, and 12.2 mg/kg of T-2 in their feed. The concentration of mT-2 in the hepatopancreas was detected by liquid chromatography-tandem mass spectrophotometry before and after trifluoroacetic acid (TFA) treatment that converted mT-2 to free T-2. A dose-dependent increase in mT-2 concentration was observed in the hepatopancreas. Dietary exposure to T-2 significantly decreased (P < 0.05) shrimp growth and survival rate compared with the controls. The malondialdehyde (MDA) concentration was significantly increased in shrimp exposed to feed with ≥2.4 mg/kg T-2 (P < 0.05). The antioxidant enzymes, superoxide dismutase (SOD) and glutathione peroxidase (GPx), total antioxidant capacity (T-AOC), and also glutathione (GSH) content increased in shrimp dosed with 2.4-4.8 mg/kg T-2 but declined at the highest dose (12.2 mg/kg), probably indicating an inability to cope with high concentrations of reactive oxygen species (ROS) as evident from a marked increase in MDA (P < 0.05) culminating in cellular toxicity. Histopathological changes in the hepatopancreas were dose dependent, with cell autophagy evident at the highest exposure dose. This is the first report in shrimp of a dose-dependent increase in ROS, SOD enzyme activity, and T-AOC at low T-2 exposures, and associated histopathological changes in the hepatopancreas, in response to dietary T-2. Received January 26, 2016; accepted October 9, 2016.

  14. Modification of smoothing in 4253H[T

    NASA Astrophysics Data System (ADS)

    Azmi, Nurul Nisa'Khairol; Adam, Mohd Bakri; Shitan, Mahendran; Ali, Norhaslinda Mohd

    2017-05-01

    Some modified non-linear smoothers particularly 4253H[T] are explained in this paper. The modifications are focused on estimating the middle point of running median for even span by applying the following types of means; geometric, harmonic, quadratic and contraharmonic. The performance of the techniques is assessed by applying it to daily price index of a bank in Malaysia that issues sukuk for funding in Islamic banking and financial business. The results show that 4253H[T] with geometric mean modification is better than others in preserving variation and curve fitting.

  15. Oligomerization of Clostridium perfringens Epsilon Toxin Is Dependent upon Caveolins 1 and 2

    PubMed Central

    Fennessey, Christine M.; Sheng, Jinsong; Rubin, Donald H.; McClain, Mark S.

    2012-01-01

    Evidence from multiple studies suggests that Clostridium perfringens ε-toxin is a pore-forming toxin, assembling into oligomeric complexes in the plasma membrane of sensitive cells. In a previous study, we used gene-trap mutagenesis to identify mammalian factors contributing to toxin activity, including caveolin-2 (CAV2). In this study, we demonstrate the importance of caveolin-2 and its interaction partner, caveolin-1 (CAV1), in ε-toxin-induced cytotoxicity. Using CAV2-specific shRNA in a toxin-sensitive human kidney cell line, ACHN, we confirmed that cells deficient in CAV2 exhibit increased resistance to ε-toxin. Similarly, using CAV1-specific shRNA, we demonstrate that cells deficient in CAV1 also exhibit increased resistance to the toxin. Immunoprecipitation of CAV1 and CAV2 from ε-toxin-treated ACHN cells demonstrated interaction of both CAV1 and -2 with the toxin. Furthermore, blue-native PAGE indicated that the toxin and caveolins were components of a 670 kDa protein complex. Although ε-toxin binding was only slightly perturbed in caveolin-deficient cells, oligomerization of the toxin was dramatically reduced in both CAV1- and CAV2-deficient cells. These results indicate that CAV1 and -2 potentiate ε-toxin induced cytotoxicity by promoting toxin oligomerization – an event which is requisite for pore formation and, by extension, cell death. PMID:23056496

  16. Oligomerization of Clostridium perfringens epsilon toxin is dependent upon caveolins 1 and 2.

    PubMed

    Fennessey, Christine M; Sheng, Jinsong; Rubin, Donald H; McClain, Mark S

    2012-01-01

    Evidence from multiple studies suggests that Clostridium perfringens ε-toxin is a pore-forming toxin, assembling into oligomeric complexes in the plasma membrane of sensitive cells. In a previous study, we used gene-trap mutagenesis to identify mammalian factors contributing to toxin activity, including caveolin-2 (CAV2). In this study, we demonstrate the importance of caveolin-2 and its interaction partner, caveolin-1 (CAV1), in ε-toxin-induced cytotoxicity. Using CAV2-specific shRNA in a toxin-sensitive human kidney cell line, ACHN, we confirmed that cells deficient in CAV2 exhibit increased resistance to ε-toxin. Similarly, using CAV1-specific shRNA, we demonstrate that cells deficient in CAV1 also exhibit increased resistance to the toxin. Immunoprecipitation of CAV1 and CAV2 from ε-toxin-treated ACHN cells demonstrated interaction of both CAV1 and -2 with the toxin. Furthermore, blue-native PAGE indicated that the toxin and caveolins were components of a 670 kDa protein complex. Although ε-toxin binding was only slightly perturbed in caveolin-deficient cells, oligomerization of the toxin was dramatically reduced in both CAV1- and CAV2-deficient cells. These results indicate that CAV1 and -2 potentiate ε-toxin induced cytotoxicity by promoting toxin oligomerization - an event which is requisite for pore formation and, by extension, cell death.

  17. Vicenin-2 inhibits Wnt/β-catenin signaling and induces apoptosis in HT-29 human colon cancer cell line.

    PubMed

    Yang, Dong; Zhang, Xiling; Zhang, Wencun; Rengarajan, Thamaraiselvan

    2018-01-01

    Colorectal cancer (CRC) is among highest prevailing cancers in the whole world, especially in western countries. For a diverse of reasons, patients prefer naturally occurring dietary substances over synthetic agents to prevent cancer. Vicenin-2 is largely available in a medicinal plant Ocimum sanctum and is an apigenin form, 6,8-di-C-glucoside, which has been reported to have a range of pharmacological values which includes antioxidant, hepatoprotective, anti-inflammatory and anti-cancer. This study was aimed to analyze the anti-proliferative effect of Vicenin-2 on human colon cancer cells via the Wnt/β-catenin signaling inhibition. MTT assay was used to assess the cell viability at different concentrations and time point. Vicenin-2 at a concentration of 50 µM (IC 50 ) decreased the phosphorylated (inactive) glycogen synthase kinase-3β, cyclin D1, and non-p-β-catenin expressions in HT-29 cells, which were evidenced through western blot analysis. Further, Vincenin-2 reduced the T-cell factor (TCF) / Leukocyte erythroid factor (LEF) reporter activity in HT-29 cells. Vicenin-2 also promoted substantial cell cycle arrest at the G 2 M phase of HT-29 cells, as well induced apoptosis in HT-29 cells, as revealed through flow cytometric analysis. Furthermore, immunoblot analysis showed that Vicenin-2 treatment enhanced the expression of Cytochrome C, Bax and caspase-3 whereas suppressed the Bcl-2 expression. Together, these results revealed that Vicenin-2 can act as a potent inhibitor of HT-29 cell proliferation and can be used as an agent against CRC.

  18. Differential actions of antiparkinson agents at multiple classes of monoaminergic receptor. III. Agonist and antagonist properties at serotonin, 5-HT(1) and 5-HT(2), receptor subtypes.

    PubMed

    Newman-Tancredi, Adrian; Cussac, Didier; Quentric, Yann; Touzard, Manuelle; Verrièle, Laurence; Carpentier, Nathalie; Millan, Mark J

    2002-11-01

    Although certain antiparkinson agents interact with serotonin (5-HT) receptors, little information is available concerning functional actions. Herein, we characterized efficacies of apomorphine, bromocriptine, cabergoline, lisuride, piribedil, pergolide, roxindole, and terguride at human (h)5-HT(1A), h5-HT(1B), and h5-HT(1D) receptors [guanosine 5'-O-(3-[(35)S]thio)triphosphate ([(35)S]GTPgammaS) binding], and at h5-HT(2A), h5-HT(2B), and h5-HT(2C) receptors (depletion of membrane-bound [(3)H]phosphatydilinositol). All drugs stimulated h5-HT(1A) receptors with efficacies (compared with 5-HT, 100%) ranging from modest (apomorphine, 35%) to high (cabergoline, 93%). At h5-HT(1B) receptors, efficacies varied from mild (terguride, 37%) to marked (cabergoline, 102%) and potencies were modest (pEC(50) values of 5.8-7.6): h5-HT(1D) sites were activated with a similar range of efficacies and greater potency (7.1-8.5). Piribedil and apomorphine were inactive at h5-HT(1B) and h5-HT(1D) receptors. At h5-HT(2A) receptors, terguride, lisuride, bromocriptine, cabergoline, and pergolide displayed potent (7.6-8.8) agonist properties (49-103%), whereas apomorphine and roxindole were antagonists and piribedil was inactive. Only pergolide (113%/8.2) and cabergoline (123%/8.6) displayed pronounced agonist properties at h5-HT(2B) receptors. At 5-HT(2C) receptors, lisuride, bromocriptine, pergolide, and cabergoline were efficacious (75-96%) agonists, apomorphine and terguride were antagonists, and piribedil was inactive. MDL100,907 and SB242,084, selective antagonists at 5-HT(2A) and 5-HT(2C) receptors, respectively, abolished these actions of pergolide, cabergoline, and bromocriptine. In conclusion, antiparkinson agents display markedly different patterns of agonist and antagonist properties at multiple 5-HT receptor subtypes. Although all show modest (agonist) activity at 5-HT(1A) sites, their contrasting actions at 5-HT(2A) and 5-HT(2C) sites may be of particular significance to their

  19. Different responses of Fe transporters in Caco2/HT29-MTX cocultures than in independent Caco-2 cell cultures

    USDA-ARS?s Scientific Manuscript database

    The human intestinal epithelium is composed of several cell types; mainly enterocytes and globet (mucin-secreting) cells. This study compares the cellular response for Fe transporters in Caco-2, HT29-MTX, and Caco-2/HT29-MTX coculture models for Fe bioavailability studies. Under culture, Caco-2 cell...

  20. EGA Protects Mammalian Cells from Clostridium difficile CDT, Clostridium perfringens Iota Toxin and Clostridium botulinum C2 Toxin

    PubMed Central

    Schnell, Leonie; Mittler, Ann-Katrin; Sadi, Mirko; Popoff, Michel R.; Schwan, Carsten; Aktories, Klaus; Mattarei, Andrea; Tehran, Domenico Azarnia; Montecucco, Cesare; Barth, Holger

    2016-01-01

    The pathogenic bacteria Clostridium difficile, Clostridium perfringens and Clostridium botulinum produce the binary actin ADP-ribosylating toxins CDT, iota and C2, respectively. These toxins are composed of a transport component (B) and a separate enzyme component (A). When both components assemble on the surface of mammalian target cells, the B components mediate the entry of the A components via endosomes into the cytosol. Here, the A components ADP-ribosylate G-actin, resulting in depolymerization of F-actin, cell-rounding and eventually death. In the present study, we demonstrate that 4-bromobenzaldehyde N-(2,6-dimethylphenyl)semicarbazone (EGA), a compound that protects cells from multiple toxins and viruses, also protects different mammalian epithelial cells from all three binary actin ADP-ribosylating toxins. In contrast, EGA did not inhibit the intoxication of cells with Clostridium difficile toxins A and B, indicating a possible different entry route for this toxin. EGA does not affect either the binding of the C2 toxin to the cells surface or the enzyme activity of the A components of CDT, iota and C2, suggesting that this compound interferes with cellular uptake of the toxins. Moreover, for C2 toxin, we demonstrated that EGA inhibits the pH-dependent transport of the A component across cell membranes. EGA is not cytotoxic, and therefore, we propose it as a lead compound for the development of novel pharmacological inhibitors against clostridial binary actin ADP-ribosylating toxins. PMID:27043629

  1. EGA Protects Mammalian Cells from Clostridium difficile CDT, Clostridium perfringens Iota Toxin and Clostridium botulinum C2 Toxin.

    PubMed

    Schnell, Leonie; Mittler, Ann-Katrin; Sadi, Mirko; Popoff, Michel R; Schwan, Carsten; Aktories, Klaus; Mattarei, Andrea; Azarnia Tehran, Domenico; Montecucco, Cesare; Barth, Holger

    2016-04-01

    The pathogenic bacteria Clostridium difficile, Clostridium perfringens and Clostridium botulinum produce the binary actin ADP-ribosylating toxins CDT, iota and C2, respectively. These toxins are composed of a transport component (B) and a separate enzyme component (A). When both components assemble on the surface of mammalian target cells, the B components mediate the entry of the A components via endosomes into the cytosol. Here, the A components ADP-ribosylate G-actin, resulting in depolymerization of F-actin, cell-rounding and eventually death. In the present study, we demonstrate that 4-bromobenzaldehyde N-(2,6-dimethylphenyl)semicarbazone (EGA), a compound that protects cells from multiple toxins and viruses, also protects different mammalian epithelial cells from all three binary actin ADP-ribosylating toxins. In contrast, EGA did not inhibit the intoxication of cells with Clostridium difficile toxins A and B, indicating a possible different entry route for this toxin. EGA does not affect either the binding of the C2 toxin to the cells surface or the enzyme activity of the A components of CDT, iota and C2, suggesting that this compound interferes with cellular uptake of the toxins. Moreover, for C2 toxin, we demonstrated that EGA inhibits the pH-dependent transport of the A component across cell membranes. EGA is not cytotoxic, and therefore, we propose it as a lead compound for the development of novel pharmacological inhibitors against clostridial binary actin ADP-ribosylating toxins.

  2. Inhibition of Shiga toxin 2 (Stx2) in apple juices and its resistance to pasteurization.

    PubMed

    Rasooly, Reuven; Do, Paula M; Levin, Carol E; Friedman, Mendel

    2010-06-01

    In the present study, we evaluated Shiga toxin (Stx2) activity in apple juices by measuring a decrease in dehydrogenase activity of Vero cells with the microculture tetrazolium (MTT) assay. Freshly prepared juice from Red Delicious apples and Golden Delicious apples inhibited the biological activity of the bacterial toxin Stx2 produced by E. coli O157:H7 strains. Studies with immunomagnetic beads bearing specific antibodies against the toxin revealed that Stx2 activity was restored when removed from the apple juice. SDS gel electrophoresis revealed no difference (P < 0.05) in the densities or molecular weights between Stx2 in either PBS or apple juices. These results suggest that Stx2 may be reversibly bound to small molecular weight constituents in the juice. The Stx2 toxin was not inactivated on exposure to heat programs (63 degrees C for 30 min, 72 degrees C for 15 s, 89 degrees C for 1 s) commonly used to pasteurize apple juice, but lost all activity when exposed to 100 degrees C for 5 min. The results suggest that pasteurization of apple juice used to inactivate E. coli O157:H7 has no effect on Stx2, and that food-compatible and safe antitoxin compounds can be used to inhibit the biological activity of the Shiga toxin.

  3. Cellular Uptake of Clostridium botulinum C2 Toxin Requires Acid Sphingomyelinase Activity.

    PubMed

    Nagahama, Masahiro; Takehara, Masaya; Takagishi, Teruhisa; Seike, Soshi; Miyamoto, Kazuaki; Kobayashi, Keiko

    2017-04-01

    Clostridium botulinum C2 toxin consists of an enzyme component (C2I) and a binding component (C2II). Activated C2II (C2IIa) binds to a cell receptor, giving rise to lipid raft-dependent oligomerization, and it then assembles with C2I. The whole toxin complex is then endocytosed into the cytosol, resulting in the destruction of the actin cytoskeleton and cell rounding. Here, we showed that C2 toxin requires acid sphingomyelinase (ASMase) activity during internalization. In this study, inhibitors of ASMase and lysosomal exocytosis blocked C2 toxin-induced cell rounding. C2IIa induced Ca 2+ influx from the extracellular medium to cells. C2 toxin-induced cell rounding was enhanced in the presence of Ca 2+ ASMase was released extracellularly when cells were incubated with C2IIa in the presence of Ca 2+ Small interfering RNA (siRNA) knockdown of ASMase reduced C2 toxin-induced cell rounding. ASMase hydrolyzes sphingomyelin to ceramide on the outer leaflet of the membrane at acidic pH. Ceramide was detected in cytoplasmic vesicles containing C2IIa. These results indicated that ASMase activity is necessary for the efficient internalization of C2 toxin into cells. Inhibitors of ASMase may confer protection against infection. Copyright © 2017 American Society for Microbiology.

  4. Fusarial toxins: secondary metabolites of Fusarium fungi.

    PubMed

    Nesic, Ksenija; Ivanovic, Snezana; Nesic, Vladimir

    2014-01-01

    Exposure to mycotoxins occurs worldwide, even though there are geographic and climatic differences in the amounts produced and occurrence of these substances.Mycotoxins are secondary chemical metabolites of different fungi. They are natural contaminants of cereals, so their presence is often inevitable. Among many genera that produce mycotoxins, Fusarium fungi are the most widespread in cereal-growing areas of the planet. Fusarium fungi produce a diversity of mycotoxin types, whose distributions are also diverse. What is produced and where it is produced is influenced primarily by environmental conditions, and crop production and storage methods. The amount of toxin produced depends on physical (viz., moisture, relative humidity, temperature, and mechanical damage), chemical (viz., carbon dioxide,oxygen, composition of substrate, insecticides and fungicides), and biological factors (viz., plant variety, stress, insects, spore load, etc.). Moisture and temperature have a major influence on mold growth rate and mycotoxin production.Among the most toxic and prevalent fusaria) toxins are the following: zearalenone,fumonisins, moniliformin and trichothecenes (T-2/HT-2 toxin, deoxynivalenol,diacetoxyscirpenol, nivalenol). Zearalenone (ZEA; ZON, F-2 toxin) isaphy to estrogenic compound, primarily a field contaminant, which exhibits estrogenic activity and has been implicated in numerous mycotoxicoses of farm animals,especially pigs. Recently, evidence suggests that ZEA has potential to stimulate the growth of human breast cancer cells. Fumonisins are also cancer-promoting metabolites,of which Fumonisin 8 I (FBI) is the most important. Moniliformin (MON) isalso highly toxic to both animals and humans. Trichothecenes are classified as gastrointestinal toxins, dermatotoxins, immunotoxins, hematotoxins, and gene toxins.T-2 and HT-2 toxin, and diacetoxyscirpenol (DAS, anguidine) are the most toxic mycotoxins among the trichothecene group. Deoxynivalenol (DON, vomitoxin) and

  5. Variation in Dopamine D2 and Serotonin 5-HT2A Receptor Genes is Associated with Working Memory Processing and Response to Treatment with Antipsychotics

    PubMed Central

    Blasi, Giuseppe; Selvaggi, Pierluigi; Fazio, Leonardo; Antonucci, Linda Antonella; Taurisano, Paolo; Masellis, Rita; Romano, Raffaella; Mancini, Marina; Zhang, Fengyu; Caforio, Grazia; Popolizio, Teresa; Apud, Jose; Weinberger, Daniel R; Bertolino, Alessandro

    2015-01-01

    Dopamine D2 and serotonin 5-HT2A receptors contribute to modulate prefrontal cortical physiology and response to treatment with antipsychotics in schizophrenia. Similarly, functional variation in the genes encoding these receptors is also associated with these phenotypes. In particular, the DRD2 rs1076560 T allele predicts a lower ratio of expression of D2 short/long isoforms, suboptimal working memory processing, and better response to antipsychotic treatment compared with the G allele. Furthermore, the HTR2A T allele is associated with lower 5-HT2A expression, impaired working memory processing, and poorer response to antipsychotics compared with the C allele. Here, we investigated in healthy subjects whether these functional polymorphisms have a combined effect on prefrontal cortical physiology and related cognitive behavior linked to schizophrenia as well as on response to treatment with second-generation antipsychotics in patients with schizophrenia. In a total sample of 620 healthy subjects, we found that subjects with the rs1076560 T and rs6314 T alleles have greater fMRI prefrontal activity during working memory. Similar results were obtained within the attentional domain. Also, the concomitant presence of the rs1076560 T/rs6314 T alleles also predicted lower behavioral accuracy during working memory. Moreover, we found that rs1076560 T carrier/rs6314 CC individuals had better responses to antipsychotic treatment in two independent samples of patients with schizophrenia (n=63 and n=54, respectively), consistent with the previously reported separate effects of these genotypes. These results indicate that DRD2 and HTR2A genetic variants together modulate physiological prefrontal efficiency during working memory and also modulate the response to antipsychotics. Therefore, these results suggest that further exploration is needed to better understand the clinical consequences of these genotype–phenotype relationships. PMID:25563748

  6. Variation in Dopamine D2 and Serotonin 5-HT2A Receptor Genes is Associated with Working Memory Processing and Response to Treatment with Antipsychotics.

    PubMed

    Blasi, Giuseppe; Selvaggi, Pierluigi; Fazio, Leonardo; Antonucci, Linda Antonella; Taurisano, Paolo; Masellis, Rita; Romano, Raffaella; Mancini, Marina; Zhang, Fengyu; Caforio, Grazia; Popolizio, Teresa; Apud, Jose; Weinberger, Daniel R; Bertolino, Alessandro

    2015-06-01

    Dopamine D2 and serotonin 5-HT2A receptors contribute to modulate prefrontal cortical physiology and response to treatment with antipsychotics in schizophrenia. Similarly, functional variation in the genes encoding these receptors is also associated with these phenotypes. In particular, the DRD2 rs1076560 T allele predicts a lower ratio of expression of D2 short/long isoforms, suboptimal working memory processing, and better response to antipsychotic treatment compared with the G allele. Furthermore, the HTR2A T allele is associated with lower 5-HT2A expression, impaired working memory processing, and poorer response to antipsychotics compared with the C allele. Here, we investigated in healthy subjects whether these functional polymorphisms have a combined effect on prefrontal cortical physiology and related cognitive behavior linked to schizophrenia as well as on response to treatment with second-generation antipsychotics in patients with schizophrenia. In a total sample of 620 healthy subjects, we found that subjects with the rs1076560 T and rs6314 T alleles have greater fMRI prefrontal activity during working memory. Similar results were obtained within the attentional domain. Also, the concomitant presence of the rs1076560 T/rs6314 T alleles also predicted lower behavioral accuracy during working memory. Moreover, we found that rs1076560 T carrier/rs6314 CC individuals had better responses to antipsychotic treatment in two independent samples of patients with schizophrenia (n=63 and n=54, respectively), consistent with the previously reported separate effects of these genotypes. These results indicate that DRD2 and HTR2A genetic variants together modulate physiological prefrontal efficiency during working memory and also modulate the response to antipsychotics. Therefore, these results suggest that further exploration is needed to better understand the clinical consequences of these genotype-phenotype relationships.

  7. Identification, expression, and pharmacology of a Cys{sub 23}-Ser{sub 23} substitution in the human 5-HT{sub 2C} receptor gene (HTR2C)

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Lappalainen, J.; Ozaki, N.; Goldman, D.

    1995-05-20

    The function of brain serotonin-2C (5-HT{sub 2C}) receptors, including behavioral and neurochemical responses to 5-HT{sub 2C} agonist challenge, has been suggested to be abnormal in individuals with neuropsychiatric disorders. Thus, it is important to identify polymorphisms and functional variants within this gene. Using SSCP analysis, the authors identified a Cys{sub 23}-Ser{sub 23} substitution (designated 5-HT{sub 2Ccys} and 5-HT{sub 2Cser}) in the first hydrophobic region of the human 5-HT{sub 2C} receptor. Allele frequencies in unrelated Caucasians were 0.13 and 0.87 for 5-HT{sub 2Cser} and 5-HT{sub 2Ccys}, respectively. DNAs from informative CEPH families were typed for this polymorphism and analyzed with respectmore » to 20 linked markers on the X chromosome. Linkage analysis placed the 5-HT{sub 2C} receptor gene (HTR2C) on Xq24. To evaluate whether this amino acid substitution causes a variant function of this receptor, recombinant human 5-HT{sub 2Ccys} and 5-HT{sub 2Cser} receptors were expressed in Xenopus oocytes and tested for responses to 5-HT using electrophysiological techniques. Concentration-response curves for 5-HT were not significantly different in oocytes expressing either form of the receptor, suggesting that the 5-HT{sub 2Ccys} and 5-HT{sub 2Cser} receptor proteins may not differ in their responses to serotonin under baseline physiological conditions. 43 refs., 3 figs., 1 tab.« less

  8. Cellular Entry of Clostridium perfringens Iota-Toxin and Clostridium botulinum C2 Toxin.

    PubMed

    Takehara, Masaya; Takagishi, Teruhisa; Seike, Soshi; Oda, Masataka; Sakaguchi, Yoshihiko; Hisatsune, Junzo; Ochi, Sadayuki; Kobayashi, Keiko; Nagahama, Masahiro

    2017-08-11

    Clostridium perfringens iota-toxin and Clostridium botulinum C2 toxin are composed of two non-linked proteins, one being the enzymatic component and the other being the binding/translocation component. These latter components recognize specific receptors and oligomerize in plasma membrane lipid-rafts, mediating the uptake of the enzymatic component into the cytosol. Enzymatic components induce actin cytoskeleton disorganization through the ADP-ribosylation of actin and are responsible for cell rounding and death. This review focuses upon the recent advances in cellular internalization of clostridial binary toxins.

  9. Cellular Entry of Clostridium perfringens Iota-Toxin and Clostridium botulinum C2 Toxin

    PubMed Central

    Takehara, Masaya; Takagishi, Teruhisa; Seike, Soshi; Oda, Masataka; Sakaguchi, Yoshihiko; Hisatsune, Junzo; Ochi, Sadayuki; Kobayashi, Keiko; Nagahama, Masahiro

    2017-01-01

    Clostridium perfringens iota-toxin and Clostridium botulinum C2 toxin are composed of two non-linked proteins, one being the enzymatic component and the other being the binding/translocation component. These latter components recognize specific receptors and oligomerize in plasma membrane lipid-rafts, mediating the uptake of the enzymatic component into the cytosol. Enzymatic components induce actin cytoskeleton disorganization through the ADP-ribosylation of actin and are responsible for cell rounding and death. This review focuses upon the recent advances in cellular internalization of clostridial binary toxins. PMID:28800062

  10. The serotonin 5-HT2C receptor and the non-addictive nature of classic hallucinogens.

    PubMed

    Canal, Clinton E; Murnane, Kevin S

    2017-01-01

    Classic hallucinogens share pharmacology as serotonin 5-HT 2A , 5-HT 2B , and 5-HT 2C receptor agonists. Unique among most other Schedule 1 drugs, they are generally non-addictive and can be effective tools in the treatment of addiction. Mechanisms underlying these attributes are largely unknown. However, many preclinical studies show that 5-HT 2C agonists counteract the addictive effects of drugs from several classes, suggesting this pharmacological property of classic hallucinogens may be significant. Drawing from a comprehensive analysis of preclinical behavior, neuroanatomy, and neurochemistry studies, this review builds rationale for this hypothesis, and also proposes a testable, neurobiological framework. 5-HT 2C agonists work, in part, by modulating dopamine neuron activity in the ventral tegmental area-nucleus accumbens (NAc) reward pathway. We argue that activation of 5-HT 2C receptors on NAc shell, GABAergic, medium spiny neurons inhibits potassium Kv1.x channels, thereby enhancing inhibitory activity via intrinsic mechanisms. Together with experiments that show that addictive drugs, such as cocaine, potentiate Kv1.x channels, thereby suppressing NAc shell GABAergic activity, this hypothesis provides a mechanism by which classic hallucinogen-mediated stimulation of 5-HT 2C receptors could thwart addiction. It also provides a potential reason for the non-addictive nature of classic hallucinogens.

  11. Metabotropic glutamate mGlu2 receptor is necessary for the pharmacological and behavioral effects induced by hallucinogenic 5-HT2A receptor agonists

    PubMed Central

    Moreno, José L.; Holloway, Terrell; Albizu, Laura; Sealfon, Stuart C.; González-Maeso, Javier

    2011-01-01

    Hallucinogenic drugs, including mescaline, psilocybin and lysergic acid diethylamide (LSD), act at serotonin 5-HT2A receptors (5-HT2ARs). Metabotropic glutamate receptor 2/3 (mGluR2/3) ligands show efficacy in modulating the responses induced by activation of 5-HT2ARs. The formation of a 5-HT2AR-mGluR2 complex suggests a functional interaction that affects the hallucinogen-regulated cellular signaling pathways. Here, we tested the cellular and behavioral effects of hallucinogenic 5-HT2AR agonists in mGluR2 knockout (mGluR2-KO) mice. Mice were intraperitoneally injected with the hallucinogens DOI (2 mg/kg) and LSD (0.24 mg/kg), or vehicle. Head-twitch behavioral response, expression of c-fos, which is induced by all 5-HT2AR agonists, and expression of egr-2, which is hallucinogen-specific, were determined in wild type and mGluR2-KO mice. [3H]Ketanserin binding displacement curves by DOI were performed in mouse frontal cortex membrane preparations. Head twitch behavior was abolished in mGluR2-KO mice. The high-affinity binding site of DOI was undetected in mGluR2-KO mice. The hallucinogen DOI induced c-fos in both wild type and mGluR2-KO mice. However, the induction of egr-2 by DOI was eliminated in mGlu2-KO mice. These findings suggest that the 5-HT2AR-mGluR2 complex is necessary for the neuropsychological responses induced by hallucinogens. PMID:21276828

  12. Beta2 toxin is not involved in in vitro cell cytotoxicity caused by human and porcine cpb2-harbouring Clostridium perfringens.

    PubMed

    Allaart, Janneke G; van Asten, Alphons J A M; Vernooij, Johannes C M; Gröne, Andrea

    2014-06-25

    Clostridium perfringens is a common cause of intestinal disease in animals and humans. Its pathogenicity is attributed to the toxins it can produce, including the beta2 toxin. The presence of cpb2, the gene encoding the beta2 toxin, has been associated with diarrhoea in neonatal piglets and humans. However, the exact role of the beta2 toxin in the development of diarrhoea is still unknown. In this study we investigated the level of cytotoxicity to porcine IPI-21 and human Caco-2 cell-lines caused by porcine and human cpb2-harbouring C. perfringens and the significance of the beta2 toxin for the induction of cell cytotoxicity. Supernatants of porcine cpb2-harbouring C. perfringens strains were cytotoxic to both cell lines. Cell cytotoxicity caused by supernatant of human cpb2-harbouring C. perfringens strains was variable among strains. However, removal of the beta2 toxin by anti-beta2 toxin antibodies or degradation of the beta2 toxin by trypsin did not reduce the cytotoxic effect of any of the supernatants. These data suggest that beta2 toxin does not play a role in the development of cell cytotoxicity in in vitro experiments. In vivo studies are necessary to definitely define the role of beta2 toxin in the development of cell cytotoxicity and subsequent diarrhoea. Copyright © 2014 Elsevier B.V. All rights reserved.

  13. MDMA Increases Excitability in the Dentate Gyrus: Role of 5HT2A Receptor Induced PGE2 Signaling

    PubMed Central

    Collins, Stuart A.; Huff, Courtney; Chiaia, Nicolas; Gudelsky, Gary A.; Yamamoto, Bryan K.

    2015-01-01

    MDMA is a widely abused psychostimulant which causes release of serotonin in various forebrain regions. Recently, we reported that MDMA increases extracellular glutamate concentrations in the dentate gyrus, via activation of 5HT2A receptors. We examined the role of prostaglandin signaling in mediating the effects of 5HT2A receptor activation on the increases in extracellular glutamate and the subsequent long-term loss of parvalbumin interneurons in the dentate gyrus caused by MDMA. Administration of MDMA into the dentate gyrus of rats increased PGE2 concentrations which was prevented by coadministration of MDL100907, a 5HT2A receptor antagonist. MDMA-induced increases in extracellular glutamate were inhibited by local administration of SC-51089, an inhibitor of the EP1 prostaglandin receptor. Systemic administration of SC-51089 during injections of MDMA prevented the decreases in parvalbumin interneurons observed 10 days later. The loss of parvalbumin immunoreactivity after MDMA exposure coincided with a decrease in paired-pulse inhibition and afterdischarge threshold in the dentate gyrus. These changes were prevented by inhibition of EP1 and 5HT2A receptors during MDMA. Additional experiments revealed an increased susceptibility to kainic acid-induced seizures in MDMA treated rats which could be prevented with SC51089 treatments during MDMA exposure. Overall, these findings suggest that 5HT2A receptors mediate MDMA-induced PGE2 signaling and subsequent increases in glutamate. This signaling mediates parvalbumin cell losses as well as physiologic changes in the dentate gyrus, suggesting that the lack of the inhibition provided by these neurons increases the excitability within the dentate gyrus of MDMA treated rats. PMID:26670377

  14. K2 killer toxin-induced physiological changes in the yeast Saccharomyces cerevisiae.

    PubMed

    Orentaite, Irma; Poranen, Minna M; Oksanen, Hanna M; Daugelavicius, Rimantas; Bamford, Dennis H

    2016-03-01

    Saccharomyces cerevisiae cells produce killer toxins, such as K1, K2 and K28, that can modulate the growth of other yeasts giving advantage for the killer strains. Here we focused on the physiological changes induced by K2 toxin on a non-toxin-producing yeast strain as well as K1, K2 and K28 killer strains. Potentiometric measurements were adjusted to observe that K2 toxin immediately acts on the sensitive cells leading to membrane permeability. This correlated with reduced respiration activity, lowered intracellular ATP content and decrease in cell viability. However, we did not detect any significant ATP leakage from the cells treated by killer toxin K2. Strains producing heterologous toxins K1 and K28 were less sensitive to K2 than the non-toxin producing one suggesting partial cross-protection between the different killer systems. This phenomenon may be connected to the observed differences in respiratory activities of the killer strains and the non-toxin-producing strain at low pH. This might also have practical consequences in wine industry; both as beneficial ones in controlling contaminating yeasts and non-beneficial ones causing sluggish fermentation. © FEMS 2016. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.

  15. Serotonergic modulation of the rat pup ultrasonic isolation call: studies with 5HT1 and 5HT2 subtype-selective agonists and antagonists.

    PubMed

    Winslow, J T; Insel, T R

    1991-01-01

    A modulatory role for serotonin has been described for the development and expression of the ultrasonic call of infant rat pups during brief maternal separations. In previous studies, serotonin reuptake inhibitors selectively reduced the rate of calling following acute administration to 9-11-day-old pups and a serotonin neurotoxin (MDMA) systematically disrupted the development of ultrasonic vocalizations but not other measures of motor development. In the current studies, we extended our investigations to include drugs with purported receptor subtype selectivities. Consistent with previous reports, acute administration of 5HT1A agonists buspirone and 8-OH-DPAT [+/-)-8-hydroxy-2-(di-N-propylamino)tetralin) reduced the rate of calling at doses which did not affect motor activity or core body temperature. The rate reducing effects of buspirone persisted up to 1 but not 2 h after injection. Administration of purported 5HT1B receptor agonists, CGS12066B (7-trifluoromethyl-4(4-methyl-1-piperazinyl)-pyrrolo[1,2-a] quinoxaline) and TFMPP (1-[3-fluoromethyl)phenyl]-piperazine) increased the rate of calling depending on the specificity of the drug for the 5HT1B receptor. d,l-Propranolol, a 5HT1 receptor antagonist, blocked the effects of both 8-OH-DPAT and TFMPP. m-CPP (1-(3-chlorophenyl)piperazine) and DOI [+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane), drugs with putative actions at 5HT1C and 5HT2 receptor sites both decreased calling but differed according to their effects on motor activity. Ritanserin, a 5HT2 and 5HT1C antagonist, produced a dose-related increase in call rate. A dose of ritanserin with no apparent intrinsic effects effectively antagonized DOI rate reducing effects but potentiated the rate reducing effects of m-CPP.(ABSTRACT TRUNCATED AT 250 WORDS)

  16. Blockade of 5-HT2A/2C-type receptors impairs learning in female rats in the course of estrous cycle.

    PubMed

    Fedotova, Yu O; Ordyan, N E

    2010-12-01

    We studied the effects of chronic administration (14 days) of agonist of 5-HT2B/2C serotonin receptors m-CPP (0.5 mg/kg subcutaneously) and agonist of 5-HT2A/2C serotonin receptors ketanserin (0.1 mg/kg intraperitoneally) on conditioned reactions in female rats in different phases of the estrous cycle. Passive avoidance (PA) paradigm and Morris water maze were used as behavioral tests. Chronic administration of m-CPP did not affect PA retrieval during the proestrus and estrus phases, but improved the dynamics of spatial learning in Morris water maze in comparison with control rats. Chronic administration of ketanserin uniformly impaired processes of spatial and nonspatial learning in female rats irrespective to the phase of the estrous cycle. A modulating role of 5-HT2A/2C and 5-HT2B/2C serotonin receptors in process of learning in female rats during the key phases of the estrous cycle was demonstrated.

  17. Metabotropic glutamate mGlu2 receptor is necessary for the pharmacological and behavioral effects induced by hallucinogenic 5-HT2A receptor agonists.

    PubMed

    Moreno, José L; Holloway, Terrell; Albizu, Laura; Sealfon, Stuart C; González-Maeso, Javier

    2011-04-15

    Hallucinogenic drugs, including mescaline, psilocybin and lysergic acid diethylamide (LSD), act at serotonin 5-HT2A receptors (5-HT2ARs). Metabotropic glutamate receptor 2/3 (mGluR2/3) ligands show efficacy in modulating the responses induced by activation of 5-HT2ARs. The formation of a 5-HT2AR-mGluR2 complex suggests a functional interaction that affects the hallucinogen-regulated cellular signaling pathways. Here, we tested the cellular and behavioral effects of hallucinogenic 5-HT2AR agonists in mGluR2 knockout (mGluR2-KO) mice. Mice were intraperitoneally injected with the hallucinogens DOI (2 mg/kg) and LSD (0.24 mg/kg), or vehicle. Head-twitch behavioral response, expression of c-fos, which is induced by all 5-HT2AR agonists, and expression of egr-2, which is hallucinogen-specific, were determined in wild type and mGluR2-KO mice. [(3)H]Ketanserin binding displacement curves by DOI were performed in mouse frontal cortex membrane preparations. Head twitch behavior was abolished in mGluR2-KO mice. The high-affinity binding site of DOI was undetected in mGluR2-KO mice. The hallucinogen DOI induced c-fos in both wild type and mGluR2-KO mice. However, the induction of egr-2 by DOI was eliminated in mGlu2-KO mice. These findings suggest that the 5-HT2AR-mGluR2 complex is necessary for the neuropsychological responses induced by hallucinogens. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  18. 5-HT2C Receptor Desensitization Moderates Anxiety in 5-HTT Deficient Mice: From Behavioral to Cellular Evidence

    PubMed Central

    Martin, Cédric BP; Martin, Vincent S.; Trigo, José M.; Chevarin, Caroline; Maldonado, Rafael; Fink, Latham H.; Cunningham, Kathryn A.; Hamon, Michel; Lanfumey, Laurence

    2015-01-01

    Background: Desensitization and blockade of 5-HT2C receptors (5-HT2CR) have long been thought to be central in the therapeutic action of antidepressant drugs. However, besides behavioral pharmacology studies, there is little in vivo data documenting antidepressant-induced 5-HT2CR desensitization in specific brain areas. Methods: Mice lacking the 5-HT reuptake carrier (5-HTT-/-) were used to model the consequences of chronic 5-HT reuptake inhibition with antidepressant drugs. The effect of this mutation on 5-HT2CR was evaluated at the behavioral (social interaction, novelty-suppressed feeding, and 5-HT2CR–induced hypolocomotion tests), the neurochemical, and the cellular (RT-qPCR, mRNA editing, and c-fos–induced expression) levels. Results: Although 5-HTT-/- mice had an anxiogenic profile in the novelty-suppressed feeding test, they displayed less 5-HT2CR–mediated anxiety in response to the agonist m-chlorophenylpiperazine in the social interaction test. In addition, 5-HT2CR–mediated inhibition of a stress-induced increase in 5-HT turnover, measured in various brain areas, was markedly reduced in 5-HTT-/- mutants. These indices of tolerance to 5-HT2CR stimulation were associated neither with altered levels of 5-HT2CR protein and mRNA nor with changes in pre-mRNA editing in the frontal cortex. However, basal c-fos mRNA production in cells expressing 5-HT2CR was higher in 5-HTT-/- mutants, suggesting an altered basal activity of these cells following sustained 5-HT reuptake carrier inactivation. Furthermore, the increased c-fos mRNA expression in 5-HT2CR–like immune-positive cortical cells observed in wild-type mice treated acutely with the 5-HT2CR agonist RO-60,0175 was absent in 5-HTT-/- mutants. Conclusions: Such blunted responsiveness of the 5-HT2CR system, observed at the cell signaling level, probably contributes to the moderation of the anxiety phenotype in 5-HTT-/- mice. PMID:25522398

  19. Actions of alpha2 adrenoceptor ligands at alpha2A and 5-HT1A receptors: the antagonist, atipamezole, and the agonist, dexmedetomidine, are highly selective for alpha2A adrenoceptors.

    PubMed

    Newman-Tancredi, A; Nicolas, J P; Audinot, V; Gavaudan, S; Verrièle, L; Touzard, M; Chaput, C; Richard, N; Millan, M J

    1998-08-01

    This study examined the activity of chemically diverse alpha2 adrenoceptor ligands at recombinant human (h) and native rat (r) alpha2A adrenoceptors compared with 5-HT1A receptors. First, in competition binding experiments at h alpha2A and h5-HT1A receptors expressed in CHO cells, several compounds, including the antagonists 1-(2-pyrimidinyl)piperazine (1-PP), (+/-)-idazoxan, benalfocin (SKF 86466), yohimbine and RX 821,002, displayed preference for h alpha2A versus h5-HT1A receptors of only 1.4-, 3.6-, 4-, 10- and 11-fold, respectively (based on differences in pKi values). Clonidine, brimonidine (UK 14304), the benzopyrrolidine fluparoxan and the guanidines guanfacine and guanabenz exhibited intermediate selectivity (22- to 31-fold) for h alpha2A receptors. Only the antagonist atipamezole and the agonist dexmedetomidine (DMT) displayed high preference for alpha2 adrenoceptors (1290- and 91-fold, respectively). Second, the compounds were tested for their ability to induce h5-HT1A receptor-mediated G-protein activation, as indicated by the stimulation of [35S]GTPgammaS binding. All except atipamezole and RX 821,002 exhibited agonist activity, with potencies which correlated with their affinity for h5-HT1A receptors. Relative efficacies (Emax values) were 25-35% for guanabenz, guanfacine, WB 4101 and benalfocin, 50-65% for 1-PP, (+/-)-idazoxan and clonidine, and over 70% for fluparoxan, oxymetazoline and yohimbine (relative to 5-HT = 100%). Yohimbine-induced [35S]GTPgammaS binding was inhibited by the selective 5-HT1A receptor antagonist WAY 100,635. In contrast, RX 821,002 was the only ligand which exhibited antagonist activity at h5-HT1A receptors, inhibiting 5-HT-stimulated [35S]GTPgammaS binding. Atipamezole, which exhibited negligeable affinity for 5-HT1A receptors, was inactive. Third, the affinities for r alpha2A differed considerably from the affinities for h alpha2A receptors whereas the affinities for r5-HT1A differed much less from the affinities for h5-HT

  20. The serotonin 5-HT2C receptor and the non-addictive nature of classic hallucinogens

    PubMed Central

    Canal, Clinton E; Murnane, Kevin S

    2017-01-01

    Classic hallucinogens share pharmacology as serotonin 5-HT2A, 5-HT2B, and 5-HT2C receptor agonists. Unique among most other Schedule 1 drugs, they are generally non-addictive and can be effective tools in the treatment of addiction. Mechanisms underlying these attributes are largely unknown. However, many preclinical studies show that 5-HT2C agonists counteract the addictive effects of drugs from several classes, suggesting this pharmacological property of classic hallucinogens may be significant. Drawing from a comprehensive analysis of preclinical behavior, neuroanatomy, and neurochemistry studies, this review builds rationale for this hypothesis, and also proposes a testable, neurobiological framework. 5-HT2C agonists work, in part, by modulating dopamine neuron activity in the ventral tegmental area—nucleus accumbens (NAc) reward pathway. We argue that activation of 5-HT2C receptors on NAc shell, GABAergic, medium spiny neurons inhibits potassium Kv1.x channels, thereby enhancing inhibitory activity via intrinsic mechanisms. Together with experiments that show that addictive drugs, such as cocaine, potentiate Kv1.x channels, thereby suppressing NAc shell GABAergic activity, this hypothesis provides a mechanism by which classic hallucinogen-mediated stimulation of 5-HT2C receptors could thwart addiction. It also provides a potential reason for the non-addictive nature of classic hallucinogens. PMID:27903793

  1. Target size analysis of serotonin 5-HT/sub 1/ and 5-HT/sub 2/ receptors in bovine brain membranes

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Nishino, N.; Tanaka, C.

    1985-09-23

    Freeze-dried crude synaptic membranes prepared from bovine cerebral cortex and striatum were exposed to high energy gamma ray from the source of /sup 60/Co. The size of serotonin 5-HT/sub 1/ receptors labeled by (/sup 3/H)serotonin and that of 5-HT/sub 2/ receptors labeled by (/sup 3/H)spiperone or (/sup 3/H)ketanserin was determined by target size analyses. The values were 57,000 daltons, 145,000 daltons and 152,000 daltons for the cerebral cortex and 56,000 daltons, 141,000 daltons and 150,000 daltons for the striatum, respectively. The estimated sizes were deduced by reference to enzyme standards with known molecular masses and which were irradiated in parallel.more » These results demonstrate that the molecular entities in situ for 5-HT/sub 1/ receptors are distinct from those for 5-HT/sub 2/ receptors, thus supporting data on the existence of two distinct populations of serotonin receptors, hitherto evidenced physiopharmacologically.« less

  2. Investigation of T-2 Mycotoxin-Induced Cytotoxicity in vitro and Protective Effects of Flavonoid Compounds

    DTIC Science & Technology

    1986-01-01

    Quercetin , a flavonoid compound was able to decrease the effect of T-2 toxin when the drug was added within an hour of mixing the T-2 toxin with the...were examined microscopically using a Neubauer hemocytometer and viability of at least 200 cells was deter- mined. Quercetin or other flavonold... quercetin and additional OMSO had a cytotoxic effect on the thymocytes. RESULTS Figure 1 shows the results of 8 separate experiments performed at 2 week

  3. Paper-based immune-affinity arrays for detection of multiple mycotoxins in cereals.

    PubMed

    Li, Li; Chen, Hongpu; Lv, Xiaolan; Wang, Min; Jiang, Xizhi; Jiang, Yifei; Wang, Heye; Zhao, Yongfu; Xia, Liru

    2018-03-01

    Mycotoxins produced by different species of fungi may coexist in cereals and feedstuffs, and could be highly toxic for humans and animals. For quantification of multiple mycotoxins in cereals, we developed a paper-based mycotoxin immune-affinity array. First, paper-based microzone arrays were fabricated by photolithography. Then, monoclonal mycotoxin antibodies were added in a copolymerization reaction with a cross-linker to form an immune-affinity monolith on the paper-based microzone array. With use of a competitive immune-response format, paper-based mycotoxin immune-affinity arrays were successfully applied to detect mycotoxins in samples. The detection limits for deoxynivalenol, zearalenone, T-2 toxin, and HT-2 toxin were 62.7, 10.8, 0.36, and 0.23 μg·kg -1 , respectively, which meet relevant requirements for these compounds in food. The recovery rates were 81-86% for deoxynivalenol, 89-117% for zearalenone, 79-86% for T-2 toxin, and 78-83% for HT-2 toxin, and showed the paper-based immune-affinity arrays had good reproducibility. In summary, the paper-based mycotoxin immune-affinity array provides a sensitive, rapid, accurate, stable, and convenient platform for detection of multiple mycotoxins in agro-foods. Graphical abstract Paper-based immune-affinity monolithic array. DON deoxynivalenol, HT-2 HT-2 toxin, T-2 T-2 toxin, PEGDA polyethylene glycol diacrylate, ZEN zearalenone.

  4. Dm5-HT2B: Pharmacological Characterization of the Fifth Serotonin Receptor Subtype of Drosophila melanogaster.

    PubMed

    Blenau, Wolfgang; Daniel, Stöppler; Balfanz, Sabine; Thamm, Markus; Baumann, Arnd

    2017-01-01

    Serotonin (5-hydroxytryptamine, 5-HT) is an important regulator of physiological and behavioral processes in both protostomes (e.g., insects) and deuterostomes (e.g., mammals). In insects, serotonin has been found to modulate the heart rate and to control secretory processes, development, circadian rhythms, aggressive behavior, as well as to contribute to learning and memory. Serotonin exerts its activity by binding to and activating specific membrane receptors. The clear majority of these receptors belong to the superfamily of G-protein-coupled receptors. In Drosophila melanogaster , a total of five genes have been identified coding for 5-HT receptors. From this family of proteins, four have been pharmacologically examined in greater detail, so far. While Dm5-HT 1A , Dm5-HT 1B , and Dm5-HT 7 couple to cAMP signaling cascades, the Dm5-HT 2A receptor leads to Ca 2+ signaling in an inositol-1,4,5-trisphosphate-dependent manner. Based on sequence similarity to homologous genes in other insects, a fifth D. melanogaster gene was uncovered coding for a Dm5-HT 2B receptor. Knowledge about this receptor's pharmacological properties is very limited. This is quite surprising because Dm5-HT 2B has been attributed to distinct physiological functions based on genetic interference with its gene expression. Mutations were described reducing the response of the larval heart to 5-HT, and specific knockdown of Dm5-HT 2B mRNA in hemocytes resulted in a higher susceptibility of the flies to bacterial infection. To gain deeper understanding of Dm5-HT 2B 's pharmacology, we evaluated the receptor's response to a series of established 5-HT receptor agonists and antagonists in a functional cell-based assay. Metoclopramide and mianserin were identified as two potent antagonists that may allow pharmacological interference with Dm5-HT 2B signaling in vitro and in vivo .

  5. Dm5-HT2B: Pharmacological Characterization of the Fifth Serotonin Receptor Subtype of Drosophila melanogaster

    PubMed Central

    Blenau, Wolfgang; Daniel, Stöppler; Balfanz, Sabine; Thamm, Markus; Baumann, Arnd

    2017-01-01

    Serotonin (5-hydroxytryptamine, 5-HT) is an important regulator of physiological and behavioral processes in both protostomes (e.g., insects) and deuterostomes (e.g., mammals). In insects, serotonin has been found to modulate the heart rate and to control secretory processes, development, circadian rhythms, aggressive behavior, as well as to contribute to learning and memory. Serotonin exerts its activity by binding to and activating specific membrane receptors. The clear majority of these receptors belong to the superfamily of G-protein-coupled receptors. In Drosophila melanogaster, a total of five genes have been identified coding for 5-HT receptors. From this family of proteins, four have been pharmacologically examined in greater detail, so far. While Dm5-HT1A, Dm5-HT1B, and Dm5-HT7 couple to cAMP signaling cascades, the Dm5-HT2A receptor leads to Ca2+ signaling in an inositol-1,4,5-trisphosphate-dependent manner. Based on sequence similarity to homologous genes in other insects, a fifth D. melanogaster gene was uncovered coding for a Dm5-HT2B receptor. Knowledge about this receptor’s pharmacological properties is very limited. This is quite surprising because Dm5-HT2B has been attributed to distinct physiological functions based on genetic interference with its gene expression. Mutations were described reducing the response of the larval heart to 5-HT, and specific knockdown of Dm5-HT2B mRNA in hemocytes resulted in a higher susceptibility of the flies to bacterial infection. To gain deeper understanding of Dm5-HT2B’s pharmacology, we evaluated the receptor’s response to a series of established 5-HT receptor agonists and antagonists in a functional cell-based assay. Metoclopramide and mianserin were identified as two potent antagonists that may allow pharmacological interference with Dm5-HT2B signaling in vitro and in vivo. PMID:28553207

  6. 5-HT2A receptor gene polymorphisms in Croatian subjects with autistic disorder.

    PubMed

    Hranilovic, Dubravka; Blazevic, Sofia; Babic, Marina; Smurinic, Maja; Bujas-Petkovic, Zorana; Jernej, Branimir

    2010-08-15

    Disturbances in the expression/function of the 5-HT2A receptor are implicated in autism. The association of the 5-HT2A receptor gene with autism was studied in the Croatian population. Distribution frequencies for alleles, genotypes and haplotypes of -1438 A/G and His452Tyr polymorphisms were compared in samples of 103 autistic and 214 control subjects. Significant overrepresentation of the G allele and the GG genotype of the -1438 A/G polymorphism was observed in group of autistic subjects, supporting the possible involvement of the 5-HT2A receptor in the development of autism. Copyright (c) 2010 Elsevier Ltd. All rights reserved.

  7. The involvement of medial septum 5-HT1 and 5-HT2 receptors on ACPA-induced memory consolidation deficit: possible role of TRPC3, TRPC6 and TRPV2.

    PubMed

    Najar, Farzaneh; Nasehi, Mohammad; Haeri-Rohani, Seyed-Ali; Zarrindast, Mohammad-Reza

    2015-11-01

    The present study evaluates the roles of serotonergic receptors of the medial septum on amnesia induced by arachidonylcyclopropylamide (ACPA; as selective cannabinoid CB1 receptor agonist) in adult male Wistar rats. Cannulae were implanted in the medial septum of the brain of the rats. The animals were trained in a passive avoidance learning apparatus, and were tested 24 hours after training for step-through latency. Results indicated that post-training medial septum administration of CP94253 (5-HT1B/1D receptor agonist) and cinancerine (as 5-HT2 receptor antagonist) reduced the step-through latency showing an amnesic response, while GR127935 (5-HT1B/1D receptor antagonist) and αm5htm (as 5-HT2A/2B/2D receptor agonist) did not alter memory consolidation by themselves. On continuing the test, the results showed that CP94253 increased and GR127935 did not alter ACPA (0.02 µg/rat)-induced memory impairment, respectively. Other data indicated that αm5htm induced a modulatory effect, while cinancerine restored ACPA-induced amnesia. Using SKF-96365 (inhibitor of transient receptor potential TRPC3/6 and TRPV2 channels) demonstrated that TRPC3, TRPC3 and TRPV2 channels have a significant role, according to our results. © The Author(s) 2015.

  8. Multiple mechanisms of serotonin 5-HT2 receptor desensitization.

    PubMed

    Rahman, S; Neuman, R S

    1993-07-20

    Desensitization of serotonin 5-HT2 receptor-mediated enhancement of the N-methyl-D-aspartate (NMDA) depolarization was studied in rat cortical neurons. Serotonin and (+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) induced long term desensitization. Staurosporine, a nonspecific protein kinase C inhibitor, potentiated the serotonin and DOI facilitation, suggesting acute desensitization was operative. In the case of DOI, long term desensitization was prevented by staurosporine. Activators of protein kinase C abolished the serotonin facilitation, an action prevented by staurosporine. Concanavalin A potentiated the facilitation at 100 microM, but not 30 microM serotonin, suggesting these receptors undergo dose dependent internalization. Calmodulin antagonists prevent long term desensitization induced by serotonin. The depolarization induced by NMDA alone was not altered by staurosporine, protein kinase C activators, concanavalin A or calmodulin antagonists. Serotonin at 100 microM, but not 30 microM, induced heterologous desensitization of phenylephrine and carbachol induced facilitation of the NMDA depolarization. We conclude that serotonin 5-HT2 receptors both induce and undergo several forms of desensitization.

  9. Receptor interaction profiles of novel N-2-methoxybenzyl (NBOMe) derivatives of 2,5-dimethoxy-substituted phenethylamines (2C drugs).

    PubMed

    Rickli, Anna; Luethi, Dino; Reinisch, Julian; Buchy, Danièle; Hoener, Marius C; Liechti, Matthias E

    2015-12-01

    N-2-methoxybenzyl-phenethylamines (NBOMe drugs) are newly used psychoactive substances with poorly defined pharmacological properties. The aim of the present study was to characterize the receptor binding profiles of a series of NBOMe drugs compared with their 2,5-dimethoxy-phenethylamine analogs (2C drugs) and lysergic acid diethylamide (LSD) in vitro. We investigated the binding affinities of 2C drugs (2C-B, 2C-C, 2C-D, 2C-E, 2C-H, 2C-I, 2C-N, 2C-P, 2C-T-2, 2C-T-4, 2C-T-7, and mescaline), their NBOMe analogs, and LSD at monoamine receptors and determined functional 5-hydroxytryptamine-2A (5-HT2A) and 5-HT2B receptor activation. Binding at and the inhibition of monoamine uptake transporters were also determined. Human cells that were transfected with the respective human receptors or transporters were used (with the exception of trace amine-associated receptor-1 [TAAR1], in which rat/mouse receptors were used). All of the compounds potently interacted with serotonergic 5-HT2A, 5-HT2B, 5-HT2C receptors and rat TAAR1 (most Ki and EC50: <1 μM). The N-2-methoxybenzyl substitution of 2C drugs increased the binding affinity at serotonergic 5-HT2A, 5-HT2C, adrenergic α1, dopaminergic D1-3, and histaminergic H1 receptors and monoamine transporters but reduced binding to 5-HT1A receptors and TAAR1. As a result, NBOMe drugs were very potent 5-HT2A receptor agonists (EC50: 0.04-0.5 μM) with high 5-HT2A/5-HT1A selectivity and affinity for adrenergic α1 receptors (Ki: 0.3-0.9 μM) and TAAR1 (Ki: 0.06-2.2 μM), similar to LSD, but not dopaminergic D1-3 receptors (most Ki:>1 μM), unlike LSD. The binding profile of NBOMe drugs predicts strong hallucinogenic effects, similar to LSD, but possibly more stimulant properties because of α1 receptor interactions. Copyright © 2015 Elsevier Ltd. All rights reserved.

  10. Identification of a cys-ser substitution in the 5-HT{sub 2C} (HTR2C) receptor gene and allelic association to violent behavior and alcoholism

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Lappalainen, J.; Ozaki, N.; Goldman, D.

    1994-09-01

    Several lines of evidence suggest that brain serotonergic functions, including behavioral and neurochemical responses to 5-HT{sub 2C} agonist, are abnormal in some individuals with alcoholism and aggressive behaviors. The aim of the present study was to identify coding sequence variants in the human 5-HT{sub 2C} receptor gene which may cause abnormal or variant function of this receptor. Using SSCP analysis, a non-conservative cys-ser substitution was found in the 5-HT{sub 2C} receptor (designated 5-HT{sub 2Ccys} and 5-HT{sub 2Cser}). The polymorphism was typed in CEPH families to genetically map the gene. To test for association of the variant to alcoholism, violent behaviormore » and serotonin function, the 5-HT{sub 2C} genotypes of 151 non-related Finnish male alcoholic violent offenders and impulsive fire setters and 127 Finnish psychiatrically interviewed healthy male volunteers were determined. CSF 5-HIAA concentrations were available for 74 alcoholic violent offenders and 25 healthy volunteers. Linkage analysis placed the 5-HT{sub 2C} gene on Xq21, a region that has been previously shown to contain genes for several mental retardation syndromes. The 5-HT{sub 2Ccys}/5-HT{sub 2Cser} genotype frequencies in alcoholic violent offenders and controls differed significantly (0.90/0.10 and 0.82/0.18, respectively, P=0.048). The association was found to be strongest in the violent offenders who did not fulfill the criteria for antisocial personality disorder (5-HT{sub 2Ccys}/5-HT{sub 2Cser} 0.93/0.07, p=0.021). No association was found between CSF 5-HIAA concentrations and 5-HT{sub 2C} genotype. These results implicate a 5-HT{sub 2C} receptor amino acid substitution in predisposition to alcohol abuse and violent behavior in a subgroup of alcoholics.« less

  11. Quinoxalin-2-carboxamides: synthesis and pharmacological evaluation as serotonin type-3 (5-HT3) receptor antagonists.

    PubMed

    Mahesh, Radhakrishnan; Devadoss, Thangaraj; Pandey, Dilip Kumar; Yadav, Shushil Kumar

    2011-10-01

    A series of quinoxalin-2-carboxamides were designed as per the pharmacophoric requirements of 5-HT(3) receptor antagonists and synthesized by condensing the carboxylic group of quinoxalin-2-carboxylic acid with various amines in the presence of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride and 1-hydroxybenzotriazole. The structures of the synthesized compounds were confirmed by physical and spectroscopic data. The carboxamides were evaluated for their 5-HT(3) receptor antagonisms in longitudinal muscle-myenteric plexus preparation from guinea pig ileum against 5-HT(3) agonist, 2-methy-5-HT. All the synthesized compounds showed 5-HT(3) receptor antagonism, (4-benzylpiperazin-1-yl)(quinoxalin-2-yl)methanone was the most potent compound among this series.

  12. Geldanamycin Enhances Retrograde Transport of Shiga Toxin in HEp-2 Cells

    PubMed Central

    Simm, Roger; Torgersen, Maria Lyngaas; Sandvig, Kirsten

    2015-01-01

    The heat shock protein 90 (Hsp90) inhibitor geldanamycin (GA) has been shown to alter endosomal sorting, diverting cargo destined for the recycling pathway into the lysosomal pathway. Here we investigated whether GA also affects the sorting of cargo into the retrograde pathway from endosomes to the Golgi apparatus. As a model cargo we used the bacterial toxin Shiga toxin, which exploits the retrograde pathway as an entry route to the cytosol. Indeed, GA treatment of HEp-2 cells strongly increased the Shiga toxin transport to the Golgi apparatus. The enhanced Golgi transport was not due to increased endocytic uptake of the toxin or perturbed recycling, suggesting that GA selectively enhances endosomal sorting into the retrograde pathway. Moreover, GA activated p38 and both inhibitors of p38 or its substrate MK2 partially counteracted the GA-induced increase in Shiga toxin transport. Thus, our data suggest that GA-induced p38 and MK2 activation participate in the increased Shiga toxin transport to the Golgi apparatus. PMID:26017782

  13. Screening the budding yeast genome reveals unique factors affecting K2 toxin susceptibility.

    PubMed

    Servienė, Elena; Lukša, Juliana; Orentaitė, Irma; Lafontaine, Denis L J; Urbonavičius, Jaunius

    2012-01-01

    Understanding how biotoxins kill cells is of prime importance in biomedicine and the food industry. The budding yeast (S. cerevisiae) killers serve as a convenient model to study the activity of biotoxins consistently supplying with significant insights into the basic mechanisms of virus-host cell interactions and toxin entry into eukaryotic target cells. K1 and K2 toxins are active at the cell wall, leading to the disruption of the plasma membrane and subsequent cell death by ion leakage. K28 toxin is active in the cell nucleus, blocking DNA synthesis and cell cycle progression, thereby triggering apoptosis. Genome-wide screens in the budding yeast S. cerevisiae identified several hundred effectors of K1 and K28 toxins. Surprisingly, no such screen had been performed for K2 toxin, the most frequent killer toxin among industrial budding yeasts. We conducted several concurrent genome-wide screens in S. cerevisiae and identified 332 novel K2 toxin effectors. The effectors involved in K2 resistance and hypersensitivity largely map in distinct cellular pathways, including cell wall and plasma membrane structure/biogenesis and mitochondrial function for K2 resistance, and cell wall stress signaling and ion/pH homeostasis for K2 hypersensitivity. 70% of K2 effectors are different from those involved in K1 or K28 susceptibility. Our work demonstrates that despite the fact that K1 and K2 toxins share some aspects of their killing strategies, they largely rely on different sets of effectors. Since the vast majority of the host factors identified here is exclusively active towards K2, we conclude that cells have acquired a specific K2 toxin effectors set. Our work thus indicates that K1 and K2 have elaborated different biological pathways and provides a first step towards the detailed characterization of K2 mode of action.

  14. Serotonin systems upregulate the expression of hypothalamic NUCB2 via 5-HT2C receptors and induce anorexia via a leptin-independent pathway in mice.

    PubMed

    Nonogaki, Katsunori; Ohba, Yukie; Sumii, Makiko; Oka, Yoshitomo

    2008-07-18

    NEFA/nucleobindin2 (NUCB2), a novel satiety molecule, is associated with leptin-independent melanocortin signaling in the central nervous system. Here, we show that systemic administration of m-chlorophenylpiperazine (mCPP), a serotonin 5-HT1B/2C receptor agonist, significantly increased the expression of hypothalamic NUCB2 in wild-type mice. The increases in hypothalamic NUCB2 expression induced by mCPP were attenuated in 5-HT2C receptor mutant mice. Systemic administration of mCPP suppressed food intake in db/db mice with leptin receptor mutation as well as lean control mice. On the other hand, the expression of hypothalamic NUCB2 and proopiomelanocortin (POMC) was significantly decreased in hyperphagic and non-obese 5-HT2C receptor mutants compared with age-matched wild-type mice. Interestingly, despite increased expression of hypothalamic POMC, hypothalamic NUCB2 expression was decreased in 5-HT2C receptor mutant mice with heterozygous mutation of beta-endorphin gene. These findings suggest that 5-HT systems upregulate the expression of hypothalamic NUCB2 via 5-HT2C receptors, and induce anorexia via a leptin-independent pathway in mice.

  15. Role of the 5-HT2A receptor in the locomotor hyperactivity produced by phenylalkylamine hallucinogens in mice

    PubMed Central

    Halberstadt, Adam L.; Powell, Susan B.; Geyer, Mark A.

    2014-01-01

    The 5-HT2A receptor mediates the effects of serotonergic hallucinogens and may play a role in the pathophysiology of certain psychiatric disorders, including schizophrenia. Given these findings, there is a need for animal models to assess the behavioral effects of 5-HT2A receptor activation. Our previous studies demonstrated that the phenylalkylamine hallucinogen and 5-HT2A/2C agonist 2,5-dimethoxy-4-iodoamphetamine (DOI) produces dose-dependent effects on locomotor activity in C57BL/6J mice, increasing activity at low to moderate doses and reducing activity at high doses. DOI did not increase locomotor activity in 5-HT2A knockout mice, indicating the effect is a consequence of 5-HT2A receptor activation. Here, we tested a series of phenylalkylamine hallucinogens in C57BL/6J mice using the Behavioral Pattern Monitor (BPM) to determine whether these compounds increase locomotor activity by activating the 5-HT2A receptor. Low doses of mescaline, 2,5-dimethoxy-4-ethylamphetamine (DOET), 2,5-dimethoxy-4-propylamphetamine (DOPR), 2,4,5-trimethoxyamphetamine (TMA-2), and the conformationally restricted phenethylamine (4-bromo-3,6-dimethoxybenzocyclobuten-1-yl)methylamine (TCB-2) increased locomotor activity. By contrast, the non-hallucinogenic phenylalkylamine 2,5-dimethoxy-4-tert-butylamphetamine (DOTB) did not alter locomotor activity at any dose tested (0.1-10 mg/kg i.p.). The selective 5-HT2A antagonist M100907 blocked the locomotor hyperactivity induced by mescaline and TCB-2. Similarly, mescaline and TCB-2 did not increase locomotor activity in 5-HT2A knockout mice. These results confirm that phenylalkylamine hallucinogens increase locomotor activity in mice and demonstrate that this effect is mediated by 5-HT2A receptor activation. Thus, locomotor hyperactivity in mice can be used to assess phenylalkylamines for 5-HT2A agonist activity and hallucinogen-like behavioral effects. These studies provide additional support for the link between 5-HT2A activation and

  16. 5-HT2C Receptor Knockdown in the Amygdala Inhibits Neuropathic-Pain-Related Plasticity and Behaviors.

    PubMed

    Ji, Guangchen; Zhang, Wei; Mahimainathan, Lenin; Narasimhan, Madhusudhanan; Kiritoshi, Takaki; Fan, Xiuzhen; Wang, Jigong; Green, Thomas A; Neugebauer, Volker

    2017-02-08

    Neuroplasticity in the amygdala drives pain-related behaviors. The central nucleus (CeA) serves major amygdala output functions and can generate emotional-affective behaviors and modulate nocifensive responses. The CeA receives excitatory and inhibitory inputs from the basolateral nucleus (BLA) and serotonin receptor subtype 5-HT 2C R in the BLA, but not CeA, has been implicated anxiogenic behaviors and anxiety disorders. Here, we tested the hypothesis that 5-HT 2C R in the BLA plays a critical role in CeA plasticity and neuropathic pain behaviors in the rat spinal nerve ligation (SNL) model. Local 5-HT 2C R knockdown in the BLA with stereotaxic injection of 5-HT 2C R shRNA AAV vector decreased vocalizations and anxiety- and depression-like behaviors and increased sensory thresholds of SNL rats, but had no effect in sham controls. Extracellular single-unit recordings of CeA neurons in anesthetized rats showed that 5-HT 2C R knockdown blocked the increase in neuronal activity (increased responsiveness, irregular spike firing, and increased burst activity) in SNL rats. At the synaptic level, 5-HT 2C R knockdown blocked the increase in excitatory transmission from BLA to CeA recorded in brain slices from SNL rats using whole-cell patch-clamp conditions. Inhibitory transmission was decreased by 5-HT 2C R knockdown in control and SNL conditions to a similar degree. The findings can be explained by immunohistochemical data showing increased expression of 5-HT 2C R in non-GABAergic BLA cells in SNL rats. The results suggest that increased 5-HT 2C R in the BLA contributes to neuropathic-pain-related amygdala plasticity by driving synaptic excitation of CeA neurons. As a rescue strategy, 5-HT 2C R knockdown in the BLA inhibits neuropathic-pain-related behaviors. SIGNIFICANCE STATEMENT Neuroplasticity in the amygdala has emerged as an important pain mechanism. This study identifies a novel target and rescue strategy to control abnormally enhanced amygdala activity in an

  17. AHR-16303B, a novel antagonist of 5-HT2 receptors and voltage-sensitive calcium channels

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Barrett, R.J.; Appell, K.C.; Kilpatrick, B.F.

    1991-01-01

    In vivo and in vitro methods were used to characterize AHR-16303B, a novel compound with antagonistic action at 5-HT2 receptors and voltage-sensitive calcium channels. The 5-HT2 receptor-antagonistic properties of AHR-16303B were demonstrated by inhibition of (a) (3H)ketanserin binding to rat cerebral cortical membranes (IC50 = 165 nM); (b) 5-hydroxytryptamine (5-HT)-induced foot edema in rats (minimum effective dose, (MED) = 0.32 mg/kg orally, p.o.); (c) 5-HT-induced vasopressor responses in spontaneously hypertensive rats (SHR) (ID50 = 0.18 mg/kg intravenously (i.v.), 1.8 mg/kg p.o.), (d) 5-HT-induced antidiuresis in rats (MED = 1 mg/kg p.o.), and (e) platelet aggregation induced by 5-HT + ADPmore » (IC50 = 1.5 mM). The calcium antagonist properties of AHR-16303B were demonstrated by inhibition of (a) (3H)nimodipine binding to voltage-sensitive calcium channels on rabbit skeletal muscle membranes (IC50 = 15 nM), (b) KCl-stimulated calcium flux into cultured PC12 cells (IC50 = 81 nM), and (c) CaCl2-induced contractions of rabbit thoracic aortic strips (pA2 = 8.84). AHR-16303B had little or no effect on binding of radioligands to dopamine2 (DA2) alpha 1, alpha 2, H1, 5-HT1 alpha, beta 2, muscarinic M1, or sigma opioid receptors; had no effect on 5-HT3 receptor-mediated vagal bradycardia; and had only minor negative inotropic, chronotropic, and dromotropic effects on isolated guinea pig atria. In conscious SHR, 30 mg/kg p.o. AHR-16303B completely prevented the vasopressor responses to i.v. 5-HT, and decreased blood pressure (BP) by 24% 3 h after dosing.« less

  18. Atypical sympathomimetic drug lerimazoline mediates contractile effects in rat aorta predominantly by 5-HT2A receptors.

    PubMed

    Rizvić, Eldina; Janković, Goran; Kostić-Rajačić, Slađana; Savić, Miroslav M

    2017-08-20

    Lerimazoline is a sympathomimetic drug that belongs to the imidazoline class of compounds, and is used as a nasal decongestant. Studies on lerimazoline are rare, and its pharmacological profile is not completely understood. Here, we analyzed the affinity of lerimazoline for dopamine receptor D2, serotonin 5-HT1A and 5-HT2A receptors and α1-adrenoceptor, and investigated lerimazoline contractile effects in isolated rat thoracic aorta. We also determined the effect of several antagonists on the contractile response to lerimazoline, including prazosin (α1-adrenoceptor antagonist), RX 821002 and rauwolscine (α2-adrenoceptor antagonists), JP 1302 (α2C-adrenoceptor antagonist), methiothepin (non-selective 5-HT receptor antagonist), SB 224289 (5-HT1B receptor antagonist), BRL 15572 (5-HT1D receptor antagonist), and ketanserin (5-HT2A receptor antagonist). Lerimazoline displayed high affinity for the 5-HT1A receptor (Ki = 162.5 nM), similar to the previously reported affinity for the 5-HT1D receptor. Binding affinity estimates (Ki) for α1, 5-HT2A, and D2 receptors were 6656, 4202 and 3437.5 nM, respectively (the literature reported Ki for 5-HT1B receptor is 3480 nM). Lerimazoline caused concentration-dependent contractions in 70% of preparations, varying in the range between 40% and 55% of the maximal contraction elicited by phenylephrine. While prazosin reduced the maximum contractile response to lerimazoline, rauwolscine showed a non-significant trend in reduction of the response. Both ketanserin (10 nM and 1 µM) and methiothepin strongly suppressed the maximum response to lerimazoline. Overall, our results suggest that 5-HT2A and, less distinctly, α1-adrenergic receptors are involved in the lerimazoline-induced contractions, which makes lerimazoline an "atypical" decongestant.

  19. Pasteurella multocida Toxin Manipulates T Cell Differentiation

    PubMed Central

    Hildebrand, Dagmar; Heeg, Klaus; Kubatzky, Katharina F.

    2015-01-01

    Pasteurella multocida causes various diseases in a broad range of wild and domestic animals. Toxigenic strains of the serotypes A and D produce an AB protein toxin named Pasteurella multocida toxin (PMT). PMT constitutively activates the heterotrimeric G protein subunits Gαq, Gα13, and Gαi through deamidation of a glutamine residue, which results in cytoskeletal rearrangements as well as increased proliferation and survival of the host cell. In human monocytes, PMT alters the lipopolysaccharide (LPS)-induced activation toward a phenotype that suppresses T cell activation. Here we describe that the toxin also modulates CD4-positive T helper (Th) cells directly. PMT amplifies the expansion of Th cells through enhanced cell cycle progression and suppression of apoptosis and manipulates the differentiation of Th subclasses through activation of Signal Transducers and Activators of Transcription (STAT) family members and induction of subtype-specific master transcription factors. A large population of toxin-treated T cells is double-positive for Foxp3 and RORγt, the transcription factors expressed by Treg and Th17 cells, respectively. This suggests that these cells could have the potential to turn into Th17 cells or suppressive Treg cells. However, in terms of function, the PMT-differentiated cells behave as inflammatory Th17 cells that produce IL-17 and trigger T cell proliferation. PMID:26635744

  20. Chloroquine derivatives block the translocation pores and inhibit cellular entry of Clostridium botulinum C2 toxin and Bacillus anthracis lethal toxin.

    PubMed

    Kreidler, Anna-Maria; Benz, Roland; Barth, Holger

    2017-03-01

    The pathogenic bacteria Clostridium botulinum and Bacillus anthracis produce the binary protein toxins C2 and lethal toxin (LT), respectively. These toxins consist of a binding/transport (B 7 ) component that delivers the separate enzyme (A) component into the cytosol of target cells where it modifies its specific substrate and causes cell death. The B 7 components of C2 toxin and LT, C2IIa and PA 63 , respectively, are ring-shaped heptamers that bind to their cellular receptors and form complexes with their A components C2I and lethal factor (LF), respectively. After receptor-mediated endocytosis of the toxin complexes, C2IIa and PA 63 insert into the membranes of acidified endosomes and form trans-membrane pores through which C2I and LF translocate across endosomal membranes into the cytosol. C2IIa and PA 63 also form channels in planar bilayer membranes, and we used this approach earlier to identify chloroquine as a potent blocker of C2IIa and PA 63 pores. Here, a series of chloroquine derivatives was investigated to identify more efficient toxin inhibitors with less toxic side effects. Chloroquine, primaquine, quinacrine, and fluphenazine blocked C2IIa and PA 63 pores in planar lipid bilayers and in membranes of living epithelial cells and macrophages, thereby preventing the pH-dependent membrane transport of the A components into the cytosol and protecting cells from intoxication with C2 toxin and LT. These potent inhibitors of toxin entry underline the central role of the translocation pores for cellular uptake of binary bacterial toxins and as relevant drug targets, and might be lead compounds for novel pharmacological strategies against severe enteric diseases and anthrax.

  1. Discovery and structure-activity relationship of (1R)-8-chloro-2,3,4,5-tetrahydro-1-methyl-1H-3-benzazepine (Lorcaserin), a selective serotonin 5-HT2C receptor agonist for the treatment of obesity.

    PubMed

    Smith, Brian M; Smith, Jeffrey M; Tsai, James H; Schultz, Jeffrey A; Gilson, Charles A; Estrada, Scott A; Chen, Rita R; Park, Douglas M; Prieto, Emily B; Gallardo, Charlemagne S; Sengupta, Dipanjan; Dosa, Peter I; Covel, Jon A; Ren, Albert; Webb, Robert R; Beeley, Nigel R A; Martin, Michael; Morgan, Michael; Espitia, Stephen; Saldana, Hazel R; Bjenning, Christina; Whelan, Kevin T; Grottick, Andrew J; Menzaghi, Frederique; Thomsen, William J

    2008-01-24

    The synthesis and SAR of a novel 3-benzazepine series of 5-HT2C agonists is described. Compound 7d (lorcaserin, APD356) was identified as one of the more potent and selective compounds in vitro (pEC50 values in functional assays measuring [(3)H]phosphoinositol turnover: 5-HT2C = 8.1; 5-HT2A = 6.8; 5-HT2B = 6.1) and was potent in an acute in vivo rat food intake model upon oral administration (ED50 at 6 h = 18 mg/kg). Lorcaserin was further characterized in a single-dose pharmacokinetic study in rat (t1/2 = 3.7 h; F = 86%) and a 28-day model of weight gain in growing Sprague-Dawley rat (8.5% decrease in weight gain observed at 36 mg/kg b.i.d.). Lorcaserin was selected for further evaluation in clinical trials for the treatment of obesity.

  2. Repeated 7-Day Treatment with the 5-HT2C Agonist Lorcaserin or the 5-HT2A Antagonist Pimavanserin Alone or in Combination Fails to Reduce Cocaine vs Food Choice in Male Rhesus Monkeys

    PubMed Central

    Banks, Matthew L; Negus, S Stevens

    2017-01-01

    Cocaine use disorder is a global public health problem for which there are no Food and Drug Administration-approved pharmacotherapies. Emerging preclinical evidence has implicated both serotonin (5-HT) 2C and 2A receptors as potential mechanisms for mediating serotonergic attenuation of cocaine abuse-related neurochemical and behavioral effects. Therefore, the present study aim was to determine whether repeated 7-day treatment with the 5-HT2C agonist lorcaserin (0.1–1.0 mg/kg per day, intramuscular; 0.032–0.1 mg/kg/h, intravenous) or the 5-HT2A inverse agonist/antagonist pimavanserin (0.32–10 mg/kg per day, intramuscular) attenuated cocaine reinforcement under a concurrent ‘choice’ schedule of cocaine and food availability in rhesus monkeys. During saline treatment, cocaine maintained a dose-dependent increase in cocaine vs food choice. Repeated pimavanserin (3.2 mg/kg per day) treatments significantly increased small unit cocaine dose choice. Larger lorcaserin (1.0 mg/kg per day and 0.1 mg/kg/h) and pimavanserin (10 mg/kg per day) doses primarily decreased rates of operant behavior. Coadministration of ineffective lorcaserin (0.1 mg/kg per day) and pimavanserin (0.32 mg/kg per day) doses also failed to significantly alter cocaine choice. These results suggest that neither 5-HT2C receptor activation nor 5-HT2A receptor blockade are sufficient to produce a therapeutic-like decrease in cocaine choice and a complementary increase in food choice. Overall, these results do not support the clinical utility of 5-HT2C agonists and 5-HT2A inverse agonists/antagonists alone or in combination as candidate anti-cocaine use disorder pharmacotherapies. PMID:27857126

  3. Repeated 7-Day Treatment with the 5-HT2C Agonist Lorcaserin or the 5-HT2A Antagonist Pimavanserin Alone or in Combination Fails to Reduce Cocaine vs Food Choice in Male Rhesus Monkeys.

    PubMed

    Banks, Matthew L; Negus, S Stevens

    2017-04-01

    Cocaine use disorder is a global public health problem for which there are no Food and Drug Administration-approved pharmacotherapies. Emerging preclinical evidence has implicated both serotonin (5-HT) 2C and 2A receptors as potential mechanisms for mediating serotonergic attenuation of cocaine abuse-related neurochemical and behavioral effects. Therefore, the present study aim was to determine whether repeated 7-day treatment with the 5-HT 2C agonist lorcaserin (0.1-1.0 mg/kg per day, intramuscular; 0.032-0.1 mg/kg/h, intravenous) or the 5-HT 2A inverse agonist/antagonist pimavanserin (0.32-10 mg/kg per day, intramuscular) attenuated cocaine reinforcement under a concurrent 'choice' schedule of cocaine and food availability in rhesus monkeys. During saline treatment, cocaine maintained a dose-dependent increase in cocaine vs food choice. Repeated pimavanserin (3.2 mg/kg per day) treatments significantly increased small unit cocaine dose choice. Larger lorcaserin (1.0 mg/kg per day and 0.1 mg/kg/h) and pimavanserin (10 mg/kg per day) doses primarily decreased rates of operant behavior. Coadministration of ineffective lorcaserin (0.1 mg/kg per day) and pimavanserin (0.32 mg/kg per day) doses also failed to significantly alter cocaine choice. These results suggest that neither 5-HT 2C receptor activation nor 5-HT 2A receptor blockade are sufficient to produce a therapeutic-like decrease in cocaine choice and a complementary increase in food choice. Overall, these results do not support the clinical utility of 5-HT 2C agonists and 5-HT 2A inverse agonists/antagonists alone or in combination as candidate anti-cocaine use disorder pharmacotherapies.

  4. Improved Detection of Vowel Envelope Frequency Following Responses Using Hotelling's T2 Analysis.

    PubMed

    Vanheusden, Frederique J; Bell, Steven L; Chesnaye, Michael A; Simpson, David M

    2018-05-11

    Objective detection of brainstem responses to natural speech stimuli is an important tool for the evaluation of hearing aid fitting, especially in people who may not be able to respond reliably in behavioral tests. Of particular interest is the envelope frequency following response (eFFR), which refers to the EEG response at the stimulus' fundamental frequency (and its harmonics), and here in particular to the response to natural spoken vowel sounds. This article introduces the frequency-domain Hotelling's T (HT2) method for eFFR detection. This method was compared, in terms of sensitivity in detecting eFFRs at the fundamental frequency (HT2_F0), to two different single-channel frequency domain methods (F test on Fourier analyzer (FA) amplitude spectra [FA-F-Test] and magnitude-squared coherence [MSC]) in detecting envelope following responses to natural vowel stimuli in simulated data and EEG data from normal-hearing subjects. Sensitivity was assessed based on the number of detections and the time needed to detect a response for a false-positive rate of 5%. The study also explored whether a single-channel, multifrequency HT2 (HT2_3F) and a multichannel, multifrequency HT2 (HT2_MC) could further improve response detection. Four repeated words were presented sequentially at 70 dB SPL LAeq through ER-2 insert earphones. The stimuli consisted of a prolonged vowel in a /hVd/ structure (where V represents different vowel sounds). Each stimulus was presented over 440 sweeps (220 condensation and 220 rarefaction). EEG data were collected from 12 normal-hearing adult participants. After preprocessing and artifact removal, eFFR detection was compared between the algorithms. For the simulation study, simulated EEG signals were generated by adding random noise at multiple signal to noise ratios (SNRs; 0 to -60dB) to the auditory stimuli as well as to a single sinusoid at the fluctuating and flattened fundamental frequency (f0). For each SNR, 1000 sets of 440 simulated epochs

  5. Molecular interactions of agonist and inverse agonist ligands at serotonin 5-HT2C G protein-coupled receptors: computational ligand docking and molecular dynamics studies validated by experimental mutagenesis results

    NASA Astrophysics Data System (ADS)

    Córdova-Sintjago, Tania C.; Liu, Yue; Booth, Raymond G.

    2015-02-01

    To understand molecular determinants for ligand activation of the serotonin 5-HT2C G protein-coupled receptor (GPCR), a drug target for obesity and neuropsychiatric disorders, a 5-HT2C homology model was built according to an adrenergic β2 GPCR (β2AR) structure and validated using a 5-HT2B GPCR crystal structure. The models were equilibrated in a simulated phosphatidyl choline membrane for ligand docking and molecular dynamics studies. Ligands included (2S, 4R)-(-)-trans-4-(3'-bromo- and trifluoro-phenyl)-N,N-dimethyl-1,2,3,4-tetrahydronaphthalene-2-amine (3'-Br-PAT and 3'-CF3-PAT), a 5-HT2C agonist and inverse agonist, respectively. Distinct interactions of 3'-Br-PAT and 3'-CF3-PAT at the wild-type (WT) 5-HT2C receptor model were observed and experimental 5-HT2C receptor mutagenesis studies were undertaken to validate the modelling results. For example, the inverse agonist 3'-CF3-PAT docked deeper in the WT 5-HT2C binding pocket and altered the orientation of transmembrane helices (TM) 6 in comparison to the agonist 3'-Br-PAT, suggesting that changes in TM orientation that result from ligand binding impact function. For both PATs, mutation of 5-HT2C residues S3.36, T3.37, and F5.47 to alanine resulted in significantly decreased affinity, as predicted from modelling results. It was concluded that upon PAT binding, 5-HT2C residues T3.37 and F5.47 in TMs 3 and 5, respectively, engage in inter-helical interactions with TMs 4 and 6, respectively. The movement of TMs 5 and 6 upon agonist and inverse agonist ligand binding observed in the 5-HT2C receptor modelling studies was similar to movements reported for the activation and deactivation of the β2AR, suggesting common mechanisms among aminergic neurotransmitter GPCRs.

  6. Disease-specific expression of the serotonin-receptor 5-HT(2C) in natural killer cells in Alzheimer's dementia.

    PubMed

    Martins, Luiza Conceição Amorim; Rocha, Natália Pessoa; Torres, Karen Cecília Lima; Dos Santos, Rodrigo Ribeiro; França, Giselle Sabrina; de Moraes, Edgar Nunes; Mukhamedyarov, Marat Alexandrovich; Zefirov, Andrey Lvovich; Rizvanov, Albert Anatolyevich; Kiyasov, Andrey Pavlovich; Vieira, Luciene Bruno; Guimarães, Melissa Monteiro; Yalvaç, Mehmet Emir; Teixeira, Antônio Lúcio; Bicalho, Maria Aparecida Camargo; Janka, Zoltán; Romano-Silva, Marco Aurélio; Palotás, András; Reis, Helton José

    2012-10-15

    Alzheimer's dementia (AD) is a degenerative brain disorder characterized mainly by cholinergic failure, but other neuro-transmitters are also deficient especially at late stages of the disease. Misfolded β-amyloid peptide has been identified as a causative agent, however inflammatory changes also play a pivotal role. Even though the most prominent pathology is seen in the cognitive functions, specific abnormalities of the central nervous system (CNS) are also reflected in the periphery, particularly in the immune responses of the body. The aim of this study was to characterize the dopaminergic and serotonergic systems in AD, which are also markedly disrupted along with the hallmark acetyl-choline dysfunction. Peripheral blood mono-nuclear cells (PBMCs) from demented patients were judged against comparison groups including individuals with late-onset depression (LOD), as well as non-demented and non-depressed subjects. Cellular sub-populations were evaluated by mono-clonal antibodies against various cell surface receptors: CD4/CD8 (T-lymphocytes), CD19 (B-lymphocytes), CD14 (monocytes), and CD56 (natural-killer (NK)-cells). The expressions of dopamine D(3) and D(4), as well as serotonin 5-HT(1A), 5-HT(2A), 5-HT(2B) and 5-HT(2C) were also assessed. There were no significant differences among the study groups with respect to the frequency of the cellular sub-types, however a unique profound increase in 5-HT(2C) receptor exclusively in NK-cells was observed in AD. The disease-specific expression of 5-HT(2C), as well as the NK-cell cyto-toxicity, has been linked with cognitive derangement in dementia. These changes not only corroborate the existence of bi-directional communication between the immune system and the CNS, but also elucidate the role of inflammatory activity in AD pathology, and may serve as potential biomarkers for less invasive and early diagnostic purposes as well. Copyright © 2012 Elsevier B.V. All rights reserved.

  7. Screening the Budding Yeast Genome Reveals Unique Factors Affecting K2 Toxin Susceptibility

    PubMed Central

    Servienė, Elena; Lukša, Juliana; Orentaitė, Irma

    2012-01-01

    Background Understanding how biotoxins kill cells is of prime importance in biomedicine and the food industry. The budding yeast (S. cerevisiae) killers serve as a convenient model to study the activity of biotoxins consistently supplying with significant insights into the basic mechanisms of virus-host cell interactions and toxin entry into eukaryotic target cells. K1 and K2 toxins are active at the cell wall, leading to the disruption of the plasma membrane and subsequent cell death by ion leakage. K28 toxin is active in the cell nucleus, blocking DNA synthesis and cell cycle progression, thereby triggering apoptosis. Genome-wide screens in the budding yeast S. cerevisiae identified several hundred effectors of K1 and K28 toxins. Surprisingly, no such screen had been performed for K2 toxin, the most frequent killer toxin among industrial budding yeasts. Principal Findings We conducted several concurrent genome-wide screens in S. cerevisiae and identified 332 novel K2 toxin effectors. The effectors involved in K2 resistance and hypersensitivity largely map in distinct cellular pathways, including cell wall and plasma membrane structure/biogenesis and mitochondrial function for K2 resistance, and cell wall stress signaling and ion/pH homeostasis for K2 hypersensitivity. 70% of K2 effectors are different from those involved in K1 or K28 susceptibility. Significance Our work demonstrates that despite the fact that K1 and K2 toxins share some aspects of their killing strategies, they largely rely on different sets of effectors. Since the vast majority of the host factors identified here is exclusively active towards K2, we conclude that cells have acquired a specific K2 toxin effectors set. Our work thus indicates that K1 and K2 have elaborated different biological pathways and provides a first step towards the detailed characterization of K2 mode of action. PMID:23227207

  8. 5-HT2A receptor antagonists improve motor impairments in the MPTP mouse model of Parkinson's disease.

    PubMed

    Ferguson, Marcus C; Nayyar, Tultul; Deutch, Ariel Y; Ansah, Twum A

    2010-01-01

    Clinical observations have suggested that ritanserin, a 5-HT(2A/C) receptor antagonist may reduce motor deficits in persons with Parkinson's Disease (PD). To better understand the potential antiparkinsonian actions of ritanserin, we compared the effects of ritanserin with the selective 5-HT(2A) receptor antagonist M100907 and the selective 5-HT(2C) receptor antagonist SB 206553 on motor impairments in mice treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). MPTP-treated mice exhibited decreased performance on the beam-walking apparatus. These motor deficits were reversed by acute treatment with L-3,4-dihydroxyphenylalanine (levodopa). Both the mixed 5-HT(2A/C) antagonist ritanserin and the selective 5-HT(2A) antagonist M100907 improved motor performance on the beam-walking apparatus. In contrast, SB 206553 was ineffective in improving the motor deficits in MPTP-treated mice. These data suggest that 5-HT(2A) receptor antagonists may represent a novel approach to ameliorate motor symptoms of Parkinson's disease. Published by Elsevier Ltd.

  9. 5-HT2A receptor antagonists improve motor impairments in the MPTP mouse model of Parkinson's disease

    PubMed Central

    Ferguson, Marcus C.; Nayyar, Tultul; Deutch, Ariel Y.; Ansah, Twum A.

    2010-01-01

    Clinical observations have suggested that ritanserin, a 5-HT2A/C receptor antagonist may reduce motor deficits in persons with Parkinson's Disease (PD). To better understand the potential antiparkinsonian actions of ritanserin, we compared the effects of ritanserin with the selective 5-HT2A receptor antagonist M100907 and the selective 5-HT2C receptor antagonist SB 206553 on motor impairments in mice treated with 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). MPTP-treated mice exhibited decreased performance on the beam-walking apparatus. These motor deficits were reversed by acute treatment with L-3,4-dihydroxyphenylalanine (levodopa). Both the mixed 5-HT2A/C antagonist ritanserin and the selective 5-HT2A antagonist M100907 improved motor performance on the beam-walking apparatus. In contrast, SB 206553 was ineffective in improving the motor deficits in MPTP-treated mice. These data suggest that 5-HT2A receptor antagonists may represent a novel approach to ameliorate motor symptoms of Parkinson's disease. PMID:20361986

  10. Increased alternate splicing of Htr2c in a mouse model for Prader-Willi syndrome leads disruption of 5HT2C receptor mediated appetite.

    PubMed

    Garfield, Alastair S; Davies, Jennifer R; Burke, Luke K; Furby, Hannah V; Wilkinson, Lawrence S; Heisler, Lora K; Isles, Anthony R

    2016-12-08

    Alternate splicing of serotonin (5-hydroxytryptamine; 5-HT) 2C receptor (5-HT 2C R) pre-RNA is negatively regulated by the small nucleolar RNA, Snord115, loss of which is observed in nearly all individuals with Prader-Willi Syndrome (PWS), a multigenic disorder characterised by hyperphagia and obesity. Given the role of the 5-HT 2C R in the regulation of ingestive behaviour we investigated the pathophysiological implications of Snord115 deficiency on 5-HT 2C R regulated appetite in a genotypically relevant PWS mouse model (PWS-IC). Specifically, we demonstrate that loss of Snord115 expression is associated with increased levels of hypothalamic truncated 5-HT 2C R pre-mRNA. The 5-HT 2C R promotes appetite suppression via engagement of the central melanocortin system. Pro-opiomelancortin (Pomc) mRNA levels within the arcuate nucleus of the hypothalamus (ARC) were reduced in PWS-IC mice. We then went on to assess the functional consequences of these molecular changes, demonstrating that PWS-IC mice are unresponsive to an anorectic doses of a 5-HT 2C R agonist and that this is associated with attenuated activation of POMC neurons within the ARC. These data provide new insight into the significance of Htr2c pre-mRNA processing to the physiological regulation of appetite and potentially the pathological manifestation of hyperphagia in PWS. Furthermore, these findings have translational relevance for individuals with PWS who may seek to control appetite with another 5-HT 2C R agonist, the new obesity treatment lorcaserin.

  11. 5HT2A receptor blockade in dorsomedial striatum reduces repetitive behaviors in BTBR mice.

    PubMed

    Amodeo, D A; Rivera, E; Cook, E H; Sweeney, J A; Ragozzino, M E

    2017-03-01

    Restricted and repetitive behaviors are a defining feature of autism, which can be expressed as a cognitive flexibility deficit or stereotyped, motor behaviors. There is limited knowledge about the underlying neuropathophysiology contributing to these behaviors. Previous findings suggest that central 5HT 2A receptor activity is altered in autism, while recent work indicates that systemic 5HT 2A receptor antagonist treatment reduces repetitive behaviors in an idiopathic model of autism. 5HT 2A receptors are expressed in the orbitofrontal cortex and striatum. These two regions have been shown to be altered in autism. The present study investigated whether 5HT 2A receptor blockade in the dorsomedial striatum or orbitofrontal cortex in the BTBR mouse strain, an idiopathic model of autism, affects the phenotype related to restricted and repetitive behaviors. Microinfusion of the 5HT 2A receptor antagonist, M100907 into the dorsomedial striatum alleviated a reversal learning impairment and attenuated grooming behavior. M100907 infusion into the orbitofrontal cortex increased perseveration during reversal learning and potentiated grooming. These findings suggest that increased 5HT 2A receptor activity in the dorsomedial striatum may contribute to behavioral inflexibility and stereotyped behaviors in the BTBR mouse. 5HT 2A receptor signaling in the orbitofrontal cortex may be critical for inhibiting a previously learned response during reversal learning and expression of stereotyped behavior. The present results suggest which brain areas exhibit abnormalities underlying repetitive behaviors in an idiopathic mouse model of autism, as well as which brain areas systemic treatment with M100907 may principally act on in BTBR mice to attenuate repetitive behaviors. © 2016 John Wiley & Sons Ltd and International Behavioural and Neural Genetics Society.

  12. Cellular uptake of Clostridium botulinum C2 toxin: membrane translocation of a fusion toxin requires unfolding of its dihydrofolate reductase domain.

    PubMed

    Haug, Gerd; Wilde, Christian; Leemhuis, Jost; Meyer, Dieter K; Aktories, Klaus; Barth, Holger

    2003-12-30

    The Clostridium botulinum C2 toxin is the prototype of the family of binary actin-ADP-ribosylating toxins. C2 toxin is composed of two separated nonlinked proteins. The enzyme component C2I ADP-ribosylates actin in the cytosol of target cells. The binding/translocation component C2II mediates cell binding of the enzyme component and its translocation from acidic endosomes into the cytosol. After proteolytic activation, C2II forms heptameric pores in endosomal membranes, and most likely, C2I translocates through these pores into the cytosol. For this step, the cellular heat shock protein Hsp90 is essential. We analyzed the effect of methotrexate on the cellular uptake of a fusion toxin in which the enzyme dihydrofolate reductase (DHFR) was fused to the C-terminus of C2I. Here, we report that unfolding of C2I-DHFR is required for cellular uptake of the toxin via the C2IIa component. The C2I-DHFR fusion toxin catalyzed ADP-ribosylation of actin in vitro and was able to intoxicate cultured cells when applied together with C2IIa. Binding of the folate analogue methotrexate favors a stable three-dimensional structure of the dihydrofolate reductase domain. Pretreatment of C2I-DHFR with methotrexate prevented cleavage of C2I-DHFR by trypsin. In the presence of methotrexate, intoxication of cells with C2I-DHFR/C2II was inhibited. The presence of methotrexate diminished the translocation of the C2I-DHFR fusion toxin from endosomal compartments into the cytosol and the direct C2IIa-mediated translocation of C2I-DHFR across cell membranes. Methotrexate had no influence on the intoxication of cells with C2I/C2IIa and did not alter the C2IIa-mediated binding of C2I-DHFR to cells. The data indicate that methotrexate prevented unfolding of the C2I-DHFR fusion toxin, and thereby the translocation of methotrexate-bound C2I-DHFR from endosomes into the cytosol of target cells is inhibited.

  13. Attenuation of nicotine's discriminative stimulus effects in rats and its locomotor activity effects in mice by serotonergic 5-HT2A/2C receptor agonists.

    PubMed

    Batman, Angela M; Munzar, Patrik; Beardsley, Patrick M

    2005-05-01

    Reports have indicated that administration of nicotine inhibits, while withdrawal of chronically administered nicotine augments effects of serotonergic 5HT2A/2C agonists. It was our objective to determine whether 5HT2A/2C agonists can modulate the discriminative stimulus effects of nicotine in rats or its locomotor activity effects in mice. Adult male Sprague-Dawley rats were trained to discriminate 0.3 mg/kg nicotine base from saline in a two-lever, fixed-ratio (FR10), food-reinforced, operant-conditioning task during daily (Monday-Friday) 15-min experimental sessions. After characterizing a dose-response curve for nicotine, we tested the ability of the 5HT(2A/2C) agonists (+/-)-1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane HCL (DOI; 0.18-1.0 mg/kg) and 1-(4-bromo-2, 5-dimethoxyphenyl)-2-aminopropane (DOB; 0.1-1.0 mg/kg), the 5HT2C agonist 6-chloro-2-(1-piperazinyl)pyrazine hydrochloride (MK 212; 0.1 mg/kg-1.0 mg/kg), and the 5HT1A agonist (+/-)-8-hydroxy-2-(di-n-propylamino)tetralin hydrobromide (8-OH-DPAT; 0.01 mg/kg-1.0 mg/kg) to modulate nicotine's discriminative stimulus effects. After finding that DOI was able to attenuate the percentage nicotine lever responding (%NLR), we tested for it to also reverse nicotine's effects on locomotor activity in mice. The 5HT2A/2C agonists-in particular DOI-dose dependently attenuated %NLR. The effects of DOI were reversed by the 5HT2A/2C antagonist ketanserin. MK 212 and 8-OH-DPAT had irregular effects among rats and only reduced %NLR to below 50% levels at doses markedly suppressing responding. DOI also dose dependently blocked nicotine's acute rate-lowering locomotor activity effects. These results indicate that activation of serotonin 5HT2A/2C receptors can blunt the discriminative stimulus and locomotor activity effects of nicotine and presents the possibility that activation of these receptors might also be able to attenuate other effects of nicotine.

  14. Retargeting the Clostridium botulinum C2 toxin to the neuronal cytosol.

    PubMed

    Pavlik, Benjamin J; Hruska, Elizabeth J; Van Cott, Kevin E; Blum, Paul H

    2016-03-30

    Many biological toxins are known to attack specific cell types, delivering their enzymatic payloads to the cytosol. This process can be manipulated by molecular engineering of chimeric toxins. Using toxins with naturally unlinked components as a starting point is advantageous because it allows for the development of payloads separately from the binding/translocation components. Here the Clostridium botulinum C2 binding/translocation domain was retargeted to neural cell populations by deleting its non-specific binding domain and replacing it with a C. botulinum neurotoxin binding domain. This fusion protein was used to deliver fluorescently labeled payloads to Neuro-2a cells. Intracellular delivery was quantified by flow cytometry and found to be dependent on artificial enrichment of cells with the polysialoganglioside receptor GT1b. Visualization by confocal microscopy showed a dissociation of payloads from the early endosome indicating translocation of the chimeric toxin. The natural Clostridium botulinum C2 toxin was then delivered to human glioblastoma A172 and synchronized HeLa cells. In the presence of the fusion protein, native cytosolic enzymatic activity of the enzyme was observed and found to be GT1b-dependent. This retargeted toxin may enable delivery of therapeutics to peripheral neurons and be of use in addressing experimental questions about neural physiology.

  15. Retargeting the Clostridium botulinum C2 toxin to the neuronal cytosol

    PubMed Central

    Pavlik, Benjamin J.; Hruska, Elizabeth J.; Van Cott, Kevin E.; Blum, Paul H.

    2016-01-01

    Many biological toxins are known to attack specific cell types, delivering their enzymatic payloads to the cytosol. This process can be manipulated by molecular engineering of chimeric toxins. Using toxins with naturally unlinked components as a starting point is advantageous because it allows for the development of payloads separately from the binding/translocation components. Here the Clostridium botulinum C2 binding/translocation domain was retargeted to neural cell populations by deleting its non-specific binding domain and replacing it with a C. botulinum neurotoxin binding domain. This fusion protein was used to deliver fluorescently labeled payloads to Neuro-2a cells. Intracellular delivery was quantified by flow cytometry and found to be dependent on artificial enrichment of cells with the polysialoganglioside receptor GT1b. Visualization by confocal microscopy showed a dissociation of payloads from the early endosome indicating translocation of the chimeric toxin. The natural Clostridium botulinum C2 toxin was then delivered to human glioblastoma A172 and synchronized HeLa cells. In the presence of the fusion protein, native cytosolic enzymatic activity of the enzyme was observed and found to be GT1b-dependent. This retargeted toxin may enable delivery of therapeutics to peripheral neurons and be of use in addressing experimental questions about neural physiology. PMID:27025362

  16. T-2 mycotoxin treatment of newborn rat pups does not significantly affect nervous system functions in adulthood.

    PubMed

    Varró, Petra; Béldi, Melinda; Kovács, Melinda; Világi, Ildikó

    2018-03-01

    T-2 toxin is primarily produced by Fusarium sp. abundant under temperate climatic conditions. Its main harmful effect is the inhibition of protein synthesis. Causing oxidative stress, it also promotes lipid peroxidation and changes plasma membrane phospholipid composition; this may lead to nervous system alterations. The aim of the present study was to examine whether a single dose of T-2 toxin administered at newborn age has any long-lasting effects on nervous system functions. Rat pups were treated on the first postnatal day with a single intraperitoneal dose of T-2 toxin (0.2 mg/bwkg). Body weight of treated pups was lower during the second and third week of life, compared to littermates; later, weight gain was recovered. At young adulthood, behavior was tested in the open field, and no difference was observed between treated and control rats. Field potential recordings from somatosensory cortex and hippocampus slices did not reveal any significant difference in neuronal network functions. In case of neocortical field EPSP, the shape was slightly different in treated pups. Long-term synaptic plasticity was also comparable in both groups. Seizure susceptibility of the slices was not different, either. In conclusion, T-2 toxin did not significantly affect basic nervous system functions at this dose.

  17. Molecular interactions between general anesthetics and the 5HT2B receptor.

    PubMed

    Matsunaga, Felipe; Gao, Lu; Huang, Xi-Ping; Saven, Jeffery G; Roth, Bryan L; Liu, Renyu

    2015-01-01

    Serotonin modulates many processes through a family of seven serotonin receptors. However, no studies have screened for interactions between general anesthetics currently in clinical use and serotonergic G-protein-coupled receptors (GPCRs). Given that both intravenous and inhalational anesthetics have been shown to target other classes of GPCRs, we hypothesized that general anesthetics might interact directly with some serotonin receptors and thus modify their function. Radioligand binding assays were performed to screen serotonin receptors for interactions with propofol and isoflurane as well as for affinity determinations. Docking calculations using the crystal structure of 5-HT2B were performed to computationally confirm the binding assay results and locate anesthetic binding sites. The 5-HT2B class of receptors interacted significantly with both propofol and isoflurane in the primary screen. The affinities for isoflurane and propofol were determined to be 7.78 and .95 μM, respectively, which were at or below the clinical concentrations for both anesthetics. The estimated free energy derived from docking calculations for propofol (-6.70 kcal/mol) and isoflurane (-5.10 kcal/mol) correlated with affinities from the binding assay. The anesthetics were predicted to dock at a pharmacologically relevant binding site of 5HT2B. The molecular interactions between propofol and isoflurane with the 5-HT2B class of receptors were discovered and characterized. This finding implicates the serotonergic GPCRs as potential anesthetic targets.

  18. A short history of the 5-HT2C receptor: from the choroid plexus to depression, obesity and addiction treatment.

    PubMed

    Palacios, Jose M; Pazos, Angel; Hoyer, Daniel

    2017-05-01

    This paper is a personal account on the discovery and characterization of the 5-HT 2C receptor (first known as the 5-HT 1C receptor) over 30 years ago and how it translated into a number of unsuspected features for a G protein-coupled receptor (GPCR) and a diversity of clinical applications. The 5-HT 2C receptor is one of the most intriguing members of the GPCR superfamily. Initially referred to as 5-HT 1C R, the 5-HT 2C R was discovered while studying the pharmacological features and the distribution of [ 3 H]mesulergine-labelled sites, primarily in the brain using radioligand binding and slice autoradiography. Mesulergine (SDZ CU-085), was, at the time, best defined as a ligand with serotonergic and dopaminergic properties. Autoradiographic studies showed remarkably strong [ 3 H]mesulergine-labelling to the rat choroid plexus. [ 3 H]mesulergine-labelled sites had pharmacological properties different from, at the time, known or purported 5-HT receptors. In spite of similarities with 5-HT 2 binding, the new binding site was called 5-HT 1C because of its very high affinity for 5-HT itself. Within the following 10 years, the 5-HT 1C R (later named 5-HT 2C ) was extensively characterised pharmacologically, anatomically and functionally: it was one of the first 5-HT receptors to be sequenced and cloned. The 5-HT 2C R is a GPCR, with a very complex gene structure. It constitutes a rarity in the GPCR family: many 5-HT 2C R variants exist, especially in humans, due to RNA editing, in addition to a few 5-HT 2C R splice variants. Intense research led to therapeutically active 5-HT 2C receptor ligands, both antagonists (or inverse agonists) and agonists: keeping in mind that a number of antidepressants and antipsychotics are 5-HT 2C R antagonists/inverse agonists. Agomelatine, a 5-HT 2C R antagonist is registered for the treatment of major depression. The agonist Lorcaserin is registered for the treatment of aspects of obesity and has further potential in addiction

  19. The serotonergic hallucinogen 5-methoxy-N,N-dimethyltryptamine disrupts cortical activity in a regionally-selective manner via 5-HT(1A) and 5-HT(2A) receptors.

    PubMed

    Riga, Maurizio S; Bortolozzi, Analia; Campa, Letizia; Artigas, Francesc; Celada, Pau

    2016-02-01

    5-Methoxy-N,N-dimethyltryptamine (5-MeO-DMT) is a natural hallucinogen, acting as a non-selective serotonin 5-HT(1A)/5-HT(2A)-R agonist. Psychotomimetic agents such as the non-competitive NMDA-R antagonist phencyclidine and serotonergic hallucinogens (DOI and 5-MeO-DMT) disrupt cortical synchrony in the low frequency range (<4 Hz) in rat prefrontal cortex (PFC), an effect reversed by antipsychotic drugs. Here we extend these observations by examining the effect of 5-MeO-DMT on low frequency cortical oscillations (LFCO, <4 Hz) in PFC, visual (V1), somatosensory (S1) and auditory (Au1) cortices, as well as the dependence of these effects on 5-HT(1A)-R and 5-HT(2A)-R, using wild type (WT) and 5-HT(2A)-R knockout (KO2A) anesthetized mice. 5-MeO-DMT reduced LFCO in the PFC of WT and KO2A mice. The effect in KO2A mice was fully prevented by the 5-HT(1A)-R antagonist WAY-100635. Systemic and local 5-MeO-DMT reduced 5-HT release in PFC mainly via 5-HT(1A)-R. Moreover, 5-MeO-DMT reduced LFCO in S1, Au1 and V1 of WT mice and only in V1 of KO2A mice, suggesting the involvement of 5-HT(1A)-R activation in the 5-MeO-DMT-induced disruption of V1 activity. In addition, antipsychotic drugs reversed 5-MeO-DMT effects in WT mice. The present results suggest that the hallucinogen action of 5-MeO-DMT is mediated by simultaneous alterations of the activity of sensory (S1, Au1, V1) and associative (PFC) cortical areas, also supporting a role of 5-HT(1A)-R stimulation in V1 and PFC, in addition to the well-known action on 5-HT(2A)-R. Moreover, the reversal by antipsychotic drugs of 5-MeO-DMT effects adds to previous literature supporting the usefulness of the present model in antipsychotic drug development. Copyright © 2015 Elsevier Ltd. All rights reserved.

  20. Effect of Emetine on T-2 Toxin-Induced Inhibition of Protein Synthesis in Mammalian Cells

    DTIC Science & Technology

    1993-01-01

    manuscript. 748 Leatheman and Mlddlbrook Vol. 266 References toxins in Human and Animal Health , ed. by J. V. Rodericks, C. W. Hesaeltine ANTONm, F., HRABAK...20: 160-163, 1973. in Human and Animal Health , ed. by J. V. Rodericks, C. W. Hesseltine and FEUERSTEIN, G., POWELL, J. A., KNOWER, A. T. AND HUNTER, K... Animal Health , ed. by J. V. Rodericks, C. W. Mechanism of action of the mycotoxin trichodermin, a 12,13 epoxytrichothe. Hesseltine and M. A. Mehlman, pp

  1. Stimulation of 5-HT2C Receptors Improves Cognitive Deficits Induced by Human Tryptophan Hydroxylase 2 Loss of Function Mutation

    PubMed Central

    Del'Guidice, Thomas; Lemay, Francis; Lemasson, Morgane; Levasseur-Moreau, Jean; Manta, Stella; Etievant, Adeline; Escoffier, Guy; Doré, François Y; Roman, François S; Beaulieu, Jean-Martin

    2014-01-01

    Polymorphisms in the gene encoding the serotonin synthesis enzyme Tph2 have been identified in mental illnesses, including bipolar disorder, major depression, autism, schizophrenia, and ADHD. Deficits in cognitive flexibility and perseverative behaviors are shared common symptoms in these disorders. However, little is known about the impact of Tph2 gene variants on cognition. Mice expressing a human TPH2 variant (Tph2-KI) were used to investigate cognitive consequences of TPH2 loss of function and pharmacological treatments. We applied a recently developed behavioral assay, the automated H-maze, to study cognitive functions in Tph2-KI mice. This assay involves the consecutive discovery of three different rules: a delayed alternation task, a non-alternation task, and a delayed reversal task. Possible contribution of locomotion, reward, and sensory perception were also investigated. The expression of loss-of-function mutant Tph2 in mice was associated with impairments in reversal learning and cognitive flexibility, accompanied by perseverative behaviors similar to those observed in human clinical studies. Pharmacological restoration of 5-HT synthesis with 5-hydroxytryptophan or treatment with the 5-HT2C receptor agonist CP809.101 reduced cognitive deficits in Tph2-KI mice and abolished perseveration. In contrast, treatment with the psychostimulant methylphenidate exacerbated cognitive deficits in mutant mice. Results from this study suggest a contribution of TPH2 in the regulation of cognition. Furthermore, identification of a role for a 5-HT2 receptor agonist as a cognition-enhancing agent in mutant mice suggests a potential avenue to explore for the personalized treatment of cognitive symptoms in humans with reduced 5-HT synthesis and TPH2 polymorphisms. PMID:24196946

  2. Detection of T-2 Toxin by a Modified Radioimmunoassay.

    DTIC Science & Technology

    1982-08-06

    Mehlman (ed.), Mycotoxins in human and animal health . Pathotox Publishers Inc., Park Forest South, Ill. 2. Lutsky, I, N. Mor, B. Yagen, and A. Z. Joffe...human and animal health . Pathotox Publishers Inc., Park Forest South, Il. 5. Wallace, E.M., S. V. Pathre, C. J. Mirocha, T. S. Robinson, and S. W

  3. Shiga toxin type 2 (Stx2), a potential agent of bioterrorism, has a short distribution and a long elimination half-life, and induces kidney and thymus lesions in rats.

    PubMed

    Liu, Yue-Nan; Wang, Sheng-Han; Li, Tao; Wang, Qin; Tu, Wei; Cai, Kun; Hou, Xiao-Jun; Tian, Ren-Mao; Gao, Xiang; Liu, Hao; Xiao, Le; Shi, Jing; Cheng, Yuan-Guo; Li, Jian-Chun; Wang, Hui

    2011-09-01

    Shiga toxin type 2, a major virulence factor produced by the Shiga toxin-producing Escherichia coli, is a potential toxin agent of bioterrorism. In this study, iodine-125 (125I) was used as an indicator to describe the in vivo Stx2 biodistribution profile. The rats were injected intravenously (i.v.) with 125I-Stx2 at three doses of 5.1-127.5 μg/kg body weight. Stx2 had a short distribution half-life (t (1/2)α, less than 6 min) and a long elimination half-life in rat. The toxicokinetics of Stx2 in rats was dose dependent and nonlinear. Stx2 concentrations in various tissues were detected at 5-min, 0.5-h, and 72-h postinjection. High radioactivity was found in the lungs, kidneys, nasal turbinates, and sometimes in the eyes, which has never been reported in previous studies. In a preliminary assessment, lesions were found in the kidney and thymus.

  4. Molecular Evolutionary Constraints that Determine the Avirulence State of Clostridium botulinum C2 Toxin.

    PubMed

    Prisilla, A; Prathiviraj, R; Chellapandi, P

    2017-04-01

    Clostridium botulinum (group-III) is an anaerobic bacterium producing C2 toxin along with botulinum neurotoxins. C2 toxin is belonged to binary toxin A family in bacterial ADP-ribosylation superfamily. A structural and functional diversity of binary toxin A family was inferred from different evolutionary constraints to determine the avirulence state of C2 toxin. Evolutionary genetic analyses revealed evidence of C2 toxin cluster evolution through horizontal gene transfer from the phage or plasmid origins, site-specific insertion by gene divergence, and homologous recombination event. It has also described that residue in conserved NAD-binding core, family-specific domain structure, and functional motifs found to predetermine its virulence state. Any mutational changes in these residues destabilized its structure-function relationship. Avirulent mutants of C2 toxin were screened and selected from a crucial site required for catalytic function of C2I and pore-forming function of C2II. We found coevolved amino acid pairs contributing an essential role in stabilization of its local structural environment. Avirulent toxins selected in this study were evaluated by detecting evolutionary constraints in stability of protein backbone structure, folding and conformational dynamic space, and antigenic peptides. We found 4 avirulent mutants of C2I and 5 mutants of C2II showing more stability in their local structural environment and backbone structure with rapid fold rate, and low conformational flexibility at mutated sites. Since, evolutionary constraints-free mutants with lack of catalytic and pore-forming function suggested as potential immunogenic candidates for treating C. botulinum infected poultry and veterinary animals. Single amino acid substitution in C2 toxin thus provides a major importance to understand its structure-function link, not only of a molecule but also of the pathogenesis.

  5. Interaction of cruciferin-based nanoparticles with Caco-2 cells and Caco-2/HT29-MTX co-cultures.

    PubMed

    Akbari, Ali; Lavasanifar, Afsaneh; Wu, Jianping

    2017-12-01

    The objective of this work was to assess the potential of Cruciferin/Calcium (Cru/Ca) and Cruciferin/Chitosan (Cru/Cs) nanoparticles for oral drug delivery. For this purpose, Cru/Ca and Cru/Cs nanoparticles were developed through cold gelation of Cruciferin, a major canola protein, and in interaction with calcium and chitosan, respectively. The extent and rate of particle uptake in Caco-2 cells and Caco-2/HT29 co-culture was then evaluated by fluorescence spectroscopy as well as flow cytometry. Through pre-incubation of Caco-2 cell monolayer with specific endocytosis inhibitors, the mechanism of cell uptake was investigated. Our results showed that the uptake of negatively-charged Cru/Ca particles to be ∼3 times higher than positively-charged Cru/Cs ones by Caco-2 cells. Presence of mucus secreted by HT29 cells in their co-culture with Caco-2 had negligible influence on the uptake and transport of both particles. In contrast to Cru/Ca particles which were dissociated in the simulated gastrointestinal conditions, digestion of Cru/Cs particles resulted in 6- and 2-fold increase in the cellular uptake and transport of encapsulated coumarin in the latter particles, respectively. While the presence of mucus in Caco-2/HT29 co-culture caused 40-50% decrease of cellular uptake and transport for coumarin encapsulated in digested Cru/Cs particles, it had no significant effect on the cell uptake and transport of coumarin associated with Cru/Ca particles after digestion. Energy-dependent mechanisms were the dominant mechanism for uptake of both undigested and digested particles. Therefore, in Caco-2/HT29 co-culture which closely simulated intestinal epithelial cells, undigested Cru/Ca and Cru/Cs particles had the ability to penetrate mucus layers, while digested Cru/Cs particles showed mucoadhesive property, and digested Cru/Ca particles were dissociated. Our results points to a potential for cruciferin based nanoparticles for oral drug delivery. The long-term objective of

  6. Yersinia enterocolitica YopT and Clostridium difficile Toxin B Induce Expression of GILZ in Epithelial Cells

    PubMed Central

    Köberle, Martin; Göppel, David; Grandl, Tanja; Gaentzsch, Peer; Manncke, Birgit; Berchtold, Susanne; Müller, Steffen; Lüscher, Bernhard; Asselin-Labat, Marie-Liesse; Pallardy, Marc; Sorg, Isabel; Langer, Simon; Barth, Holger; Zumbihl, Robert; Autenrieth, Ingo B.; Bohn, Erwin

    2012-01-01

    Glucocorticoid induced-leucine zipper (GILZ) has been shown to be induced in cells by different stimuli such as glucocorticoids, IL-10 or deprivation of IL-2. GILZ has anti-inflammatory properties and may be involved in signalling modulating apoptosis. Herein we demonstrate that wildtype Yersinia enterocolitica which carry the pYV plasmid upregulated GILZ mRNA levels and protein expression in epithelial cells. Infection of HeLa cells with different Yersinia mutant strains revealed that the protease activity of YopT, which cleaves the membrane-bound form of Rho GTPases was sufficient to induce GILZ expression. Similarly, Clostridium difficile toxin B, another bacterial inhibitor of Rho GTPases induced GILZ expression. YopT and toxin B both increased transcriptional activity of the GILZ promoter in HeLa cells. GILZ expression could not be linked to the inactivation of an individual Rho GTPase by these toxins. However, forced expression of RhoA and RhoB decreased basal GILZ promoter activity. Furthermore, MAPK activation proved necessary for profound GILZ induction by toxin B. Promoter studies and gel shift analyses defined binding of upstream stimulatory factor (USF) 1 and 2 to a canonical c-Myc binding site (E-box) in the GILZ promoter as a crucial step of its trans-activation. In addition we could show that USF-1 and USF-2 are essential for basal as well as toxin B induced GILZ expression. These findings define a novel way of GILZ promoter trans-activation mediated by bacterial toxins and differentiate it from those mediated by dexamethasone or deprivation of IL-2. PMID:22792400

  7. PKM2 Subcellular Localization Is Involved in Oxaliplatin Resistance Acquisition in HT29 Human Colorectal Cancer Cell Lines

    PubMed Central

    Ginés, Alba; Bystrup, Sara; Ruiz de Porras, Vicenç; Guardia, Cristina; Musulén, Eva; Martínez-Cardús, Anna; Manzano, José Luis; Layos, Laura; Abad, Albert; Martínez-Balibrea, Eva

    2015-01-01

    Chemoresistance is the main cause of treatment failure in advanced colorectal cancer (CRC). However, molecular mechanisms underlying this phenomenon remain to be elucidated. In a previous work we identified low levels of PKM2 as a putative oxaliplatin-resistance marker in HT29 CRC cell lines and also in patients. In order to assess how PKM2 influences oxaliplatin response in CRC cells, we silenced PKM2 using specific siRNAs in HT29, SW480 and HCT116 cells. MTT test demonstrated that PKM2 silencing induced resistance in HT29 and SW480 cells and sensitivity in HCT116 cells. Same experiments in isogenic HCT116 p53 null cells and double silencing of p53 and PKM2 in HT29 cells failed to show an influence of p53. By using trypan blue stain and FITC-Annexin V/PI tests we detected that PKM2 knockdown was associated with an increase in cell viability but not with a decrease in apoptosis activation in HT29 cells. Fluorescence microscopy revealed PKM2 nuclear translocation in response to oxaliplatin in HCT116 and HT29 cells but not in OXA-resistant HTOXAR3 cells. Finally, by using a qPCR Array we demonstrated that oxaliplatin and PKM2 silencing altered cell death gene expression patterns including those of BMF, which was significantly increased in HT29 cells in response to oxaliplatin, in a dose and time-dependent manner, but not in siPKM2-HT29 and HTOXAR3 cells. BMF gene silencing in HT29 cells lead to a decrease in oxaliplatin-induced cell death. In conclusion, our data report new non-glycolytic roles of PKM2 in response to genotoxic damage and proposes BMF as a possible target gene of PKM2 to be involved in oxaliplatin response and resistance in CRC cells. PMID:25955657

  8. Identification and expression analyses of a novel serotonin receptor gene, 5-HT2β, in the field cricket, Gryllus bimaculatus.

    PubMed

    Watanabe, T; Aonuma, H

    2012-01-01

    Biogenic amine serotonin (5-HT) modulates various aspects of behaviors such as aggressive behavior and circadian behavior in the cricket. In our previous report, in order to elucidate the molecular basis of the cricket 5-HT system, we identified three genes involved in 5-HT biosynthesis, as well as four 5-HT receptor genes (5-HT1A, 5-HT1B, 5-HT2α, and 5-HT7) expressed in the brain of the field cricket Gryllus bimaculatus DeGeer [7]. In the present study, we identified Gryllus 5-HT2β gene, an additional 5-HT receptor gene expressed in the cricket brain, and examined its tissue-specific distribution and embryonic stage-dependent expression. Gryllus 5-HT2β gene was ubiquitously expressed in the all examined adult tissues, and was expressed during early embryonic development, as well as during later stages. This study suggests functional differences between two 5-HT2 receptors in the cricket.

  9. Modulatory effect of the 5-HT1A agonist buspirone and the mixed non-hallucinogenic 5-HT1A/2A agonist ergotamine on psilocybin-induced psychedelic experience.

    PubMed

    Pokorny, Thomas; Preller, Katrin H; Kraehenmann, Rainer; Vollenweider, Franz X

    2016-04-01

    The mixed serotonin (5-HT) 1A/2A/2B/2C/6/7 receptor agonist psilocybin dose-dependently induces an altered state of consciousness (ASC) that is characterized by changes in sensory perception, mood, thought, and the sense of self. The psychological effects of psilocybin are primarily mediated by 5-HT2A receptor activation. However, accumulating evidence suggests that 5-HT1A or an interaction between 5-HT1A and 5-HT2A receptors may contribute to the overall effects of psilocybin. Therefore, we used a double-blind, counterbalanced, within-subject design to investigate the modulatory effects of the partial 5-HT1A agonist buspirone (20mg p.o.) and the non-hallucinogenic 5-HT2A/1A agonist ergotamine (3mg p.o.) on psilocybin-induced (170 µg/kg p.o.) psychological effects in two groups (n=19, n=17) of healthy human subjects. Psychological effects were assessed using the Altered State of Consciousness (5D-ASC) rating scale. Buspirone significantly reduced the 5D-ASC main scale score for Visionary Restructuralization (VR) (p<0.001), which was mostly driven by a reduction of the VR item cluster scores for elementary and complex visual hallucinations. Further, buspirone also reduced the main scale score for Oceanic Boundlessness (OB) including derealisation and depersonalisation phenomena at a trend level (p=0.062), whereas ergotamine did not show any effects on the psilocybin-induced 5D-ASC main scale scores. The present finding demonstrates that buspirone exerts inhibitory effects on psilocybin-induced effects, presumably via 5-HT1A receptor activation, an interaction between 5-HT1A and 5-HT2A receptors, or both. The data suggest that the modulation of 5-HT1A receptor activity may be a useful target in the treatment of visual hallucinations in different psychiatric and neurological diseases. Copyright © 2016 Elsevier B.V. and ECNP. All rights reserved.

  10. [ASSOCIATION BETWEEN FOUR SEROTONIC GENES POLYMORPHISM (5HTTL, 5HT1A, 5HT2A, AND MAOA) AND PERSONALITY TRAITS IN WRESTLERS AND CONTROL GROUP].

    PubMed

    Butovskaya, P R; Lazebnij, O E; Fekhretdionva, D I; Vasil'ev, V A; Prosikova, E A; Lysenko, V V; Udina, I G; Butovskaya, M L

    2015-01-01

    This study presents the data on the polymorphisms of the serotonin system genes (5-HTTL, 5-HT1A, 5-HT2A, and MAOA) in male and female wrestlers and in the control group. The population genetics analysis of the 5HTTL gene showed the highest frequency of the SS genotype 5-HTTLPR in sportsmen (p = 0.04), as well as the trend toward higher frequency of united genotypes of the locus of 5-HTTLPR VNTR and SNP rs25531--SASA (p = 0.06) in comparison with the control group. As for the polymorphisms for other genes 5-HT1A (rs6295), 5-HT2A (rs6311), and MAOA (VNTR), we found no significant differences between the groups tested. Using the NEO PI-R questionnaire we analyzed the possible correlations between the genotypes and the psychological traits in our samples. It was demonstrated that the athletic success in elite sportsmen was associated with lower openness to experience and higher conscientiousness. The interaction effect of the gender and 5-HT2A on the self-rating for openness to experience, interaction effect of the level of the sport success and 5-HT2A, and the interaction effect of the gender and 5-HT1A genotype on self-reported conscientiousness were observed as a trend.

  11. Serotonin 5-HT2C receptor-mediated inhibition of the M-current in hypothalamic POMC neurons.

    PubMed

    Roepke, T A; Smith, A W; Rønnekleiv, O K; Kelly, M J

    2012-06-01

    Hypothalamic proopiomelanocortin (POMC) neurons are controlled by many central signals, including serotonin. Serotonin increases POMC activity and reduces feeding behavior via serotonion [5-hydroxytryptamine (5-HT)] receptors by modulating K(+) currents. A potential K(+) current is the M-current, a noninactivating, subthreshold outward K(+) current. Previously, we found that M-current activity was highly reduced in fasted vs. fed states in neuropeptide Y neurons. Because POMC neurons also respond to energy states, we hypothesized that fasting may alter the M-current and/or its modulation by serotonergic input to POMC neurons. Using visualized-patch recording in neurons from fed male enhanced green fluorescent protein-POMC transgenic mice, we established that POMC neurons expressed a robust M-current (102.1 ± 6.7 pA) that was antagonized by the selective KCNQ channel blocker XE-991 (40 μM). However, the XE-991-sensitive current in POMC neurons did not differ between fed and fasted states. To determine if serotonin suppresses the M-current via the 5-HT(2C) receptor, we examined the effects of the 5-HT(2A)/5-HT(2C) receptor agonist 2,5-dimethoxy-4-iodoamphetamine (DOI) on the M-current. Indeed, DOI attenuated the M-current by 34.5 ± 6.9% and 42.0 ± 5.3% in POMC neurons from fed and fasted male mice, respectively. In addition, the 5-HT(1B)/5-HT(2C) receptor agonist m-chlorophenylpiperazine attenuated the M-current by 42.4 ± 5.4% in POMC neurons from fed male mice. Moreover, the selective 5-HT(2C) receptor antagonist RS-102221 abrogated the actions of DOI in suppressing the M-current. Collectively, these data suggest that although M-current expression does not differ between fed and fasted states in POMC neurons, serotonin inhibits the M-current via activation of 5-HT(2C) receptors to increase POMC neuronal excitability and, subsequently, reduce food intake.

  12. Selective verbal and spatial memory impairment after 5-HT1A and 5-HT2A receptor blockade in healthy volunteers pre-treated with an SSRI.

    PubMed

    Wingen, M; Kuypers, K P C; Ramaekers, J G

    2007-07-01

    Serotonergic neurotransmission has been implicated in memory impairment. It is unclear however if memory performance is mediated through general 5-HT availability, through specific 5-HT receptors or both. The aim of the present study was to assess the contribution of 5-HT reuptake inhibition and specific blockade of 5-HT(1A) and 5-HT(2A) receptors to memory impairment. The study was conducted according to a randomized, double-blind, placebo-controlled, four-way cross-over design including 16 healthy volunteers. The treatment consisted of oral administration of escitalopram 20 mg + placebo, escitalopram 20 mg + ketanserin 50 mg, escitalopram 20 mg + pindolol 10 mg and placebo on 4 separate days with a washout period of minimum 7 days. Different memory tasks were performed including verbal memory, spatial working memory and reversal learning. Escitalopram showed an impairing effect on immediate verbal recall which nearly reached statistical significance. No effects of escitalopram were found on other types of memory. In combination with pindolol, immediate verbal recall was significantly impaired. Escitalopram in combination with ketanserin impaired spatial working memory significantly. No effects were found on reversal learning. Selective impairment of immediate verbal recall after a 5-HT(1A) partial agonist and selective impairment of spatial working memory performance after 5-HT(2A) receptor antagonist, both in combination with a selective serotonergic reuptake inhibitor (escitalopram), suggests that 5-HT(1A) and 5-HT(2A) receptors are distinctly involved in verbal and spatial memory.

  13. Comparative effect of lurasidone and blonanserin on cortical glutamate, dopamine, and acetylcholine efflux: role of relative serotonin (5-HT)2A and DA D2 antagonism and 5-HT1A partial agonism.

    PubMed

    Huang, Mei; Panos, John J; Kwon, Sunoh; Oyamada, Yoshihiro; Rajagopal, Lakshmi; Meltzer, Herbert Y

    2014-03-01

    Atypical antipsychotic drugs (AAPDs) have been suggested to be more effective in improving cognitive impairment in schizophrenia than typical APDs, a conclusion supported by differences in receptor affinities and neurotransmitter efflux in the cortex and the hippocampus. More potent serotonin (5-HT)2A than dopamine (DA) D2 receptors antagonism, and direct or indirect 5-HT1A agonism, characterize almost all AAPDs. Blonanserin, an AAPD, has slightly greater affinity for D2 than 5-HT2A receptors. Using microdialysis and ultra performance liquid chromatography-mass spectrometry/mass spectrometry, we compared the abilities of the typical APD, haloperidol, three AAPDs, blonanserin, lurasidone, and olanzapine, and a selective 5-HT1A partial agonist, tandospirone, and all, except haloperidol, were found to ameliorate the cognitive deficits produced by the N-methyl-d-aspartate antagonist, phencyclidine, altering the efflux of neurotransmitters and metabolites in the rat cortex and nucleus accumbens. Blonanserin, lurasidone, olanzapine, and tandospirone, but not haloperidol, increased the efflux of cortical DA and its metabolites, homovanillic acid and 3,4-dihydroxyphenylacetic acid. Olanzapine and lurasidone increased the efflux of acetylcholine; lurasidone increased glutamate as well. None of the compounds significantly altered the efflux of 5-HT or its metabolite, 5-hydroxyindole acetic acid, or GABA, serine, and glycine. The ability to increase cortical DA efflux was the only shared effect of the compounds which ameliorates the deficit in cognition in rodents following phencyclidine. © 2013 International Society for Neurochemistry.

  14. Intrahippocampal LSD accelerates learning and desensitizes the 5-HT(2A) receptor in the rabbit, Romano et al.

    PubMed

    Romano, Anthony G; Quinn, Jennifer L; Li, Luchuan; Dave, Kuldip D; Schindler, Emmanuelle A; Aloyo, Vincent J; Harvey, John A

    2010-10-01

    Parenteral injections of d-lysergic acid diethylamide (LSD), a serotonin 5-HT(2A) receptor agonist, enhance eyeblink conditioning. Another hallucinogen, (±)-1(2, 5-dimethoxy-4-iodophenyl)-2-aminopropane hydrochloride (DOI), was shown to elicit a 5-HT(2A)-mediated behavior (head bobs) after injection into the hippocampus, a structure known to mediate trace eyeblink conditioning. This study aims to determine if parenteral injections of the hallucinogens LSD, d,l-2,5-dimethoxy-4-methylamphetamine, and 5-methoxy-dimethyltryptamine elicit the 5-HT(2A)-mediated behavior of head bobs and whether intrahippocampal injections of LSD would produce head bobs and enhance trace eyeblink conditioning. LSD was infused into the dorsal hippocampus just prior to each of eight conditioning sessions. One day after the last infusion of LSD, DOI was infused into the hippocampus to determine whether there had been a desensitization of the 5-HT(2A) receptor as measured by a decrease in DOI-elicited head bobs. Acute parenteral or intrahippocampal LSD elicited a 5-HT(2A) but not a 5-HT(2C)-mediated behavior, and chronic administration enhanced conditioned responding relative to vehicle controls. Rabbits that had been chronically infused with 3 or 10 nmol per side of LSD during Pavlovian conditioning and then infused with DOI demonstrated a smaller increase in head bobs relative to controls. LSD produced its enhancement of Pavlovian conditioning through an effect on 5-HT(2A) receptors located in the dorsal hippocampus. The slight, short-lived enhancement of learning produced by LSD appears to be due to the development of desensitization of the 5-HT(2A) receptor within the hippocampus as a result of repeated administration of its agonist (LSD).

  15. Mutagenesis Analysis Reveals Distinct Amino Acids of the Human Serotonin 5-HT2C Receptor Underlying the Pharmacology of Distinct Ligands.

    PubMed

    Liu, Yue; Canal, Clinton E; Cordova-Sintjago, Tania C; Zhu, Wanying; Booth, Raymond G

    2017-01-18

    While exploring the structure-activity relationship of 4-phenyl-2-dimethylaminotetralins (PATs) at serotonin 5-HT 2C receptors, we discovered that relatively minor modification of PAT chemistry impacts function at 5-HT 2C receptors. In HEK293 cells expressing human 5-HT 2C-INI receptors, for example, (-)-trans-3'-Br-PAT and (-)-trans-3'-Cl-PAT are agonists regarding Gα q -inositol phosphate signaling, whereas (-)-trans-3'-CF 3 -PAT is an inverse agonist. To investigate the ligand-receptor interactions that govern this change in function, we performed site-directed mutagenesis of 14 amino acids of the 5-HT 2C receptor based on molecular modeling and reported G protein-coupled receptor crystal structures, followed by molecular pharmacology studies. We found that S3.36, T3.37, and F5.47 in the orthosteric binding pocket are critical for affinity (K i ) of all PATs tested, we also found that F6.44, M6.47, C7.45, and S7.46 are primarily involved in regulating EC/IC 50 functional potencies of PATs. We discovered that when residue S5.43, N6.55, or both are mutated to alanine, (-)-trans-3'-CF 3 -PAT switches from inverse agonist to agonist function, and when N6.55 is mutated to leucine, (-)-trans-3'-Br-PAT switches from agonist to inverse agonist function. Notably, most point-mutations that affected PAT pharmacology did not significantly alter affinity (K D ) of the antagonist radioligand [ 3 H]mesulergine, but every mutation tested negatively impacted serotonin binding. Also, amino acid mutations differentially affected the pharmacology of other commercially available 5-HT 2C ligands tested. Collectively, the data show that functional outcomes shared by different ligands are mediated by different amino acids and that some 5-HT 2C receptor residues important for pharmacology of one ligand are not necessarily important for another ligand.

  16. 5-HT2 receptor blockade exhibits 5-HT vasodilator effects via nitric oxide, prostacyclin and ATP-sensitive potassium channels in rat renal vasculature.

    PubMed

    García-Pedraza, J A; García, M; Martín, M L; Rodríguez-Barbero, A; Morán, A

    2016-04-01

    The aim of this study was to determine whether orally sarpogrelate (selective 5-HT2 antagonist) treatment (30 mg/kg/day; 14 days) could modify 5-HT renal vasoconstrictor responses, characterizing 5-HT receptors and mediator mechanisms involved in serotonergic responses in the in situ autoperfused rat kidney. Intra-arterial (i.a.) injections of 5-HT (0.00000125 to 0.1 μg/kg) decreased renal perfusion pressure (RPP) but did not affect the mean blood pressure (MBP). i.a. agonists 5-CT (5-HT1/7), CGS-12066B (5-HT1B), L-694,247 (5-HT1D) or AS-19 (5-HT7) mimicked renal 5-HT vasodilator effect. However, neither 8-OH-DPAT (5-HT1A) nor 1-phenylbiguanide (5-HT3) modified RPP. Moreover: (i) GR-55562 (5-HT1B antagonist) and L-NAME (nitric oxide synthase [NOS] inhibitor) blocked CGS-12066B-induced vasodilator response, (ii) LY310762 (5-HT1D antagonist) and indomethacin (non-selective cyclooxygenase inhibitor) blocked L-694,247-induced vasodilator response; (iii) SB-258719 (5-HT7 antagonist) and glibenclamide (ATP-sensitive K+ channel blocker) blocked AS-19-induced vasodilator response; and (iv) 5-HT- or 5-CT-elicited renal vasodilation was significantly blocked by the mixture of GR-55562 + LY310762 + SB-258719. Furthermore, eNOS and iNOS proteins and prostacyclin levels are overexpressed in sarpogrelate-treated rats. Our data suggest that 5-HT exerts renal vasodilator effect in the in situ autoperfused sarpogrelate-treated rat kidney, mediated by 5-HT1D, 5-HT1B and 5-HT7 receptors, involving cyclooxygenase-derived prostacyclin, nitric oxide synthesis/release and ATP-sensitive K+ channels, respectively.

  17. Fusarium mycotoxin content of UK organic and conventional oats.

    PubMed

    Edwards, S G

    2009-07-01

    Every year between 2002 and 2005 approximately 100 samples of oats from fields of known agronomy were analysed by GC/MS for 10 trichothecenes: deoxynivalenol (DON), nivalenol, 3-acetylDON, 15-acetylDON, fusarenone X, T-2 toxin (T2), HT-2 toxin (HT2), diacetoxyscirpenol, neosolaniol and T-2 triol. Samples were also analysed for moniliformin and zearalenone by HPLC. Of the 10 trichothecenes analysed from 458 harvest samples of oat only three, 15-acetylDON, fusarenone X and diacetoxyscirpenol, were not detected. Moniliformin and zearalenone were absent or rarely detected, respectively. HT2 and T2 were the most frequently detected fusarium mycotoxins, present above the limit of quantification (10 microg kg(-1)) in 92 and 84% of samples, respectively, and were usually present at the highest concentrations. The combined mean and median for HT2 and T2 (HT2 + T2) was 570 and 213 microg kg(-1), respectively. There were good correlations between concentrations of HT2 and all other type A trichothecenes detected (T2, T2 triol and neosolaniol). Year and region had a significant effect on HT2 + T2 concentration. There was also a highly significant difference between HT2 + T2 content in organic and conventional samples, with the predicted mean for organic samples five times lower than that of conventional samples. This is the largest difference reported for any mycotoxin level in organic and conventional cereals. No samples exceeded the legal limits for DON or zearalenone in oats intended for human consumption. Legislative limits for HT2 and T2 are currently under consideration by the European Commission. Depending on the limits set for unprocessed oats intended for human consumption, the levels detected here could have serious consequences for the UK oat-processing industry.

  18. Evaluation of Performance and Potential Clinical Impact of ProSpecT Shiga Toxin Escherichia coli Microplate Assay for Detection of Shiga Toxin-Producing E. coli in Stool Samples

    PubMed Central

    Gavin, Patrick J.; Peterson, Lance R.; Pasquariello, Anna C.; Blackburn, Joanna; Hamming, Mark G.; Kuo, Kuo J.; Thomson, Richard B.

    2004-01-01

    Shiga toxin-producing Escherichia coli bacteria (STEC) are emerging pathogens capable of producing sporadic and epidemic diarrhea, hemorrhagic colitis, and potentially life-threatening hemolytic-uremic syndrome. Although the presence of E. coli O157 can be readily detected in stool by sorbitol-MacConkey agar culture (SMAC), STEC non-O157 serotypes cannot. In contrast to culture, testing for the presence of Shiga toxins 1 and 2 in stool detects both O157 and non-O157 STEC serotypes capable of causing disease. Over two consecutive summers, we evaluated the performance of the ProSpecT Shiga toxin E. coli Microplate assay (Alexon-Trend, Ramsey, Minn.), an enzyme immunoassay for the detection of Shiga toxins 1 and 2, on all stools submitted for culture of enteric pathogens, and the potential clinical impact of Shiga toxin detection. Twenty-nine stool specimens were STEC positive by ProSpecT assay. Twenty-seven of 29 STEC-positive isolates were confirmed by SMAC and serotyping or by a second enzyme immunoassay and PCR (positive predictive value, 93%). Thirteen of 27 confirmed Shiga toxin-producing strains were serotype O157. The remaining 14 strains represented 8 other serotypes. The ProSpecT assay was 100% sensitive and specific for detection of E. coli O157 in stool (7 of 7) compared to SMAC. In addition, the ProSpecT assay detected twice as many STEC as SMAC. Fifty-two percent of confirmed STEC-positive stools were nonbloody. Thus, in our population, screening strategies that test only visibly bloody stools for STEC would miss a majority of cases. Eleven (41%) STEC-positive patients were hospitalized, and eight (30%) developed severe disease (two developed hemolytic-uremic syndrome, and six developed hemorrhagic colitis). Prior to detection of STEC infection, seven (26%) and eight patients (30%) underwent unnecessary diagnostic procedures or received potentially deleterious empirical treatment, respectively. We propose that establishing a specific diagnosis of STEC

  19. Potential natural exposure of endangered red-crowned crane (Grus japonensis) to mycotoxins aflatoxin B1, deoxynivalenol, zearalenone, T-2 toxin, and ochratoxin A*

    PubMed Central

    Liu, Da-wei; Liu, Hong-yi; Zhang, Hai-bin; Cao, Ming-chang; Sun, Yong; Wu, Wen-da; Lu, Chang-hu

    2016-01-01

    A survey was conducted to determine whether mycotoxins were present in the foods consumed by red-crowned cranes (Grus japonensis) in the Yancheng Biosphere Reserve, China. Collected in the reserve’s core, buffer, and experimental zones during overwintering periods of 2013 to 2015, a total of 113 food samples were analyzed for aflatoxin B1, deoxynivalenol, zearalenone, T-2 toxin, and ochratoxin A using high performance liquid chromatography (HPLC). The contamination incidences vary among different zones and the mycotoxins levels of different food samples also presented disparity. Average mycotoxin concentration from rice grain was greater than that from other food types. Among mycotoxin-positive samples, 59.3% were simultaneously contaminated with more than one toxin. This study demonstrated for the first time that red-crowned cranes were exposed to mycotoxins in the Yancheng Biosphere Reserve and suggested that artificial wetlands could not be considered good habitats for the birds in this reserve, especially rice fields. PMID:26834016

  20. Serotonin (5-HT)2A/2C receptor agonist (2,5-dimethoxy-4-idophenyl)-2-aminopropane hydrochloride (DOI) improves voiding efficiency in the diabetic rat.

    PubMed

    Tu, Hongjian; Cao, Nailong; Gu, Baojun; Si, Jiemin; Chen, Zhong; Andersson, Karl-Erik

    2015-07-01

    To examine the effects of the serotonin (5-HT)2A/2C receptor agonist (2,5-dimethoxy-4-idophenyl)-2-aminopropane hydrochloride (DOI) on micturition in rats with diabetes mellitus (DM). Female Sprague-Dawley rats (n = 16) were divided into two groups: rats with Type 1 DM and age-matched control rats. DM was induced by i.p. injection of streptozotocin (65 mg/kg) and detailed cystometrogram (CMG) studies were performed 8 weeks post-injection in all rats under urethane anaesthesia. The selective 5-HT2A antagonist ketanserin was administered after each DOI dose-response curve was plotted. All drugs were administered i.v. Compared with controls, comprehensive urodynamic studies showed that DM rats had a higher bladder capacity and post-void residual urine volume (PVR), and a markedly lower voiding efficiency. In DM rats, DOI (0.01-0.3 mg/kg) induced significant dose-dependent increases in micturition volume and reductions in PVR, resulting in greater voiding efficiency. CMG measurements showed a dose-dependent increase in high-frequency oscillation (HFO) activity, evidenced by an increased duration of HFOs per voiding. This correlated with the improved voiding efficiency. Ketanserin (0.1 mg/kg) partially or completely reversed the DOI-induced changes. The HFOs observed in the present study seem to correlate with external urethral sphincter bursting activity during voiding. Bladder voiding efficiency was reduced in DM rats. The 5-HT2A receptor agonist can enhance HFO activity and improves voiding efficiency, and so may represent a new strategy to improve voiding efficiency after DM in experimental studies. © 2014 The Authors BJU International © 2014 BJU International Published by John Wiley & Sons Ltd.

  1. Nitrate-Melt Synthesized HT-LiCoO2 as a Superior Cathode-Material for Lithium-Ion Batteries

    PubMed Central

    Sathiya, Mariyappan; Prakash, Annigere S.; Ramesha, Kannadka; Shukla, Ashok K.

    2009-01-01

    An electrochemically-active high-temperature form of LiCoO2 (HT-LiCoO2) is prepared by thermally decomposing its constituent metal-nitrates at 700 ºC. The synthetic conditions have been optimized to achieve improved performance with the HT-LiCoO2 cathode in Li-ion batteries. For this purpose, the synthesized materials have been characterized by powder X-ray diffraction, scanning electron microscopy, and galvanostatic charge-discharge cycling. Cathodes comprising HT-LiCoO2 exhibit a specific capacity of 140 mAhg-1 with good capacity-retention over several charge-discharge cycles in the voltage range between 3.5 V and 4.2 V, and can sustain improved rate capability in contrast to a cathode constituting LiCoO2 prepared by conventional ceramic method. The nitrate-melt-decomposition method is also found effective for synthesizing Mg-/Al- doped HT-LiCoO2; these also are investigated as cathode materials for Li-ion batteries.

  2. Postnatal Day 2 to 11 Constitutes a 5-HT-Sensitive Period Impacting Adult mPFC Function

    PubMed Central

    Rebello, Tahilia J.; Yu, Qinghui; Goodfellow, Nathalie M.; Caffrey Cagliostro, Martha K.; Teissier, Anne; Morelli, Emanuela; Demireva, Elena Y.; Chemiakine, Alexei; Rosoklija, Gorazd B.; Dwork, Andrew J.; Lambe, Evelyn K.; Ansorge, Mark S.

    2014-01-01

    Early-life serotonin [5-hydroxytryptamine (5-HT)] signaling modulates brain development, which impacts adult behavior, but 5-HT-sensitive periods, neural substrates, and behavioral consequences remain poorly understood. Here we identify the period ranging from postnatal day 2 (P2) to P11 as 5-HT sensitive, with 5-HT transporter (5-HTT) blockade increasing anxiety- and depression-like behavior, and impairing fear extinction learning and memory in adult mice. Concomitantly, P2–P11 5-HTT blockade causes dendritic hypotrophy and reduced excitability of infralimbic (IL) cortex pyramidal neurons that normally promote fear extinction. By contrast, the neighboring prelimbic (PL) pyramidal neurons, which normally inhibit fear extinction, become more excitable. Excitotoxic IL but not PL lesions in adult control mice reproduce the anxiety-related phenotypes. These findings suggest that increased 5-HT signaling during P2–P11 alters adult mPFC function to increase anxiety and impair fear extinction, and imply a differential role for IL and PL neurons in regulating affective behaviors. Together, our results support a developmental mechanism for the etiology and pathophysiology of affective disorders and fear-related behaviors. PMID:25209278

  3. Structure-Activity Relationships of Constrained Phenylethylamine Ligands for the Serotonin 5-HT2 Receptors

    PubMed Central

    Isberg, Vignir; Paine, James; Leth-Petersen, Sebastian; Kristensen, Jesper L.; Gloriam, David E.

    2013-01-01

    Serotonergic ligands have proven effective drugs in the treatment of migraine, pain, obesity, and a wide range of psychiatric and neurological disorders. There is a clinical need for more highly 5-HT2 receptor subtype-selective ligands and the most attention has been given to the phenethylamine class. Conformationally constrained phenethylamine analogs have demonstrated that for optimal activity the free lone pair electrons of the 2-oxygen must be oriented syn and the 5-oxygen lone pairs anti relative to the ethylamine moiety. Also the ethyl linker has been constrained providing information about the bioactive conformation of the amine functionality. However, combined 1,2-constriction by cyclization has only been tested with one compound. Here, we present three new 1,2-cyclized phenylethylamines, 9–11, and describe their synthetic routes. Ligand docking in the 5-HT2B crystal structure showed that the 1,2-heterocyclized compounds can be accommodated in the binding site. Conformational analysis showed that 11 can only bind in a higher-energy conformation, which would explain its absent or low affinity. The amine and 2-oxygen interactions with D3.32 and S3.36, respectively, can form but shift the placement of the core scaffold. The constraints in 9–11 resulted in docking poses with the 4-bromine in closer vicinity to 5.46, which is polar only in the human 5-HT2A subtype, for which 9–11 have the lowest affinity. The new ligands, conformational analysis and docking expand the structure-activity relationships of constrained phenethylamines and contributes towards the development of 5-HT2 receptor subtype-selective ligands. PMID:24244317

  4. Structure-activity relationships of constrained phenylethylamine ligands for the serotonin 5-HT2 receptors.

    PubMed

    Isberg, Vignir; Paine, James; Leth-Petersen, Sebastian; Kristensen, Jesper L; Gloriam, David E

    2013-01-01

    Serotonergic ligands have proven effective drugs in the treatment of migraine, pain, obesity, and a wide range of psychiatric and neurological disorders. There is a clinical need for more highly 5-HT2 receptor subtype-selective ligands and the most attention has been given to the phenethylamine class. Conformationally constrained phenethylamine analogs have demonstrated that for optimal activity the free lone pair electrons of the 2-oxygen must be oriented syn and the 5-oxygen lone pairs anti relative to the ethylamine moiety. Also the ethyl linker has been constrained providing information about the bioactive conformation of the amine functionality. However, combined 1,2-constriction by cyclization has only been tested with one compound. Here, we present three new 1,2-cyclized phenylethylamines, 9-11, and describe their synthetic routes. Ligand docking in the 5-HT2B crystal structure showed that the 1,2-heterocyclized compounds can be accommodated in the binding site. Conformational analysis showed that 11 can only bind in a higher-energy conformation, which would explain its absent or low affinity. The amine and 2-oxygen interactions with D3.32 and S3.36, respectively, can form but shift the placement of the core scaffold. The constraints in 9-11 resulted in docking poses with the 4-bromine in closer vicinity to 5.46, which is polar only in the human 5-HT2A subtype, for which 9-11 have the lowest affinity. The new ligands, conformational analysis and docking expand the structure-activity relationships of constrained phenethylamines and contributes towards the development of 5-HT2 receptor subtype-selective ligands.

  5. The relationship between the daily dose, the plasma concentration of blonanserin, and its plasma anti-dopamine D2 and anti-serotonin 5-HT2A activity.

    PubMed

    Suzuki, Hidenobu; Gen, Keishi

    2010-01-01

    Blonanserin (BNS) possesses anti-serotonin 5-HT(2A) activity in addition to anti-dopamine D(2) activity, which is characteristic of second-generation antipsychotics, little information is available on its pharmacologic profile in vivo. We investigated the BNS daily dose, plasma concentration, plasma anti-D(2) activity, and plasma anti-5-HT(2A) activity in schizophrenia in a total of 14 subjects. Blood samples were taken 14 days after the BNS dose was fixed, and the plasma concentration was measured by means of high-performance liquid chromatographic (HPLC) method. In addition, the plasma anti-D(2) activity and anti-5-HT(2A) activity were measured by means of radioreceptor assays in which [(3)H]-spiperone and [(3)H]-ketanserin were used. The results revealed a statistically significant correlation between the daily dose and the plasma concentration (p = 0.04). Statistically significant correlations were also observed between the plasma concentration and the anti-D(2) activity and between the plasma concentration and the anti-5-HT(2A) activity (p = 0.003 and 0.04). It is therefore believed that both the anti-D(2) activity in plasma and the anti-5-HT(2A) activity in plasma are regulated almost solely by the unchanged principal. Moreover, the mean plasma serotonin/dopamine (S/D) ratio was 0.9 and BNS exhibited both anti-D(2) activity and also anti-5-HT(2A) activity in vivo, as well, so it was clear that the in vitro pharmacological profile was retained in vivo.

  6. Transport Study of Egg-Derived Antihypertensive Peptides (LKP and IQW) Using Caco-2 and HT29 Coculture Monolayers.

    PubMed

    Xu, Qingbiao; Fan, Hongbing; Yu, Wenlin; Hong, Hui; Wu, Jianping

    2017-08-30

    The objective of this study was to investigate the mechanisms of the transport of antihypertensive tripeptides LKP (Leu-Lys-Pro) and IQW (Ile-Gln-Trp) derived from egg white using a coculture system of Caco-2 and HT29 cell monolayers. The results revealed that LKP and IQW have no cytotoxicity to the cell viability after 2 h incubation, could be transported intact across coculture monolayers (apparent permeability coefficient: (18.11 ± 1.57) × 10 -8 and (13.21 ± 1.12) × 10 -8 cm/s, respectively), and were resistant to peptidase secreted by enterocytes. In addition, the transports were significantly inhibited by dipeptide Gly-Pro (P < 0.05), a competitive substance of peptide transporter 1 (PepT1). The transports from apical to basolateral side were significantly higher than that of the reverse direction (P < 0.05). These results suggest that PepT1 is involved in LKP and IQW transports. The transports were also significantly decreased by theaflavin-3'-O-gallate (P < 0.05), an enhancer of tight junction (TJ) and increased by cytochalasin D (P < 0.05), a disruptor of TJ but not influenced by wortamanin, a transcytosis inhibitor, suggesting that passive paracellular route via TJs is also involved in LKP and IQW transports but not transcytosis. In addition, siRNA was also used to knockdown the expression of PepT1 and significantly inhibited the transport (P < 0.05), confirming that PepT1 is involved in transport process. Therefore, both passive paracellular route via TJ and active route via PepT1 coexist in the transport of antihypertensive LKP and IQW across Caco-2/HT29 coculture monolayers.

  7. Potentiation of 5-methoxy-N,N-dimethyltryptamine-induced hyperthermia by harmaline and the involvement of activation of 5-HT1A and 5-HT2A receptors.

    PubMed

    Jiang, Xi-Ling; Shen, Hong-Wu; Yu, Ai-Ming

    2015-02-01

    5-Methoxy-N,N-dimethyltryptamine (5-MeO-DMT) and harmaline are serotonin (5-HT) analogs often abused together, which alters thermoregulation that may indicate the severity of serotonin toxicity. Our recent studies have revealed that co-administration of monoamine oxidase inhibitor harmaline leads to greater and prolonged exposure to 5-HT agonist 5-MeO-DMT that might be influenced by cytochrome P450 2D6 (CYP2D6) status. This study was to define the effects of harmaline and 5-MeO-DMT on thermoregulation in wild-type and CYP2D6-humanized (Tg-CYP2D6) mice, as well as the involvement of 5-HT receptors. Animal core body temperatures were monitored noninvasively in the home cages after implantation of telemetry transmitters and administration of drugs. Harmaline (5 and 15 mg/kg, i.p.) alone was shown to induce hypothermia that was significantly affected by CYP2D6 status. In contrast, higher doses of 5-MeO-DMT (10 and 20 mg/kg) alone caused hyperthermia. Co-administration of harmaline (2, 5 or 15 mg/kg) remarkably potentiated the hyperthermia elicited by 5-MeO-DMT (2 or 10 mg/kg), which might be influenced by CYP2D6 status at certain dose combination. Interestingly, harmaline-induced hypothermia was only attenuated by 5-HT1A receptor antagonist WAY-100635, whereas 5-MeO-DMT- and harmaline-5-MeO-DMT-induced hyperthermia could be suppressed by either WAY-100635 or 5-HT2A receptor antagonists (MDL-100907 and ketanserin). Moreover, stress-induced hyperthermia under home cage conditions was not affected by WAY-100635 but surprisingly attenuated by MDL-100907 and ketanserin. Our results indicate that co-administration of monoamine oxidase inhibitor largely potentiates 5-MeO-DMT-induced hyperthermia that involves the activation of both 5-HT1A and 5-HT2A receptors. These findings shall provide insights into development of anxiolytic drugs and new strategies to relieve the lethal hyperthermia in serotonin toxicity.

  8. Potentiation of 5-methoxy-N,N-dimethyltryptamine-induced hyperthermia by harmaline and the involvement of activation of 5-HT1A and 5-HT2A receptors

    PubMed Central

    Jiang, Xi-Ling; Shen, Hong-Wu; Yu, Ai-Ming

    2014-01-01

    5-Methoxy-N,N-dimethyltryptamine (5-MeO-DMT) and harmaline are serotonin (5-HT) analogs often abused together, which alters thermoregulation that may indicate the severity of serotonin toxicity. Our recent studies have revealed that co-administration of monoamine oxidase inhibitor harmaline leads to greater and prolonged exposure to 5-HT agonist 5-MeO-DMT that might be influenced by cytochrome P450 2D6 (CYP2D6) status. This study was to define the effects of harmaline and 5-MeO-DMT on thermoregulation in wild-type and CYP2D6-humanized (Tg-CYP2D6) mice, as well as the involvement of 5-HT receptors. Animal core body temperatures were monitored noninvasively in the home cages after implantation of telemetry transmitters and administration of drugs. Harmaline (5 and 15 mg/kg, i.p.) alone was shown to induce hypothermia that was significantly affected by CYP2D6 status. In contrast, higher doses of 5-MeO-DMT (10 and 20 mg/kg) alone caused hyperthermia. Co-administration of harmaline (2, 5 or 15 mg/kg) remarkably potentiated the hyperthermia elicited by 5-MeO-DMT (2 or 10 mg/kg), which might be influenced by CYP2D6 status at certain dose combination. Interestingly, harmaline-induced hypothermia was only attenuated by 5-HT1A receptor antagonist WAY-100635, whereas 5-MeO-DMT- and harmaline-5-MeO-DMT-induced hyperthermia could be suppressed by either WAY-100635 or 5-HT2A receptor antagonists (MDL-100907 and ketanserin). Moreover, stress-induced hyperthermia under home cage conditions was not affected by WAY-100635 but surprisingly attenuated by MDL-100907 and ketanserin. Our results indicate that co-administration of monoamine oxidase inhibitor largely potentiates 5-MeO-DMT-induced hyperthermia that involves the activation of both 5-HT1A and 5-HT2A receptors. These findings shall provide insights into development of anxiolytic drugs and new strategies to relieve the lethal hyperthermia in serotonin toxicity. PMID:25446678

  9. Chronic cannabis promotes pro-hallucinogenic signaling of 5-HT2A receptors through Akt/mTOR pathway.

    PubMed

    Ibarra-Lecue, Inés; Mollinedo-Gajate, Irene; Meana, J Javier; Callado, Luis F; Diez-Alarcia, Rebeca; Urigüen, Leyre

    2018-04-27

    Long-term use of potent cannabis during adolescence increases the risk of developing schizophrenia later in life, but to date, the mechanisms involved remain unknown. Several findings suggest that the functional selectivity of serotonin 2A receptor (5-HT2AR) through inhibitory G-proteins is involved in the molecular mechanisms responsible for psychotic symptoms. Moreover, this receptor is dysregulated in the frontal cortex of schizophrenia patients. In this context, studies involving cannabis exposure and 5-HT2AR are scarce. Here, we tested in mice the effect of an early chronic Δ 9 -tetrahydrocannabinol (THC) exposure on cortical 5-HT2AR expression, as well as on its in vivo and in vitro functionality. Long-term exposure to THC induced a pro-hallucinogenic molecular conformation of the 5-HT2AR and exacerbated schizophrenia-like responses, such as prepulse inhibition disruption. Supersensitive coupling of 5-HT2AR toward inhibitory Gαi1-, Gαi3-, Gαo-, and Gαz-proteins after chronic THC exposure was observed, without changes in the canonical Gαq/11-protein pathway. In addition, we found that inhibition of Akt/mTOR pathway by rapamycin blocks the changes in 5-HT2AR signaling pattern and the supersensitivity to schizophrenia-like effects induced by chronic THC. The present study provides the first evidence of a mechanistic explanation for the relationship between chronic cannabis exposure in early life and increased risk of developing psychosis-like behaviors in adulthood.

  10. The human serotonin 5-HT{sub 2C} receptor: Complete cDNA, genomic structure, and alternatively spliced variant

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Xie, Enzhong; Zhu, Lingyu; Zhao, Lingyun

    1996-08-01

    The complete 4775-nt cDNA encoding the human serotonin 5-HT{sub 2C} receptor (5-HT{sub 2C}R), a G-protein-coupled receptor, has been isolated. It contains a 1377-nt coding region flanked by a 728-nt 5{prime}-untranslated region and a 2670-nt 3{prime}-untranslated region. By using the cloned 5-HT{sub 2C}R cDNA probe, the complete human gene for this receptor has been isolated and shown to contain six exons and five introns spanning at least 230 kb of DNA. The coding region of the human 5-HT{sub 2C}R gene is interrupted by three introns, and the positions of the intron/exon junctions are conserved between the human and the rodent genes.more » In addition, an alternatively spliced 5-HT{sub 2C}R RNA that contains a 95-nt deletion in the region coding for the second intracellular loop and the fourth transmembrane domain of the receptor has been identified. This deletion leads to a frameshift and premature termination so that the short isoform RNA encodes a putative protein of 248 amino acids. The ratio for the short isoform over the 5-HT{sub 2C}R RNA was found to be higher in choroid plexus tumor than in normal brain tissue, suggesting the possibility of differential regulation of the 5-HT{sub 2C}R gene in different neural tissues or during tumorigenesis. Transcription of the human 5-HT{sub 2C}R gene was found to be initiated at multiple sites. No classical TATA-box sequence was found at the appropriate location, and the 5{prime}-flanking sequence contains many potential transcription factor-binding sites. A 7.3-kb 5{prime}-flanking 5-HT{sub 2C}R DNA directed the efficient expression of a luciferase reported gene in SK-N-SH and IMR32 neuroblastoma cells, indicating that is contains a functional promoter. 69 refs., 8 figs., 1 tab.« less

  11. Relationship of obstructive sleep apnea syndrome with the 5-HT2A receptor gene in Brazilian patients.

    PubMed

    de Carvalho, Thiago Bittencourt Ottoni; Suman, Marcela; Molina, Fernando Drimel; Piatto, Vânia Belintani; Maniglia, José Victor

    2013-03-01

    Serotonin (5-HT) regulates a variety of visceral and physiological functions, including sleep. Polymorphisms in the 5-HT2A receptor gene can alter its transcription, affecting the number of receptors in the serotoninergic system, contributing to obstructive sleep apnea syndrome (OSAS). The aim of this study was to determine the prevalence of the 102T-C and -1438G-A polymorphisms in the 5-HTR2A gene in Brazilian patients with and without OSAS. A cross-sectional study performed at the Otorhinolaryngology and Sleep Disorder Out Clinics, São José do Rio Preto Medical School, FAMERP. One hundred patients were examined as index cases and 100 persons as controls, of both genders to both groups. DNA was extracted from peripheral blood leukocytes, and the sites that encompassed both polymorphisms were amplified by PCR-RFLP. There was a significant prevalence of the male gender in index cases compared with the control group gender (p < 0.0001). There was no significant genotypic difference in the 102T-C polymorphism between the case and control groups (p = 1.000). The AA genotype of the -1438G-A polymorphism was more prevalent in the patients with OSAS compared with the controls (OR, 2.3; CI 95% 1.20-4.38; p = 0.01). There was no difference in the prevalence of the 102T-C polymorphism between patients with OSAS and the control group. Serotoninergic system dysfunction appeared to be related to OSAS. The -1438G-A polymorphism and OSAS are related in this studied Brazilian population.

  12. Gq/5-HT2c receptor signals activate a local GABAergic inhibitory feedback circuit to modulate serotonergic firing and anxiety in mice.

    PubMed

    Spoida, Katharina; Masseck, Olivia A; Deneris, Evan S; Herlitze, Stefan

    2014-04-29

    Serotonin 2c receptors (5-HT2c-Rs) are drug targets for certain mental disorders, including schizophrenia, depression, and anxiety. 5-HT2c-Rs are expressed throughout the brain, making it difficult to link behavioral changes to circuit specific receptor expression. Various 5-HT-Rs, including 5-HT2c-Rs, are found in the dorsal raphe nucleus (DRN); however, the function of 5-HT2c-Rs and their influence on the serotonergic signals mediating mood disorders remain unclear. To investigate the role of 5-HT2c-Rs in the DRN in mice, we developed a melanopsin-based optogenetic probe for activation of Gq signals in cellular domains, where 5-HT2c-Rs are localized. Our results demonstrate that precise temporal control of Gq signals in 5-HT2c-R domains in GABAergic neurons upstream of 5-HT neurons provides negative feedback regulation of serotonergic firing to modulate anxiety-like behavior in mice.

  13. Gq/5-HT2c receptor signals activate a local GABAergic inhibitory feedback circuit to modulate serotonergic firing and anxiety in mice

    PubMed Central

    Spoida, Katharina; Masseck, Olivia A.; Deneris, Evan S.; Herlitze, Stefan

    2014-01-01

    Serotonin 2c receptors (5-HT2c-Rs) are drug targets for certain mental disorders, including schizophrenia, depression, and anxiety. 5-HT2c-Rs are expressed throughout the brain, making it difficult to link behavioral changes to circuit specific receptor expression. Various 5-HT-Rs, including 5-HT2c-Rs, are found in the dorsal raphe nucleus (DRN); however, the function of 5-HT2c-Rs and their influence on the serotonergic signals mediating mood disorders remain unclear. To investigate the role of 5-HT2c-Rs in the DRN in mice, we developed a melanopsin-based optogenetic probe for activation of Gq signals in cellular domains, where 5-HT2c-Rs are localized. Our results demonstrate that precise temporal control of Gq signals in 5-HT2c-R domains in GABAergic neurons upstream of 5-HT neurons provides negative feedback regulation of serotonergic firing to modulate anxiety-like behavior in mice. PMID:24733892

  14. 5-HT2C receptor involvement in the control of persistence in the reinforced spatial alternation animal model of obsessive-compulsive disorder.

    PubMed

    Papakosta, Vassiliki-Maria; Kalogerakou, Stamatina; Kontis, Dimitris; Anyfandi, Eleni; Theochari, Eirini; Boulougouris, Vasileios; Papadopoulos, Sokrates; Panagis, George; Tsaltas, Eleftheria

    2013-04-15

    The serotonergic system is implicated in the pathophysiology of obsessive-compulsive disorder (OCD). However, the distinct role of serotonin (5-HT) receptor subtypes remains unclear. This study investigates the contribution of 5-HT2A and 5-HT2C receptors in the modulation of persistence in the reinforced spatial alternation model of OCD. Male Wistar rats were assessed for spontaneous and pharmacologically induced (by m-chlorophenylpiperazine: mCPP) directional persistence in the reinforced alternation OCD model. Systemic administration of mCPP (non-specific 5-HT agonist, 2.5mg/kg), M100907 (selective 5-HT2A receptor antagonist, 0.08 mg/kg), SB242084 (selective 5-HT2C receptor antagonist, 0.5 mg/kg) and vehicle was used. Experiment 1 investigated M100907 and SB242084 effects in animals spontaneously exhibiting high and low persistence during the early stages of alternation training. Experiment 2 investigated M100900 and SB242084 effects on mCPP-induced persistence. Under the regime used in Experiment 1, 5-HT2A or 5-HT2C receptor antagonism did not affect spontaneous directional persistence in either high or low persistence groups. In Experiment 2, 5-HT2C but not 5-HT2A receptor antagonism significantly reduced, but did not abolish, mCPP-induced directional persistence. These findings suggest that 5-HT2C but not 5-HT2A receptors contribute to the modulation of mCPP-induced persistent behaviour, raising the possibility that the use of 5-HT2C antagonists may have a therapeutic value in OCD. Copyright © 2013 Elsevier B.V. All rights reserved.

  15. Immunodetection of T-2 Metabolites in Rat Urines after Dermal, Oral, or Intramuscular Exposure to T-2 Toxin

    DTIC Science & Technology

    1988-07-25

    laboratory studies, but not useful for evaluation of natural exposure. Several conventional immunoassay techniques based on tritiated ligands (Fontelo. et...1978. Natural occurrence of trichothecenes (nivaienol, deoxynivalenol, T-2) and zearalenone in corn. ExperiLentia 34, 1333-1334. LEE. S. and CHU, F.S

  16. Structure-activity relationship of 5-chloro-2-methyl-3-(1,2,3,6-tetrahydropyridin-4-yl)-1H-indole analogues as 5-HT(6) receptor agonists.

    PubMed

    Mattsson, Cecilia; Svensson, Peder; Boettcher, Henning; Sonesson, Clas

    2013-05-01

    To further investigate the structure-activity relationship (SAR) of the 5-hydroxytryptamine type 6 (5-HT6) receptor agonist 5-chloro-2-methyl-3-(1,2,3,6-tetrahydropyridin-4-yl)-1H-indole (EMD386088, 6), a series of 2-methyl-3-(1,2,3,6-tetrahydropyridin-4-yl)-1H-indoles were synthesized, and in vitro affinity to, and functional activity at 5-HT6 receptors was tested. We focused on substituents made at the indole N(1)-, 2- and 5-positions and these were found to not only influence the affinity at 5-HT6 receptors but also the intrinsic activity leading to antagonists, partial agonists and full agonists. In order for a compound to demonstrate potent 5-HT6 receptor agonist properties, the indole N(1) should be unsubstituted, an alkyl group such as 2-methyl is needed and finally halogen substituents in the indole 5-position (fluoro, chloro or, bromo) were essential requirements. However, the introduction of a benzenesulfonyl group at N(1)-position switched the full agonist 6 to be a 5-HT6 receptor antagonist (30). A few compounds within the 2-methyl-3-(1,2,3,6-tetrahydropyridin-4-yl)-1H-indoles were also screened for off-targets and generally they displayed low affinity for other 5-HT subtypes and serotonin transporter protein (SERT). Copyright © 2013 Elsevier Masson SAS. All rights reserved.

  17. MDMA-Induced Dissociative State not Mediated by the 5-HT2A Receptor.

    PubMed

    Puxty, Drew J; Ramaekers, Johannes G; de la Torre, Rafael; Farré, Magí; Pizarro, Neus; Pujadas, Mitona; Kuypers, Kim P C

    2017-01-01

    Previous research has shown that a single dose of MDMA induce a dissociative state, by elevating feelings of depersonalization and derealization. Typically, it is assumed that action on the 5-HT 2A receptor is the mechanism underlying these psychedelic experiences. In addition, other studies have shown associations between dissociative states and biological parameters (heart rate, cortisol), which are elevated by MDMA. In order to investigate the role of the 5-HT 2 receptor in the MDMA-induced dissociative state and the association with biological parameters, a placebo-controlled within-subject study was conducted including a single oral dose of MDMA (75 mg), combined with placebo or a single oral dose of the 5-HT 2 receptor blocker ketanserin (40 mg). Twenty healthy recreational MDMA users filled out a dissociative states scale (CADSS) 90 min after treatments, which was preceded and followed by assessment of a number of biological parameters (cortisol levels, heart rate, MDMA blood concentrations). Findings showed that MDMA induced a dissociative state but this effect was not counteracted by pre-treatment with ketanserin. Heart rate was the only biological parameter that correlated with the MDMA-induced dissociative state, but an absence of correlation between these measures when participants were pretreated with ketanserin suggests an absence of directional effects of heart rate on dissociative state. It is suggested that the 5-HT 2 receptor does not mediate the dissociative effects caused by a single dose of MDMA. Further research is needed to determine the exact neurobiology underlying this effect and whether these effects contribute to the therapeutic potential of MDMA.

  18. Pharmacological Properties and Discriminative Stimulus Effects of a Novel and Selective 5-HT2 Receptor Agonist AL-38022A [(S)-2-(8,9-dihydro-7H-pyrano[2,3-g]indazol-1-yl)-1-methylethylamine

    PubMed Central

    May, Jesse A.; Sharif, Najam A.; Chen, Hwang-Hsing; Liao, John C.; Kelly, Curtis R.; Glennon, Richard A.; Young, Richard; Li, Jun-Xu; Rice, Kenner C.; France, Charles P.

    2013-01-01

    AL-38022A is a novel synthetic serotonergic (5-HT) ligand that exhibited high affinity for each of the 5-HT2 receptor subtypes (Ki ≤ 2.2 nM), but a significantly lower (>100-fold less) affinity for other 5-HT receptors. In addition, AL-38022A displayed a very low affinity for a broad array of other receptors, neurotransmitter transport sites, ion channels, and second messenger elements, making it a relatively selective agent. AL-38022A potently stimulated functional responses via native and cloned rat (EC50 range: 1.9 – 22.5 nM) and human (EC50 range: 0.5 – 2.2 nM) 5-HT2 receptor subtypes including [Ca2+]i mobilization and tissue contractions with apparently similar potencies and intrinsic activities and was a full agonist at all 5-HT2 receptor subtypes. The CNS activity of AL-38022A was assessed by evaluating its discriminative stimulus effects in both a rat and a monkey drug discrimination paradigm using DOM as the training drug. AL-38022A fully generalized to the DOM stimulus in each of these studies; in monkeys MDL 100907 antagonized both DOM and AL-38022A. The pharmacological profile of AL-38022A suggests that it could be a useful tool in defining 5-HT2 receptor signaling and receptor characterization where 5-HT may function as a neurotransmitter. PMID:18718483

  19. Comparisons of Native Shiga Toxins (Stxs) Type 1 and 2 with Chimeric Toxins Indicate that the Source of the Binding Subunit Dictates Degree of Toxicity

    PubMed Central

    Russo, Lisa M.; Melton-Celsa, Angela R.; Smith, Michael J.; O'Brien, Alison D.

    2014-01-01

    Shiga toxin (Stx)-producing E. coli (STEC) cause food-borne outbreaks of hemorrhagic colitis. The main virulence factor expressed by STEC, Stx, is an AB5 toxin that has two antigenically distinct forms, Stx1a and Stx2a. Although Stx1a and Stx2a bind to the same receptor, globotriaosylceramide (Gb3), Stx2a is more potent than Stx1a in mice, whereas Stx1a is more cytotoxic than Stx2a in cell culture. In this study, we used chimeric toxins to ask what the relative contribution of individual Stx subunits is to the differential toxicity of Stx1a and Stx2a in vitro and in vivo. Chimeric stx1/stx2 operons were generated by PCR such that the coding regions for the A2 and B subunits of one toxin were combined with the coding region for the A1 subunit of the heterologous toxin. The toxicities of purified Stx1a, Stx2a, and the chimeric Stxs were determined on Vero and HCT-8 cell lines, while polarized HCT-8 cell monolayers grown on permeable supports were used to follow toxin translocation. In all in vitro assays, the activity of the chimeric toxin correlated with that of the parental toxin from which the B subunit originated. The origin of the native B subunit also dictated the 50% lethal dose of toxin after intraperitoneal intoxication of mice; however, the chimeric Stxs exhibited reduced oral toxicity and pH stability compared to Stx1a and Stx2a. Taken together, these data support the hypothesis that the differential toxicity of the chimeric toxins for cells and mice is determined by the origin of the B subunit. PMID:24671194

  20. Activation of serotonin 5-HT2C receptor suppresses behavioral sensitization and naloxone-precipitated withdrawal symptoms in morphine-dependent mice

    PubMed Central

    Zhang, Gongliang; Wu, Xian; Zhang, Yong-Mei; Liu, Huan; Jiang, Qin; Pang, Gang; Tao, Xinrong; Dong, Liuyi; Stackman, Robert W.

    2015-01-01

    Opioid abuse and dependence have evolved into an international epidemic as a significant clinical and social problem with devastating consequences. Repeated exposure to the opioid, for example morphine, can induce profound, long-lasting behavioral sensitization and physical dependence, which are thought to reflect neuroplasticity in neural circuitry. Central serotonin (5-HT) neurotransmission participates in the development of dependence on and the expression of withdrawal from morphine. Serotonin 5-HT2C receptor (5-HT2CR) agonists suppress psychostimulant nicotine or cocaine-induced behavioral sensitization and drug-seeking behavior; however, the impact of 5-HT2CR agonists on behaviors relevant to opioid abuse and dependence has not been reported. In the present study, the effects of 5-HT2CR activation on the behavioral sensitization and naloxone-precipitated withdrawal symptoms were examined in mice underwent repeated exposure to morphine. Male mice received morphine (10 mg/kg, s.c.) to develop behavioral sensitization. Lorcaserin, a 5-HT2CR agonist, prevented the induction and expression, but not the development, of morphine-induced behavioral sensitization. Another cohort of mice received increasing doses of morphine over a 7-day period to induce morphine-dependence. Pretreatment of lorcaserin, or the positive control clonidine (an alpha 2-adrenoceptor agonist), ameliorated the naloxone-precipitated withdrawal symptoms. SB 242084, a selective 5-HT2CR antagonist, prevented the lorcaserin-mediated suppression of behavioral sensitization and withdrawal. Chronic morphine treatment was associated with an increase in the expression of 5-HT2CR protein in the ventral tegmental area, locus coeruleus and nucleus accumbens. These findings suggest that 5-HT2CR can modulate behavioral sensitization and withdrawal in morphine-dependent mice, and the activation of 5-HT2CR may represent a new avenue for the treatment of opioid addiction. PMID:26432939

  1. Structural constraints-based evaluation of immunogenic avirulent toxins from Clostridium botulinum C2 and C3 toxins as subunit vaccines.

    PubMed

    Prisilla, A; Prathiviraj, R; Sasikala, R; Chellapandi, P

    2016-10-01

    Clostridium botulinum (group-III) is an anaerobic bacterium producing C2 and C3 toxins in addition to botulinum neurotoxins in avian and mammalian cells. C2 and C3 toxins are members of bacterial ADP-ribosyltransferase superfamily, which modify the eukaryotic cell surface proteins by ADP-ribosylation reaction. Herein, the mutant proteins with lack of catalytic and pore forming function derived from C2 (C2I and C2II) and C3 toxins were computationally evaluated to understand their structure-function integrity. We have chosen many structural constraints including local structural environment, folding process, backbone conformation, conformational dynamic sub-space, NAD-binding specificity and antigenic determinants for screening of suitable avirulent toxins. A total of 20 avirulent mutants were identified out of 23 mutants, which were experimentally produced by site-directed mutagenesis. No changes in secondary structural elements in particular to α-helices and β-sheets and also in fold rate of all-β classes. Structural stability was maintained by reordered hydrophobic and hydrogen bonding patterns. Molecular dynamic studies suggested that coupled mutations may restrain the binding affinity to NAD(+) or protein substrate upon structural destabilization. Avirulent toxins of this study have stable energetic backbone conformation with a common blue print of folding process. Molecular docking studies revealed that avirulent mutants formed more favorable hydrogen bonding with the side-chain of amino acids near to conserved NAD-binding core, despite of restraining NAD-binding specificity. Thus, structural constraints in the avirulent toxins would determine their immunogenic nature for the prioritization of protein-based subunit vaccine/immunogens to avian and veterinary animals infected with C. botulinum. Copyright © 2016 Elsevier B.V. All rights reserved.

  2. Novel 5-HT6 receptor antagonists/D2 receptor partial agonists targeting behavioral and psychological symptoms of dementia.

    PubMed

    Kołaczkowski, Marcin; Marcinkowska, Monika; Bucki, Adam; Śniecikowska, Joanna; Pawłowski, Maciej; Kazek, Grzegorz; Siwek, Agata; Jastrzębska-Więsek, Magdalena; Partyka, Anna; Wasik, Anna; Wesołowska, Anna; Mierzejewski, Paweł; Bienkowski, Przemyslaw

    2015-03-06

    We describe a novel class of designed multiple ligands (DMLs) combining serotonin 5-HT6 receptor (5-HT6R) antagonism with dopamine D2 receptor (D2R) partial agonism. Prototype hybrid molecules were designed using docking to receptor homology models. Diverse pharmacophore moieties yielded 3 series of hybrids with varying in vitro properties at 5-HT6R and D2R, and at M1 receptor and hERG channel antitargets. 4-(piperazin-1-yl)-1H-indole derivatives showed highest antagonist potency at 5-HT6R, with 7-butoxy-3,4-dihydroquinolin-2(1H)-one and 2-propoxybenzamide derivatives having promising D2R partial agonism. 2-(3-(4-(1-(phenylsulfonyl)-1H-indol-4-yl)piperazin-1-yl)propoxy)benzamide (47) exhibited nanomolar affinity at both 5-HT6R and D2R and was evaluated in rat models. It displayed potent antidepressant-like and anxiolytic-like activity in the Porsolt and Vogel tests, respectively, more pronounced than that of a reference selective 5-HT6R antagonist or D2R partial agonist. In addition, 47 also showed antidepressant-like activity (Porsolt's test) and anxiolytic-like activity (open field test) in aged (>18-month old) rats. In operant conditioning tests, 47 enhanced responding for sweet reward in the saccharin self-administration test, consistent with anti-anhedonic properties. Further, 47 facilitated extinction of non-reinforced responding for sweet reward, suggesting potential procognitive activity. Taken together, these studies suggest that DMLs combining 5-HT6R antagonism and D2R partial agonism may successfully target affective disorders in patients from different age groups without a risk of cognitive deficits. Copyright © 2014 Elsevier Masson SAS. All rights reserved.

  3. Do imipramine and dihydroergosine possess two components - one stimulating 5-HT sub 1 and the other inhibiting 5-HT sub 2 receptors

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Pericic, D.; Mueck-Seler, D.

    1990-01-01

    The mechanisms by which imipramine and dihydroergosine stimulate the 5-HT syndrome in rats and inhibit the head-twitch response in rats and mice were studied. Imipramine- and dihydroergosine-included stimulation of the 5-HT syndrome was inhibited stereoselectively by propranolol, a high affinity ligand for 5-HT{sub 1} receptor sites, but not by ritanserin, a specific 5-HT{sub 2} receptor antagonist. (-) -Propranolol potentiated the inhibitory effect of imipramine, but not of dihydroergosine on the head-twitch response, while ritanserin was without effect. As expected, 8-OH-DPAT, a selective 5-HT{sub 1A} receptor agonist, stimulated, and 5-HT{sub 1B} agonists CGS 12066B and 1-(trifluoromethylphenyl) piperazine (TFMPP) failed to stimulatemore » the 5-HT syndrome induced in rats by pargyline and 5-HTP administration. A higher dose of ritanserin inhibited the syndrome. While 8-OH-DPAT alone produced all behavioral components of the 5-HT syndrome, dihydroergosine or imipramine alone even at very high doses never produced tremor or a more intensive forepaw padding as seen when these drugs were given in combination with pargyline and 5-HTP. A single administration of (-)-propranolol also inhibited the head-twitch response. This effect lasted in mice longer that after ritanserin administration. In in vitro experiments dihydroergosine expressed approximately twenty-fold higher affinity for {sup 3}H-ketanserin binding sites than imipramine.« less

  4. Molecular dynamics simulations and docking studies on 3D models of the heterodimeric and homodimeric 5-HT(2A) receptor subtype.

    PubMed

    Bruno, Agostino; Beato, Claudia; Costantino, Gabriele

    2011-04-01

    G-protein coupled receptors may exist as functional homodimers, heterodimers and even as higher aggregates. In this work, we investigate the 5-HT(2A) receptor, which is a known target for antipsychotic drugs. Recently, 5-HT(2A) has been shown to form functional homodimers and heterodimers with the mGluR2 receptor. The objective of this study is to build up 3D models of the 5-HT(2A)/mGluR2 heterodimer and of the 5-HT(2A)-5-HT(2A) homodimer, and to evaluate the impact of the dimerization interface on the shape of the 5-HT(2A) binding pocket by using molecular dynamics simulations and docking studies. The heterodimer, homodimer and monomeric 5-HT(2A) receptors were simulated by molecular dynamics for 40 ns each. The trajectories were clustered and representative structures of six clusters for each system were generated. Inspection of the these representative structures clearly indicate an effect of the dimerization interface on the topology of the binding pocket. Docking studies allowed to generate receiver operating characteristic curves for a set of 5-HT(2A) ligands, indicating that different complexes prefer different classes of 5-HT(2A) ligands. This study clearly indicates that the presence of a dimerization interface must explicitly be considered when studying G-protein coupled receptors known to exist as dimers. Molecular dynamics simulation and cluster analysis are appropriate tools to study the phenomenon.

  5. Tph2 gene deletion enhances amphetamine-induced hypermotility: effect of 5-HT restoration and role of striatal noradrenaline release.

    PubMed

    Carli, Mirjana; Kostoula, Chrysaugi; Sacchetti, Giuseppina; Mainolfi, Pierangela; Anastasia, Alessia; Villani, Claudia; Invernizzi, Roberto William

    2015-11-01

    Variants of tryptophan hydroxylase-2 (Tph2), the gene encoding enzyme responsible for the synthesis of brain serotonin (5-HT), have been associated with neuropsychiatric disorders, substance abuse and addiction. This study assessed the effect of Tph2 gene deletion on motor behavior and found that motor activity induced by 2.5 and 5 mg/kg amphetamine was enhanced in Tph2(-/-) mice. Using the in vivo microdialysis technique we found that the ability of amphetamine to stimulate noradrenaline (NA) release in the striatum was reduced by about 50% in Tph2(-/-) mice while the release of dopamine (DA) was not affected. Tph2 deletion did not affect the release of NA and DA in the prefrontal cortex. The role of endogenous 5-HT in enhancing the effect of amphetamine was confirmed showing that treatment with the 5-HT precursor 5-hydroxytryptophan (10 mg/kg) restored tissue and extracellular levels of brain 5-HT and the effects of amphetamine on striatal NA release and motor activity in Tph2(-/-) mice. Treatment with the NA precursor dihydroxyphenylserine (400 mg/kg) was sufficient to restore the effect of amphetamine on striatal NA release and motor activity in Tph2(-/-) mice. These findings indicate that amphetamine-induced hyperactivity is attenuated by endogenous 5-HT through the inhibition of striatal NA release. Tph2(-/-) mice may be a useful preclinical model to assess the role of 5-HT-dependent mechanisms in the action of psychostimulants. Acute sensitivity to the motor effects of amphetamine has been associated to increased risk of psychostimulant abuse. Here, we show that deletion of Tph2, the gene responsible for brain 5-HT synthesis, enhances the motor effect of amphetamine in mice through the inhibition of striatal NA release. This suggests that Tph2(-/-) mice is a useful preclinical model to assess the role of 5-HT-dependent mechanisms in psychostimulants action. Tph2, tryptophan hydroxylase-2. © 2015 International Society for Neurochemistry.

  6. Deoxynivalenol, zearalenone and T-2 in grain based swine feed in Hungary.

    PubMed

    Tima, Helga; Rácz, Anita; Guld, Zsuzsanna; Mohácsi-Farkas, Csilla; Kiskó, Gabriella

    2016-12-01

    Fusarium genera can produce trichothecenes like deoxynivalenol (DON), zearalenone (ZEN) and T-2 toxin, which can occur in feed cereal grains. Enzyme-linked immunosorbent assays (ELISA) tests of different Hungarian swine feedstuff proved that these mycotoxins were present. In this survey, 45 feed samples from 3 significant Hungarian swine feedstuff manufacturers were tested. ELISA methodology validation showed mean recovery rates in ranges from 85.3% to 98.1%, with intermediate precision of 86.9-96.9% and variation coefficients of 3.4-5.7% and 5.9-7.1%, respectively. The results showed that among Fusarium toxins, generally DON was present in the highest concentration, followed by T-2 and finally ZEN in all tested swine feeds. Each of the mycotoxins was found above the limit of detection in all swine feedstuffs. Boars feed's DON (average ± standard deviation was 872 ± 139 µg kg -1 ) and ZEN (172 ± 18 µg kg -1 ) results of one of the manufacturers were above the guidance values. It indicates the necessity for efficient monitoring of DON, ZEN and T-2 mycotoxins in swine feeds.

  7. Serotonin 5-HT2 Receptor Interactions with Dopamine Function: Implications for Therapeutics in Cocaine Use Disorder

    PubMed Central

    Cunningham, Kathryn A.

    2015-01-01

    Cocaine exhibits prominent abuse liability, and chronic abuse can result in cocaine use disorder with significant morbidity. Major advances have been made in delineating neurobiological mechanisms of cocaine abuse; however, effective medications to treat cocaine use disorder remain to be discovered. The present review will focus on the role of serotonin (5-HT; 5-hydroxytryptamine) neurotransmission in the neuropharmacology of cocaine and related abused stimulants. Extensive research suggests that the primary contribution of 5-HT to cocaine addiction is a consequence of interactions with dopamine (DA) neurotransmission. The literature on the neurobiological and behavioral effects of cocaine is well developed, so the focus of the review will be on cocaine with inferences made about other monoamine uptake inhibitors and releasers based on mechanistic considerations. 5-HT receptors are widely expressed throughout the brain, and several different 5-HT receptor subtypes have been implicated in mediating the effects of endogenous 5-HT on DA. However, the 5-HT2A and 5-HT2C receptors in particular have been implicated as likely candidates for mediating the influence of 5-HT in cocaine abuse as well as to traits (e.g., impulsivity) that contribute to the development of cocaine use disorder and relapse in humans. Lastly, new approaches are proposed to guide targeted development of serotonergic ligands for the treatment of cocaine use disorder. PMID:25505168

  8. MDMA-Induced Dissociative State not Mediated by the 5-HT2A Receptor

    PubMed Central

    Puxty, Drew J.; Ramaekers, Johannes G.; de la Torre, Rafael; Farré, Magí; Pizarro, Neus; Pujadas, Mitona; Kuypers, Kim P. C.

    2017-01-01

    Previous research has shown that a single dose of MDMA induce a dissociative state, by elevating feelings of depersonalization and derealization. Typically, it is assumed that action on the 5-HT2A receptor is the mechanism underlying these psychedelic experiences. In addition, other studies have shown associations between dissociative states and biological parameters (heart rate, cortisol), which are elevated by MDMA. In order to investigate the role of the 5-HT2 receptor in the MDMA-induced dissociative state and the association with biological parameters, a placebo-controlled within-subject study was conducted including a single oral dose of MDMA (75 mg), combined with placebo or a single oral dose of the 5-HT2 receptor blocker ketanserin (40 mg). Twenty healthy recreational MDMA users filled out a dissociative states scale (CADSS) 90 min after treatments, which was preceded and followed by assessment of a number of biological parameters (cortisol levels, heart rate, MDMA blood concentrations). Findings showed that MDMA induced a dissociative state but this effect was not counteracted by pre-treatment with ketanserin. Heart rate was the only biological parameter that correlated with the MDMA-induced dissociative state, but an absence of correlation between these measures when participants were pretreated with ketanserin suggests an absence of directional effects of heart rate on dissociative state. It is suggested that the 5-HT2 receptor does not mediate the dissociative effects caused by a single dose of MDMA. Further research is needed to determine the exact neurobiology underlying this effect and whether these effects contribute to the therapeutic potential of MDMA. PMID:28744219

  9. Antibodies derived from a toxoid MEFA (multiepitope fusion antigen) show neutralizing activities against heat-labile toxin (LT), heat-stable toxins (STa, STb), and Shiga toxin 2e (Stx2e) of porcine enterotoxigenic Escherichia coli (ETEC).

    PubMed

    Rausch, Dana; Ruan, Xiaosai; Nandre, Rahul; Duan, Qiangde; Hashish, Emad; Casey, Thomas A; Zhang, Weiping

    2017-04-01

    Enterotoxigenic Escherichia coli (ETEC) strains are the main cause of diarrhea in pigs. Pig diarrhea especially post-weaning diarrhea remains one of the most important swine diseases. ETEC bacterial fimbriae including K88, F18, 987P, K99 and F41 promote bacterial attachment to intestinal epithelial cells and facilitate ETEC colonization in pig small intestine. ETEC enterotoxins including heat-labile toxin (LT) and heat-stable toxins type Ia (porcine-type STa) and type II (STb) stimulate fluid hyper-secretion, leading to watery diarrhea. Blocking bacteria colonization and/or neutralizing enterotoxicity of ETEC toxins are considered effective prevention against ETEC diarrhea. In this study, we applied the MEFA (multiepitope fusion antigen) strategy to create toxoid MEFAs that carried antigenic elements of ETEC toxins, and examined for broad antitoxin immunogenicity in a murine model. By embedding STa toxoid STa P12F (NTFYCCELCCNFACAGCY), a STb epitope (KKDLCEHY), and an epitope of Stx2e A subunit (QSYVSSLN) into the A1 peptide of a monomeric LT toxoid (LT R192G ), two toxoid MEFAs, 'LT R192G -STb-Stx2e-STa P12F ' and 'LT R192G -STb-Stx2e-3xSTa P12F ' which carried three copies of STa P12F , were constructed. Mice intraperitoneally immunized with each toxoid MEFA developed IgG antibodies to all four toxins. Induced antibodies showed in vitro neutralizing activities against LT, STa, STb and Stx2e toxins. Moreover, suckling piglets born by a gilt immunized with 'LT R192G -STb-Stx2e-3xSTa P12F ' were protected when challenged with ETEC strains, whereas piglets born by a control gilt developed diarrhea. Results from this study showed that the toxoid MEFA induced broadly antitoxin antibodies, and suggested potential application of the toxoid MEFA for developing a broad-spectrum vaccine against ETEC diarrhea in pigs. Copyright © 2016 Elsevier B.V. All rights reserved.

  10. Molecular Dynamics Simulation Reveals Specific Interaction Sites between Scorpion Toxins and Kv1.2 Channel: Implications for Design of Highly Selective Drugs

    PubMed Central

    Yuan, Shouli; Gao, Bin

    2017-01-01

    The Kv1.2 channel plays an important role in the maintenance of resting membrane potential and the regulation of the cellular excitability of neurons, whose silencing or mutations can elicit neuropathic pain or neurological diseases (e.g., epilepsy and ataxia). Scorpion venom contains a variety of peptide toxins targeting the pore region of this channel. Despite a large amount of structural and functional data currently available, their detailed interaction modes are poorly understood. In this work, we choose four Kv1.2-targeted scorpion toxins (Margatoxin, Agitoxin-2, OsK-1, and Mesomartoxin) to construct their complexes with Kv1.2 based on the experimental structure of ChTx-Kv1.2. Molecular dynamics simulation of these complexes lead to the identification of hydrophobic patches, hydrogen-bonds, and salt bridges as three essential forces mediating the interactions between this channel and the toxins, in which four Kv1.2-specific interacting amino acids (D353, Q358, V381, and T383) are identified for the first time. This discovery might help design highly selective Kv1.2-channel inhibitors by altering amino acids of these toxins binding to the four channel residues. Finally, our results provide new evidence in favor of an induced fit model between scorpion toxins and K+ channel interactions. PMID:29104247

  11. Chemical synthesis, 3D structure, and ASIC binding site of the toxin mambalgin-2.

    PubMed

    Schroeder, Christina I; Rash, Lachlan D; Vila-Farrés, Xavier; Rosengren, K Johan; Mobli, Mehdi; King, Glenn F; Alewood, Paul F; Craik, David J; Durek, Thomas

    2014-01-20

    Mambalgins are a novel class of snake venom components that exert potent analgesic effects mediated through the inhibition of acid-sensing ion channels (ASICs). The 57-residue polypeptide mambalgin-2 (Ma-2) was synthesized by using a combination of solid-phase peptide synthesis and native chemical ligation. The structure of the synthetic toxin, determined using homonuclear NMR, revealed an unusual three-finger toxin fold reminiscent of functionally unrelated snake toxins. Electrophysiological analysis of Ma-2 on wild-type and mutant ASIC1a receptors allowed us to identify α-helix 5, which borders on the functionally critical acidic pocket of the channel, as a major part of the Ma-2 binding site. This region is also crucial for the interaction of ASIC1a with the spider toxin PcTx1, thus suggesting that the binding sites for these toxins substantially overlap. This work lays the foundation for structure-activity relationship (SAR) studies and further development of this promising analgesic peptide. Copyright © 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  12. Structure-5-HT/D2 Receptor Affinity Relationship in a New Group of 1-Arylpiperazynylalkyl Derivatives of 8-Dialkylamino-3,7-dimethyl-1H-purine-2,6(3H,7H)-dione.

    PubMed

    Żmudzki, Paweł; Satała, Grzegorz; Chłoń-Rzepa, Grażyna; Bojarski, Andrzej J; Kazek, Grzegorz; Siwek, Agata; Gryboś, Anna; Głuch-Lutwin, Monika; Wesołowska, Anna; Pawłowski, Maciej

    2016-10-01

    In our previous papers, we have reported that some 8-amino-1,3-dimethyl-1H-purine-2,6(3H,7H)-dione derivatives possessed high affinity and displayed agonistic, partial agonistic, or antagonistic activity for serotonin 5-HT 1A and dopamine D 2 receptors. In order to examine further the influence of the substituent in the position 8 of the purine moiety and the influence of the xanthine core on the affinity for serotonin 5-HT 1A , 5-HT 2A , 5-HT 6 , 5-HT 7 , and dopamine D 2 receptors, two series of 1-arylpiperazynylalkyl derivatives of 8-amino-3,7-dimethyl-1H-purine-2,6(3H,7H)-dione were synthesized. All the final compounds were investigated in in vitro competition binding experiments for the serotonin 5-HT 1A , 5-HT 2A , 5-HT 6 , 5-HT 7 , and dopamine D 2 receptors. The structure-affinity relationships for this group of compounds were discussed. For selected compounds, the functional assays for the 5-HT 1A and D 2 receptors were carried out. The results of the assays indicated that these groups of derivatives possessed antagonistic activity for 5-HT 1A receptors and agonistic, partial agonistic, or antagonistic activity for D 2 receptors. In total, 26 new compounds were synthesized, 20 of which were tested in in vitro binding experiments and 5 were tested in in vitro functional assays. © 2016 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  13. Simultaneous acquisition for T2 -T2 Exchange and T1 -T2 correlation NMR experiments

    NASA Astrophysics Data System (ADS)

    Montrazi, Elton T.; Lucas-Oliveira, Everton; Araujo-Ferreira, Arthur G.; Barsi-Andreeta, Mariane; Bonagamba, Tito J.

    2018-04-01

    The NMR measurements of longitudinal and transverse relaxation times and its multidimensional correlations provide useful information about molecular dynamics. However, these experiments are very time-consuming, and many researchers proposed faster experiments to reduce this issue. This paper presents a new way to simultaneously perform T2 -T2 Exchange and T1 -T2 correlation experiments by taking the advantage of the storage time and the two steps phase cycling used for running the relaxation exchange experiment. The data corresponding to each step is either summed or subtracted to produce the T2 -T2 and T1 -T2 data, enhancing the information obtained while maintaining the experiment duration. Comparing the results from this technique with traditional NMR experiments it was possible to validate the method.

  14. Serotonin hyperinnervation and upregulated 5-HT2A receptor expression and motor-stimulating function in nigrostriatal dopamine-deficient Pitx3 mutant mice.

    PubMed

    Li, Li; Qiu, Guozhen; Ding, Shengyuan; Zhou, Fu-Ming

    2013-01-23

    The striatum receives serotonin (5-hydroxytryptamine, 5-HT) innervation and expresses 5-HT2A receptors (5-HT2ARs) and other 5-HT receptors, raising the possibility that the striatal 5-HT system may undergo adaptive changes after chronic severe dopamine (DA) loss and contribute to the function and dysfunction of the striatum. Here we show that in transcription factor Pitx3 gene mutant mice with a selective, severe DA loss in the dorsal striatum mimicking the DA denervation in late Parkinson's disease (PD), both the 5-HT innervation and the 5-HT2AR mRNA expression were increased in the dorsal striatum. Functionally, while having no detectable motor effect in wild type mice, the 5-HT2R agonist 2,5-dimethoxy-4-iodoamphetamine increased both the baseline and l-dopa-induced normal ambulatory and dyskinetic movements in Pitx3 mutant mice, whereas the selective 5-HT2AR blocker volinanserin had the opposite effects. These results demonstrate that Pitx3 mutant mice are a convenient and valid mouse model to study the compensatory 5-HT upregulation following the loss of the nigrostriatal DA projection and that the upregulated 5-HT2AR function in the DA deficient dorsal striatum may enhance both normal and dyskinetic movements. Copyright © 2012 Elsevier B.V. All rights reserved.

  15. Maternal influenza viral infection causes schizophrenia-like alterations of 5-HT2A and mGlu2 receptors in the adult offspring

    PubMed Central

    Moreno, José L.; Kurita, Mitsumasa; Holloway, Terrell; López, Javier; Cadagan, Richard; Martínez-Sobrido, Luis; García-Sastre, Adolfo; González-Maeso, Javier

    2011-01-01

    Epidemiological studies indicate that maternal influenza viral infection increases the risk for schizophrenia in the adult offspring. The serotonin and glutamate systems are suspected in the etiology of schizophrenia, as well as in the mechanism of action of antipsychotic drugs. The effects of hallucinogens, such as psilocybin and mescaline, require the serotonin 5-HT2A receptor, and induce schizophrenia-like psychosis in humans. In addition, metabotropic glutamate receptor mGlu2/3 agonists show promise as a new treatment for schizophrenia. Here, we investigated the level of expression and behavioral function of 5-HT2A and mGlu2 receptors in a mouse model of maternal influenza viral infection. We show that spontaneous locomotor activity is diminished by maternal infection with the mouse-adapted influenza A/WSN/33 (H1N1) virus. The behavioral responses to hallucinogens and glutamate antipsychotics are both affected by maternal exposure to influenza virus, with increased head-twitch response to hallucinogens and diminished antipsychotic-like effect of the glutamate agonist. In frontal cortex of mice born to influenza virus-infected mothers, the 5-HT2A receptor is up-regulated and the mGlu2 receptor is down-regulated, an alteration that may be involved in the behavioral changes observed. Additionally, we find that the cortical 5-HT2A receptor-dependent signaling pathways are significantly altered in the offspring of infected mothers, showing higher c-fos, egr-1 and egr-2 expression in response to the hallucinogenic drug DOI. Identifying a biochemical alteration that parallels the behavioral changes observed in a mouse model of prenatal viral infection may facilitate targeting therapies for treatment and prevention of schizophrenia. PMID:21289196

  16. The recovery of 5-HT transporter and 5-HT immunoreactivity in injured rat spinal cord.

    PubMed

    Saruhashi, Yasuo; Matsusue, Yoshitaka; Fujimiya, Mineko

    2009-09-01

    Experimental spinal cord injury. To determine the role of serotonin (5-HT) and 5-HT transporter in recovery from spinal cord injury. We examined 5-HT and 5-HT transporter of spinal cord immunohistologically and assessed locomotor recovery after extradural compression at the thoracic (T8) spinal cord in 21 rats. Eighteen rats had laminectomy and spinal cord injury, while the remaining three rats received laminectomy only. All rats were evaluated every other day for 4 weeks, using a 0-14 point scale open field test. Extradural compression markedly reduced mean hindlimbs scores from 14 to 1.5 +/- 2.0 (mean +/- standard error of mean). The rats recovered apparently normal walking by 4 weeks. The animals were perfused with fixative 1-3 days, 1, 2 and 4 weeks (three rats in each) after a spinal cord injury. The 5-HT transporter immunohistological study revealed a marked reduction of 5-HT transporter-containing terminals by 1 day after injury. By 4 weeks after injury, 5-HT transporter immunoreactive terminals returned to the control level. The 5-HT immunohistological study revealed a reduction of 5-HT-containing terminals by 1 week after injury. By 4 weeks after injury, 5-HT immunoreactive fibers and terminals returned to the control level. We estimated the recovery of 5-HT transporter and 5-HT neural elements in lumbosacral ventral horn by ranking 5-HT transporter and 5-HT staining intensity and counting 5-HT and 5-HT transporter terminals. The return of 5-HT transporter and 5-HT immunoreactivity of the lumbosacral ventral horn correlated with locomotor recovery, while 5-HT transporter showed closer relationship with locomotor recovery than 5-HT. The presence of 5-HT transporter indicates that the 5-HT fibers certainly function. This study shows that return of the function of 5-HT fibers predict the time course and extent of locomotory recovery after thoracic spinal cord injury.

  17. Repeated lysergic acid diethylamide in an animal model of depression: Normalisation of learning behaviour and hippocampal serotonin 5-HT2 signalling.

    PubMed

    Buchborn, Tobias; Schröder, Helmut; Höllt, Volker; Grecksch, Gisela

    2014-06-01

    A re-balance of postsynaptic serotonin (5-HT) receptor signalling, with an increase in 5-HT1A and a decrease in 5-HT2A signalling, is a final common pathway multiple antidepressants share. Given that the 5-HT1A/2A agonist lysergic acid diethylamide (LSD), when repeatedly applied, selectively downregulates 5-HT2A, but not 5-HT1A receptors, one might expect LSD to similarly re-balance the postsynaptic 5-HT signalling. Challenging this idea, we use an animal model of depression specifically responding to repeated antidepressant treatment (olfactory bulbectomy), and test the antidepressant-like properties of repeated LSD treatment (0.13 mg/kg/d, 11 d). In line with former findings, we observe that bulbectomised rats show marked deficits in active avoidance learning. These deficits, similarly as we earlier noted with imipramine, are largely reversed by repeated LSD administration. Additionally, bulbectomised rats exhibit distinct anomalies of monoamine receptor signalling in hippocampus and/or frontal cortex; from these, only the hippocampal decrease in 5-HT2 related [(35)S]-GTP-gamma-S binding is normalised by LSD. Importantly, the sham-operated rats do not profit from LSD, and exhibit reduced hippocampal 5-HT2 signalling. As behavioural deficits after bulbectomy respond to agents classified as antidepressants only, we conclude that the effect of LSD in this model can be considered antidepressant-like, and discuss it in terms of a re-balance of hippocampal 5-HT2/5-HT1A signalling. © The Author(s) 2014.

  18. The serotonin hallucinogen 5-MeO-DMT alters cortico-thalamic activity in freely moving mice: Regionally-selective involvement of 5-HT1A and 5-HT2A receptors.

    PubMed

    Riga, Maurizio S; Lladó-Pelfort, Laia; Artigas, Francesc; Celada, Pau

    2017-12-06

    5-MeO-DMT is a natural hallucinogen acting as serotonin 5-HT 1A /5-HT 2A receptor agonist. Its ability to evoke hallucinations could be used to study the neurobiology of psychotic symptoms and to identify new treatment targets. Moreover, recent studies revealed the therapeutic potential of serotonin hallucinogens in treating mood and anxiety disorders. Our previous results in anesthetized animals show that 5-MeO-DMT alters cortical activity via 5-HT 1A and 5-HT 2A receptors. Here, we examined 5-MeO-DMT effects on oscillatory activity in prefrontal (PFC) and visual (V1) cortices, and in mediodorsal thalamus (MD) of freely-moving wild-type (WT) and 5-HT 2A -R knockout (KO2A) mice. We performed local field potential multi-recordings evaluating the power at different frequency bands and coherence between areas. We also examined the prevention of 5-MeO-DMT effects by the 5-HT 1A -R antagonist WAY-100635. 5-MeO-DMT affected oscillatory activity more in cortical than in thalamic areas. More marked effects were observed in delta power in V1 of KO2A mice. 5-MeO-DMT increased beta band coherence between all examined areas. In KO2A mice, WAY100635 prevented most of 5-MeO-DMT effects on oscillatory activity. The present results indicate that hallucinatory activity of 5-MeO-DMT is likely mediated by simultaneous alteration of prefrontal and visual activities. The prevention of these effects by WAY-100635 in KO2A mice supports the potential usefulness of 5-HT 1A receptor antagonists to treat visual hallucinations. 5-MeO-DMT effects on PFC theta activity and cortico-thalamic coherence may be related to its antidepressant activity. Copyright © 2017. Published by Elsevier Ltd.

  19. LSD and DOB: interaction with 5-HT2A receptors to inhibit NMDA receptor-mediated transmission in the rat prefrontal cortex.

    PubMed

    Arvanov, V L; Liang, X; Russo, A; Wang, R Y

    1999-09-01

    Both the phenethylamine hallucinogen (-)-1-2, 5-dimethoxy-4-bromophenyl-2-aminopropane (DOB), a selective serotonin 5-HT2A,2C receptor agonist, and the indoleamine hallucinogen D-lysergic acid diethylamide (LSD, which binds to 5-HT1A, 1B, 1D, 1E, 1F, 2A, 2C, 5, 6, 7, dopamine D1 and D2, and alpha1 and alpha2 adrenergic receptors), but not their non-hallucinogenic congeners, inhibited N-methyl-D-aspartate (NMDA)-induced inward current and NMDA receptor-mediated synaptic responses evoked by electrical stimulation of the forceps minor in pyramidal cells of the prefrontal cortical slices. The inhibitory effect of hallucinogens was mimicked by 5-HT in the presence of selective 5-HT1A and 5-HT3 receptor antagonists. The inhibitory action of DOB, LSD and 5-HT on the NMDA transmission was blocked by the 5-HT2A receptor antagonists R-(+)-alpha-(2, 3-dimethoxyphenil)-1-[4-fluorophenylethyl]-4-piperidineme thanol (M100907) and ketanserin. However, at low concentrations, when both LSD and DOB by themselves only partially depressed the NMDA response, they blocked the inhibitory effect of 5-HT, suggesting a partial agonist action. Whereas N-(4-aminobutyl)-5-chloro-2-naphthalenesulphonamide (W-7, a calmodulin antagonist) and N-[2-[[[3-(4'-chlorophenyl)- 2-propenyl]methylamino]methyl]phenyl]-N-(2-hydroxyethyl)-4'-methoxy-b enzenesulphonamide phosphate (KN-93, a Ca2+/CaM-KII inhibitor), but not the negative control 2-[N-4'methoxybenzenesulphonyl]amino-N-(4'-chlorophenyl)-2-propeny l-N -methylbenzylamine phosphate (KN-92), blocked the inhibitory action of LSD and DOB, the selective protein kinase C inhibitor chelerythrine was without any effect. We conclude that phenethylamine and indoleamine hallucinogens may exert their hallucinogenic effect by interacting with 5-HT2A receptors via a Ca2+/CaM-KII-dependent signal transduction pathway as partial agonists and modulating the NMDA receptors-mediated sensory, perceptual, affective and cognitive processes.

  20. Nucleus accumbens shell excitability is decreased by methamphetamine self-administration and increased by 5-HT2C receptor inverse agonism and agonism

    PubMed Central

    Graves, Steven M.; Clark, Mary J.; Traynor, John R.; Hu, Xiu-Ti; Napier, T. Celeste

    2014-01-01

    Methamphetamine profoundly increases brain monoamines and is a widely abused psychostimulant. The effects of methamphetamine self-administration on neuron function are not known for the nucleus accumbens, a brain region involved in addictive behaviors, including drug-seeking. One therapeutic target showing preclinical promise at attenuating psychostimulant-seeking is 5-HT2C receptors; however, the effects of 5-HT2C receptor ligands on neuronal physiology are unclear. 5-HT2C receptor agonism decreases psychostimulant-mediated behaviors, and the putative 5-HT2C receptor inverse agonist, SB 206553, attenuates methamphetamine-seeking in rats. To ascertain the effects of methamphetamine, and 5-HT2C receptor inverse agonism and agonism, on neuronal function in the nucleus accumbens, we evaluated methamphetamine, SB 206553, and the 5-HT2C receptor agonist and Ro 60-0175, on neuronal excitability within the accumbens shell subregion using whole-cell current-clamp recordings in forebrain slices ex vivo. We reveal that methamphetamine self-administration decreased generation of evoked action potentials. In contrast, SB 206553 and Ro 60-0175 increased evoked spiking, effects that were prevented by the 5-HT2C receptor antagonist, SB 242084. We also assessed signaling mechanisms engaged by 5-HT2C receptors, and determined that accumbal 5-HT2C receptors stimulated Gq, but not Gi/o. These findings demonstrate that methamphetamine-induced decreases in excitability of neurons within the nucleus accumbens shell were abrogated by both 5-HT2C inverse agonism and agonism, and this effect likely involved activation of Gq–mediated signaling pathways. PMID:25229719

  1. Casein kinase 2 (CK2) increases survivin expression via enhanced β-catenin–T cell factor/lymphoid enhancer binding factor-dependent transcription

    PubMed Central

    Tapia, J. C.; Torres, V. A.; Rodriguez, D. A.; Leyton, L.; Quest, A. F. G.

    2006-01-01

    Increased expression of casein kinase 2 (CK2) is associated with hyperproliferation and suppression of apoptosis in cancer. Mutations in the tumor suppressor APC (adenomatous polyposis coli) are frequent in colon cancer and often augment β-catenin–T cell factor (Tcf)/lymphoid enhancer binding factor (Lef)-dependent transcription of genes such as c-myc and cyclin-D1. CK2 has also been implicated recently in the regulation of β-catenin stability. To identify mechanisms by which CK2 promotes survival, effects of the specific CK2 inhibitors 4,5,6,7-tetrabromobenzotriazole (TBB) and 2-dimethylamino-4,5,6,7-tetrabromo-1H-benzimidazole were assessed. TBB and 2-dimethylamino-4,5,6,7-tetrabromo-1H-benzimidazole significantly decreased proliferation and increased apoptosis of HT29(US) colon cancer cells. RT-PCR and immunoblot analysis revealed that both inhibitors decreased survivin mRNA and protein levels in HT29(US) cells. Similar effects were observed with TBB in human DLD-1 and SW-480 colorectal cells as well as ZR-75 breast cancer cells and HEK-293T embryonic kidney cells. Expression of GFP–CK2α in HEK-293T cells resulted in β-catenin–Tcf/Lef-dependent up-regulation of survivin and increased resistance to anticancer drugs. Augmented β-catenin–Tcf/Lef-dependent transcription and resistance to apoptosis observed upon GFP–CK2α expression were abolished by TBB. Alternatively, HEK-293T cells expressing GFP–survivin were resistant to TBB-induced apoptosis. Finally, siRNA-mediated down-regulation of CK2α in HEK-293T cells coincided with reduced β-catenin and survivin levels. Taken together, these results suggest that CK2 kinase activity promotes survival by increasing survivin expression via β-catenin–Tcf/Lef-mediated transcription. Hence, selective CK2 inhibition or down-regulation in tumors may provide an attractive opportunity for the development of novel cancer therapies. PMID:17005722

  2. Glycan Encapsulated Gold Nanoparticles Selectively Inhibit Shiga Toxins 1 and 2

    PubMed Central

    Kulkarni, Ashish A.; Fuller-Schaefer, Cynthia; Korman, Henry; Weiss, Alison A.; Iyer, Suri S.

    2011-01-01

    Shiga toxins (Stx) released by Escherichia coli O157:H7 and Shigella dysentriae, cause life-threatening conditions that include hemolytic-uremic syndrome (HUS), kidney failure and neurological complications. Cellular entry is mediated by the B subunit of the AB5 toxin, which recognizes cell surface glycolipids present in lipid raft like structures. We developed gold glyconanoparticles that present a multivalent display similar to the cell surface glycolipids to compete for these toxins. These highly soluble glyconanoparticles were nontoxic to the Vero monkey kidney cell line and protected Vero cells from Stx-mediated toxicity in a dose dependent manner. The inhibition is highly dependent on the structure and density of the glycans; selective inhibition of Stx1 and the more clinically relevant Stx2 was achieved. Interestingly, natural variants of Stx2, Stx2c and Stx2d, possessing minimal amino acid variation in the receptor binding site of the B subunit or changes in the A subunit were not neutralized by either the Stx1- or Stx2-specific gold glyconanoparticles. Our results suggest that tailored glyconanoparticles that mimic the natural display of glycans in lipid rafts could serve as potential therapeutics for Stx1 and Stx2. However, a few amino acid changes in emerging Stx2 variants can change receptor specificity, and further research is needed to develop receptor mimics for the emerging variants of Stx2. PMID:20669970

  3. Diphtheria toxin treatment of Pet-1-Cre floxed diphtheria toxin receptor mice disrupts thermoregulation without affecting respiratory chemoreception.

    PubMed

    Cerpa, V; Gonzalez, A; Richerson, G B

    2014-10-24

    In genetically-modified Lmx1b(f/f/p) mice, selective deletion of LMX1B in Pet-1 expressing cells leads to failure of embryonic development of serotonin (5-HT) neurons. As adults, these mice have a decreased hypercapnic ventilatory response and abnormal thermoregulation. This mouse model has been valuable in defining the normal role of 5-HT neurons, but it is possible that developmental compensation reduces the severity of observed deficits. Here we studied mice genetically modified to express diphtheria toxin receptors (DTR) on Pet-1 expressing neurons (Pet-1-Cre/floxed DTR or Pet1/DTR mice). These mice developed with a normal complement of 5-HT neurons. As adults, systemic treatment with 2-35μg of diphtheria toxin (DT) reduced the number of tryptophan hydroxylase-immunoreactive (TpOH-ir) neurons in the raphe nuclei and ventrolateral medulla by 80%. There were no effects of DT on minute ventilation (VE) or the ventilatory response to hypercapnia or hypoxia. At an ambient temperature (TA) of 24°C, all Pet1/DTR mice dropped their body temperature (TB) below 35°C after DT treatment, but the latency was shorter in males than females (3.0±0.37 vs. 4.57±0.29days, respectively; p<0.001). One week after DT treatment, mice were challenged by dropping TA from 37°C to 24°C, which caused TB to decrease more in males than in females (29.7±0.31°C vs. 33.0±1.3°C, p<0.01). We conclude that the 20% of 5-HT neurons that remain after DT treatment in Pet1/DTR mice are sufficient to maintain normal baseline breathing and a normal response to CO2, while those affected include some essential for thermoregulation, in males more than females. In comparison to models with deficient embryonic development of 5-HT neurons, acute deletion of 5-HT neurons in adults leads to a greater defect in thermoregulation, suggesting that significant developmental compensation can occur. Copyright © 2014 IBRO. Published by Elsevier Ltd. All rights reserved.

  4. 5-hydroxytryptamine level and 5-HT2A receptor mRNA expression in the guinea pigs eyes with spectacle lens-induced myopia

    PubMed Central

    Yang, Ji-Wen; Xu, Yan-Chun; Sun, Lin; Tian, Xiao-Dan

    2010-01-01

    AIM To investigate 5-hydroxytryptamine (5-HT) function and 5-HT receptor 2A (5-HT2A) mRNA expression in the formation of lens-induced myopia (LIM). METHODS Lens-induced myopia construction method was applied to generate myopia on guinea pig right eye (LIM eye). RESULTS LIM eyes formed significant myopia with longer axial length. 5-HT level in retina, choroids and sclera from LIM eyes was significantly higher than that in control group. 5-HT2A mRNA expression was also significantly up-regulated. CONCLUSION Refraction lens could induce myopia in guinea pig and 5-HT may play an important role in the formation of myopia by binding with 5-HT2A receptor. PMID:22553578

  5. Structure and action of the binary C2 toxin from Clostridium botulinum.

    PubMed

    Schleberger, Christian; Hochmann, Henrike; Barth, Holger; Aktories, Klaus; Schulz, Georg E

    2006-12-08

    C2 toxin from Clostridium botulinum is composed of the enzyme component C2-I, which ADP-ribosylates actin, and the binding and translocation component C2-II, responsible for the interaction with eukaryotic cell receptors and the following endocytosis. Three C2-I crystal structures at resolutions of up to 1.75 A are presented together with a crystal structure of C2-II at an appreciably lower resolution and a model of the prepore formed by fragment C2-IIa. The C2-I structure was determined at pH 3.0 and at pH 6.1. The structural differences are small, indicating that C2-I does not unfold, even at a pH value as low as 3.0. The ADP-ribosyl transferase activity of C2-I was determined for alpha and beta/gamma-actin and related to that of Iota toxin and of mutant S361R of C2-I that introduced the arginine observed in Iota toxin. The substantial activity differences between alpha and beta/gamma-actin cannot be explained by the protein structures currently available. The structure of the transport component C2-II at pH 4.3 was established by molecular replacement using a model of the protective antigen of anthrax toxin at pH 6.0. The C-terminal receptor-binding domain of C2-II could not be located but was present in the crystals. It may be mobile. The relative orientation and positions of the four other domains of C2-II do not differ much from those of the protective antigen, indicating that no large conformational changes occur between pH 4.3 and pH 6.0. A model of the C2-IIa prepore structure was constructed based on the corresponding assembly of the protective antigen. It revealed a surprisingly large number of asparagine residues lining the pore. The interaction between C2-I and C2-IIa and the translocation of C2-I into the target cell are discussed.

  6. Paroxetine-induced reduction of sexual incentive motivation in female rats is not modified by 5-HT1B or 5-HT2C antagonists.

    PubMed

    Kaspersen, Helge; Agmo, Anders

    2012-03-01

    Clinical data show that paroxetine causes sexual dysfunction in a substantial proportion of women taking this compound. This work was conducted to determine whether chronic paroxetine reduces sexual incentive motivation in female rats and whether this compound can modify any aspect of paced mating. The role of the 5-HT(1B) and 5-HT(2C) receptors in any potential effects was also evaluated. Ovariectomized female rats were implanted with osmotic minipumps releasing 10 mg/kg per day of paroxetine or vehicle for 28 days. Tests for sexual incentive motivation and paced mating were performed just before implantation and at regular intervals thereafter. The females were primed with estradiol benzoate (25 μg/rat) and progesterone (1 mg/rat) before each of these tests. On days 25-27 of treatment, the females were injected with the 5-HT(1B) antagonist GR125,743 (5 mg/kg), the 5-HT(2C) antagonist SB206,553 (5 mg/kg) and vehicle in counterbalanced order. Preinjection time was 30 min. Paroxetine reduced sexual incentive motivation on day 20 of treatment without affecting any aspect of paced mating. None of the antagonists modified the inhibitory effect of paroxetine on sexual incentive motivation. In the group chronically treated with vehicle, SB206,553 reduced proceptive behaviors in the paced mating test. No other effect was obtained. The effects of paroxetine seen in female rats are similar to those observed in women, suggesting that disturbances of sexual incentive motivation in rats are predictive of sexual dysfunction in women. The 5-HT(1B) and 5-HT(2C) receptors do not seem to be of any importance for paroxetine's inhibitory effect.

  7. Yeast β-1,6-Glucan Is a Primary Target for the Saccharomyces cerevisiae K2 Toxin

    PubMed Central

    Lukša, Juliana; Podoliankaitė, Monika; Vepštaitė, Iglė; Strazdaitė-Žielienė, Živilė; Urbonavičius, Jaunius

    2015-01-01

    Certain Saccharomyces cerevisiae strains secrete different killer proteins of double-stranded-RNA origin. These proteins confer a growth advantage to their host by increasing its survival. K2 toxin affects the target cell by binding to the cell surface, disrupting the plasma membrane integrity, and inducing ion leakage. In this study, we determined that K2 toxin saturates the yeast cell surface receptors in 10 min. The apparent amount of K2 toxin, bound to a single cell of wild type yeast under saturating conditions, was estimated to be 435 to 460 molecules. It was found that an increased level of β-1,6-glucan directly correlates with the number of toxin molecules bound, thereby impacting the morphology and determining the fate of the yeast cell. We observed that the binding of K2 toxin to the yeast surface receptors proceeds in a similar manner as in case of the related K1 killer protein. It was demonstrated that the externally supplied pustulan, a poly-β-1,6-glucan, but not the glucans bearing other linkage types (such as laminarin, chitin, and pullulan) efficiently inhibits the K2 toxin killing activity. In addition, the analysis of toxin binding to the intact cells and spheroplasts confirmed that majority of K2 protein molecules attach to the β-1,6-glucan, rather than the plasma membrane-localized receptors. Taken together, our results reveal that β-1,6-glucan is a primary target of K2 toxin and is important for the execution of its killing property. PMID:25710965

  8. Genome-wide siRNA screen identifies UNC50 as a regulator of Shiga toxin 2 trafficking

    PubMed Central

    Iles, Lakesla R.; Bartholomeusz, Geoffrey

    2017-01-01

    Shiga toxins 1 and 2 (STx1 and STx2) undergo retrograde trafficking to reach the cytosol. Early endosome-to-Golgi transport allows the toxins to evade degradation in lysosomes. Targeting this trafficking step has therapeutic promise, but the mechanism of trafficking for the more potent toxin STx2 is unclear. To identify host factors required for early endosome-to-Golgi trafficking of STx2, we performed a viability-based genome-wide siRNA screen in HeLa cells. 564, 535, and 196 genes were found to be required for toxicity induced by STx1 only, STx2 only, and both toxins, respectively. We focused on validating endosome/Golgi-localized hits specific for STx2 and found that depletion of UNC50 blocked early endosome-to-Golgi trafficking and induced lysosomal degradation of STx2. UNC50 acted by recruiting GBF1, an ADP ribosylation factor–guanine nucleotide exchange factor (ARF-GEF), to the Golgi. These results provide new information about STx2 trafficking mechanisms and may advance efforts to generate therapeutically viable toxin-trafficking inhibitors. PMID:28883040

  9. Genome-wide siRNA screen identifies UNC50 as a regulator of Shiga toxin 2 trafficking.

    PubMed

    Selyunin, Andrey S; Iles, Lakesla R; Bartholomeusz, Geoffrey; Mukhopadhyay, Somshuvra

    2017-10-02

    Shiga toxins 1 and 2 (STx1 and STx2) undergo retrograde trafficking to reach the cytosol. Early endosome-to-Golgi transport allows the toxins to evade degradation in lysosomes. Targeting this trafficking step has therapeutic promise, but the mechanism of trafficking for the more potent toxin STx2 is unclear. To identify host factors required for early endosome-to-Golgi trafficking of STx2, we performed a viability-based genome-wide siRNA screen in HeLa cells. 564, 535, and 196 genes were found to be required for toxicity induced by STx1 only, STx2 only, and both toxins, respectively. We focused on validating endosome/Golgi-localized hits specific for STx2 and found that depletion of UNC50 blocked early endosome-to-Golgi trafficking and induced lysosomal degradation of STx2. UNC50 acted by recruiting GBF1, an ADP ribosylation factor-guanine nucleotide exchange factor (ARF-GEF), to the Golgi. These results provide new information about STx2 trafficking mechanisms and may advance efforts to generate therapeutically viable toxin-trafficking inhibitors. © 2017 Selyunin et al.

  10. Production of trichothecene mycotoxins by Australian Fusarium species.

    PubMed

    McLachlan, A; Shaw, K J; Hocking, A D; Pitt, J I; Nguyen, T H

    1992-01-01

    Australian isolates of Fusarium species were grown on potato dextrose agar. Trichothecenes produced by these species were extracted by ethyl acetate followed by methanol and a silica gel column was used to clean-up the extract. The extracted samples were derivatized by acetylation with trifluoroacetic anhydride and the derivatives analysed by gas chromatography/mass spectrometry (GC/MS). Multiple ion detection was used to trace ions characteristic of the trichothecenes expected to be present. Quantitation of those found was based on a known mass of pentabromophenol that was added as an internal standard. Eight species of Fusarium (nineteen strains) were surveyed, of which three species, F. acuminatum, F. equiseti and F. sporotrichioides, produced the trichothecenes scirpentriol, diacetoxyscirpenol, neosolaniol, HT-2 toxin, T-2 toxin, T-2 tetraol and deoxynivalenol. Wheat samples were inoculated with four different species of Fusarium, F. acuminatum, F. equiseti, F. graminearum and F. sporotrichioides, and in these samples diacetoxyscirpenol, neosolaniol, HT-2 toxin and T-2 toxin were found.

  11. PhcrTx2, a New Crab-Paralyzing Peptide Toxin from the Sea Anemone Phymanthus crucifer

    PubMed Central

    Garateix, Anoland; Salceda, Emilio; Zaharenko, André Junqueira; Pons, Tirso; Santos, Yúlica; Arreguín, Roberto; Ständker, Ludger; Forssmann, Wolf-Georg; Tytgat, Jan; Vega, Rosario

    2018-01-01

    Sea anemones produce proteinaceous toxins for predation and defense, including peptide toxins that act on a large variety of ion channels of pharmacological and biomedical interest. Phymanthus crucifer is commonly found in the Caribbean Sea; however, the chemical structure and biological activity of its toxins remain unknown, with the exception of PhcrTx1, an acid-sensing ion channel (ASIC) inhibitor. Therefore, in the present work, we focused on the isolation and characterization of new P. crucifer toxins by chromatographic fractionation, followed by a toxicity screening on crabs, an evaluation of ion channels, and sequence analysis. Five groups of toxic chromatographic fractions were found, and a new paralyzing toxin was purified and named PhcrTx2. The toxin inhibited glutamate-gated currents in snail neurons (maximum inhibition of 35%, IC50 4.7 µM), and displayed little or no influence on voltage-sensitive sodium/potassium channels in snail and rat dorsal root ganglion (DRG) neurons, nor on a variety of cloned voltage-gated ion channels. The toxin sequence was fully elucidated by Edman degradation. PhcrTx2 is a new β-defensin-fold peptide that shares a sequence similarity to type 3 potassium channels toxins. However, its low activity on the evaluated ion channels suggests that its molecular target remains unknown. PhcrTx2 is the first known paralyzing toxin in the family Phymanthidae. PMID:29414882

  12. In vitro and in vivo binding of (E)- and (Z)-N-(iodoallyl)spiperone to dopamine D sub 2 and serotonin 5-HT sub 2 neuroreceptors

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Lever, J.R.; Scheffel, U.A.; Stathis, M.

    1990-01-01

    Apparent affinities (K{sub i}) of (E)- and (Z)-N-(iodoallyl)spiperone ((E)- and (Z)- NIASP) for dopamine D{sub 2} and serotonin 5-HT{sub 2} receptors were determined in competition binding assays. (Z)-NIASP (K{sub i} 0.35 nM, D{sub 2}; K{sub i} 1.75 nM, 5-HT{sub 2}) proved slightly more potent and selective for D{sub 2} sites in vitro than (E)-NIASP (K{sub i} 0.72 nM, D{sub 2}; K{sub i} 1.14 nM, 5-HT{sub 2}). In vivo, radioiodinated (E)- and (Z)-({sup 125}I)-NIASP showed regional distributions in mouse brain which are consonant with prolonged binding to dopamine D{sub 2} receptors accompanied by a minor serotonergic component of shorter duration. Stereoselective,more » dose-dependent blockade of (E)-({sup 125}I)-NIASP uptake was found for drugs binding to dopamine D{sub 2} sites, while drugs selective for serotonin 5-HT{sub 2}, {alpha}{sub 1}-adrenergic and dopamine D{sub 1} receptors did not inhibit radioligand binding 2 hr postinjection. Specific binding in striatal tissue was essentially irreversible over the time course of the study, and (E)-({sup 125}I)-NIASP gave a striatal to cerebellar tissue radioactivity concentration of 16.9 to 1 at 6 hr postinjection. Thus, (E)-({sup 125}I)-NIASP binds with high selectivity and specificity to dopamine D{sub 2} sites in vivo.« less

  13. Molecular modeling and docking studies of human 5-hydroxytryptamine 2A (5-HT2A) receptor for the identification of hotspots for ligand binding.

    PubMed

    Kanagarajadurai, Karuppiah; Malini, Manoharan; Bhattacharya, Aditi; Panicker, Mitradas M; Sowdhamini, Ramanathan

    2009-12-01

    The serotonergic system has been implicated in emotional and cognitive function. In particular, 5-HT(2A) (5-hydroxytrytamine receptor 2A) is attributed to a number of disorders like schizophrenia, depression, eating disorders and anxiety. 5-HT(2A), being a GPCR (G-protein coupled receptor), is important in the pharmaceutical industry as a proven target for these disorders. Despite their extensive clinical importance, the structural studies of this protein is lacking due to difficulties in determining its crystal structure. We have performed sequence analysis and molecular modeling of 5-HT(2A) that has revealed a set of conserved residues and motifs considered to play an important role in maintaining structural integrity and function of the receptor. The analysis also revealed a set of residues specific to the receptor which distinguishes them from other members of the subclass and their orthologs. Further, starting from the model structure of human 5-HT(2A) receptor, docking studies were attempted to envisage how it might interact with eight of its ligands (such as serotonin, dopamine, DOI, LSD, haloperidol, ketanserin, risperidone and clozapine). The binding studies of dopamine to 5-HT(2A) receptor can bring up better understanding in the etiology of a number of neurological disorders involving both these two receptors. Our sequence analysis and study of interactions of this receptor with other ligands reveal additional residue hotspots such as Asn 363 and Tyr 370. The function of these residues can be further analyzed by rational design of site-directed mutagenesis. Two distinct binding sites are identified which could play important roles in ligand binding and signaling.

  14. Tolerance to LSD and DOB induced shaking behaviour: differential adaptations of frontocortical 5-HT(2A) and glutamate receptor binding sites.

    PubMed

    Buchborn, Tobias; Schröder, Helmut; Dieterich, Daniela C; Grecksch, Gisela; Höllt, Volker

    2015-03-15

    Serotonergic hallucinogens, such as lysergic acid diethylamide (LSD) and dimethoxy-bromoamphetamine (DOB), provoke stereotype-like shaking behaviour in rodents, which is hypothesised to engage frontocortical glutamate receptor activation secondary to serotonin2A (5-HT2A) related glutamate release. Challenging this hypothesis, we here investigate whether tolerance to LSD and DOB correlates with frontocortical adaptations of 5-HT2A and/or overall-glutamate binding sites. LSD and DOB (0.025 and 0.25 mg/kg, i.p.) induce a ketanserin-sensitive (0.5 mg/kg, i.p., 30-min pretreatment) increase in shaking behaviour (including head twitches and wet dog shakes), which with repeated application (7× in 4 ds) is undermined by tolerance. Tolerance to DOB, as indexed by DOB-sensitive [(3)H]spiroperidol and DOB induced [(35)S]GTP-gamma-S binding, is accompanied by a frontocortical decrease in 5-HT2A binding sites and 5-HT2 signalling, respectively; glutamate-sensitive [(3)H]glutamate binding sites, in contrast, remain unchanged. As to LSD, 5-HT2 signalling and 5-HT2A binding, respectively, are not or only marginally affected, yet [(3)H]glutamate binding is significantly decreased. Correlation analysis interrelates tolerance to DOB to the reduced 5-HT2A (r=.80) as well as the unchanged [(3)H]glutamate binding sites (r=.84); tolerance to LSD, as opposed, shares variance with the reduction in [(3)H]glutamate binding sites only (r=.86). Given that DOB and LSD both induce tolerance, one correlating with 5-HT2A, the other with glutamate receptor adaptations, it might be inferred that tolerance can arise at either level. That is, if a hallucinogen (like LSD in our study) fails to induce 5-HT2A (down-)regulation, glutamate receptors (activated postsynaptic to 5-HT2A related glutamate release) might instead adapt and thus prevent further overstimulation of the cortex. Copyright © 2014 Elsevier B.V. All rights reserved.

  15. Differential inhibition of N and P/Q Ca2+ currents by 5-HT1A and 5-HT1D receptors in spinal neurons of Xenopus larvae

    PubMed Central

    Sun, Qian-Quan; Dale, Nicholas

    1998-01-01

    In whole-cell patch clamp recordings made from non-sensory neurons acutely isolated from the spinal cord of Xenopus (stage 40–42) larvae, two forms of inhibition of the high voltage-activated (HVA) Ca2+ currents were produced by 5-HT. One was voltage dependent and associated with both slowing of the activation kinetics and shifting of the voltage dependence of the HVA currents. This inhibition was relieved by strong depolarizing prepulses. A second form of inhibition was neither associated with slowing of the activation kinetics nor relieved by depolarizing prepulses and was thus voltage independent. In all neurons examined, 5-HT (1 μM) reversibly reduced 34 ± 1.6 % (n = 102) of the HVA Ca2+ currents. In about 40 % of neurons, the inhibition was totally voltage independent. In another 5 %, the inhibition was totally voltage dependent. In the remaining neurons, inhibition was only partially (by around 40 %) relieved by a large depolarizing prepulse, suggesting that in these, the inhibition consisted of both voltage-dependent and -independent components. By using selective channel blockers, we found that 5-HT acted on both N- and P/Q-type channels. However, whereas the inhibition of P/Q-type currents was only voltage independent, the inhibition of N-type currents had both voltage-dependent and -independent components. The effects of 5-HT on HVA Ca2+ currents were mediated by 5-HT1A and 5-HT1D receptors. The 5-HT1A receptors not only preferentially caused voltage-independent inhibition, but did so by acting mainly on the ω-agatoxin-IVA-sensitive Ca2+ channels. In contrast, the 5-HT1D receptor produced both voltage-dependent and -independent inhibition and was preferentially coupled to ω-conotoxin-GVIA sensitive channels. This complexity of modulation may allow fine tuning of transmitter release and calcium signalling in the spinal circuitry of Xenopus larvae. PMID:9625870

  16. Novel spirotetracyclic zwitterionic dual H(1)/5-HT(2A) receptor antagonists for the treatment of sleep disorders.

    PubMed

    Gianotti, Massimo; Botta, Maurizio; Brough, Stephen; Carletti, Renzo; Castiglioni, Emiliano; Corti, Corrado; Dal-Cin, Michele; Delle Fratte, Sonia; Korajac, Denana; Lovric, Marija; Merlo, Giancarlo; Mesic, Milan; Pavone, Francesca; Piccoli, Laura; Rast, Slavko; Roscic, Maja; Sava, Anna; Smehil, Mario; Stasi, Luigi; Togninelli, Andrea; Wigglesworth, Mark J

    2010-11-11

    Histamine H(1) and serotonin 5-HT(2A) receptors mediate two different mechanisms involved in sleep regulation: H(1) antagonists are sleep inducers, while 5-HT(2A) antagonists are sleep maintainers. Starting from 9'a, a novel spirotetracyclic compound endowed with good H(1)/5-HT(2A) potency but poor selectivity, very high Cli, and a poor P450 profile, a specific optimization strategy was set up. In particular, we investigated the possibility of introducing appropriate amino acid moieties to optimize the developability profile of the series. Following this zwitterionic approach, we were able to identify several advanced leads (51, 65, and 73) with potent dual H(1)/5-HT(2A) activity and appropriate developability profiles. These compounds exhibited efficacy as hypnotic agents in a rat telemetric sleep model with minimal effective doses in the range 3-10 mg/kg po.

  17. ChLae1 and ChVel1 Regulate T-toxin Production, Virulence, Oxidative Stress Response, and Development of the Maize Pathogen Cochliobolus heterostrophus

    PubMed Central

    Wu, Dongliang; Oide, Shinichi; Zhang, Ning; Choi, May Yee; Turgeon, B. Gillian

    2012-01-01

    LaeA and VeA coordinate secondary metabolism and differentiation in response to light signals in Aspergillus spp. Their orthologs, ChLae1 and ChVel1, were identified in the maize pathogen Cochliobolus heterostrophus, known to produce a wealth of secondary metabolites, including the host selective toxin, T-toxin. Produced by race T, T-toxin promotes high virulence to maize carrying Texas male sterile cytoplasm (T-cms). T-toxin production is significantly increased in the dark in wild type (WT), whereas Chvel1 and Chlae1 mutant toxin levels are much reduced in the dark compared to WT. Correspondingly, expression of T-toxin biosynthetic genes (Tox1) is up-regulated in the dark in WT, while dark-induced expression is much reduced/minimal in Chvel1 and Chlae1 mutants. Toxin production and Tox1 gene expression are increased in ChVEL1 overexpression (OE) strains grown in the dark and in ChLAE1 strains grown in either light or dark, compared to WT. These observations establish ChLae1 and ChVel1 as the first factors known to regulate host selective toxin production. Virulence of Chlae1 and Chvel1 mutants and OE strains is altered on both T-cms and normal cytoplasm maize, indicating that both T-toxin mediated super virulence and basic pathogenic ability are affected. Deletion of ChLAE1 or ChVEL1 reduces tolerance to H2O2. Expression of CAT3, one of the three catalase genes, is reduced in the Chvel1 mutant. Chlae1 and Chvel1 mutants also show decreased aerial hyphal growth, increased asexual sporulation and female sterility. ChLAE1 OE strains are female sterile, while ChVEL1 OE strains are more fertile than WT. ChLae1 and ChVel1 repress expression of 1,8-dihydroxynaphthalene (DHN) melanin biosynthesis genes, and, accordingly, melanization is enhanced in Chlae1 and Chvel1 mutants, and reduced in OE strains. Thus, ChLae1 and ChVel1 positively regulate T-toxin biosynthesis, pathogenicity and super virulence, oxidative stress responses, sexual development, and aerial hyphal

  18. The effects of benzofury (5-APB) on the dopamine transporter and 5-HT2-dependent vasoconstriction in the rat.

    PubMed

    Dawson, Patrick; Opacka-Juffry, Jolanta; Moffatt, James D; Daniju, Yusuf; Dutta, Neelakshi; Ramsey, John; Davidson, Colin

    2014-01-03

    5-APB, commonly marketed as 'benzofury' is a new psychoactive substance and erstwhile 'legal high' which has been implicated in 10 recent drug-related deaths in the UK. This drug was available on the internet and in 'head shops' and was one of the most commonly sold legal highs up until its recent UK temporary ban (UK Home Office). Despite its prominence, very little is known about its pharmacology. This study was undertaken to examine the pharmacology of 5-APB in vitro. We hypothesised that 5-APB would activate the dopamine and 5-HT systems which may underlie its putative stimulant and hallucinogenic effects. Autoradiographic studies showed that 5-APB displaced both [(125)I] RTI-121 and [(3)H] ketanserin from rat brain tissue suggesting affinity at the dopamine transporter and 5-HT2 receptor sites respectively. Voltammetric studies in rat accumbens brain slices revealed that 5-APB slowed dopamine reuptake, and at high concentrations caused reverse transport of dopamine. 5-APB also caused vasoconstriction of rat aorta, an effect antagonised by the 5-HT2A receptor antagonist ketanserin, and caused contraction of rat stomach fundus, which was reversed by the 5-HT2B receptor antagonist RS-127445. These data show that 5-APB interacts with the dopamine transporter and is an agonist at the 5-HT2A and 5-HT2B receptors in the rat. Thus 5-APB's pharmacology is consistent with it having both stimulant and hallucinogenic properties. In addition, 5-APB's activity at the 5-HT2B receptor may cause cardiotoxicity. © 2013.

  19. Rikkunshito and isoliquiritigenin counteract 5-HT-induced 2C receptor-mediated activation of pro-opiomelanocortin neurons in the hypothalamic arcuate nucleus.

    PubMed

    Arai, Takeshi; Maejima, Yuko; Muroya, Shinji; Yada, Toshihiko

    2013-08-01

    Anorexia deteriorates the quality of life in patients with anorexia nervosa, stress disorders, gastrointestinal disorders, and cancer. Pro-opiomelanocortin (POMC) neurons in the hypothalamic arcuate nucleus (ARC), serotonin (5-HT) and its 2C receptor (5-HT2CR) are implicated in anorexia. Rikkunshito, a traditional Japanese medicine, has been used to treat anorexia and gastrointestinal disorders. The present study aimed to clarify whether rikkunshito influences the 5-HT action on ARC POMC neurons. We isolated single neurons from the ARC of adult rats and measured cytosolic Ca²⁺ concentration ([Ca²⁺](i)) by fura-2 microfluorometry combined with immunocytochemical identification of POMC neurons. Administration of 5-HT increased [Ca²⁺](i) in ARC neurons, and 80% of the 5-HT-responsive neurons were immunoreactive to POMC. Rikkunshito concentration-dependently and 5-HT2CR antagonist SB242084 significantly suppressed 5-HT-induced [Ca²⁺](i) increases. The rikkunshito-suppressed neurons highly overlapped SB242084-suppressed neurons. Isoliquiritigenin, an ingredient of rikkunshito, suppressed 5-HT-induced [Ca²⁺](i) increases to a lesser extent than rikkunshito. These results demonstrate that rikkunshito counteracts 5-HT-induced 5-HT2CR-mediated Ca²⁺ signaling in ARC POMC neurons, and that isoliquiritigenin may serve as an active component of rikkunshito. The ability of rikkunshito to antagonize 5-HT action in ARC POMC neurons could underlie the rikkunshito's action to attenuate anorexia induced by excessive 5-HT release and/or action associated with psychiatric diseases, gastrointestinal disorders, cancer, and anti-cancer medicines. Copyright © 2013 Elsevier Ltd. All rights reserved.

  20. Further investigation into the mechanism of tachykinin NK(2) receptor-triggered serotonin release from guinea-pig proximal colon.

    PubMed

    Kojima, Shu-Ichi; Ikeda, Masashi; Kamikawa, Yuichiro

    2009-05-01

    The effects of the monoamine oxidase A (MAO-A) inhibitor clorgyline, the L-type calcium-channel blocker nicardipine, the syntaxin inhibitor botulinum toxin type C, and the potent thiol-oxidant phenylarsine oxide (PAO) on the selective tachykinin NK(2)-receptor agonist [beta-Ala(8)]-neurokinin A(4-10) [betaAla-NKA-(4-10)]-evoked 5-hydroxytryptamine (5-HT) outflow from colonic enterochromaffin (EC) cells was investigated in vitro using isolated guinea-pig proximal colon. The betaAla-NKA-(4-10)-evoked outflow of 5-HT from clorgyline-treated colonic strips was markedly higher than that from clorgyline-untreated colonic strips. The betaAla-NKA-(4-10)-evoked 5-HT outflow from the clorgyline-treated colonic strips was sensitive to nicardipine or botulinum toxin type C. Moreover, PAO concentration-dependently suppressed the betaAla-NKA-(4-10)-evoked 5-HT outflow from the clorgyline-treated colonic strips. The suppressant action of PAO was reversed by the reducing agent dithiothrietol, but was not blocked by the protein tyrosine kinase inhibitor genistein. These results suggest that the tachykinin NK(2) receptor-triggered 5-HT release from guinea-pig colonic EC cells is mediated by syntaxin-related exocytosis mechanisms and that colonic mucosa MAO-A activity has the important function of modulating the tachykinin NK(2) receptor-triggered 5-HT release. It also appears that PAO-mediated sulfhydryl oxidation plays a role in modulating the tachykinin NK(2) receptor-triggered 5-HT release through a mechanism independent of inhibition of protein tyrosine phosphatase activity.

  1. 5-HT2A serotonin receptor agonist DOI alleviates cytotoxicity in neuroblastoma cells: role of the ERK pathway.

    PubMed

    Marinova, Zoya; Walitza, Susanne; Grünblatt, Edna

    2013-07-01

    Disturbances of serotonergic signaling, including the serotonin 2A (5-HT2A) receptor, have been implicated in neuropsychiatric and neurodegenerative disorders. The aim of the present study was to characterize the effect of a 5-HT2A receptor agonist on cytotoxicity in a neuronal cell line and address the involved mechanism. HTR2A mRNA and protein expression in human neuroblastoma SK-N-SH cells was confirmed. Cells were subjected to serum deprivation and cell viability was monitored continuously with xCELLigence. In a dose-response study the 5-HT2A agonist (±)-2,5-dimethoxy-4-iodoamphetamine hydrochloride (DOI) (25 nM to 5 μM) protected against serum deprivation cytotoxicity. The selective 5-HT2A receptor antagonist MDL 11,939, the general protein tyrosine kinase inhibitor genistein, and the extracellular signal-regulated kinase (ERK) pathway MEK inhibitor U0126, all attenuated DOI's protective effect. An antibody array suggested that 1 μM DOI affected phosphorylation of several tyrosine kinases. Western blot further confirmed that DOI transiently increased ERK phosphorylation, indicating its activation. Finally, protective concentrations of DOI increased cellular mitochondrial mass, an effect prevented by pretreatment with U0126. In conclusion, our results suggest that DOI protects SK-N-SH cells against serum deprivation through ERK pathway activation. They imply 5-HT2A receptor modulation as a potential target for neuroprotection. Copyright © 2013 Elsevier Inc. All rights reserved.

  2. Paroxetine and Low-dose Risperidone Induce Serotonin 5-HT1A and Dopamine D2 Receptor Heteromerization in the Mouse Prefrontal Cortex.

    PubMed

    Kolasa, Magdalena; Solich, Joanna; Faron-Górecka, Agata; Żurawek, Dariusz; Pabian, Paulina; Łukasiewicz, Sylwia; Kuśmider, Maciej; Szafran-Pilch, Kinga; Szlachta, Marta; Dziedzicka-Wasylewska, Marta

    2018-05-01

    Recently, it has been shown that serotonin 5-HT 1A receptor interacts with dopamine D2 receptor in vitro. However, the existence of 5-HT 1A -D2 heteromers in native tissue remains unexplored. In the present study, we investigated 5-HT 1A -D2 receptor heteromerization in mice treated acutely or chronically with paroxetine (10 mg/kg) or risperidone (0.05 mg/kg). Receptor heteromerization was visualized and quantified in the mouse brain by in situ proximity ligation assay (PLA). Additionally, we aimed to determine the cellular localization of 5-HT 1A -D2 receptor heteromers in mouse adult primary neuronal cells by immunofluorescent staining with markers for astrocytes (GFAP) and neurons (NeuN and MAP2). The results from the current study demonstrated that 5-HT 1A and D2 receptor co-localization and heteromerization occurred in the mouse prefrontal cortex. Counterstaining after PLA confirmed neuronal (pyramidal and GABAergic) as well as astrocytal localization of 5-HT 1A -D2 receptor heteromers. Chronic administration of paroxetine or risperidone increased the level of 5-HT 1A -D2 receptor heteromers in the prefrontal cortex. These changes were not accompanied by any changes in the expression of mRNAs (measured by in situ hybridization) or densities of 5-HT 1A and D2 receptors (quantified by receptor autoradiography with [3H]8-OH-DPAT and [3H]domperidone, respectively), what all indicated that paroxetine and risperidone facilitated 5-HT 1A -D2 heteromer formation independently of the receptor expression. In vitro homogenous time-resolved FRET (HTRF) study confirmed the ability of tested drugs to influence the human 5-HT 1A -D2 heteromer formation. The obtained data indicate that the increase in 5-HT 1A -D2 receptor heteromerization is a common molecular characteristic of paroxetine and low-dose risperidone treatment. Copyright © 2018 IBRO. Published by Elsevier Ltd. All rights reserved.

  3. Yeast β-1,6-glucan is a primary target for the Saccharomyces cerevisiae K2 toxin.

    PubMed

    Lukša, Juliana; Podoliankaitė, Monika; Vepštaitė, Iglė; Strazdaitė-Žielienė, Živilė; Urbonavičius, Jaunius; Servienė, Elena

    2015-04-01

    Certain Saccharomyces cerevisiae strains secrete different killer proteins of double-stranded-RNA origin. These proteins confer a growth advantage to their host by increasing its survival. K2 toxin affects the target cell by binding to the cell surface, disrupting the plasma membrane integrity, and inducing ion leakage. In this study, we determined that K2 toxin saturates the yeast cell surface receptors in 10 min. The apparent amount of K2 toxin, bound to a single cell of wild type yeast under saturating conditions, was estimated to be 435 to 460 molecules. It was found that an increased level of β-1,6-glucan directly correlates with the number of toxin molecules bound, thereby impacting the morphology and determining the fate of the yeast cell. We observed that the binding of K2 toxin to the yeast surface receptors proceeds in a similar manner as in case of the related K1 killer protein. It was demonstrated that the externally supplied pustulan, a poly-β-1,6-glucan, but not the glucans bearing other linkage types (such as laminarin, chitin, and pullulan) efficiently inhibits the K2 toxin killing activity. In addition, the analysis of toxin binding to the intact cells and spheroplasts confirmed that majority of K2 protein molecules attach to the β-1,6-glucan, rather than the plasma membrane-localized receptors. Taken together, our results reveal that β-1,6-glucan is a primary target of K2 toxin and is important for the execution of its killing property. Copyright © 2015, American Society for Microbiology. All Rights Reserved.

  4. Fate and Distribution of 3H-Labeled T-2 Mycotoxin in Guinea Pigs

    DTIC Science & Technology

    1984-08-03

    HPLC as well as TLC. . . of T-2 sycotox in guinea pigs. To establish the toxicity of an im injection of T-2 mycotoxin in the guinea pig, si:: ;A...that remained at the origin with p- glucuronidase , two additional less-polar radiolabeled peaks were smnarated by HPLC. This suggests that at least one...all tissues within 30 min - ’af.ter, an a injection" of, thr toxin. The rapi& reationff label suggests that toxic effects could begin shortly after

  5. Responses of Oat Grains to Fusarium poae and F. langsethiae Infections and Mycotoxin Contaminations

    PubMed Central

    Martin, Charlotte; Schöneberg, Torsten; Morisoli, Romina; Bertossa, Mario; Bucheli, Thomas D.; Mauch-Mani, Brigitte; Mascher, Fabio

    2018-01-01

    Recent increases of Fusarium head blight (FHB) disease caused by infections with F. poae (FP) and F. langsethiae (FL) have been observed in oats. These pathogens are producers of nivalenol (NIV) and T-2/HT-2 toxin (T-2/HT-2), respectively, which are now considered major issues for cereal food and feed safety. To date, the impact of FP and FL on oat grains has not yet been identified, and little is known about oat resistance elements against these pathogens. In the present study, the impact of FL and FP on oat grains was assessed under different environmental conditions in field experiments with artificial inoculations. The severity of FP and FL infection on grains were compared across three field sites, and the resistance against NIV and T-2/HT2 accumulation was assessed for seven oat genotypes. Grain weight, β-glucan content, and protein content were compared between infected and non-infected grains. Analyses of grain infection showed that FL was able to cause infection on the grain only in the field site with the highest relative humidity, whereas FP infected grains in all field sites. The FP infection of grains resulted in NIV contamination (between 30–500 μg/kg). The concentration of NIV in grains was not conditioned by environmental conditions. FL provoked an average contamination of grains with T-2/HT-2 (between 15–132 μg/kg). None of the genotypes was able to fully avoid toxin accumulation. The general resistance of oat grains against toxin accumulation was weak, and resistance against NIV accumulation was strongly impacted by the interaction between the genotype and the environment. Only the genotype with hull-less grains showed partial resistance to both NIV and T-2/HT-2 contamination. FP and FL infections could change the β-glucan content in grains, depending on the genotypes and environmental conditions. FP and FL did not have a significant impact on the thousand kernel weight (TKW) and protein content. Hence, resistance against toxin

  6. Sleep Homeostatic and Waking Behavioral Phenotypes in Egr3-Deficient Mice Associated with Serotonin Receptor 5-HT2 Deficits.

    PubMed

    Grønli, Janne; Clegern, William C; Schmidt, Michelle A; Nemri, Rahmi S; Rempe, Michael J; Gallitano, Amelia L; Wisor, Jonathan P

    2016-12-01

    The expression of the immediate early gene early growth response 3 ( Egr3 ) is a functional marker of brain activity including responses to novelty, sustained wakefulness, and sleep. We examined the role of this gene in regulating wakefulness and sleep. Electroencephalogram/electromyogram (EEG/EMG) were recorded in Egr3 -/- and wild-type (WT) mice during 24 h baseline, 6 h sleep disruption and 6 h recovery. Serotonergic signaling was assessed with 6 h EEG/EMG recordings after injections of nonselective 5-HT2 antagonist (clozapine), selective 5-HT2 antagonists (5-HT2A; MDL100907 and 5-HT2BC; SB206553) and a cocktail of both selective antagonists, administered in a randomized order to each animal. Egr3 -/- mice did not exhibit abnormalities in the timing of wakefulness and slow wave sleep (SWS); however, EEG dynamics in SWS (suppressed 1-3 Hz power) and in quiet wakefulness (elevated 3-8 Hz and 15-35 Hz power) differed in comparison to WT-mice. Egr3 -/- mice showed an exaggerated response to sleep disruption as measured by active wakefulness, but with a blunted increase in homeostatic sleep drive (elevated 1-4 Hz power) relative to WT-mice. Egr3 -/-mice exhibit greatly reduced sedative effects of clozapine at the electroencephalographic level. In addition, clozapine induced a previously undescribed dissociated state (low amplitude, low frequency EEG and a stable, low muscle tone) lasting up to 2 h in WT-mice. Egr3 -/- mice did not exhibit this phenomenon. Selective 5-HT2A antagonist, alone or in combination with selective 5-HT2BC antagonist, caused EEG slowing coincident with behavioral quiescence in WT-mice but not in Egr3 -/- mice. Egr3 has an essential role in regulating cortical arousal, wakefulness, and sleep, presumably by its regulation of 5-HT2 receptors. © 2016 Associated Professional Sleep Societies, LLC.

  7. Lorcaserin and adiposopathy: 5-HT2c agonism as a treatment for 'sick fat' and metabolic disease.

    PubMed

    Bays, Harold E

    2009-11-01

    Agonists of 5-hydroxytryptamine (5-HT; serotonin) receptors promote loss of excessive body fat (adiposity) and improve metabolic parameters associated with adiposity-induced adipose tissue dysfunction (adiposopathy or 'sick fat'). By improving adipose tissue pathogenic endocrine and immune responses in overweight patients, 5-HT receptor agonists may improve metabolic disease. Lorcaserin (APD-356) is a selective 5-HT2c receptor agonist that promotes weight loss. Probably owing to its selectivity for the 5-HT2c receptor, clinical trial evidence supports that lorcaserin does not adversely affect heart valves or pulmonary artery pressure. This review examines: the mechanisms by which serotonergic pathways improve adiposity and adiposopathy; historical data and perspective regarding the efficacy and safety of prior 5-HT agonists; speculation regarding future paradigms in treating adiposopathy; and why lorcaserin may prove to be a safe and generally well-tolerated agent that not only improves the weight of patients, but also improves the health of patients.

  8. Identification and pathogenomic analysis of an Escherichia coli strain producing a novel Shiga toxin 2 subtype

    USDA-ARS?s Scientific Manuscript database

    Shiga toxin (Stx) is the key virulent factor in Shiga toxin-producing Escherichia coli (STEC). To date, three Stx1 subtypes and Seven Stx2 subtypes have been described in E. coli, which were found to differ in receptor preference and toxin potency. Here, we identified a novel Stx2 subtype designated...

  9. Hallucinogen-like effects of 2-([2-(4-cyano-2,5-dimethoxyphenyl) ethylamino]methyl)phenol (25CN-NBOH), a novel N-benzylphenethylamine with 100-fold selectivity for 5-HT2A receptors, in mice

    PubMed Central

    Gray, Bradley W.; Bailey, Jessica M.; Smith, Douglas; Hansen, Martin; Kristensen, Jesper L.

    2014-01-01

    Rationale 2-([2-(4-cyano-2,5-dimethoxyphenyl)ethylamino]methyl)phenol (25CN-NBOH) is structurally similar to N-benzyl substituted phenethylamine hallucinogens currently emerging as drugs of abuse. 25CN-NBOH exhibits dramatic selectivity for 5-HT2A receptors in vitro, but has not been behaviorally characterized. Objective 25CN-NBOH was compared to the traditional phenethylamine hallucinogen R(−)-2,5-dimethoxy-4-iodoamphetamine (DOI) using mouse models of drug-elicited head twitch behavior and drug discrimination. Methods Drug-elicited head twitches were quantified for 10 min following administration of various doses of either DOI or 25CN-NBOH, with and without pretreatments of 0.01 mg/kg 5-HT2A antagonist M100907 or 3.0 mg/kg 5-HT2C antagonist RS102221. The capacity of 25CN-NBOH to attenuate DOI-elicited head twitch was also investigated. Mice were trained to discriminate DOI or M100907 from saline, and 25CN-NBOH was tested for generalization. Results 25CN-NBOH induced a head twitch response in the mouse that was lower in magnitude than that of DOI, blocked by M100907, but not altered by RS102221. DOI-elicited head twitch was dose-dependently attenuated by 25CN-NBOH pretreatment. 25CN-NBOH produced an intermediate degree of generalization (55%) for the DOI training dose, and these interoceptive effects were attenuated by M100907. Finally, 25CN-NBOH did not generalize to M100907 at any dose, but ketanserin fully substituted in these animals. Conclusions 25CN-NBOH was behaviorally active, but less effective than DOI in two mouse models of hallucinogenic effects. The effectiveness with which M100907 antagonized the behavioral actions of 25CN-NBOH strongly suggests that the 5-HT2A receptor is an important site of agonist action for this compound in vivo. PMID:25224567

  10. Dual ligands targeting dopamine D2 and serotonin 5-HT1A receptors as new antipsychotical or anti-Parkinsonian agents.

    PubMed

    Ye, Na; Song, Zilan; Zhang, Ao

    2014-01-01

    Psychiatric disorders like schizophrenia and neurodegenerative diseases like Parkinson's disease are associated with poly-factorial pathogenic mechanisms, with several neurotransmitter systems closely involved. In addition to the cerebral dopaminergic (DA) system, the serotoninergic (5-HT) system also plays a crucial role in regulating psychoemotional, cognitive and motor functions in the central nervous system (CNS). Among the large 5-HT receptor family, accumulating data have revealed new insights into the therapeutic benefit of the 5-HT1A receptor in treating various CNS disorders, especially schizophrenia and Parkinson's disease. The present review discusses the advance of dual agents with mixed actions at the dopamine D2 and serotonin 5-HT1A receptors in the treatment of these diseases. Aripiprazole was the only marketed drug with dual D2 and 5-HT1A profile. It is a partial D2 and 5-HT1A receptor agonist and has been prescribed as an atypical antipsychotical drug. Two other drugs Cariprazine and Pardoprunox are being investigated in clinic. Most of the other candidate compounds, including Bifeprunox, Sarizotan, Mazapertine succinate, PF-217830, and Adoprazine were discontinued due to either non-optimal pharmacokinetic properties or insufficient therapeutical efficacy. Although much effort has been done to highlight the advantages of the 5-HT1A and D2 dual approach, it has to be pointed out that many of these drugs showed poly-pharmacological profile by targeting many other receptors and/or transporters besides the D2 and 5-HT1A receptors. In this regard, 'pure' compounds exclusively acting on the D2 and 5-HT1A receptors are highly needed to further validate this approach. Meanwhile, safety concerns and in vivo pharmacokinetic alerts should also be implanted to the drug design art early.

  11. A Method for Simultaneous Determination of 20 Fusarium Toxins in Cereals by High-Resolution Liquid Chromatography-Orbitrap Mass Spectrometry with a Pentafluorophenyl Column

    PubMed Central

    Tamura, Masayoshi; Mochizuki, Naoki; Nagatomi, Yasushi; Harayama, Koichi; Toriba, Akira; Hayakawa, Kazuichi

    2015-01-01

    A high-resolution liquid chromatography-Orbitrap mass spectrometry (LC-Orbitrap MS) method was developed for simultaneous determination of 20 Fusarium toxins (nivalenol, fusarenon-X, deoxynivalenol, 3-acetyl deoxynivalenol, 15-acetyl deoxynivalenol, HT-2 toxin, T-2 toxin, neosolaniol, diacetoxyscirpenol, fumonisin B1, fumonisin B2, fumonisin B3, fumonisin A1, fumonisin A2, fumonisin A3, zearalenone, α-zearalenol, β-zearalenol, α-zearalanol, and β-zearalanol) in cereals. The separation of 20 Fusarium toxins with good peak shapes was achieved using a pentafluorophenyl column, and Orbitrap MS was able to detect accurately from cereal matrix components within ±0.77 ppm. The samples were prepared using a QuEChERS kit for extraction and a multifunctional cartridge for purification. The linearity, repeatability, and recovery of the method were >0.9964, 0.8%–14.7%, and 71%–106%, respectively. Using this method, an analysis of 34 commercially available cereals detected the presence of deoxynivalenol, 15-acetyl deoxynivalenol, fumonisin B1, fumonisin B2, fumonisin B3, fumonisn A1, fumonisin A2, fumonisin A3, and zearalenone in corn samples with high concentration and frequency. Trichothecenes was detected from wheat samples with high frequency; in particular, the concentration of deoxynivalenol was high. Conversely, α-zearalenol, β-zearalenol, α-zearalanol, and β-zearalanol were not detected in any of the samples. PMID:26008230

  12. A Conserved Structural Motif Mediates Retrograde Trafficking of Shiga Toxin Types 1 and 2.

    PubMed

    Selyunin, Andrey S; Mukhopadhyay, Somshuvra

    2015-12-01

    Shiga toxin-producing Escherichia coli (STEC) produce two types of Shiga toxin (STx): STx1 and STx2. The toxin A-subunits block protein synthesis, while the B-subunits mediate retrograde trafficking. STEC infections do not have definitive treatments, and there is growing interest in generating toxin transport inhibitors for therapy. However, a comprehensive understanding of the mechanisms of toxin trafficking is essential for drug development. While STx2 is more toxic in vivo, prior studies focused on STx1 B-subunit (STx1B) trafficking. Here, we show that, compared with STx1B, trafficking of the B-subunit of STx2 (STx2B) to the Golgi occurs with slower kinetics. Despite this difference, similar to STx1B, endosome-to-Golgi transport of STx2B does not involve transit through degradative late endosomes and is dependent on dynamin II, epsinR, retromer and syntaxin5. Importantly, additional experiments show that a surface-exposed loop in STx2B (β4-β5 loop) is required for its endosome-to-Golgi trafficking. We previously demonstrated that residues in the corresponding β4-β5 loop of STx1B are required for interaction with GPP130, the STx1B-specific endosomal receptor, and for endosome-to-Golgi transport. Overall, STx1B and STx2B share a common pathway and use a similar structural motif to traffic to the Golgi, suggesting that the underlying mechanisms of endosomal sorting may be evolutionarily conserved. © 2015 John Wiley & Sons A/S. Published by John Wiley & Sons Ltd.

  13. The diphtheria toxin transmembrane domain as a pH sensitive membrane anchor for human interleukin-2 and murine interleukin-3.

    PubMed

    Liger, D; Nizard, P; Gaillard, C; vanderSpek, J C; Murphy, J R; Pitard, B; Gillet, D

    1998-11-01

    We have constructed two fusion proteins T-hIL-2 and T-mIL-3 in which human interleukin-2 (hIL-2) or murine interleukin-3 (mIL-3) are fused to the C-terminus of the diphtheria toxin transmembrane domain (T domain). Two additional fusion proteins, T-(Gly4-Ser)2-hIL-2 and T-(Gly4-Ser)2-mIL-3, were derived by introduction of the (Gly4-Ser)2 spacer between the T domain and cytokine components. Recognition of the hIL-2 receptor or the mIL-3 receptor by the corresponding recombinant proteins was demonstrated by their capacity to stimulate cytokine-dependent cell lines. All proteins retained the capacity of the T domain to insert into phospholipid membranes at acidic pH. Finally, anchoring of both cytokines to the membrane of lipid vesicles or living cells was assessed by specific antibody recognition. Our results show that the T domain fused to the N-terminus of a given protein can function as a pH sensitive membrane anchor for that protein.

  14. Induction of Apoptosis of 2,4′,6-Trihydroxybenzophenone in HT-29 Colon Carcinoma Cell Line

    PubMed Central

    Lay, Ma Ma; Karsani, Saiful Anuar

    2014-01-01

    2,4′,6-Trihydroxy-4-methoxybenzophenone was isolated from the ethyl acetate fraction of Phaleria macrocarpa (Scheff.) Boerl. fruits. It was found to inhibit cell proliferation in HT-29 human colon carcinoma cell line but caused little damage to WRL-68 normal human liver and MRC-5 normal human fibroblast lung cell lines. The compound was found to sharply affect the viability of HT-29 cells in a dose- and time-dependent manner. HT-29 cells treated with the compound showed morphological changes under microscopic examination such as cell shrinkage, membrane blebbing, DNA fragmentation, and the occurrence of apoptotic nuclei. The percentage of early apoptotic, late apoptotic, and dead or necrotic cells was determined by flow cytometry using annexin V-FTIC/PI staining. In addition, flow cytometry showed that, when the HT-29 cells were treated with 115 µM of the compound, it resulted in G0/G1 phase arrest in a time-dependent manner. Western blot revealed an upregulation of PUMA, Bak, Bcl-2, and Mcl-1 proteins suggesting that the compound induced apoptosis in HT-29 cells by regulating these proteins. PMID:24579081

  15. Hallucinogen-like effects of N,N-dipropyltryptamine (DPT): possible mediation by serotonin 5-HT1A and 5-HT2A receptors in rodents

    PubMed Central

    Fantegrossi, William E.; Reissig, Chad J.; Katz, Elyse B.; Yarosh, Haley L.; Rice, Kenner C.; Winter, Jerrold C.

    2008-01-01

    N,N-dipropyltryptamine (DPT) is a synthetic tryptamine hallucinogen which has been used psychotherapeutically in humans, but has been studied preclinically only rarely. In the present studies, DPT was tested in a drug-elicited head twitch assay in mice, and in rats trained to discriminate lysergic acid diethylamide (LSD), N,N-dimethyl-4-phosphoryloxytryptamine (psilocybin), or 3,4-methylenedioxymethamphetamine (MDMA). A separate group of rats was also trained to recognize DPT itself as a discriminative stimulus, and in all cases, the behavioral effects of DPT were challenged with the selective serotonin (5-HT)2A antagonist M100907, the 5-HT1A selective antagonist WAY-100635, or their combination. In the head twitch assay, DPT elicited dose-dependent effects, producing a biphasic dose-effect curve. WAY-100635 produced a parallel rightward shift in the dose-effect curve for head twitches, indicative of surmountable antagonism, but the antagonist effects of M100907 were functionally insurmountable. DPT produced partial to full substitution when tested in rats trained to discriminate LSD, psilocybin or MDMA, and served as a discriminative stimulus. In all cases, the antagonist effects of M100907 were more profound than were those of WAY-100635. DPT is thus active in two rodent models relevant to 5-HT2 agonist activity. The effectiveness with which M100907 antagonizes the behavioral actions of this compound strongly suggests that the 5-HT2A receptor is an important site of action for DPT, but the modulatory actions of WAY-100635 also imply a 5-HT1A-mediated component to the actions of this compound. PMID:17905422

  16. Sleep Homeostatic and Waking Behavioral Phenotypes in Egr3-Deficient Mice Associated with Serotonin Receptor 5-HT2 Deficits

    PubMed Central

    Grønli, Janne; Clegern, William C.; Schmidt, Michelle A.; Nemri, Rahmi S.; Rempe, Michael J.; Gallitano, Amelia L.; Wisor, Jonathan P.

    2016-01-01

    Study Objective: The expression of the immediate early gene early growth response 3 (Egr3) is a functional marker of brain activity including responses to novelty, sustained wakefulness, and sleep. We examined the role of this gene in regulating wakefulness and sleep. Methods: Electroencephalogram/electromyogram (EEG/EMG) were recorded in Egr3-/- and wild-type (WT) mice during 24 h baseline, 6 h sleep disruption and 6 h recovery. Serotonergic signaling was assessed with 6 h EEG/EMG recordings after injections of nonselective 5-HT2 antagonist (clozapine), selective 5-HT2 antagonists (5-HT2A; MDL100907 and 5-HT2BC; SB206553) and a cocktail of both selective antagonists, administered in a randomized order to each animal. Results: Egr3-/- mice did not exhibit abnormalities in the timing of wakefulness and slow wave sleep (SWS); however, EEG dynamics in SWS (suppressed 1–3 Hz power) and in quiet wakefulness (elevated 3–8 Hz and 15–35 Hz power) differed in comparison to WT-mice. Egr3-/- mice showed an exaggerated response to sleep disruption as measured by active wakefulness, but with a blunted increase in homeostatic sleep drive (elevated 1–4 Hz power) relative to WT-mice. Egr3-/-mice exhibit greatly reduced sedative effects of clozapine at the electroencephalographic level. In addition, clozapine induced a previously undescribed dissociated state (low amplitude, low frequency EEG and a stable, low muscle tone) lasting up to 2 h in WT-mice. Egr3-/- mice did not exhibit this phenomenon. Selective 5-HT2A antagonist, alone or in combination with selective 5-HT2BC antagonist, caused EEG slowing coincident with behavioral quiescence in WT-mice but not in Egr3-/- mice. Conclusion: Egr3 has an essential role in regulating cortical arousal, wakefulness, and sleep, presumably by its regulation of 5-HT2 receptors. Citation: Grønli J, Clegern WC, Schmidt MA, Nemri RS, Rempe MJ, Gallitano AL, Wisor JP. Sleep homeostatic and waking behavioral phenotypes in Egr3-deficient

  17. Blockade of serotonin 5-HT2A receptors potentiates dopamine D2 activation-induced disruption of pup retrieval on an elevated plus maze, but has no effect on D2 blockade-induced one.

    PubMed

    Nie, Lina; Di, Tianqi; Li, Yu; Cheng, Peng; Li, Ming; Gao, Jun

    2018-06-23

    Appetitive aspect of rat maternal behavior, such as pup retrieval, is motivationally driven and sensitive to dopamine disturbances. Activation or blockade of dopamine D 2 receptors causes a similar disruption of pup retrieval, which may also reflect an increase in maternal anxiety and/or a disruption of executive function. Recent work indicates that serotonin 5-HT 2A receptors also play an important role in rat maternal behavior. Given the well-known modulation of 5-HT 2A on the mesolimbic and mesocortical dopamine functions, the present study examined the extent to which blockade of 5-HT 2A receptors on dopamine D 2 -mediated maternal effects using a pup retrieval on the elevated plus maze (EPM) test. Sprague-Dawley postpartum female rats were acutely injected with quinpirole (a D 2 agonist, 0.10 and 0.25 mg/kg, sc), or haloperidol (a D 2 antagonist, 0.1 or 0.2 mg/kg, sc), in combination of MDL100907 (a 5-HT 2A receptor antagonist, 1.0 mg/kg, sc, 30 min before quinpirole or haloperidol injection) or saline and tested at 30, 90 and 240 min after quinpirole or haloperidol injection on postpartum days 3 and 7. Quinpirole and haloperidol decreased the number of pup retrieved (an index of maternal motivation) and sequential retrieval score (an index of executive function), prolonged the pup retrieval latencies, reduced the percentage of time spent on the open arms (an index of maternal anxiety), and decreased the distance travelled on the maze in a dose-dependent and time-dependent fashion. MDL100907 treatment by itself had no effect on pup retrieval, but it exacerbated the quinpirole-induced disruption of pup retrieval, but had no effect on the haloperidol-induced one. These findings suggest a complex interactive effect between 5-HT 2A and D 2 receptors on one or several maternal processes (maternal motivation, anxiety and executive function), and support the idea that one molecular mechanism by which 5-HT 2A receptors mediate maternal behavior is through

  18. Rapid desensitization and resensitization of 5-HT sub 2 receptor mediated phosphatidyl inositol hydrolysis by serotonin agonists in quiescent calf aortic smooth muscle cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Pauwels, P.J.; Van Gompel, P.; Leysen, J.E.

    1990-01-01

    Agonist regulation of 5-hydroxytryptamine{sub 2} (5-HT{sub 2}) receptors was studied in calf aortic smooth muscle cultures incubated in a quiescent, defined synthetic medium that does not stimulate cell proliferation, but that provides cells with supplements that maintain cell viability. In these cells, 5-hydroxytryptamine (5-HT)-induced ({sup 3}H)inositol phosphates accumulation showed the characteristics of a 5-HT{sub 2} receptor coupled transducing system according to the inhibition of the response by 5-HT{sub 2} antagonists at nanomolar concentrations. The 5-HT{sub 2} receptor coupled response became rapidly desensitized during continued incubation with 5-HT and 1-(2,5-dimethoxy-4-methylphenyl)-2-aminopropane (DOM); nearly full desensitization was obtained in two hours with 10more » {mu}M 5-HT and DOM pretreatment. The recovery of the response had a half-live of 5 hours after 2 hours pretreatment and of 9.5 to 12.5 hours after 24 to 96 hours agonist pretreatment. The DOM-induced desensitization of the 5-HT{sub 2} receptor coupled response was fully blocked by 0.1 {mu}M cinanserin. Cinanserin alone did not induce desensitization or up-regulation of the 5-HT{sub 2} receptor coupled response at 0.1 {mu}M.« less

  19. Structure, Function and Evolution of Clostridium botulinum C2 and C3 Toxins: Insight to Poultry and Veterinary Vaccines.

    PubMed

    Chellapandi, Paulchamy; Prisilla, Arokiyasamy

    2017-01-01

    Clostridium botulinum group III strains are able to produce cytotoxins, C2 toxin and C3 exotoxin, along with botulinum neurotoxin types C and D. C2 toxin and C3 exotoxin produced by this organism are the most important members of bacterial ADP-ribosyltransferase superfamily. Both toxins have distinct pathophysiological functions in the avian and mammalian hosts. The members of this superfamily transfer an ADP-ribose moiety of NAD+ to specific eukaryotic target proteins. The present review describes the structure, function and evolution aspects of these toxins with a special emphasis to the development of veterinary vaccines. C2 toxin is a binary toxin that consists of a catalytic subunit (C2I) and a translocation subunit (C2II). C2I component is structurally and functionally similar to the VIP2 and iota A toxin whereas C2II component shows a significant homology with the protective antigen from anthrax toxin and iota B. Unlike C2 toxin, C3 toxin is devoid of translocation/binding subunit. Extensive studies on their sequence-structure-function link spawn additional efforts to understand the catalytic mechanisms and target recognition. Structural and functional relationships with them are often determined by using evolutionary constraints as valuable biological measures. Enzyme-deficient mutants derived from these toxins have been used as drug/protein delivery systems in eukaryotic cells. Thus, current knowledge on their molecular diversity is a well-known perspective to design immunotoxin or subunit vaccine for C. botulinum infection. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  20. Central administration of a 5-HT2 receptor agonist and antagonist: lack of effect on rapid eye movement sleep and pgo waves.

    PubMed

    Sanford, L D; Hunt, W K; Ross, R J; Pack, A I; Morrison, A R

    1998-01-01

    Serotonin (5-HT) has a role in regulating behavioral state and controlling the production of ponto-geniculo-occipital (PGO) waves, though the exact mechanism of action is not known. The most prevailing explanation is that 5-HT exerts its influence on behavioral state and PGO waves by inhibiting and disinhibiting cholinergic cells in the pedunculopontine tegmentum (PPT) and laterodorsal tegmentum (LDT), which have been implicated in their generation. Recent work in rats has demonstrated 5-HT2 receptors on most cholinergic cells in PPT/LDT. We microinfused the relatively specific 5-HT2 agonist, DOI (1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane), the relatively specific 5-HT2 antagonist, ketanserin, and the nonspecific 5-HT antagonist, methysergide, locally into the peribrachial region of PPT in cats and monitored behavioral state and PGO waves. Neither drug significantly affected behavioral state or PGO wave activity. These results suggest that 5-HT2 receptors associated with cholinergic cells are minimally involved in the control of behavioral state and, together with the recent findings of others, suggest that 5-HT may not modulate PGO wave generation via direct action on cholinergic neurons in PPT/LDT, a departure from the long-held but minimally-tested view.

  1. Degradation Mechanism of Cyanobacterial Toxin Cylindrospermopsin by Hydroxyl Radicals in Homogeneous UV/H2O2 Process

    EPA Science Inventory

    The degradation of cylindrospermopsin (CYN), a widely distributed and highly toxic cyanobacterial toxin (cyanotoxin), remains poorly elucidated. In this study, the mechanism of CYN destruction by UV-254 nm/H2O2 advanced oxidation process (AOP) was investigated by mass spectrometr...

  2. Inhibition of Cocaine and 3,4-Methylenedioxypyrovalerone (MDPV) Self-Administration by Lorcaserin Is Mediated by 5-HT2C Receptors in Rats.

    PubMed

    Gannon, Brenda M; Sulima, Agnieszka; Rice, Kenner C; Collins, Gregory T

    2018-03-01

    Lorcaserin is a serotonin (5-HT) 2C receptor-preferring agonist approved by the US Food and Drug Administration to treat obesity. Lorcaserin decreases cocaine self-administration in rats and monkeys. Although this effect is partially inhibited by a 5-HT 2C receptor antagonist (SB242084), lorcaserin also has effects at 5-HT 2A and 5-HT 1A receptors, and the relative contribution of these receptors to its anti-cocaine effects has not been investigated. The goals of this study were to determine 1) the potency and effectiveness of lorcaserin to decrease self-administration of cocaine and 3,4-methylenedioxypyrovalerone (MDPV), a common "bath salts" constituent; and 2) the receptor(s) mediating the effects of lorcaserin on cocaine and MDPV self-administration. Male Sprague-Dawley rats ( n = 6) were trained to self-administer MDPV under a progressive ratio schedule of reinforcement and maintained under this schedule with daily access to 0.32 mg/kg per infusion of cocaine or 0.032 mg/kg per infusion of MDPV. Dose-response curves for the effects of lorcaserin on cocaine and MDPV self-administration were generated by administering lorcaserin (0.1-5.6 mg/kg) 25 minutes before the start of the session. To assess the effects of 5-HT 2C (SB242084, 0.1 mg/kg), 5-HT 2A (MDL100907, 0.1 mg/kg), and 5-HT 1A (WAY100635, 0.178 mg/kg) receptor antagonists, they were administered 15 minutes before lorcaserin. Lorcaserin decreased cocaine and MDPV self-administration with equal potency. Antagonism of 5-HT 2C (but not 5-HT 1A or 5-HT 2A ) receptors blocked the effects of lorcaserin on cocaine and MDPV self-administration. Taken together, these data provide additional support for further development of 5-HT 2C receptor agonists, such as lorcaserin, for the treatment of stimulant abuse. U.S. Government work not protected by U.S. copyright.

  3. Inhibition of Cocaine and 3,4-Methylenedioxypyrovalerone (MDPV) Self-Administration by Lorcaserin Is Mediated by 5-HT2C Receptors in Rats

    PubMed Central

    Gannon, Brenda M.; Sulima, Agnieszka; Rice, Kenner C.

    2018-01-01

    Lorcaserin is a serotonin (5-HT)2C receptor-preferring agonist approved by the US Food and Drug Administration to treat obesity. Lorcaserin decreases cocaine self-administration in rats and monkeys. Although this effect is partially inhibited by a 5-HT2C receptor antagonist (SB242084), lorcaserin also has effects at 5-HT2A and 5-HT1A receptors, and the relative contribution of these receptors to its anti-cocaine effects has not been investigated. The goals of this study were to determine 1) the potency and effectiveness of lorcaserin to decrease self-administration of cocaine and 3,4-methylenedioxypyrovalerone (MDPV), a common “bath salts” constituent; and 2) the receptor(s) mediating the effects of lorcaserin on cocaine and MDPV self-administration. Male Sprague-Dawley rats (n = 6) were trained to self-administer MDPV under a progressive ratio schedule of reinforcement and maintained under this schedule with daily access to 0.32 mg/kg per infusion of cocaine or 0.032 mg/kg per infusion of MDPV. Dose-response curves for the effects of lorcaserin on cocaine and MDPV self-administration were generated by administering lorcaserin (0.1–5.6 mg/kg) 25 minutes before the start of the session. To assess the effects of 5-HT2C (SB242084, 0.1 mg/kg), 5-HT2A (MDL100907, 0.1 mg/kg), and 5-HT1A (WAY100635, 0.178 mg/kg) receptor antagonists, they were administered 15 minutes before lorcaserin. Lorcaserin decreased cocaine and MDPV self-administration with equal potency. Antagonism of 5-HT2C (but not 5-HT1A or 5-HT2A) receptors blocked the effects of lorcaserin on cocaine and MDPV self-administration. Taken together, these data provide additional support for further development of 5-HT2C receptor agonists, such as lorcaserin, for the treatment of stimulant abuse. PMID:29217539

  4. Decreased Incentive Motivation Following Knockout or Acute Blockade of the Serotonin Transporter: Role of the 5-HT2C Receptor.

    PubMed

    Browne, Caleb J; Fletcher, Paul J

    2016-09-01

    Acute pharmacological elevation of serotonin (5-hydroxytryptamine; 5-HT) activity decreases operant responding for primary reinforcers, suggesting that 5-HT reduces incentive motivation. The mechanism by which 5-HT alters incentive motivation is unknown, but parallel evidence that 5-HT2C receptor agonists also reduce responding for primary reinforcers implicates this receptor as a potential candidate. These experiments examined whether chronic and acute disruptions of serotonin transporter (SERT) activity altered incentive motivation, and whether the 5-HT2C receptor mediated the effects of elevated 5-HT on behavior. To assess incentive motivation, we measured responding for three different reinforcers: a primary reinforcer (saccharin), a conditioned reinforcer (CRf), and an unconditioned sensory reinforcer (USRf). In the chronic condition, responding was compared between SERT knockout (SERT-KO) mice and their wild-type littermates. In the acute condition, responding was examined in wild-type mice following treatment with 10 or 20 mg/kg citalopram, or its vehicle. The ability of the selective 5-HT2C antagonist SB 242084 to prevent the effects of SERT-KO and citalopram on responding was subsequently examined. Both SERT-KO and citalopram reduced responding for saccharin, a CRf, and a USRf. Treatment with SB 242084 enhanced responding for a CRf and a USRf in SERT-KO mice and blocked the effects of citalopram on CRf and USRf responding. However, SB 242084 was unable to prevent the effects of SERT-KO or citalopram on responding for saccharin. These results support a powerful inhibitory function for 5-HT in the control of incentive motivation, and indicate that the 5-HT2C receptor mediates these effects of 5-HT in a reinforcer-dependent manner.

  5. The Antidepressant 5-HT2A Receptor Antagonists Pizotifen and Cyproheptadine Inhibit Serotonin-Enhanced Platelet Function

    PubMed Central

    Lin, Olivia A.; Karim, Zubair A.; Vemana, Hari Priya; Espinosa, Enma V. P.; Khasawneh, Fadi T.

    2014-01-01

    There is considerable interest in defining new agents or targets for antithrombotic purposes. The 5-HT2A receptor is a G-protein coupled receptor (GPCR) expressed on many cell types, and a known therapeutic target for many disease states. This serotonin receptor is also known to regulate platelet function. Thus, in our FDA-approved drug repurposing efforts, we investigated the antiplatelet activity of cyproheptadine and pizotifen, two antidepressant 5-HT2A Receptor antagonists. Our results revealed that cyproheptadine and pizotifen reversed serotonin-enhanced ADP-induced platelet aggregation in vitro and ex vivo. And the inhibitory effects of these two agents were found to be similar to that of EMD 281014, a 5-HT2A Receptor antagonist under development. In separate experiments, our studies revealed that these 5-HT2A receptor antagonists have the capacity to reduce serotonin-enhanced ADP-induced elevation in intracellular calcium levels and tyrosine phosphorylation. Using flow cytometry, we also observed that cyproheptadine, pizotifen, and EMD 281014 inhibited serotonin-enhanced ADP-induced phosphatidylserine (PS) exposure, P-selectin expression, and glycoprotein IIb-IIIa activation. Furthermore, using a carotid artery thrombosis model, these agents prolonged the time for thrombotic occlusion in mice in vivo. Finally, the tail-bleeding time was investigated to assess the effect of cyproheptadine and pizotifen on hemostasis. Our findings indicated prolonged bleeding time in both cyproheptadine- and pizotifen-treated mice. Notably, the increases in occlusion and bleeding times associated with these two agents were comparable to that of EMD 281014, and to clopidogrel, a commonly used antiplatelet drug, again, in a fashion comparable to clopidogrel and EMD 281014. Collectively, our data indicate that the antidepressant 5-HT2A antagonists, cyproheptadine and pizotifen do exert antiplatelet and thromboprotective effects, but similar to clopidogrel and EMD 281014, their

  6. Lymphocyte receptors for pertussis toxin

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Clark, C.G.; Armstrong, G.D.

    1990-12-01

    We have investigated human T-lymphocyte receptors for pertussis toxin by affinity isolation and photoaffinity labeling procedures. T lymphocytes were obtained from peripheral human blood, surface iodinated, and solubilized in Triton X-100. The iodinated mixture was then passed through pertussis toxin-agarose, and the fractions were analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Autoradiography of the fixed, dried gels revealed several bands in the pertussis toxin-bound fraction that were not observed in fractions obtained from histone or fetuin-agarose. Further investigations employed a photoaffinity labeling reagent, sulfosuccinimidyl 2-(p-azido-salicylamido)-1,3'-dithiopropionate, to identify pertussis toxin receptors in freshly isolated peripheral blood monocytic cells, T lymphocytes, andmore » Jurkat cells. In all three cell systems, the pertussis toxin affinity probe specifically labeled a single protein species with an apparent molecular weight of 70,000 that was not observed when the procedure was performed in the presence of excess unmodified pertussis toxin. A protein comparable in molecular weight to the one detected by the photoaffinity labeling technique was also observed among the species that bound to pertussis toxin-agarose. The results suggest that pertussis toxin may bind to a 70,000-Da receptor in human T lymphocytes.« less

  7. The 5-HT2A/1A agonist psilocybin disrupts modal object completion associated with visual hallucinations.

    PubMed

    Kometer, Michael; Cahn, B Rael; Andel, David; Carter, Olivia L; Vollenweider, Franz X

    2011-03-01

    Recent findings suggest that the serotonergic system and particularly the 5-HT2A/1A receptors are implicated in visual processing and possibly the pathophysiology of visual disturbances including hallucinations in schizophrenia and Parkinson's disease. To investigate the role of 5-HT2A/1A receptors in visual processing the effect of the hallucinogenic 5-HT2A/1A agonist psilocybin (125 and 250 μg/kg vs. placebo) on the spatiotemporal dynamics of modal object completion was assessed in normal volunteers (n = 17) using visual evoked potential recordings in conjunction with topographic-mapping and source analysis. These effects were then considered in relation to the subjective intensity of psilocybin-induced visual hallucinations quantified by psychometric measurement. Psilocybin dose-dependently decreased the N170 and, in contrast, slightly enhanced the P1 component selectively over occipital electrode sites. The decrease of the N170 was most apparent during the processing of incomplete object figures. Moreover, during the time period of the N170, the overall reduction of the activation in the right extrastriate and posterior parietal areas correlated positively with the intensity of visual hallucinations. These results suggest a central role of the 5-HT2A/1A-receptors in the modulation of visual processing. Specifically, a reduced N170 component was identified as potentially reflecting a key process of 5-HT2A/1A receptor-mediated visual hallucinations and aberrant modal object completion potential. Copyright © 2011 Society of Biological Psychiatry. Published by Elsevier Inc. All rights reserved.

  8. Novel 4-Substituted-N,N-dimethyltetrahydronaphthalen-2-amines: Synthesis, Affinity, and In Silico Docking Studies at Serotonin 5-HT2-type and Histamine H1 G Protein-Coupled Receptors

    PubMed Central

    Sakhuja, Rajeev; Kondabolu, Krishnakanth; Córdova-Sintjago, Tania; Travers, Sean; Vincek, Adam S.; Kim, Myong Sang; Abboud, Khalil A.; Fang, Lijuan; Sun, Zhuming; Canal, Clinton E.; Booth, Raymond G.

    2015-01-01

    Syntheses were undertaken of derivatives of (2S, 4R)-(−)-trans-4-phenyl-N,N-dimethyl-1,2,3,4-tetrahydronaphthalen-2-amine (4-phenyl-2-dimethylaminotetralin, PAT), a stereospecific agonist at the serotonin 5-HT2C G protein-coupled receptor (GPCR), with inverse agonist activity at 5-HT2A and 5-HT2B GPCRs. Molecular changes were made at the PAT C(4)-position, while preserving N, N-dimethyl substitution at the 2-position as well as trans-stereochemistry, structural features previously shown to be optimal for 5-HT2 binding. Affinities of analogs were determined at recombinant human 5-HT2 GPCRs in comparison to the phylogenetically closely-related histamine H1 GPCR, and in silico ligand docking studies were conducted at receptor molecular models to help interpret pharmacological results and guide future ligand design. In most cases, C(4)-substituted PAT analogs exhibited the same stereoselectivity ([−]-trans > [+]-trans) as the parent PAT across 5-HT2 and H1 GPCRs, albeit, with variable receptor selectivity. 4-(4′-substituted)-PAT analogs, however, demonstrated reversed stereoselectivity ([2S, 4R]-[+]-trans > [2S, 4R]-[−]-trans), with absolute configuration confirmed by single X-ray crystallographic data for the 4-(4′-Cl)-PAT analog. Pharmacological affinity results and computational results herein support further PAT drug development studies and provide a basis for predicting and interpreting translational results, including, for (+)-trans-4-(4′-Cl)-PAT and (−)-trans-4-(3′-Br)-PAT that were previously shown to be more potent and efficacious than their corresponding enantiomers in rodent models of psychoses, psychostimulant-induced behaviors, and compulsive feeding (‘binge-eating’). PMID:25703249

  9. Amino acid residues involved in membrane insertion and pore formation of Clostridium botulinum C2 toxin.

    PubMed

    Lang, Alexander E; Neumeyer, Tobias; Sun, Jianjun; Collier, R John; Benz, Roland; Aktories, Klaus

    2008-08-12

    The actin-ADP-ribosylating Clostridium botulinum C2 toxin consists of the enzymatic component C2I and the binding component C2II. C2II forms heptameric channels involved in translocation of the enzymatic component into the target cell. On the basis of the heptameric toxin channel, we studied functional consequences of mutagenesis of amino acid residues probably lining the lumen of the toxin channel. Substitution of glutamate-399 of C2II with alanine blocked channel formation and cytotoxicity of the holotoxin. Although cytotoxicity and rounding up of cells by C2I were completely blocked by exchange of phenylalanine-428 with alanine, the mutation increased potassium conductance caused by C2II in artificial membranes by about 2-3-fold over that of wild-type toxin. In contrast to its effects on single-channel potassium conductance in artificial membranes, the F428A mutation delayed the kinetics of pore formation in lipid vesicles and inhibited the activity of C2II in promoting (86)Rb (+) release from preloaded intact cells after pH shift of the medium. Moreover, F428A C2II exhibited delayed and diminished formation of C2II aggregates at low pH, indicating major changes of the biophysical properties of the toxin. The data indicate that phenylalanine-428 of C2II plays a major role in conformational changes occurring during pore formation of the binding component of C2II.

  10. Oral Intoxication of Mice with Shiga Toxin Type 2a (Stx2a) and Protection by Anti-Stx2a Monoclonal Antibody 11E10

    PubMed Central

    Russo, L. M.; Melton-Celsa, A. R.; Smith, M. A.; Smith, M. J.

    2014-01-01

    Shiga toxin (Stx)-producing Escherichia coli (STEC) strains cause food-borne outbreaks of hemorrhagic colitis and, less commonly, a serious kidney-damaging sequela called the hemolytic uremic syndrome (HUS). Stx, the primary virulence factor expressed by STEC, is an AB5 toxin with two antigenically distinct forms, Stx1a and Stx2a. Although both toxins have similar biological activities, Stx2a is more frequently produced by STEC strains that cause HUS than is Stx1a. Here we asked whether Stx1a and Stx2a act differently when delivered orally by gavage. We found that Stx2a had a 50% lethal dose (LD50) of 2.9 μg, but no morbidity occurred after oral intoxication with up to 157 μg of Stx1a. We also compared several biochemical and histological parameters in mice intoxicated orally versus intraperitoneally with Stx2a. We discovered that both intoxication routes caused similar increases in serum creatinine and blood urea nitrogen, indicative of kidney damage, as well as electrolyte imbalances and weight loss in the animals. Furthermore, kidney sections from Stx2a-intoxicated mice revealed multifocal, acute tubular necrosis (ATN). Of particular note, we detected Stx2a in kidney sections from orally intoxicated mice in the same region as the epithelial cell type in which ATN was detected. Lastly, we showed reduced renal damage, as determined by renal biomarkers and histopathology, and full protection of orally intoxicated mice with monoclonal antibody (MAb) 11E10 directed against the toxin A subunit; conversely, an irrelevant MAb had no therapeutic effect. Orally intoxicated mice could be rescued by MAb 11E10 6 h but not 24 h after Stx2a delivery. PMID:24379294

  11. 5-HT2A Receptor Binding in the Frontal Cortex of Parkinson's Disease Patients and Alpha-Synuclein Overexpressing Mice: A Postmortem Study

    PubMed Central

    Rasmussen, Nadja Bredo; Olesen, Mikkel Vestergaard; Plenge, Per; Klein, Anders Bue; Westin, Jenny E.; Fog, Karina

    2016-01-01

    The 5-HT2A receptor is highly involved in aspects of cognition and executive function and seen to be affected in neurodegenerative diseases like Alzheimer's disease and related to the disease pathology. Even though Parkinson's disease (PD) is primarily a motor disorder, reports of impaired executive function are also steadily being associated with this disease. Not much is known about the pathophysiology behind this. The aim of this study was thereby twofold: (1) to investigate 5-HT2A receptor binding levels in Parkinson's brains and (2) to investigate whether PD associated pathology, alpha-synuclein (AS) overexpression, could be associated with 5-HT2A alterations. Binding density for the 5-HT2A-specific radioligand [3H]-MDL 100.907 was measured in membrane suspensions of frontal cortex tissue from PD patients. Protein levels of AS were further measured using western blotting. Results showed higher AS levels accompanied by increased 5-HT2A receptor binding in PD brains. In a separate study, we looked for changes in 5-HT2A receptors in the prefrontal cortex in 52-week-old transgenic mice overexpressing human AS. We performed region-specific 5-HT2A receptor binding measurements followed by gene expression analysis. The transgenic mice showed lower 5-HT2A binding in the frontal association cortex that was not accompanied by changes in gene expression levels. This study is one of the first to look at differences in serotonin receptor levels in PD and in relation to AS overexpression. PMID:27579212

  12. The effects of the selective 5-HT(2C) receptor antagonist SB 242084 on learned helplessness in male Fischer 344 rats.

    PubMed

    Strong, Paul V; Greenwood, Benjamin N; Fleshner, Monika

    2009-05-01

    Rats exposed to an uncontrollable stressor demonstrate a constellation of behaviors such as exaggerated freezing and deficits in shuttle box escape learning. These behaviors in rats have been called learned helplessness and have been argued to model human stress-related mood disorders. Learned helplessness is thought to be caused by hyperactivation of serotonin (5-HT) neurons in the dorsal raphe nucleus (DRN) and a subsequent exaggerated release of 5-HT in DRN projection sites. Blocking 5-HT(2C) receptors in the face of an increase in serotonin can alleviate anxiety behaviors in some animal models. However, specific 5-HT receptor subtypes involved in learned helplessness remain unknown. The current experiments tested the hypothesis that 5-HT(2C) receptor activation is necessary and sufficient for the expression of learned helplessness. The selective 5-HT(2C) receptor antagonist SB 242084 (1.0 mg/kg) administered i.p. to adult male Fischer 344 rats prior to shuttle box behavioral testing, but not before stress, blocked stress-induced deficits in escape learning but had no effect on the exaggerated shock-elicited freezing. The selective 5-HT(2C) receptor agonist CP-809101 was sufficient to produce learned helplessness-like behaviors in the absence of prior stress and these effects were blocked by pretreatment with SB 242084. Results implicate the 5-HT(2C) receptor subtype in mediating the shuttle box escape deficits produced by exposure to uncontrollable stress and suggest that different postsynaptic 5-HT receptor subtypes underlie the different learned helplessness behaviors.

  13. Determination of Fusarium toxins in functional vegetable milks applying salting-out-assisted liquid-liquid extraction combined with ultra-high-performance liquid chromatography tandem mass spectrometry.

    PubMed

    Hamed, Ahmed M; Arroyo-Manzanares, Natalia; García-Campaña, Ana M; Gámiz-Gracia, Laura

    2017-11-01

    Vegetable milks are considered as functional foods due to their physiological benefits. Although the consumption of these products has significantly increased, they have received little attention in legislation with regard to contaminants. However, they may contain mycotoxins resulting from the use of contaminated raw materials. In this work, ultra-high-performance liquid chromatography tandem mass spectrometry has been proposed for the determination of the most relevant Fusarium toxins (fumonisin B 1 and B 2 , HT-2 and T-2 toxins, zearalenone, deoxynivalenol and fusarenon-X) in different functional beverages based on cereals, legumes and seeds. Sample treatment consisted of a simple salting-out-assisted liquid-liquid extraction with no further clean-up. The method provided limits of quantification between 3.2 and 57.7 µg L -1 , recoveries above 80% and precision with RSD lower than 12%. The method was also applied for studying the occurrence of these mycotoxins in market samples of vegetable functional beverages and deoxynivalenol was found in three oat-based commercial drinks.

  14. 5HT-2C receptor polymorphism in suicide victims. Association studies in German and Slavic populations.

    PubMed

    Stefulj, Jasminka; Büttner, Andreas; Kubat, Milovan; Zill, Peter; Balija, Melita; Eisenmenger, Wolfgang; Bondy, Brigitta; Jernej, Branimir

    2004-08-01

    Sustainable observations suggest that suicidal behaviour by itself may have biological correlates, among which those related to the serotonergic synapse hold the key position. Based on the association of suicide and serotonergic dysfunction, it was proposed that genetic mechanisms affecting suicidal behaviour could be related to the alterations of the genes encoding the elements of 5HT synapse. The present study tested the association of the polymorphism in the serotonin 2C (5HT-2C) receptor coding region (Cys23Ser) with suicide commitment. Study was based on two independent samples, one of German (284 suicide victims versus 297 controls) and other of Slavic/Croatian (118 suicide victims versus 275 controls) ethnicity. No significant differences in allele or genotype frequencies between victims and controls were demonstrated. Results did not provide supporting evidence for the potential involvement of the investigated variants of 5HT-2C receptor in the susceptibility to suicide.

  15. Cognitive Impairment Induced by Delta9-tetrahydrocannabinol Occurs through Heteromers between Cannabinoid CB1 and Serotonin 5-HT2A Receptors

    PubMed Central

    Lanfumey, Laurence; Cordomí, Arnau; Pastor, Antoni; de La Torre, Rafael; Gasperini, Paola; Navarro, Gemma; Howell, Lesley A.; Pardo, Leonardo; Lluís, Carmen; Canela, Enric I.; McCormick, Peter J.; Maldonado, Rafael; Robledo, Patricia

    2015-01-01

    Activation of cannabinoid CB1 receptors (CB1R) by delta9-tetrahydrocannabinol (THC) produces a variety of negative effects with major consequences in cannabis users that constitute important drawbacks for the use of cannabinoids as therapeutic agents. For this reason, there is a tremendous medical interest in harnessing the beneficial effects of THC. Behavioral studies carried out in mice lacking 5-HT2A receptors (5-HT2AR) revealed a remarkable 5-HT2AR-dependent dissociation in the beneficial antinociceptive effects of THC and its detrimental amnesic properties. We found that specific effects of THC such as memory deficits, anxiolytic-like effects, and social interaction are under the control of 5-HT2AR, but its acute hypolocomotor, hypothermic, anxiogenic, and antinociceptive effects are not. In biochemical studies, we show that CB1R and 5-HT2AR form heteromers that are expressed and functionally active in specific brain regions involved in memory impairment. Remarkably, our functional data shows that costimulation of both receptors by agonists reduces cell signaling, antagonist binding to one receptor blocks signaling of the interacting receptor, and heteromer formation leads to a switch in G-protein coupling for 5-HT2AR from Gq to Gi proteins. Synthetic peptides with the sequence of transmembrane helices 5 and 6 of CB1R, fused to a cell-penetrating peptide, were able to disrupt receptor heteromerization in vivo, leading to a selective abrogation of memory impairments caused by exposure to THC. These data reveal a novel molecular mechanism for the functional interaction between CB1R and 5-HT2AR mediating cognitive impairment. CB1R-5-HT2AR heteromers are thus good targets to dissociate the cognitive deficits induced by THC from its beneficial antinociceptive properties. PMID:26158621

  16. Cognitive Impairment Induced by Delta9-tetrahydrocannabinol Occurs through Heteromers between Cannabinoid CB1 and Serotonin 5-HT2A Receptors.

    PubMed

    Viñals, Xavier; Moreno, Estefanía; Lanfumey, Laurence; Cordomí, Arnau; Pastor, Antoni; de La Torre, Rafael; Gasperini, Paola; Navarro, Gemma; Howell, Lesley A; Pardo, Leonardo; Lluís, Carmen; Canela, Enric I; McCormick, Peter J; Maldonado, Rafael; Robledo, Patricia

    2015-07-01

    Activation of cannabinoid CB1 receptors (CB1R) by delta9-tetrahydrocannabinol (THC) produces a variety of negative effects with major consequences in cannabis users that constitute important drawbacks for the use of cannabinoids as therapeutic agents. For this reason, there is a tremendous medical interest in harnessing the beneficial effects of THC. Behavioral studies carried out in mice lacking 5-HT2A receptors (5-HT2AR) revealed a remarkable 5-HT2AR-dependent dissociation in the beneficial antinociceptive effects of THC and its detrimental amnesic properties. We found that specific effects of THC such as memory deficits, anxiolytic-like effects, and social interaction are under the control of 5-HT2AR, but its acute hypolocomotor, hypothermic, anxiogenic, and antinociceptive effects are not. In biochemical studies, we show that CB1R and 5-HT2AR form heteromers that are expressed and functionally active in specific brain regions involved in memory impairment. Remarkably, our functional data shows that costimulation of both receptors by agonists reduces cell signaling, antagonist binding to one receptor blocks signaling of the interacting receptor, and heteromer formation leads to a switch in G-protein coupling for 5-HT2AR from Gq to Gi proteins. Synthetic peptides with the sequence of transmembrane helices 5 and 6 of CB1R, fused to a cell-penetrating peptide, were able to disrupt receptor heteromerization in vivo, leading to a selective abrogation of memory impairments caused by exposure to THC. These data reveal a novel molecular mechanism for the functional interaction between CB1R and 5-HT2AR mediating cognitive impairment. CB1R-5-HT2AR heteromers are thus good targets to dissociate the cognitive deficits induced by THC from its beneficial antinociceptive properties.

  17. Acquisition session length modulates consolidation effects produced by 5-HT2C ligands in a mouse autoshaping-operant procedure.

    PubMed

    Walker, Ellen A; Foley, John J

    2010-03-01

    Although the neurotransmitter, 5-hydroxytryptamine (serotonin, 5-HT), has been implicated as a mediator of learning and memory, the specific role of 5-HT receptors in rodents requires further delineation. In this study, 5-HT2C receptor ligands of varying relative intrinsic efficacies were tested in a mouse learning and memory model called autoshaping-operant. On day 1, mice were placed in experimental chambers and presented with a tone on a variable interval schedule. The tone remained on for 6 s or until a nose-poke response occurred to produce a dipper with Ensure solution. Mice were then injected with saline, MK212 (full agonist), m-chlorophenylpiperazine (partial agonist), mianserin, and SB206 553 (inverse agonists), and methysergide and (+)-2-bromo lysergic acid diethylamide (+)-hydrogen tartrate (neutral antagonists). Each compound was injected after either 1 or 2-h acquisition sessions on day 1 to investigate the role of acquisition session length on consolidation. Day 1 injection of the 5-HT2C inverse agonist mianserin produced greater retrieval impairments of the autoshaped operant response on day 2 than any other agent tested. Furthermore, decreasing the length of the acquisition session to 1h significantly increased the difficulty of the autoshaping task further modulating the consolidation effects produced by the 5-HT2C ligands tested.

  18. The hallucinogen d-lysergic diethylamide (LSD) decreases dopamine firing activity through 5-HT1A, D2 and TAAR1 receptors.

    PubMed

    De Gregorio, Danilo; Posa, Luca; Ochoa-Sanchez, Rafael; McLaughlin, Ryan; Maione, Sabatino; Comai, Stefano; Gobbi, Gabriella

    2016-11-01

    d-lysergic diethylamide (LSD) is a hallucinogenic drug that interacts with the serotonin (5-HT) system binding to 5-HT 1 and 5-HT 2 receptors. Little is known about its potential interactions with the dopamine (DA) neurons of the ventral tegmental area (VTA). Using in-vivo electrophysiology in male adult rats, we evaluated the effects of cumulative doses of LSD on VTA DA neuronal activity, compared these effects to those produced on 5-HT neurons in the dorsal raphe nucleus (DRN), and attempted to identify the mechanism of action mediating the effects of LSD on VTA DA neurons. LSD, at low doses (5-20μg/kg, i.v.) induced a significant decrease of DRN 5-HT firing activity through 5-HT 2A and D 2 receptors. At these low doses, LSD did not alter VTA DA neuronal activity. On the contrary, at higher doses (30-120μg/kg, i.v.), LSD dose-dependently decreased VTA DA firing activity. The depletion of 5-HT with p-chlorophenylalanine did not modulate the effects of LSD on DA firing activity. The inhibitory effects of LSD on VTA DA firing activity were prevented by the D 2 receptor antagonist haloperidol (50μg/kg, i.v.) and by the 5-HT 1A receptor antagonist WAY-100,635 (500μg/kg, i.v.). Notably, pretreatment with the trace amine-associate receptor 1 (TAAR 1 ) antagonist EPPTB (5mg/kg, i.v.) blocked the inhibitory effect of LSD on VTA DA neurons. These results suggest that LSD at high doses strongly affects DA mesolimbic neuronal activity in a 5-HT independent manner and with a pleiotropic mechanism of action involving 5-HT 1A, D 2 and TAAR 1 receptors. Copyright © 2016 Elsevier Ltd. All rights reserved.

  19. Characterizing RecA-Independent Induction of Shiga toxin2-Encoding Phages by EDTA Treatment

    PubMed Central

    Imamovic, Lejla; Muniesa, Maite

    2012-01-01

    Background The bacteriophage life cycle has an important role in Shiga toxin (Stx) expression. The induction of Shiga toxin-encoding phages (Stx phages) increases toxin production as a result of replication of the phage genome, and phage lysis of the host cell also provides a means of Stx toxin to exit the cell. Previous studies suggested that prophage induction might also occur in the absence of SOS response, independently of RecA. Methodology/Principal Findings The influence of EDTA on RecA-independent Stx2 phage induction was assessed, in laboratory lysogens and in EHEC strains carrying Stx2 phages in their genome, by Real-Time PCR. RecA-independent mechanisms described for phage λ induction (RcsA and DsrA) were not involved in Stx2 phage induction. In addition, mutations in the pathway for the stress response of the bacterial envelope to EDTA did not contribute to Stx2 phage induction. The effect of EDTA on Stx phage induction is due to its chelating properties, which was also confirmed by the use of citrate, another chelating agent. Our results indicate that EDTA affects Stx2 phage induction by disruption of the bacterial outer membrane due to chelation of Mg2+. In all the conditions evaluated, the pH value had a decisive role in Stx2 phage induction. Conclusions/Significance Chelating agents, such as EDTA and citrate, induce Stx phages, which raises concerns due to their frequent use in food and pharmaceutical products. This study contributes to our understanding of the phenomenon of induction and release of Stx phages as an important factor in the pathogenicity of Shiga toxin-producing Escherichia coli (STEC) and in the emergence of new pathogenic strains. PMID:22393404

  20. Comparison of Systemic Toxicity between Botulinum Toxin Subtypes A1 and A2 in Mice and Rats.

    PubMed

    Torii, Yasushi; Goto, Yoshitaka; Nakahira, Shinji; Kozaki, Shunji; Kaji, Ryuji; Ginnaga, Akihiro

    2015-06-01

    The adverse events caused by botulinum toxin type A (subtype A1) product, thought to be after-effects of toxin diffusion after high-dose administration, have become serious issues. A preparation showing less diffusion in the body than existing drugs has been sought. We have attempted to produce neurotoxin derived from subtype A2 (A2NTX) with an amino acid sequence different from that of neurotoxin derived from subtype A1 (A1NTX). In this study, to investigate whether A2NTX has the potential to resolve these issues, we compared the safety of A2NTX, a progenitor toxin derived from subtype A1 (A1 progenitor toxin) and A1NTX employing the intramuscular lethal dose 50% (im LD50) in mice and rats and the compound muscle action potential (CMAP) in rats. Mouse im LD50 values for A1 progenitor toxin and A2NTX were 93 and 166 U/kg, respectively, and the rat im LD50 values were 117 and 153 U/kg, respectively. In the rat CMAP test, the dose on the contralateral side, which caused a 50% reduction in the CMAP amplitude, that is, CMAP-TD50 , was calculated as 19.0, 16.6 and 28.7 U/kg for A1 progenitor toxin, A1NTX and A2NTX, respectively. The results indicate that A2NTX is safer than A1 progenitor toxin and A1NTX. © 2014 Nordic Association for the Publication of BCPT (former Nordic Pharmacological Society).

  1. Conferring Virulence: Structure and Function of the chimeric A2B5 Typhoid Toxin

    PubMed Central

    Song, Jeongmin; Gao, Xiang; Galán, Jorge E.

    2013-01-01

    Salmonella Typhi differs from most other salmonellae in that it causes a life-threatening systemic infection known as typhoid fever1. The molecular bases for its unique clinical presentation are unknown2. Here we found that in an animal model, the systemic administration of typhoid toxin, a unique virulence factor of S. Typhi, reproduces many of the acute symptoms of typhoid fever. We identified specific carbohydrate moieties on specific surface glycoproteins that serve as receptors for typhoid toxin, which explains its broad cell target specificity. We present the atomic structure of typhoid toxin, which shows an unprecedented A2B5 organization with two covalently-linked A subunits non-covalently-associated to a pentameric B subunit. The structure provides insight into the toxin’s receptor-binding specificity and delivery mechanisms and reveals how the activities of two powerful toxins have been coopted into a single, unique toxin that can induce many of the symptoms characteristic of typhoid fever. These findings may lead to the development of potentially life-saving therapeutics against typhoid fever. PMID:23842500

  2. Dramatic changes in muscle contractile and structural properties after 2 botulinum toxin injections.

    PubMed

    Minamoto, Viviane B; Suzuki, Kentaro P; Bremner, Shannon N; Lieber, Richard L; Ward, Samuel R

    2015-10-01

    Botulinum toxin is frequently administered serially to maintain therapeutic muscle paralysis, but the effect of repeated doses on muscle function are largely unknown. This study characterized the muscle response to 2 onabotulinum toxin (BoNT) injections separated by 3 months. Animal subjects received a single toxin injection (n = 8), 2 BoNT injections separated by 3 months (n = 14), or 1 BoNT and 1 saline injection separated by 3 months (n = 8). The functional effect of 2 serial injections was exponentially greater than the effect of a single injection. While both groups treated with a single BoNT injection had decreased torque in the injected leg by approximately 50% relative to contralateral legs, the double BoNT injected group had decreased torque by over 95% relative to the preinjection level. Both single and double BoNT injections produced clear signs of fiber-type grouping. These experiments demonstrate a disproportionately greater effect of repeated BoNT injections. © 2015 Wiley Periodicals, Inc.

  3. Role of serotonin 5-HT2A receptors in the development of cardiac hypertrophy in response to aortic constriction in mice.

    PubMed

    Lairez, O; Cognet, T; Schaak, S; Calise, D; Guilbeau-Frugier, C; Parini, A; Mialet-Perez, J

    2013-06-01

    Serotonin, in addition to its fundamental role as a neurotransmitter, plays a critical role in the cardiovascular system, where it is thought to be involved in the development of cardiac hypertrophy and failure. Indeed, we recently found that mice with deletion of monoamine oxidase A had enhanced levels of blood and cardiac 5-HT, which contributed to exacerbation of hypertrophy in a model of experimental pressure overload. 5-HT2A receptors are expressed in the heart and mediate a hypertrophic response to 5-HT in cardiac cells. However, their role in cardiac remodeling in vivo and the signaling pathways associated are not well understood. In the present study, we evaluated the effect of a selective 5-HT2A receptor antagonist, M100907, on the development of cardiac hypertrophy induced by transverse aortic constriction (TAC). Cardiac 5-HT2A receptor expression was transiently increased after TAC, and was recapitulated in cardiomyocytes, as observed with 5-HT2A in situ labeling by immunohistochemistry. Selective blockade of 5-HT2A receptors prevented the development of cardiac hypertrophy, as measured by echocardiography, cardiomyocyte area and heart weight-to-body weight ratio. Interestingly, activation of calmodulin kinase (CamKII), which is a core mechanism in cardiac hypertrophy, was reduced in cardiac samples from M100907-treated TAC mice compared to vehicle-treated mice. In addition, phosphorylation of histone deacetylase 4 (HDAC4), a downstream partner of CamKII was significantly diminished in M100907-treated TAC mice. Thus, our results show that selective blockade of 5-HT2A receptors has beneficial effect in the development of cardiac hypertrophy through inhibition of the CamKII/HDAC4 pathway.

  4. 5-HT2A receptors control body temperature in mice during LPS-induced inflammation via regulation of NO production.

    PubMed

    Voronova, Irina P; Khramova, Galina M; Kulikova, Elizabeth A; Petrovskii, Dmitrii V; Bazovkina, Daria V; Kulikov, Alexander V

    2016-01-01

    G protein-coupled 5-HT2A receptors are involved in the regulation of numerous normal and pathological physiological functions. At the same time, its involvement in the regulation of body temperature (Tb) in normal conditions is obscure. Here we study the effect of the 5-HT2A receptor activation or blockade on Tb in sick animals. The experiments were carried out on adult C57BL/6 mouse males. Systemic inflammation and sickness were produced by lipopolysaccharide (LPS, 0.1mg/kg, ip), while the 5-HT2A receptor was stimulated or blocked through the administration of the receptor agonist DOI or antagonist ketanserin (1mg/kg), respectively. LPS, DOI or ketanserin alone produced no effect on Tb. However, administration of LPS together with a peripheral or central ketanserin injection reduced Tb (32.2°C). Ketanserin reversed the LPS-induced expression of inducible NO synthase in the brain. Consequently, an involvement of NO in the mechanism of the hypothermic effect of ketanserin in sick mice was hypothesized. Administration of LPS together with NO synthase inhibitor, l-nitro-arginine methyl ester (60mg/kg, ip) resulted in deep (28.5°C) and prolonged (8h) hypothermia, while administration of l-nitro-arginine methyl ester alone produced no effect on Tb. Thus, 5-HT2A receptors play a key role in Tb control in sick mice. Blockade of this GPCR produces hypothermia in mice with systemic inflammation via attenuation of LPS-induced NO production. These results indicate an unexpected role of 5-HT2A receptors in inflammation and NO production and have a considerable biological impact on understanding the mechanism of animal adaptation to pathogens and parasites. Moreover, adverse side effects of 5-HT2A receptor antagonists in patients with inflammation may be expected. Copyright © 2015 Elsevier Ltd. All rights reserved.

  5. Loss of the imprinted snoRNA mbii-52 leads to increased 5htr2c pre-RNA editing and altered 5HT2CR-mediated behaviour.

    PubMed

    Doe, Christine M; Relkovic, Dinko; Garfield, Alastair S; Dalley, Jeffrey W; Theobald, David E H; Humby, Trevor; Wilkinson, Lawrence S; Isles, Anthony R

    2009-06-15

    The Prader-Willi syndrome (PWS) genetic interval contains several brain-expressed small nucleolar (sno)RNA species that are subject to genomic imprinting. In vitro studies have shown that one of these snoRNA molecules, h/mbii-52, negatively regulates editing and alternative splicing of the serotonin 2C receptor (5htr2c) pre-RNA. However, the functional consequences of loss of h/mbii-52 and subsequent increased post-transcriptional modification of 5htr2c are unknown. 5HT2CRs are important in controlling aspects of cognition and the cessation of feeding, and disruption of their function may underlie some of the psychiatric and feeding abnormalities seen in PWS. In a mouse model for PWS lacking expression of mbii-52 (PWS-IC+/-), we show an increase in editing, but not alternative splicing, of the 5htr2c pre-RNA. This change in post-transcriptional modification is associated with alterations in a number of 5HT2CR-related behaviours, including impulsive responding, locomotor activity and reactivity to palatable foodstuffs. In a non-5HT2CR-related behaviour, marble burying, loss of mbii-52 was without effect. The specificity of the behavioural effects to changes in 5HT2CR function was further confirmed using drug challenges. These data illustrate, for the first time, the physiological consequences of altered RNA editing of 5htr2c linked to mbii-52 loss that may underlie specific aspects of the complex PWS phenotype and point to an important functional role for this imprinted snoRNA.

  6. Radiosynthesis and in vivo evaluation of a series of substituted 11C-phenethylamines as 5-HT (2A) agonist PET tracers.

    PubMed

    Ettrup, Anders; Hansen, Martin; Santini, Martin A; Paine, James; Gillings, Nic; Palner, Mikael; Lehel, Szabolcs; Herth, Matthias M; Madsen, Jacob; Kristensen, Jesper; Begtrup, Mikael; Knudsen, Gitte M

    2011-04-01

    Positron emission tomography (PET) imaging of serotonin 2A (5-HT(2A)) receptors with agonist tracers holds promise for the selective labelling of 5-HT(2A) receptors in their high-affinity state. We have previously validated [(11)C]Cimbi-5 and found that it is a 5-HT(2A) receptor agonist PET tracer. In an attempt to further optimize the target-to-background binding ratio, we modified the chemical structure of the phenethylamine backbone and carbon-11 labelling site of [(11)C]Cimbi-5 in different ways. Here, we present the in vivo validation of nine novel 5-HT(2A) receptor agonist PET tracers in the pig brain. Each radiotracer was injected intravenously into anaesthetized Danish Landrace pigs, and the pigs were subsequently scanned for 90 min in a high-resolution research tomography scanner. To evaluate 5-HT(2A) receptor binding, cortical nondisplaceable binding potentials (BP(ND)) were calculated using the simplified reference tissue model with the cerebellum as a reference region. After intravenous injection, all compounds entered the brain and distributed preferentially into the cortical areas, in accordance with the known 5-HT(2A) receptor distribution. The largest target-to-background binding ratio was found for [(11)C]Cimbi-36 which also had a high brain uptake compared to its analogues. The cortical binding of [(11)C]Cimbi-36 was decreased by pretreatment with ketanserin, supporting 5-HT(2A) receptor selectivity in vivo. [(11)C]Cimbi-82 and [(11)C]Cimbi-21 showed lower cortical BP(ND), while [(11)C]Cimbi-27, [(11)C]Cimbi-29, [(11)C]Cimbi-31 and [(11)C]Cimbi-88 gave rise to cortical BP(ND) similar to that of [(11)C]Cimbi-5. [(11)C]Cimbi-36 is currently the most promising candidate for investigation of 5-HT(2A) receptor agonist binding in the living human brain with PET.

  7. Pharmacological evaluation of halogenated and non-halogenated arylpiperazin-1-yl-ethyl-benzimidazoles as D(2) and 5-HT(2A) receptor ligands.

    PubMed

    Tomić, Mirko; Vasković, Djurdjica; Tovilović, Gordana; Andrić, Deana; Penjišević, Jelena; Kostić-Rajačić, Sladjana

    2011-05-01

    Five groups of previously synthesized and initially screened non-substituted and 4-halogenated arylpiperazin-1-yl-ethyl-benzimidazoles were estimated for their in-vitro binding affinities at the rat D(2) , 5-HT(2A) , and α(1) -adrenergic receptors. Among all these compounds, 2-methoxyphenyl and 2-chlorophenyl piperazines demonstrate the highest affinities for the tested receptors. The effects of 4-halogenation of benzimidazoles reveal that substitution with bromine may greatly increase the affinity of the compounds for the studied receptors, while the effect of substitution with chlorine is less remarkable. Most of the tested components show 5-HT(2A)/D(2) pK(i) binding ratios slightly above or less than 1, while only 4-chloro-6-(2-{4-[3-(trifluoromethyl)phenyl]piperazin-1-yl}ethyl)-1H-benzimidazole expresses an appropriate higher binding ratio (1.14), which was indicated for atypical neuroleptics. This compound exhibits a non-cataleptic action in rats and prevents d-amphetamine-induced hyperlocomotion in mice, which suggest its atypical antipsychotic potency. Copyright © 2011 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  8. Sub-Lethal Dose of Shiga Toxin 2 from Enterohemorrhagic Escherichia coli Affects Balance and Cerebellar Cytoarchitecture

    PubMed Central

    Pinto, Alipio; Cangelosi, Adriana; Geoghegan, Patricia A.; Tironi-Farinati, Carla; Brener, Gabriela J.; Goldstein, Jorge

    2016-01-01

    Shiga toxin producing Escherichia coli may damage the central nervous system before or concomitantly to manifested hemolytic–uremic syndrome symptoms. The cerebellum is frequently damaged during this syndrome, however, the deleterious effects of Shiga toxin 2 has never been integrally reported by ultrastructural, physiological and behavioral means. The aim of this study was to determine the cerebellar compromise after intravenous administration of a sub-lethal dose of Shiga toxin 2 by measuring the cerebellar blood–brain barrier permeability, behavioral task of cerebellar functionality (inclined plane test), and ultrastructural analysis (transmission electron microscope). Intravenous administration of vehicle (control group), sub-lethal dose of 0.5 and 1 ηg of Stx2 per mouse were tested for behavioral and ultrastructural studies. A set of three independent experiments were performed for each study (n = 6). Blood–brain barrier resulted damaged and consequently its permeability was significantly increased. Lower scores obtained in the inclined plane task denoted poor cerebellar functionality in comparison to their controls. The most significant lower score was obtained after 5 days of 1 ηg of toxin administration. Transmission electron microscope micrographs from the Stx2-treated groups showed neurons with a progressive neurodegenerative condition in a dose dependent manner. As sub-lethal intravenous Shiga toxin 2 altered the blood brain barrier permeability in the cerebellum the toxin penetrated the cerebellar parenchyma and produced cell damaged with significant functional implications in the test balance. PMID:26904009

  9. Loss of GluN2D subunit results in social recognition deficit, social stress, 5-HT2C receptor dysfunction, and anhedonia in mice.

    PubMed

    Yamamoto, Hideko; Kamegaya, Etsuko; Hagino, Yoko; Takamatsu, Yukio; Sawada, Wakako; Matsuzawa, Maaya; Ide, Soichiro; Yamamoto, Toshifumi; Mishina, Masayoshi; Ikeda, Kazutaka

    2017-01-01

    The N-methyl-d-aspartate (NMDA) receptor channel is involved in various physiological functions, including learning and memory. The GluN2D subunit of the NMDA receptor has low expression in the mature brain, and its role is not fully understood. In the present study, the effects of GluN2D subunit deficiency on emotional and cognitive function were investigated in GluN2D knockout (KO) mice. We found a reduction of motility (i.e., a depressive-like state) in the tail suspension test and a reduction of sucrose preference (i.e., an anhedonic state) in GluN2D KO mice that were group-housed with littermates. Despite apparently normal olfactory function and social interaction, GluN2D KO mice exhibited a decrease in preference for social novelty, suggesting a deficit in social recognition or memory. Golgi-Cox staining revealed a reduction of the complexity of dendritic trees in the accessory olfactory bulb in GluN2D KO mice, suggesting a deficit in pheromone processing pathway activation, which modulates social recognition. The deficit in social recognition may result in social stress in GluN2D KO mice. Isolation housing is a procedure that has been shown to reduce stress in mice. Interestingly, 3-week isolation and treatment with agomelatine or the 5-hydroxytryptamine-2C (5-HT 2C ) receptor antagonist SB242084 reversed the anhedonic-like state in GluN2D KO mice. In contrast, treatment with the 5-HT 2C receptor agonist CP809101 induced depressive- and anhedonic-like states in isolated GluN2D KO mice. These results suggest that social stress that is caused by a deficit in social recognition desensitizes 5-HT 2c receptors, followed by an anhedonic- and depressive-like state, in GluN2D KO mice. The GluN2D subunit of the NMDA receptor appears to be important for the recognition of individuals and development of normal emotionality in mice. 5-HT 2C receptor antagonism may be a therapeutic target for treating social stress-induced anhedonia. This article is part of the Special

  10. Obtaining T1-T2 distribution functions from 1-dimensional T1 and T2 measurements: The pseudo 2-D relaxation model

    NASA Astrophysics Data System (ADS)

    Williamson, Nathan H.; Röding, Magnus; Galvosas, Petrik; Miklavcic, Stanley J.; Nydén, Magnus

    2016-08-01

    We present the pseudo 2-D relaxation model (P2DRM), a method to estimate multidimensional probability distributions of material parameters from independent 1-D measurements. We illustrate its use on 1-D T1 and T2 relaxation measurements of saturated rock and evaluate it on both simulated and experimental T1-T2 correlation measurement data sets. Results were in excellent agreement with the actual, known 2-D distribution in the case of the simulated data set. In both the simulated and experimental case, the functional relationships between T1 and T2 were in good agreement with the T1-T2 correlation maps from the 2-D inverse Laplace transform of the full 2-D data sets. When a 1-D CPMG experiment is combined with a rapid T1 measurement, the P2DRM provides a double-shot method for obtaining a T1-T2 relationship, with significantly decreased experimental time in comparison to the full T1-T2 correlation measurement.

  11. 5-HT2A SEROTONIN RECEPTOR BIOLOGY: Interacting proteins, kinases and paradoxical regulation

    PubMed Central

    Roth, Bryan L

    2011-01-01

    5-hydroxytryptamine2A (5-HT2A) serotonin receptors are important pharmacological targets for a large number of central nervous system and peripheral serotonergic medications. In this review article I summarize work mainly from my lab regarding serotonin receptor anatomy, pharmacology, signaling and regulation. I highlight the role of serotonin receptor interacting proteins and the emerging paradigm of G-protein coupled receptor functional selectivity. PMID:21288474

  12. Synthesis and serotonergic activity of substituted 2, N-benzylcarboxamido-5-(2-ethyl-1-dioxoimidazolidinyl)-N, N-dimethyltryptamine derivatives: novel antagonists for the vascular 5-HT(1B)-like receptor.

    PubMed

    Moloney, G P; Martin, G R; Mathews, N; Milne, A; Hobbs, H; Dodsworth, S; Sang, P Y; Knight, C; Williams, M; Maxwell, M; Glen, R C

    1999-07-15

    The synthesis and vascular 5-HT(1B)-like receptor activity of a novel series of substituted 2, N-benzylcarboxamido-5-(2-ethyl-1-dioxoimidazolidinyl)-N, N-dimethyltryptamine derivatives are described. Modifications to the 5-ethylene-linked heterocycle and to substituents on the 2-benzylamide side chain have been explored. Several compounds were identified which exhibited affinity at the vascular 5-HT(1B)-like receptor of pK(B) > 7.0, up to 100-fold selectivity over alpha(1)-adrenoceptor affinity and 5-HT(2A) receptor affinity, and which exhibited a favorable pharmacokinetic profile. N-Benzyl-3-[2-(dimethylamino)ethyl]-5-[2-(4,4-dimethyl-2, 5-dioxo-1-imidazolidinyl)ethyl]-1H-indole-2-carboxamide (23) was identified as a highly potent, silent (as judged by the inability of angiotensin II to unmask 5-HT(1B)-like receptor-mediated agonist activity in the rabbit femoral artery), and competitive vascular 5-HT(1B)-like receptor antagonist with a plasma elimination half-life of approximately 4 h in dog plasma and with good oral bioavailability. The selectivity of compounds from this series for the vascular 5-HT(1B)-like receptors over other receptor subtypes is discussed as well as a proposed mode of binding to the receptor pharmacophore. It has been proposed that the aromatic ring of the 2, N-benzylcarboxamide group can occupy an aromatic binding site rather than the indole ring. The resulting conformation allows an amine-binding site to be occupied by the ethylamine nitrogen and a hydrogen-bonding site to be occupied by one of the hydantoin carbonyls. The electronic nature of the 2,N-benzylcarboxamide aromatic group as well as the size of substituents on this aromatic group is crucial for producing potent and selective antagonists. The structural requirement on the 3-ethylamine side chain incorporating the protonatable nitrogen is achieved by the bulky 2, N-benzylcarboxamide group and its close proximity to the 3-side chain.

  13. Interaction of brain 5-HT synthesis deficiency, chronic stress and sex differentially impact emotional behavior in Tph2 knockout mice.

    PubMed

    Gutknecht, Lise; Popp, Sandy; Waider, Jonas; Sommerlandt, Frank M J; Göppner, Corinna; Post, Antonia; Reif, Andreas; van den Hove, Daniel; Strekalova, Tatyana; Schmitt, Angelika; Colaςo, Maria B N; Sommer, Claudia; Palme, Rupert; Lesch, Klaus-Peter

    2015-07-01

    While brain serotonin (5-HT) function is implicated in gene-by-environment interaction (GxE) impacting the vulnerability-resilience continuum in neuropsychiatric disorders, it remains elusive how the interplay of altered 5-HT synthesis and environmental stressors is linked to failure in emotion regulation. Here, we investigated the effect of constitutively impaired 5-HT synthesis on behavioral and neuroendocrine responses to unpredictable chronic mild stress (CMS) using a mouse model of brain 5-HT deficiency resulting from targeted inactivation of the tryptophan hydroxylase-2 (Tph2) gene. Locomotor activity and anxiety- and depression-like behavior as well as conditioned fear responses were differentially affected by Tph2 genotype, sex, and CMS. Tph2 null mutants (Tph2(-/-)) displayed increased general metabolism, marginally reduced anxiety- and depression-like behavior but strikingly increased conditioned fear responses. Behavioral modifications were associated with sex-specific hypothalamic-pituitary-adrenocortical (HPA) system alterations as indicated by plasma corticosterone and fecal corticosterone metabolite concentrations. Tph2(-/-) males displayed increased impulsivity and high aggressiveness. Tph2(-/-) females displayed greater emotional reactivity to aversive conditions as reflected by changes in behaviors at baseline including increased freezing and decreased locomotion in novel environments. However, both Tph2(-/-) male and female mice were resilient to CMS-induced hyperlocomotion, while CMS intensified conditioned fear responses in a GxE-dependent manner. Our results indicate that 5-HT mediates behavioral responses to environmental adversity by facilitating the encoding of stress effects leading to increased vulnerability for negative emotionality.

  14. Chloroquine Analog Interaction with C2- and Iota-Toxin in Vitro and in Living Cells.

    PubMed

    Kronhardt, Angelika; Beitzinger, Christoph; Barth, Holger; Benz, Roland

    2016-08-10

    C2-toxin from Clostridium botulinum and Iota-toxin from Clostridium perfringens belong both to the binary A-B-type of toxins consisting of two separately secreted components, an enzymatic subunit A and a binding component B that facilitates the entry of the corresponding enzymatic subunit into the target cells. The enzymatic subunits are in both cases actin ADP-ribosyltransferases that modify R177 of globular actin finally leading to cell death. Following their binding to host cells' receptors and internalization, the two binding components form heptameric channels in endosomal membranes which mediate the translocation of the enzymatic components Iota a and C2I from endosomes into the cytosol of the target cells. The binding components form ion-permeable channels in artificial and biological membranes. Chloroquine and related 4-aminoquinolines were able to block channel formation in vitro and intoxication of living cells. In this study, we extended our previous work to the use of different chloroquine analogs and demonstrate that positively charged aminoquinolinium salts are able to block channels formed in lipid bilayer membranes by the binding components of C2- and Iota-toxin. Similarly, these molecules protect cultured mammalian cells from intoxication with C2- and Iota-toxin. The aminoquinolinium salts did presumably not interfere with actin ADP-ribosylation or receptor binding but blocked the pores formed by C2IIa and Iota b in living cells and in vitro. The blocking efficiency of pores formed by Iota b and C2IIa by the chloroquine analogs showed interesting differences indicating structural variations between the types of protein-conducting nanochannels formed by Iota b and C2IIa.

  15. Chloroquine Analog Interaction with C2- and Iota-Toxin in Vitro and in Living Cells

    PubMed Central

    Kronhardt, Angelika; Beitzinger, Christoph; Barth, Holger; Benz, Roland

    2016-01-01

    C2-toxin from Clostridium botulinum and Iota-toxin from Clostridium perfringens belong both to the binary A-B-type of toxins consisting of two separately secreted components, an enzymatic subunit A and a binding component B that facilitates the entry of the corresponding enzymatic subunit into the target cells. The enzymatic subunits are in both cases actin ADP-ribosyltransferases that modify R177 of globular actin finally leading to cell death. Following their binding to host cells’ receptors and internalization, the two binding components form heptameric channels in endosomal membranes which mediate the translocation of the enzymatic components Iota a and C2I from endosomes into the cytosol of the target cells. The binding components form ion-permeable channels in artificial and biological membranes. Chloroquine and related 4-aminoquinolines were able to block channel formation in vitro and intoxication of living cells. In this study, we extended our previous work to the use of different chloroquine analogs and demonstrate that positively charged aminoquinolinium salts are able to block channels formed in lipid bilayer membranes by the binding components of C2- and Iota-toxin. Similarly, these molecules protect cultured mammalian cells from intoxication with C2- and Iota-toxin. The aminoquinolinium salts did presumably not interfere with actin ADP-ribosylation or receptor binding but blocked the pores formed by C2IIa and Iota b in living cells and in vitro. The blocking efficiency of pores formed by Iota b and C2IIa by the chloroquine analogs showed interesting differences indicating structural variations between the types of protein-conducting nanochannels formed by Iota b and C2IIa. PMID:27517960

  16. Evaluation of ELISA tests specific for Shiga toxin 1 and 2 in food and water samples

    USDA-ARS?s Scientific Manuscript database

    Two enzyme-linked immunosorbent assay (ELISA) kits were evaluated for their effectiveness in detecting and differentiating between Shiga toxin 1 and 2 (Stx1 and Stx2) produced by Shiga toxin-producing E. coli (STEC) inoculated into food and water samples. Each kit incorporated monoclonal antibodies ...

  17. New superhindered polydentate polyphosphine ligands P(CH2CH2P(t)Bu2)3, PhP(CH2CH2P(t)Bu2)2, P(CH2CH2CH2P(t)Bu2)3, and their ruthenium(II) chloride complexes.

    PubMed

    Gilbert-Wilson, Ryan; Field, Leslie D; Bhadbhade, Mohan M

    2012-03-05

    The synthesis and characterization of the extremely hindered phosphine ligands, P(CH(2)CH(2)P(t)Bu(2))(3) (P(2)P(3)(tBu), 1), PhP(CH(2)CH(2)P(t)Bu(2))(2) (PhP(2)P(2)(tBu), 2), and P(CH(2)CH(2)CH(2)P(t)Bu(2))(3) (P(3)P(3)(tBu), 3) are reported, along with the synthesis and characterization of ruthenium chloro complexes RuCl(2)(P(2)P(3)(tBu)) (4), RuCl(2)(PhP(2)P(2)(tBu)) (5), and RuCl(2)(P(3)P(3)(tBu)) (6). The bulky P(2)P(3)(tBu) (1) and P(3)P(3)(tBu) (3) ligands are the most sterically encumbered PP(3)-type ligands so far synthesized, and in all cases, only three phosphorus donors are able to bind to the metal center. Complexes RuCl(2)(PhP(2)P(2)(tBu)) (5) and RuCl(2)(P(3)P(3)(tBu)) (6) were characterized by crystallography. Low temperature solution and solid state (31)P{(1)H} NMR were used to demonstrate that the structure of RuCl(2)(P(2)P(3)(tBu)) (4) is probably analogous to that of RuCl(2)(PhP(2)P(2)(tBu)) (5) which had been structurally characterized.

  18. Activation of serotonin 5-HT(2C) receptor suppresses behavioral sensitization and naloxone-precipitated withdrawal symptoms in heroin-treated mice.

    PubMed

    Wu, Xian; Pang, Gang; Zhang, Yong-Mei; Li, Guangwu; Xu, Shengchun; Dong, Liuyi; Stackman, Robert W; Zhang, Gongliang

    2015-10-21

    Abuse and dependence to heroin has evolved into a global epidemic as a significant clinical and societal problem with devastating consequences. Repeated exposure to heroin can induce long-lasting behavioral sensitization and withdrawal. Pharmacological activation of 5-HT2C receptors (5-HT2CRs) suppresses psychostimulant-induced drug-seeking and behavioral sensitization. The present study examined the effect of a selective 5-HT2CR agonist lorcaserin on behavioral sensitization and naloxone-precipitated withdrawal symptoms in heroin-treated mice. Male mice received heroin (1.0 mg/kg, s.c.) twice a day for 3 days and then drug treatment was suspended for 5 days. On day 9, a challenge dose of heroin (1.0 mg/kg) was administered to examine the expression of behavioral sensitization. Lorcaserin administered during the development, withdrawal or expression stage suppressed heroin-induced behavioral sensitization on day 9. Another cohort of mice received increasing doses of heroin over a 4.5-day period. Lorcaserin, or the positive control clonidine (an α2-adrenoceptor agonist) suppressed naloxone-precipitated withdrawal symptoms in heroin-treated mice. These findings suggest that activation of 5-HT2CRs suppresses behavioral sensitization and withdrawal in heroin-treated mice. Thus, pharmacological activation of 5-HT2CRs may represent a new avenue for the treatment of heroin addiction. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  19. Activation of serotonin 5-HT2C receptor suppresses behavioral sensitization and naloxone-precipitated withdrawal symptoms in heroin-treated mice

    PubMed Central

    Li, Guangwu; Xu, Shengchun; Dong, Liuyi; Stackman, Robert W.; Zhang, Gongliang

    2015-01-01

    Abuse and dependence to heroin has evolved into a global epidemic as a significant clinical and societal problem with devastating consequences. Repeated exposure to heroin can induce long-lasting behavioral sensitization and withdrawal. Pharmacological activation of 5-HT2C receptors (5-HT2CRs) suppresses psychostimulant-induced drug-seeking and behavioral sensitization. The present study examined the effect of a selective 5-HT2CR agonist lorcaserin on behavioral sensitization and naloxone-precipitated withdrawal symptoms in heroin-treated mice. Male mice received heroin (1.0 mg/kg, s.c.) twice a day for 3 days and then drug treatment was suspended for 5 days. On day 9, a challenge dose of heroin (1.0 mg/kg) was administered to examine the expression of behavioral sensitization. Lorcaserin administered during the development, withdrawal or expression stage suppressed heroin-induced behavioral sensitization on day 9. Another cohort of mice received increasing doses of heroin over a 4.5-day period. Lorcaserin, or the positive control clonidine (an α2-adrenoceptor agonist) suppressed naloxone-precipitated withdrawal symptoms in heroin-treated mice. These findings suggest that activation of 5-HT2CRs suppresses behavioral sensitization and withdrawal in heroin-treated mice. Thus, pharmacological activation of 5-HT2CRs may represent a new avenue for the treatment of heroin addiction. PMID:26375926

  20. Derivatives of 2-(dipropylamino)tetralin: effect of the C8-substituent on the interaction with 5-HT1A receptors.

    PubMed

    Liu, Y; Yu, H; Svensson, B E; Cortizo, L; Lewander, T; Hacksell, U

    1993-12-24

    A series of 2-(dipropylamino)tetralin derivatives in which the C8 substituent is varied has been prepared and evaluated pharmacologically to explore the importance of the C8 substituent in the interaction of 2-aminotetralin-based ligands with serotonin (5-HT1A) receptors. Enantiopure derivatives were prepared by facile palladium-catalyzed reactions of the triflates of the enantiomers of 8-hydroxy-2-(dipropylamino)tetralin (8-OH-DPAT, 1). The affinity of the compounds for the 5-HT1A receptors was evaluated by competition experiments with [3H]-8-OH-DPAT in rat hippocampal and cortical tissue. In addition, the compounds were evaluated for central 5-HT and dopamine receptor stimulating activity in vivo by use of biochemical and behavioral assays in rats. With the exception of the carboxy-substituted derivative which is devoid of 5-HT1A receptor affinity, the compounds have moderate to high affinities (K(i) values range from 0.7 to 130 nM) for 5-HT1A receptors. Surprisingly, several of the derivatives do not produce any apparent effects in vivo although they have fairly high 5-HT1A receptor affinities. However, the methoxycarbonyl- and acetyl-substituted derivatives are potent 5-HT1A receptor agonists in vivo and exhibit in vitro affinities in the same range as the enantiomers of 1.

  1. Activation of superficial dorsal horn neurons in the mouse by a PAR-2 agonist and 5-HT: potential role in itch.

    PubMed

    Akiyama, Tasuku; Merrill, Austin W; Carstens, Mirela Iodi; Carstens, E

    2009-05-20

    Itch, an unpleasant sensation associated with the desire to scratch, is symptomatic of dermatologic and systemic disorders that often resist antihistamine treatment. Histamine-independent itch mediators include serotonin (5-HT) and agonists of the protease-activated receptor-2 (PAR-2). We used behavior, Fos immunohistochemistry, and electrophysiology to investigate if these mediators activate spinal dorsal horn neurons in a manner consistent with itch. Intradermal (i.d.) injection of the PAR-2 agonist SLIGRL-NH(2) in the rostral back evoked bouts of directed hindlimb scratches over 20-30 min. Hindpaw injection of SLIGRL-NH(2) produced Fos staining in superficial dorsal horn which was then targeted for single-unit recording. Small id microinjections of SLIGRL-NH(2) or 5-HT identified responsive single units in the superficial dorsal horn of mice anesthetized with pentobarbital. Thirty-eight units characterized as wide dynamic range, nociceptive specific, or mechanically insensitive exhibited significantly increased firing after i.d. SLIGRL-NH(2) for 9 min, to partial (25%) tachyphylaxis with repeated injection. A majority additionally responded to 5-HT (70%), mustard oil (79%), and capsaicin (71%). Seven units isolated with the 5-HT search stimulus exhibited significant and prolonged responses to 5-HT with tachyphylaxis to repeated injections. The majority also responded to SLIGRL-NH(2), mustard oil, and capsaicin. The prolonged responses of superficial dorsal horn neurons to SLIGRL-NH(2) and 5-HT suggest a role in signaling itch. However, their responsiveness to algogens is inconsistent with itch specificity. Alternatively, such neurons may signal itch, whereas noxious stimulus levels recruit these and a larger population of pruritogen-insensitive cells to signal pain which masks or occludes the itch signal.

  2. The roles of 5-HT1A and 5-HT2 receptors in the effects of 5-MeO-DMT on locomotor activity and prepulse inhibition in rats.

    PubMed

    Krebs-Thomson, Kirsten; Ruiz, Erbert M; Masten, Virginia; Buell, Mahalah; Geyer, Mark A

    2006-12-01

    The hallucinogen 5-methoxy-N,N-dimethyltryptamine (5-MeO-DMT) is structurally similar to other indoleamine hallucinogens such as LSD. The present study examined the effects of 5-MeO-DMT in rats using the Behavioral Pattern Monitor (BPM), which enables analyses of patterns of locomotor activity and exploration, and the prepulse inhibition of startle (PPI) paradigm. A series of interaction studies using the serotonin (5-HT)(1A) antagonist WAY-100635 (1.0 mg/kg), the 5-HT(2A) antagonist M100907 (1.0 mg/kg), and the 5-HT(2C) antagonist SER-082 (0.5 mg/kg) were performed to assess the respective contributions of these receptors to the behavioral effects of 5-MeO-DMT (0.01, 0.1, and 1.0 mg/kg) in the BPM and PPI paradigms. 5-MeO-DMT decreased locomotor activity, investigatory behavior, the time spent in the center of the BPM chamber, and disrupted PPI. All of these effects were antagonized by WAY-100635 pretreatment. M100907 pretreatment failed to attenuate any of these effects, while SER-082 pretreatment only antagonized the PPI disruption produced by 5-MeO-DMT. While the prevailing view was that the activation of 5-HT(2) receptors is solely responsible for hallucinogenic drug effects, these results support a role for 5-HT(1A) receptors in the effects of the indoleamine hallucinogen 5-MeO-DMT on locomotor activity and PPI in rats.

  3. New immuno-PCR assay for detection of low concentrations of shiga toxin 2 and its variants.

    PubMed

    Zhang, Wenlan; Bielaszewska, Martina; Pulz, Matthias; Becker, Karsten; Friedrich, Alexander W; Karch, Helge; Kuczius, Thorsten

    2008-04-01

    Shiga toxin (Stx)-producing Escherichia coli (STEC) strains secrete toxins that are major virulence factors and diagnostic targets, but some STEC strains secrete Stx in amounts that cannot be detected using conventional cell cytotoxicity or immunological assays. Therefore, there is an urgent need for more-sensitive Stx detection methods. We describe the development of an assay that can detect low concentrations of Stx2 and its variants. An immuno-PCR Stx2 assay was developed based on an enzyme immunoassay (EIA) combining antibody capture and DNA amplification to increase the signal. The immuno-PCR assay detected 10 pg/ml of purified Stx2, compared to 1 ng/ml Stx2 detected by commercial EIA. Consequently, immuno-PCR detected Stx2 and its variants in STEC strains that produce the toxins at levels that are nondetectable by using the EIA, as well as the Stx2 in EIA-negative enriched stool cultures from patients. Our data demonstrate that the immuno-PCR developed here is a highly sensitive and specific method for the detection of trace amounts of Stx2 and Stx2 variants. It is therefore suitable for use by clinical microbiological laboratories to improve the toxin detection in clinical samples.

  4. New Immuno-PCR Assay for Detection of Low Concentrations of Shiga Toxin 2 and Its Variants▿

    PubMed Central

    Zhang, Wenlan; Bielaszewska, Martina; Pulz, Matthias; Becker, Karsten; Friedrich, Alexander W.; Karch, Helge; Kuczius, Thorsten

    2008-01-01

    Shiga toxin (Stx)-producing Escherichia coli (STEC) strains secrete toxins that are major virulence factors and diagnostic targets, but some STEC strains secrete Stx in amounts that cannot be detected using conventional cell cytotoxicity or immunological assays. Therefore, there is an urgent need for more-sensitive Stx detection methods. We describe the development of an assay that can detect low concentrations of Stx2 and its variants. An immuno-PCR Stx2 assay was developed based on an enzyme immunoassay (EIA) combining antibody capture and DNA amplification to increase the signal. The immuno-PCR assay detected 10 pg/ml of purified Stx2, compared to 1 ng/ml Stx2 detected by commercial EIA. Consequently, immuno-PCR detected Stx2 and its variants in STEC strains that produce the toxins at levels that are nondetectable by using the EIA, as well as the Stx2 in EIA-negative enriched stool cultures from patients. Our data demonstrate that the immuno-PCR developed here is a highly sensitive and specific method for the detection of trace amounts of Stx2 and Stx2 variants. It is therefore suitable for use by clinical microbiological laboratories to improve the toxin detection in clinical samples. PMID:18272709

  5. The effects of a selective 5-HT2 receptor antagonist (ICI 170,809) on platelet aggregation and pupillary responses in healthy volunteers.

    PubMed Central

    Millson, D S; Jessup, C L; Swaisland, A; Haworth, S; Rushton, A; Harry, J D

    1992-01-01

    1. ICI 170,809 (2-(2-dimethylamino-2-methylpropylthio)-3-phenylquinoline hydrochloride) is a potent 5-hydroxytryptamine (5-HT) type 2 postsynaptic receptor antagonist. 2. Effects of ICI 170,809 as single oral doses (3, 7, 15 and 30 mg) or placebo were studied on the duration of antagonism for the ex vivo platelet aggregatory response to 5-HT and to the pupillary light constrictor response in eight healthy male volunteers. 3. Pupillary dark adapted responses to a 0.5 s light stimulus were measured using a portable infrared pupillometer, for up to 24 h after dosing. 4. The in vitro platelet 5-HT aggregation response was reduced by ICI 170,809, with depression of the dose-response curve to 5-HT at all concentrations of 5-HT and with no evidence for a parallel shift. 5. The ex vivo platelet 5-HT response demonstrated a dose related significant (P less than 0.02) decrease in aggregation reaching a maximum at 2 h after dosing with the effect persisting for at least 8 h after dosing with the 7 and 15 mg doses. 6. Resting pupil diameter (RPD), and light induced pupillary responses in the dark adapted pupil, showed a significant (P less than 0.01) dose related reduction with significant (P less than 0.05) effects still present with the 15 and 30 mg doses at 8 h after dosing. 7. We conclude that, changes in both ex vivo platelet aggregation to 5-HT and dark adapted pupil size, are significantly correlated (P less than 0.0001) with log plasma concentrations (ng ml-1) of ICI 170,809, enabling the assessment of 5-HT2-receptor antagonism in man. PMID:1576048

  6. Molecular role of TGF-beta, secreted from a new type of CD4+ suppressor T cell, NY4.2, in the prevention of autoimmune IDDM in NOD mice.

    PubMed

    Han, H S; Jun, H S; Utsugi, T; Yoon, J W

    1997-06-01

    A new type of CD4+ T cell clone (NY4.2) isolated from pancreatic islet-infiltrated lymphocytes of acutely diabetic non-obese diabetic (NOD) mice prevents the development of insulin-dependent diabetes mellitus (IDDM) in NOD mice, as well as the recurrence of autoimmune diabetes in syngeneic islet-transplanted NOD mice. It has been demonstrated that the cytokine TGF-beta, secreted from the cells of this clone, is the substance which prevents autoimmune IDDM. This investigation was initiated to determine the molecular role TGF-beta plays in the prevention of autoimmune IDDM by determining its effect on IL-2-induced signal transduction in Con A-activated NOD mouse splenocytes and HT-2 cells. First, we determined whether TGF-beta, secreted from NY4.2 T cells, inhibits IL-2-dependent T cell proliferation in HT-2 cells (IL-2-dependent T cell line) and NOD splenocytes. We found that TGF-beta suppresses IL-2-dependent T cell proliferation. Second, we determined whether TGF-beta inhibits the activation of Janus kinases (JAKs), as well as signal transducers and activators of transcription (STAT) proteins, involved in an IL-2-induced signalling pathway that normally leads to the proliferation of T cells. We found that TGF-beta inhibited tyrosine phosphorylation of JAK1, JAK3, STAT3 and STAT5 in Con A blasts from NOD splenocytes and HT-2 cells. Third, we examined whether TGF-beta inhibits the cooperation between STAT proteins and mitogen-activated protein kinase (MAPK), especially extracellular signal-regulated kinase 2 (ERK2). We found that TGF-beta inhibited the association of STAT3 and STAT5 with ERK2 in Con A blasts from NOD splenocytes and HT-2 cells. On the basis of these observations, we conclude that TGF-beta may interfere with signal transduction via inhibition of the IL-2-induced JAK/STAT pathway and inhibition of the association of STAT proteins with ERK2 in T cells from NOD splenocytes, resulting in the inhibition of IL-2-dependent T cell proliferation. TGF

  7. Selective effects of 5-HT2C receptor modulation on performance of a novel valence-probe visual discrimination task and probabilistic reversal learning in mice.

    PubMed

    Phillips, Benjamin U; Dewan, Sigma; Nilsson, Simon R O; Robbins, Trevor W; Heath, Christopher J; Saksida, Lisa M; Bussey, Timothy J; Alsiö, Johan

    2018-04-22

    Dysregulation of the serotonin (5-HT) system is a pathophysiological component in major depressive disorder (MDD), a condition closely associated with abnormal emotional responsivity to positive and negative feedback. However, the precise mechanism through which 5-HT tone biases feedback responsivity remains unclear. 5-HT2C receptors (5-HT2CRs) are closely linked with aspects of depressive symptomatology, including abnormalities in reinforcement processes and response to stress. Thus, we aimed to determine the impact of 5-HT2CR function on response to feedback in biased reinforcement learning. We used two touchscreen assays designed to assess the impact of positive and negative feedback on probabilistic reinforcement in mice, including a novel valence-probe visual discrimination (VPVD) and a probabilistic reversal learning procedure (PRL). Systemic administration of a 5-HT2CR agonist and antagonist resulted in selective changes in the balance of feedback sensitivity bias on these tasks. Specifically, on VPVD, SB 242084, the 5-HT2CR antagonist, impaired acquisition of a discrimination dependent on appropriate integration of positive and negative feedback. On PRL, SB 242084 at 1 mg/kg resulted in changes in behaviour consistent with reduced sensitivity to positive feedback. In contrast, WAY 163909, the 5-HT2CR agonist, resulted in changes associated with increased sensitivity to positive feedback and decreased sensitivity to negative feedback. These results suggest that 5-HT2CRs tightly regulate feedback sensitivity bias in mice with consequent effects on learning and cognitive flexibility and specify a framework for the influence of 5-HT2CRs on sensitivity to reinforcement.

  8. 5-HT2a receptor in mPFC influences context-guided reconsolidation of object memory in perirhinal cortex.

    PubMed

    Morici, Juan Facundo; Miranda, Magdalena; Gallo, Francisco Tomás; Zanoni, Belén; Bekinschtein, Pedro; Weisstaub, Noelia V

    2018-05-02

    Context-dependent memories may guide adaptive behavior relaying in previous experience while updating stored information through reconsolidation. Retrieval can be triggered by partial and shared cues. When the cue is presented, the most relevant memory should be updated. In a contextual version of the object recognition task, we examined the effect of medial PFC (mPFC) serotonin 2a receptor (5-HT2aR) blockade during retrieval in reconsolidation of competing objects memories. We found that mPFC 5-HT2aR controls retrieval and reconsolidation of object memories in the perirhinal cortex (PRH), but not in the dorsal hippocampus in rats. Also, reconsolidation of objects memories in PRH required a functional interaction between the ventral hippocampus and the mPFC. Our results indicate that in the presence of conflicting information at retrieval, mPFC 5-HT2aR may facilitate top-down context-guided control over PRH to control the behavioral response and object memory reconsolidation. © 2018, Morici et al.

  9. 5-HT2a receptor in mPFC influences context-guided reconsolidation of object memory in perirhinal cortex

    PubMed Central

    Morici, Juan Facundo; Miranda, Magdalena; Gallo, Francisco Tomás; Zanoni, Belén; Bekinschtein, Pedro

    2018-01-01

    Context-dependent memories may guide adaptive behavior relaying in previous experience while updating stored information through reconsolidation. Retrieval can be triggered by partial and shared cues. When the cue is presented, the most relevant memory should be updated. In a contextual version of the object recognition task, we examined the effect of medial PFC (mPFC) serotonin 2a receptor (5-HT2aR) blockade during retrieval in reconsolidation of competing objects memories. We found that mPFC 5-HT2aR controls retrieval and reconsolidation of object memories in the perirhinal cortex (PRH), but not in the dorsal hippocampus in rats. Also, reconsolidation of objects memories in PRH required a functional interaction between the ventral hippocampus and the mPFC. Our results indicate that in the presence of conflicting information at retrieval, mPFC 5-HT2aR may facilitate top-down context-guided control over PRH to control the behavioral response and object memory reconsolidation. PMID:29717980

  10. Effect of Lactobacillus plantarum Tennozu-SU2 on Salmonella Typhimurium Infection in Human Enterocyte-Like HT-29-Luc Cells and BALB/c Mice.

    PubMed

    Hirano, Shino; Yokota, Yasushi; Eda, Mika; Kuda, Takashi; Shikano, Ayane; Takahashi, Hajime; Kimura, Bon

    2017-03-01

    The probiotic properties and inhibitory effect on Salmonella Typhimurium adhesion on human enterocyte-like HT-29-Luc cells of three Lactobacillus plantarum strains isolated from fermented fish, beach sand and a coastal plant were determined. Compared with the type strain L. plantarum NBRC 15891 T , which was isolated from pickled cabbage, L. plantarum Tennozu-SU2 isolated from the acorn of a coastal tree showed high autoaggregation in de Man, Rogosa and Sharpe (MRS) broth and an antagonistic effect against S. Typhimurium in brain heart infusion (BHI) broth. Furthermore, heat-killed L. plantarum Tennozu-SU2 cells inhibited S. Typhimurium adhesion on HT-29-Luc cells. Both live and heat-killed L. plantarum Tennozu-SU2 cells showed an inhibitory effect on gut colonisation in BALB/c mice, as assessed by viable Salmonella count in faecal samples and by invasion into liver and spleen tissues. The properties shown in this study suggest that L. plantarum Tennozu-SU2 is useful as a starter and probiotic bacteria in functional food material.

  11. A Structure-Function Analysis of Shiga-Like Toxin Type 2 of Enterohemorrhagic Escherichia Coli

    DTIC Science & Technology

    1990-05-07

    like toxins are summarized in Table 2. The genes coding for both SLT-I and SLT-II are borne on coliphage , and toxin expression by E. coli occurs as...A stock | | ’ • * suspension of toxin-converting W coliphage was prepared by inducing the phage from | "fif the E. coli C600(933W) lysogen with...mitomycin C as described previously (Marques et al., 1987). An appropriate amount of the W coliphage stock was added to an l| exponential culture of

  12. A globotetraosylceramide (Gb₄) receptor-based ELISA for quantitative detection of Shiga toxin 2e.

    PubMed

    Togashi, Katsuhiro; Sasaki, Shiho; Sato, Wataru

    2015-08-01

    Currently, no simple assays are available for routine quantitative detection of Escherichia coli-produced Shiga toxin 2e (Stx2e) that causes porcine edema disease. Here, we present a novel quantitative detection method for Stx2e based on the measurement of Stx2e binding to the specific globotetraosylceramide (Gb4) receptor by ELISA (Gb4-ELISA). No cross-reactivity was found with the other Shiga toxins Stx1 and Stx2, indicating high specificity. When the recombinant Stx2e B subunit (Stx2eB) was used, the absorbance measured by Gb4-ELISA increased linearly with Stx2eB concentration in the range of 20-2,500 ng/ml. The Gb4-ELISA method can be easily performed, suggesting that it would be a useful diagnostic tool for porcine edema disease.

  13. Interaction of Clostridium perfringens delta toxin with erythrocyte and liposome membranes and relation with the specific binding to the ganglioside GM2.

    PubMed

    Jolivet-Reynaud, C; Hauttecoeur, B; Alouf, J E

    1989-01-01

    The specific interaction of the cytolytic Clostridium perfringens delta toxin with membrane GM2 was indicated by: (i) characterization of this glycolipid in the membrane of sheep and goat erythrocytes, which are lysed by the toxin, whereas GM2 was undetectable in insensitive rabbit erythrocytes, (ii) demonstration of 125I-toxin binding to GM2, by autoradiography, following incubation with thin-layer chromatograms containing separated neuroblastoma gangliosides, and (iii) toxin fixation by phospholipid-cholesterol unilamellar vesicles containing either sheep gangliosides or GM2. In order to investigate the intramembrane events leading to membrane disruption following toxin binding, the photoreactive probe 12(4-azido-2-nitrophenoxy)stearoyl 1-14C glucosamine, which inserts into the outer layer and labels integral membrane proteins, was used to establish whether delta toxin penetrates into target cell membrane. No toxin labeling was found, suggesting that toxin action takes place at the membrane surface. This contention is supported by the observation that despite toxin binding, GM2 liposomes did not release entrapped 14C-glucose. Treatment of toxin with carboxypeptidases, but not aminopeptidases, abolished both toxin binding capacity onto erythrocytes and its combination with antitoxin neutralizing antibodies, suggesting that the carboxy terminal end of the toxin is critical for binding to cell membrane.

  14. Expression of Receptors for Tetanus Toxin and Monoclonal Antibody A2B5 by Pancreatic Islet Cells

    NASA Astrophysics Data System (ADS)

    Eisenbarth, G. S.; Shimizu, K.; Bowring, M. A.; Wells, S.

    1982-08-01

    Studies of the reaction of antibody A2B5 and tetanus toxin with pancreatic islet cells, islet cell tumors, and other human amine precursor uptake and decarboxylation (APUD) tumors are described. By indirect immunofluorescence, antibody A2B5 and tetanus toxin were shown to specifically bind to the plasma membrane of human, rat, chicken, and mouse islet cells. The binding of antibody A2B5 to the cell surface of living islet cells has allowed isolation of these cells from a suspension of pancreatic cells by using a fluorescence-activated cell sorter. In studies designed to determine whether tetanus toxin and antibody A2B5 bound to the same surface antigen, A2B5 and tetanus toxin did not compete for binding to normal islet cells, a human islet cell tumor, or a rat islet cell tumor. In addition to binding to islet cell tumors, antibody A2B5 reacts with frozen sections, isolated cells, and cell lines of neural, neural crest, and APUD origin.

  15. Lysogeny with Shiga Toxin 2-Encoding Bacteriophages Represses Type III Secretion in Enterohemorrhagic Escherichia coli

    PubMed Central

    Xu, Xuefang; McAteer, Sean P.; Tree, Jai J.; Shaw, Darren J.; Wolfson, Eliza B. K.; Beatson, Scott A.; Roe, Andrew J.; Allison, Lesley J.; Chase-Topping, Margo E.; Mahajan, Arvind; Tozzoli, Rosangela; Woolhouse, Mark E. J.; Morabito, Stefano; Gally, David L.

    2012-01-01

    Lytic or lysogenic infections by bacteriophages drive the evolution of enteric bacteria. Enterohemorrhagic Escherichia coli (EHEC) have recently emerged as a significant zoonotic infection of humans with the main serotypes carried by ruminants. Typical EHEC strains are defined by the expression of a type III secretion (T3S) system, the production of Shiga toxins (Stx) and association with specific clinical symptoms. The genes for Stx are present on lambdoid bacteriophages integrated into the E. coli genome. Phage type (PT) 21/28 is the most prevalent strain type linked with human EHEC infections in the United Kingdom and is more likely to be associated with cattle shedding high levels of the organism than PT32 strains. In this study we have demonstrated that the majority (90%) of PT 21/28 strains contain both Stx2 and Stx2c phages, irrespective of source. This is in contrast to PT 32 strains for which only a minority of strains contain both Stx2 and 2c phages (28%). PT21/28 strains had a lower median level of T3S compared to PT32 strains and so the relationship between Stx phage lysogeny and T3S was investigated. Deletion of Stx2 phages from EHEC strains increased the level of T3S whereas lysogeny decreased T3S. This regulation was confirmed in an E. coli K12 background transduced with a marked Stx2 phage followed by measurement of a T3S reporter controlled by induced levels of the LEE-encoded regulator (Ler). The presence of an integrated Stx2 phage was shown to repress Ler induction of LEE1 and this regulation involved the CII phage regulator. This repression could be relieved by ectopic expression of a cognate CI regulator. A model is proposed in which Stx2-encoding bacteriophages regulate T3S to co-ordinate epithelial cell colonisation that is promoted by Stx and secreted effector proteins. PMID:22615557

  16. Effects of Toxins on Arachidonic Acid Metabolism in Cultured Rat Pulmonary Alveolar Macrophages

    DTIC Science & Technology

    1988-12-28

    response to’toixin exposure, and fluocinolone may protect against .T-2 toxicosis. Some natural toxins are potent a nd powerful inflammtatory agents (1,2...macrophages in the inflammatory response to natural toxins, we examined the effect of T-2, microcystin-LR known inflammatory agents, and also included...effective in inducing the release of arachidonic acid metabolites, probably due to non-inflammatory nature of the toxin. We observed a large increase in

  17. An immunocapture/scintillation proximity analysis of G alpha q/11 activation by native serotonin (5-HT)2A receptors in rat cortex: blockade by clozapine and mirtazapine.

    PubMed

    Mannoury La Cour, C; Chaput, C; Touzard, M; Millan, M J

    2009-02-01

    Though transduction mechanisms recruited by heterologously expressed 5-HT(2A) receptors have been extensively studied, their interaction with specific subtypes of G-protein remains to be directly evaluated in cerebral tissue. Herein, as shown by an immunocapture/scintillation proximity analysis, 5-HT, the prototypical 5-HT(2A) agonist, DOI, and Ro60,0175 all enhanced [(35)S]GTPgammaS binding to G alpha q/11 in rat cortex with pEC(50) values of 6.22, 7.24 and 6.35, respectively. No activation of G o or G s/olf was seen at equivalent concentrations of DOI. Stimulation of G alpha q/11 by 5-HT (30 microM) and DOI (30 microM) was abolished by the selective 5-HT(2A) vs. 5-HT(2C)/5-HT(2B) antagonists, ketanserin (pK(B) values of 9.11 and 8.88, respectively) and MDL100,907 (9.82 and 9.68). By contrast, 5-HT-induced [(35)S]GTPgammaS binding to G alpha q/11 was only weakly inhibited by the preferential 5-HT(2C) receptor antagonists, RS102,221 (6.94) and SB242,084 (7.39), and the preferential 5-HT(2B) receptor antagonist, LY266,097 (6.66). The antipsychotic, clozapine, which had marked affinity for 5-HT(2A) receptors, blocked the recruitment of G alpha q/11 by 5-HT and DOI with pK(B) values of 8.54 and 8.14, respectively. Its actions were mimicked by the "atypical" antidepressant and 5-HT(2A) receptor antagonist, mirtazapine, which likewise blocked 5-HT and DOI-induced G alpha q/11 protein activation with pK(B) values of 7.90 and 7.76, respectively. In conclusion, by use of an immunocapture/scintillation proximity strategy, this study shows that native 5-HT(2A) receptors in rat frontal cortex specifically recruit G alpha q/11 and that this action is blocked by clozapine and mirtazapine. Quantification of 5-HT(2A) receptor-mediated G alpha q/11 activation in frontal cortex should prove instructive in characterizing the actions of diverse classes of psychotropic agent. 2008 Wiley-Liss, Inc.

  18. Strain differences in basal and post-citalopram extracellular 5-HT in the mouse medial prefrontal cortex and dorsal hippocampus: relation with tryptophan hydroxylase-2 activity.

    PubMed

    Calcagno, E; Canetta, A; Guzzetti, S; Cervo, L; Invernizzi, R W

    2007-11-01

    We used the microdialysis technique to compare basal extracellular serotonin (5-HT) and the response to citalopram in different strains of mice with functionally different allelic forms of tryptophan hydroxylase-2 (TPH-2), the rate-limiting enzyme in brain 5-HT synthesis. DBA/2J, DBA/2N and BALB/c mice carrying the 1473G allele of TPH-2 had less dialysate 5-HT in the medial prefrontal cortex and dorsal hippocampus (DH) (20-40% reduction) than C57BL/6J and C57BL/6N mice carrying the 1473C allele. Extracellular 5-HT estimated by the zero-net flux method confirmed the result of conventional microdialysis. Citalopram, 1.25, 5 and 20 mg/kg, dose-dependently raised extracellular 5-HT in the medial prefrontal cortex of C57BL/6J mice, with maximum effect at 5 mg/kg, but had significantly less effect in DBA/2J and BALB/c mice and in the DH of DBA/2J mice. A tryptophan (TRP) load enhanced basal extracellular 5-HT in the medial prefrontal cortex of DBA/2J mice but did not affect citalopram's ability to raise cortical and hippocampal extracellular 5-HT. The impairment of 5-HT synthesis quite likely accounts for the reduction of basal 5-HT and the citalopram-induced rise in mice carrying the mutated enzyme. These findings might explain why DBA/2 and BALB/c mice do not respond to citalopram in the forced swimming test. Although TRP could be a useful strategy to improve the antidepressant effect of citalopram (Cervo et al. 2005), particularly in subjects with low 5-HT synthesis, the contribution of serotonergic and non-serotonergic mechanisms to TRP's effect remains to be elucidated.

  19. Reversal of sibutramine-induced anorexia with a selective 5-HT(2C) receptor antagonist.

    PubMed

    Higgs, Suzanne; Cooper, Alison J; Barnes, Nicholas M

    2011-04-01

    The monoamine reuptake inhibitor sibutramine reduces food intake but the receptor subtypes mediating the effects of sibutramine on feeding remain to be clearly identified. The involvement of the 5-HT(2C) receptor subtype in the satiety-enhancing effects of sibutramine was investigated by examining the effects of co-administration of sibutramine with the selective 5-HT(2C) receptor antagonist SB 242084 Microstructural analyses of licking for a glucose solution by non-deprived, male rats were performed over a range of doses of sibutramine to identify a selective satiety-enhancing dose (experiment 1). Similar analyses were performed after administration of a vehicle control, two doses of SB 242084 alone or two doses of SB 242084 in combination with sibutramine (experiment 2). Sibutramine at doses of 1-3 mg/kg selectively reduced glucose consumption via a reduction in the number of bouts of licking. Non-selective effects to increase latency to lick were only observed at the higher dose of 6 mg/kg. Co-administration of sibutramine (3 mg/kg) with SB 242084 (1 or 3 mg/kg) reversed the effect of sibutramine on bout number whereas either dose of SB 242084 alone had no significant effect. We confirm behaviourally selective effects of sibutramine on feeding and provide further support for the satiety-enhancing effects of sibutramine. Our data also provide evidence for the involvement of the 5-HT(2C) receptor in the satiety-enhancing effects of sibutramine although additional targets may have an impact, and further investigation of the molecular mechanisms underlying the efficacy of sibutramine as an anorectic is warranted.

  20. Enhanced Charge Separation in Ternary P3HT/PCBM/CuInS2 Nanocrystals Hybrid Solar Cells

    PubMed Central

    Lefrançois, Aurélie; Luszczynska, Beata; Pepin-Donat, Brigitte; Lombard, Christian; Bouthinon, Benjamin; Verilhac, Jean-Marie; Gromova, Marina; Faure-Vincent, Jérôme; Pouget, Stéphanie; Chandezon, Frédéric; Sadki, Saïd; Reiss, Peter

    2015-01-01

    Geminate recombination of bound polaron pairs at the donor/acceptor interface is one of the major loss mechanisms in organic bulk heterojunction solar cells. One way to overcome Coulomb attraction between opposite charge carriers and to achieve their full dissociation is the introduction of high dielectric permittivity materials such as nanoparticles of narrow band gap semiconductors. We selected CuInS2 nanocrystals of 7.4 nm size, which present intermediate energy levels with respect to poly(3-hexylthiophene) (P3HT) and Phenyl-C61-butyric acid methyl ester (PCBM). Efficient charge transfer from P3HT to nanocrystals takes place as evidenced by light-induced electron spin resonance. Charge transfer between nanocrystals and PCBM only occurs after replacing bulky dodecanethiol (DDT) surface ligands with shorter 1,2-ethylhexanethiol (EHT) ligands. Solar cells containing in the active layer a ternary blend of P3HT:PCBM:CuInS2-EHT nanocrystals in 1:1:0.5 mass ratio show strongly improved short circuit current density and a higher fill factor with respect to the P3HT:PCBM reference device. Complementary measurements of the absorption properties, external quantum efficiency and charge carrier mobility indicate that enhanced charge separation in the ternary blend is at the origin of the observed behavior. The same trend is observed for blends using the glassy polymer poly(triarylamine) (PTAA). PMID:25588811

  1. Chronic fluoxetine treatment accelerates kindling epileptogenesis in mice independently of 5-HT2A receptors.

    PubMed

    Li, Crystal; Silva, Juliana; Ozturk, Ezgi; Dezsi, Gabriella; O'Brien, Terence J; Renoir, Thibault; Jones, Nigel C

    2018-06-01

    Patients with epilepsy often have mood disorders, and these are commonly treated with antidepressant drugs. Although these drugs are often successful in mitigating depressive symptoms, how they affect the epileptogenic processes has been little studied. Recent evidence has demonstrated that treatment with selective serotonin reuptake inhibitor (SSRI) antidepressant drugs adversely promotes epileptogenesis, which may be of great concern considering the number of patients exposed to these drugs. This study investigated 5-HT 2A receptor signaling as a potential mechanism driving the pro-epileptogenic effects of the prototypical SSRI fluoxetine. Male homozygous 5-HT 2A receptor knockout mice or wild-type littermates (n = 9-14/group) were treated with continuous fluoxetine (10 mg kg -1 d -1 , sc) or vehicle and subjected to electrical kindling of the amygdala. Compared to vehicle, fluoxetine treatment accelerated kindling epileptogenesis (P < .001), but there was no effect of genotype (P = .75), or any treatment x genotype interaction observed (P = .90). Of interest, fluoxetine treatment increased afterdischarge thresholds in both genotypes (P = .007). We conclude that treatment with fluoxetine promotes epileptogenesis in mice, but this effect is not mediated by 5-HT 2A receptors. This suggests that antidepressants may accelerate the onset of acquired epilepsy in patients who have experienced epileptogenic cerebral insults. Wiley Periodicals, Inc. © 2018 International League Against Epilepsy.

  2. Serotonin transporter, 5-HT1A receptor, and behavior in DBA/2J mice in comparison with four inbred mouse strains.

    PubMed

    Popova, Nina K; Naumenko, Vladimir S; Tibeikina, Marina A; Kulikov, Alexander V

    2009-12-01

    Prepulse inhibition (PPI), the reduction in acoustic startle produced when it is preceded by a weak prepulse stimulus, is impaired in schizophrenic patients. The DBA/2J mouse strain displayed deficient PPI and is therefore suggested as an experimental animal model for the loss of sensorimotor gating in schizophrenia. Brain serotonin (5-HT) has been implicated in the pathophysiology of several psychiatric disorders, including major depressive disorder and schizophrenia. In the present study, behavior, 5-HT transporter (5-HTT) mRNA level, 5-HT(1A) receptor mRNA level, and 5-HT(1A) receptor density in the brain regions were studied in DBA/2J mice in comparison with four inbred mouse strains (CBA/Lac, C57BL/6, BALB/c, and ICR). A decrease in 5-HTT mRNA level in the midbrain and a reduced density of 5-HT(1A) receptors in the frontal cortex without significant changes in 5-HT(1A) receptor mRNA level in DBA/2J mice were found. It was shown that, along with decreased PPI, DBA/2J mice demonstrated considerably reduced immobility in the tail suspension test and in the forced swim test. No significant interstrain differences in intermale aggression, or in light-dark box and elevated plus-maze tests, were found. The results suggested the involvement of decreased 5-HTT gene expression and 5-HT(1A) receptor density in genetically defined PPI deficiency and showed a lack of any association between PPI deficiency and predisposition to aggressive, anxiety, and depressive-like behaviors. Copyright 2009 Wiley-Liss, Inc.

  3. Emergence of Escherichia coli encoding Shiga toxin 2f in human Shiga toxin-producing E. coli (STEC) infections in the Netherlands, January 2008 to December 2011.

    PubMed

    Friesema, I; van der Zwaluw, K; Schuurman, T; Kooistra-Smid, M; Franz, E; van Duynhoven, Y; van Pelt, W

    2014-05-01

    The Shiga toxins of Shiga toxin-producing Escherichia coli (STEC) can be divided into Shiga toxin 1 (Stx1) and Shiga toxin 2 (Stx2) with several sub-variants. Variant Stx2f is one of the latest described, but has been rarely associated with symptomatic human infections. In the enhanced STEC surveillance in the Netherlands, 198 STEC O157 cases and 351 STEC non-O157 cases, including 87 stx2f STEC isolates, were reported between 2008 and 2011. Most stx2f strains belonged to the serogroups O63:H6 (n=47, 54%), O113:H6 (n=12, 14%) and O125:H6 (n=12, 14%). Of the 87 stx2f isolates, 84 (97%) harboured the E. coli attaching and effacing (eae) gene, but not the enterohaemorrhagic E. coli haemolysin (hly) gene. stx2f STEC infections show milder symptoms and a less severe clinical course than STEC O157 infections. Almost all infections with stx2f (n=83, 95%) occurred between June and December, compared to 170/198 (86%) of STEC O157 and 173/264 (66%) of other STEC non-O157. stx2f STEC infections in the Netherlands are more common than anticipated, and form a distinct group within STEC with regard to virulence genes and the relatively mild disease.

  4. Aromatic interactions impact ligand binding and function at serotonin 5-HT2C G protein-coupled receptors: receptor homology modelling, ligand docking, and molecular dynamics results validated by experimental studies

    NASA Astrophysics Data System (ADS)

    Córdova-Sintjago, Tania; Villa, Nancy; Fang, Lijuan; Booth, Raymond G.

    2014-02-01

    The serotonin (5-hydroxytryptamine, 5-HT) 5-HT2 G protein-coupled receptor (GPCR) family consists of types 2A, 2B, and 2C that share ∼75% transmembrane (TM) sequence identity. Agonists for 5-HT2C receptors are under development for psychoses; whereas, at 5-HT2A receptors, antipsychotic effects are associated with antagonists - in fact, 5-HT2A agonists can cause hallucinations and 5-HT2B agonists cause cardiotoxicity. It is known that 5-HT2A TM6 residues W6.48, F6.51, and F6.52 impact ligand binding and function; however, ligand interactions with these residues at the 5-HT2C receptor have not been reported. To predict and validate molecular determinants for 5-HT2C-specific activation, results from receptor homology modelling, ligand docking, and molecular dynamics simulation studies were compared with experimental results for ligand binding and function at wild type and W6.48A, F6.51A, and F6.52A point-mutated 5-HT2C receptors.

  5. 5-HT(2C) receptor RNA editing in the amygdala of C57BL/6J, DBA/2J, and BALB/cJ mice.

    PubMed

    Hackler, Elizabeth A; Airey, David C; Shannon, Caitlin C; Sodhi, Monsheel S; Sanders-Bush, Elaine

    2006-05-01

    Post-transcriptional RNA editing of the G-protein coupled 5-hydroxytryptamine-2C (5-HT(2C)) receptor predicts an array of 24 receptor isoforms, some of which are characterized by reduced constitutive activity and potency to initiate intracellular signaling. The amygdala is integral to anxiety, fear, and related psychiatric diseases. Activation of 5-HT(2C) receptors within the amygdala is anxiogenic. Here, we describe the RNA editing profiles from amygdala of two inbred mouse strains (BALB/cJ and DBA/2J) known to be more anxious than a third (C57BL/6J). We confirmed the strain anxiety differences using light<-->dark exploration, and we discovered that BALB/cJ and DBA/2J are each characterized by a higher functioning RNA editing profile than C57BL/6J. BALB/cJ and DBA/2J exhibit a roughly two-fold reduction in C site editing, and a corresponding two-fold reduction in the edited isoform VSV. C57BL/6J is characterized by a relative decrease in the unedited highly functional isoform INI. We estimated the heritability of editing at the C site to be approximately 40%. By sequencing genomic DNA, we found complete conservation between C57BL/6J, BALB/cJ, DBA/2J and 37 other inbred strains for the RNA edited region of Htr2c, suggesting Htr2c DNA sequence does not influence variation in Htr2c RNA editing between inbred strains of mice. We did, however, discover that serotonin turnover is reduced in BALB/cJ and DBA/2J, consistent with emerging evidence that synaptic serotonin levels regulate RNA editing. These results encourage further study of the causes and consequences of 5-HT(2C) receptor RNA editing in the amygdala of mice.

  6. Preclinical safety assessment of the 5-HT2A receptor agonist PET radioligand [ 11C]Cimbi-36.

    PubMed

    Ettrup, Anders; Holm, Søren; Hansen, Martin; Wasim, Muhammad; Santini, Martin Andreas; Palner, Mikael; Madsen, Jacob; Svarer, Claus; Kristensen, Jesper Langgaard; Knudsen, Gitte Moos

    2013-08-01

    [11C]Cimbi-36 was recently developed as an agonist radioligand for brain imaging of serotonin 2A receptors (5-HT2A) with positron emission tomography (PET). This may be used to quantify the high-affinity state of 5-HT2A receptors and may have the potential to quantify changes in cerebral 5-HT levels in vivo. We here investigated safety aspects related to clinical use of [11C]Cimbi-36, including radiation dosimetry and in vivo pharmacology. [11C]Cimbi-36 was injected in rats or pigs, and radiation dosimetry was examined by ex vivo dissection or with PET scanning, respectively. Based on animal data, the Organ Level INternal Dose Assessment software was used to estimate extrapolated human dosimetry for [11C]Cimbi-36. The 5-HT2A receptor agonist actions of [11C]Cimbi-36 in vivo pharmacological effects in mice elicited by increasing doses of Cimbi-36 were assessed with the head-twitch response (HTR). The effective dose as extrapolated from both rat and pig data was low, 7.67 and 4.88 μSv/MBq, respectively. In addition, the estimated absorbed radiation dose to human target organs did not exceed safety levels. Administration of 0.5 mg/kg Cimbi-36 leads to significant HTR compared to saline, whereas 0.05 mg/kg Cimbi-36 (doses much larger than those given in conjunction with a PET scan) did not elicit a significant HTR. Administration of tracer doses of [11C]Cimbi-36 does not seem to be associated with unusual radiation burden or adverse clinical effects.

  7. Development of a serum-free co-culture of human intestinal epithelium cell-lines (Caco-2/HT29-5M21)

    PubMed Central

    Nollevaux, Géraldine; Devillé, Christelle; El Moualij, Benaïssa; Zorzi, Willy; Deloyer, Patricia; Schneider, Yves-Jacques; Peulen, Olivier; Dandrifosse, Guy

    2006-01-01

    Background The absorptive and goblet cells are the main cellular types encountered in the intestine epithelium. The cell lineage Caco-2 is a model commonly used to reproduce the features of the bowel epithelium. However, there is a strong debate regarding the value of Caco-2 cell culture to mimick in vivo situation. Indeed, some authors report in Caco-2 a low paracellular permeability and an ease of access of highly diffusible small molecules to the microvilli, due to an almost complete lack of mucus. The HT29-5M21 intestinal cell lineage is a mucin-secreting cellular population. A co-culture system carried out in a serum-free medium and comprising both Caco-2 and HT29-5M21 cells was developed. The systematic use of a co-culture system requires the characterization of the monolayer under a given experimental procedure. Results In this study, we investigated the activity and localization of the alkaline phosphatase and the expression of IAP and MUC5AC genes to determine a correlation between these markers and the cellular composition of a differentiated monolayer obtained from a mixture of Caco-2 and HT29-5M21 cells. We observed that the culture conditions used (serum-free medium) did not change the phenotype of each cell type, and produced a reproducible model. The alkaline phosphatase expression characterizing Caco-2 cells was influenced by the presence of HT29-5M21 cells. Conclusion The culture formed by 75% Caco-2 and 25% HT29-5M21 produce a monolayer containing the two main cell types of human intestinal epithelium and characterized by a reduced permeability to macromolecules. PMID:16670004

  8. The 5-HT{sub 2A} serotoninergic receptor is expressed in the MCF-7 human breast cancer cell line and reveals a mitogenic effect of serotonin

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Sonier, Brigitte; Arseneault, Madeleine; Institut National de la Recherche Scientifique-Institut Armand-Frappier, Montreal, Que.

    2006-05-19

    Serotonin (5-hydroxytryptamine, 5-HT) has been described as a mitogen in a variety of cell types and carcinomas. It exerts its mitogenic effect by interacting with a wide range of 5-HT receptor types. Certain studies suggest that some selective serotonin re-uptake inhibitors promote breast cancer in animals and humans. This study attempts to clarify the role of serotonin in promoting the growth of neoplastic mammary cells. Expression of the 5-HT{sub 2A} serotoninergic receptor subtype in MCF-7 cells was determined by RT-PCR, Western blotting, and immunofluorescence analysis. The mitogenic effect of 5-HT on MCF-7 cells was determined by means of the MTTmore » proliferation assay. We have demonstrated that the 5-HT{sub 2A} receptor subtype is fully expressed in the MCF-7 human breast cancer cell line, in terms of encoding mRNA and receptor protein. Automated sequencing has confirmed that the 5-HT{sub 2A} receptor present in this cell line is identical to the 5-HT{sub 2A} receptor found in human platelets and in human cerebral cortex. Furthermore, this receptor was found by immunofluorescence to be on the plasma membrane. MTT proliferation assays revealed that 5-HT and DOI, a selective 5-HT{sub 2A} receptor subtype agonist, stimulated MCF-7 cell. These results indicate that 5-HT plays a mitogenic role in neoplastic mammary cells. Our data also indicate that 5-HT exerts this positive growth effect on MCF-7 cells through, in part, the 5-HT{sub 2A} receptor subtype, which is fully expressed in this cell line.« less

  9. Top-down proteomic identification of Shiga toxin 2 subtypes from Shiga toxin-producing Escherichia coli by Matrix-Assisted Laser Desorption Ionization-Tandem Time of Flight mass spectrometry

    USDA-ARS?s Scientific Manuscript database

    We have analyzed 26 Shiga toxin-producing Escherichia coli (STEC) strains for Shiga toxin 2 (Stx2) production using matrix-assisted laser desorption/ionization time-of-flight-time-of-flight tandem mass spectrometry (MALDI-TOF-TOF-MS/MS) and top-down proteomic analysis. STEC strains were induced to ...

  10. Activation of lumbosacral 5-HT2C receptors induces bursts of rhythmic activity in sympathetic nerves to the vas deferens in male rats

    PubMed Central

    Stafford, Stuart A; Tang, Kim; Coote, John H

    2006-01-01

    We previously demonstrated that p-chloroamphetamine (PCA) intravenously (i.v.) evokes a specific patterned bursting response in the vas deferens nerve (VDN) of anaesthetised male rats that is associated with contraction of the vas deferens, and ejaculation and contraction of the bulbospongiosus muscles. The present study used selective 5-HT agonists to induce similar rhythmic bursting responses in the VDN in order to reveal the 5-HT receptor subtypes involved. The 5-HT2C receptor agonist (1.0 mg kg−1 Ro600175 i.v.) evoked the characteristic bursting pattern responses in the VDN. The 5-HT1A receptor agonist (1.0 mg kg−1 8-OH-DPAT i.v.) failed to elicit any responses. However, 8-OH-DPAT coadministered in combination with Ro600175 induced a potentiation of the responses. Responses were also evoked in rats with a mid-thoracic spinalisation, with a more predictable response being observed following the combination of agonists. This suggests an action of both agonists in the lumbosacral spinal cord. Responses were blocked by 0.5 mg kg−1 SB206553 i.v. (5-HT2B/C receptor antagonist) or 0.5 mg kg−1 WAY100635 i.v. (5-HT1A receptor antagonist), but not 0.1 or 1.0 mg kg−1 SB269970 i.v. (5-HT7 receptor antagonist). We suggest that activation of 5-HT2C and 5-HT1A receptor subtypes synergistically elicits contraction of the vas deferens through the activation of sympathetic preganglionic neurones in the spinal cord. These data support the idea of a proejaculatory action of 5-HT2C receptors in the lumbosacral spinal cord, suggesting a descending 5-HT excitatory pathway in addition to a 5-HT inhibitory pathway. An excitatory action of 8-OH-DPAT at lumbosacral sites is also evident. PMID:16799648

  11. Cannabis Users Show Enhanced Expression of CB1-5HT2A Receptor Heteromers in Olfactory Neuroepithelium Cells.

    PubMed

    Galindo, Liliana; Moreno, Estefanía; López-Armenta, Fernando; Guinart, Daniel; Cuenca-Royo, Aida; Izquierdo-Serra, Mercè; Xicota, Laura; Fernandez, Cristina; Menoyo, Esther; Fernández-Fernández, José M; Benítez-King, Gloria; Canela, Enric I; Casadó, Vicent; Pérez, Víctor; de la Torre, Rafael; Robledo, Patricia

    2018-01-02

    Cannabinoid CB1 receptors (CB 1 R) and serotonergic 2A receptors (5HT 2A R) form heteromers in the brain of mice where they mediate the cognitive deficits produced by delta-9-tetrahydrocannabinol. However, it is still unknown whether the expression of this heterodimer is modulated by chronic cannabis use in humans. In this study, we investigated the expression levels and functionality of CB 1 R-5HT 2A R heteromers in human olfactory neuroepithelium (ON) cells of cannabis users and control subjects, and determined their molecular characteristics through adenylate cyclase and the ERK 1/2 pathway signaling studies. We also assessed whether heteromer expression levels correlated with cannabis consumption and cognitive performance in neuropsychological tests. ON cells from controls and cannabis users expressed neuronal markers such as βIII-tubulin and nestin, displayed similar expression levels of genes related to cellular self-renewal, stem cell differentiation, and generation of neural crest cells, and showed comparable Na + currents in patch clamp recordings. Interestingly, CB 1 R-5HT 2A R heteromer expression was significantly increased in cannabis users and positively correlated with the amount of cannabis consumed, and negatively with age of onset of cannabis use. In addition, a negative correlation was found between heteromer expression levels and attention and working memory performance in cannabis users and control subjects. Our findings suggest that cannabis consumption regulates the formation of CB 1 R-5HT 2A R heteromers, and may have a key role in cognitive processing. These heterodimers could be potential new targets to develop treatment alternatives for cognitive impairments.

  12. Chronic treatment with LY341495 decreases 5-HT(2A) receptor binding and hallucinogenic effects of LSD in mice.

    PubMed

    Moreno, José L; Holloway, Terrell; Rayannavar, Vinayak; Sealfon, Stuart C; González-Maeso, Javier

    2013-03-01

    Hallucinogenic drugs, such as lysergic acid diethylamide (LSD), mescaline and psilocybin, alter perception and cognitive processes. All hallucinogenic drugs have in common a high affinity for the serotonin 5-HT(2A) receptor. Metabotropic glutamate 2/3 (mGlu2/3) receptor ligands show efficacy in modulating the cellular and behavioral responses induced by hallucinogenic drugs. Here, we explored the effect of chronic treatment with the mGlu2/3 receptor antagonist 2S-2-amino-2-(1S,2S-2-carboxycyclopropan-1-yl)-3-(xanth-9-yl)-propionic acid (LY341495) on the hallucinogenic-like effects induced by LSD (0.24mg/kg). Mice were chronically (21 days) treated with LY341495 (1.5mg/kg), or vehicle, and experiments were carried out one day after the last injection. Chronic treatment with LY341495 down-regulated [(3)H]ketanserin binding in somatosensory cortex of wild-type, but not mGlu2 knockout (KO), mice. Head-twitch behavior, and expression of c-fos, egr-1 and egr-2, which are responses induced by hallucinogenic 5-HT(2A) agonists, were found to be significantly decreased by chronic treatment with LY341495. These findings suggest that repeated blockade of the mGlu2 receptor by LY341495 results in reduced 5-HT(2A) receptor-dependent hallucinogenic effects of LSD. Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.

  13. Chronic treatment with LY341495 decreases 5-HT2A receptor binding and hallucinogenic effects of LSD in mice

    PubMed Central

    Moreno, José L.; Holloway, Terrell; Rayannavar, Vinayak; Sealfon, Stuart C.; González-Maeso, Javier

    2013-01-01

    Hallucinogenic drugs, such as lysergic acid diethylamide (LSD), mescaline and psilocybin, alter perception and cognitive processes. All hallucinogenic drugs have in common a high affinity for the serotonin 5-HT2A receptor. Metabotropic glutamate 2/3 (mGlu2/3) receptor ligands show efficacy in modulating the cellular and behavioral responses induced by hallucinogenic drugs. Here, we explored the effect of chronic treatment with the mGlu2/3 receptor antagonist 2S-2-amino-2-(1S,2S-2-carboxycyclopropan-1-yl)-3-(xanth-9-yl)-propionic acid (LY341495) on the hallucinogenic-like effects induced by LSD (0.24 mg/kg). Mice were chronically (21 days) treated with LY341495 (1.5 mg/kg), or vehicle, and experiments were carried out one day after the last injection. Chronic treatment with LY341495 down-regulated [3H]ketanserin binding in somatosensory cortex of wild-type, but not mGlu2 knockout (KO), mice. Head-twitch behavior, and expression of c-fos, egr-1 and egr-2, which are responses induced by hallucinogenic 5-HT2A agonists, were found to be significantly decreased by chronic treatment with LY341495. These findings suggest that repeated blockade of the mGlu2 receptor by LY341495 results in reduced 5-HT2A receptor-dependent hallucinogenic effects of LSD. PMID:23333599

  14. Updated survey of Fusarium species and toxins in Finnish cereal grains.

    PubMed

    Hietaniemi, Veli; Rämö, Sari; Yli-Mattila, Tapani; Jestoi, Marika; Peltonen, Sari; Kartio, Mirja; Sieviläinen, Elina; Koivisto, Tauno; Parikka, Päivi

    2016-05-01

    The aim of the project was to produce updated information during 2005-14 on the Fusarium species found in Finnish cereal grains, and the toxins produced by them, as the last comprehensive survey study of Fusarium species and their toxins in Finland was carried out at the turn of the 1960s and the 1970s. Another aim was to use the latest molecular and chemical methods to investigate the occurrence and correlation of Fusarium species and their mycotoxins in Finland. The most common Fusarium species found in Finland in the FinMyco project 2005 and 2006 were F. avenaceum, F. culmorum, F. graminearum, F. poae, F. sporotrichioides and F. langsethiae. F. avenaceum was the most dominant species in barley, spring wheat and oat samples. The occurrence of F. culmorum and F. graminearum was high in oats and barley. Infection by Fusarium fungi was the lowest in winter cereal grains. The incidence of Fusarium species in 2005 was much higher than in 2006 due to weather conditions. F. langsethiae has become much more common in Finland since 2001. F. graminearum has also risen in the order of importance. A highly significant correlation was found between Fusarium graminearum DNA and deoxynivalenol (DON) levels in Finnish oats, barley and wheat. When comparing the FinMyco data in 2005-06 with the results of the Finnish safety monitoring programme for 2005-14, spring cereals were noted as being more susceptible to infection by Fusarium fungi and the formation of toxins. The contents of T-2 and HT-2 toxins and the frequency of exceptionally high DON concentrations all increased in Finland during 2005-14. Beauvericin (BEA), enniatins (ENNs) and moniliformin (MON) were also very common contaminants of Finnish grains in 2005-06. Climate change is leading to warmer weather, and this may indicate more changes in Finnish Fusarium mycobiota and toxin contents and profiles in the near future.

  15. Inhibition of Clostridium difficile toxin A and B by 1,2-cyclohexanedione modification of an arginine residue.

    PubMed

    Balfanz, J; Rautenberg, P

    1989-12-29

    Toxin A (enterotoxin) and toxin B (cytotoxin) of Clostridium difficile were both inactivated by the arginine specific reagent 1,2-cyclohexanedione. Molecular stability during the inactivation process was demonstrated by SDS-PAGE analysis showing the same migration rates for modified and unmodified forms of the 230 kDa toxin A and of the 250 kDa toxin B. Cytotoxicity of both toxins as well as mouse lethality of the enterotoxin were drastically decreased as a result of the arginine modification. The reaction followed pseudo-first-order kinetics. Analysis of the data suggested that modification of a single arginine residue was sufficient to abolish the activity of both toxins.

  16. Design, synthesis and structure-activity relationship of novel quinoxalin-2-carboxamides as 5-HT3 receptor antagonists for the management of depression.

    PubMed

    Mahesh, Radhakrishnan; Devadoss, Thangaraj; Pandey, Dilip Kumar; Bhatt, Shvetank; Yadav, Shushil Kumar

    2010-11-15

    A novel series of quinoxalin-2-carboxamides were designed based on the ligand-based approach, employing a three-point pharmacophore model; it consists of an aromatic residue and a linking carbonyl group and a basic nitrogen. The target new chemical entities were synthesized from the key intermediate, quinoxalin-2-carboxylic acid, by coupling it with various amines in the presence of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC·HCl) and 1-hydroxybenzotriazole (HOBt). The obtained compounds' structures were confirmed by spectral data. The target new chemical entities were evaluated for their 5-HT(3) receptor antagonisms in longitudinal muscle myenteric plexus preparation from guinea pig ileum against 5-HT(3) agonist, 2-methyl-5-HT, which was expressed in the form of pA(2) value. All the synthesized compounds showed antagonism towards 5-HT(3) receptor; based on this result, a structure-activity relationship was derived, which reveals that the aromatic residue in 5-HT(3) receptor antagonists may have hydrophobic interaction with 5-HT(3) receptor. Regardless of their antagonistic potentials, all the synthesized molecules were screened for their anti-depressant potentials by using forced swim test in mice model; interestingly none of the tested compounds affect the locomotion of mice in the tested dose levels. Compounds with significant pA(2) values exhibited good anti-depressant-like activity as compared to the vehicle-treated group. Copyright © 2010 Elsevier Ltd. All rights reserved.

  17. Molecular Basis of Differential B-Pentamer Stability of Shiga Toxins 1 and 2

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Conrady, Deborah G.; Flagler, Michael J.; Friedmann, David R.

    2012-06-27

    Escherichia coli strain O157:H7 is a major cause of food poisoning that can result in severe diarrhea and, in some cases, renal failure. The pathogenesis of E. coli O157:H7 is in large part due to the production of Shiga toxin (Stx), an AB{sub 5} toxin that consists of a ribosomal RNA-cleaving A-subunit surrounded by a pentamer of receptor-binding B subunits. There are two major isoforms, Stx1 and Stx2, which differ dramatically in potency despite having 57% sequence identity. Animal studies and epidemiological studies show Stx2 is associated with more severe disease. Although the molecular basis of this difference is unknown,more » data suggest it is associated with the B-subunit. Mass spectrometry studies have suggested differential B-pentamer stability between Stx1 and Stx2. We have examined the relative stability of the B-pentamers in solution. Analytical ultracentrifugation using purified B-subunits demonstrates that Stx2B, the more deadly isoform, shows decreased pentamer stability compared to Stx1B (EC{sub 50} = 2.3 {micro}M vs. EC{sub 50} = 0.043 {micro}M for Stx1B). X-ray crystal structures of Stx1B and Stx2B identified a glutamine in Stx2 (versus leucine in Stx1) within the otherwise strongly hydrophobic interface between B-subunits. Interchanging these residues switches the stability phenotype of the B-pentamers of Stx1 and Stx2, as demonstrated by analytical ultracentrifugation and circular dichroism. These studies demonstrate a profound difference in stability of the B-pentamers in Stx1 and Stx2, illustrate the mechanistic basis for this differential stability, and provide novel reagents to test the basis for differential pathogenicity of these toxins.« less

  18. 5-(Furan-2-yl)-4-(3,4,5-trimethoxyphenyl)-3H-1,2-dithiol-3-one oxime (6f), a new synthetic compound, causes human fibrosarcoma HT-1080 cell apoptosis by disrupting tubulin polymerisation and inducing G2/M arrest.

    PubMed

    Zuo, Daiying; Pang, Lili; Shen, Jiwei; Guan, Qi; Bai, Zhaoshi; Zhang, Huijuan; Li, Yao; Lu, Guodong; Zhang, Weige; Wu, Yingliang

    2017-06-01

    In the current study, we synthesized a series of new compounds targeting tubulin and tested their anti-proliferative activities. Among these new synthetic com-pounds, 5-(furan-2-yl)-4-(3,4,5-trimethoxyphenyl)-3H-1,2-dithiol-3-one oxime (6f) exhibited significant anti-proliferative activity against different human cancer cell lines including human gastric adenocarcinoma SGC-7901, human non-small cell lung cancer A549, and human fibrosarcoma HT-1080. As a result, 6f was selected to further test the sensitivity to different cancer cell lines including human cervical cancer cell line HeLa, human breast cancer cell line MCF-7, non-small cell lung cancer cell line A549, human liver carcinoma cell line HepG-2, human oral squamous cell carcinoma cell lines KB, SGC-7901 and HT-1080. Among these cell lines, HT-1080 and HeLa are the most sensitive. Therefore, HT-1080 was selected to further explore the properties of anti-proliferative activity and the underlying mechanisms. Our data proved that 6f exhibited strong anti-proliferative effects against HT-1080 cells in a time- and dose-dependent manner. We showed that the growth inhibitory effect of 6f in HT-1080 cells was related with microtubule depolymerisation. Molecular docking studies revealed that 6f interacted and bound efficiently with the colchicine-binding site of tubulin. In addition, 6f treatment induced G2/M cell cycle arrest dose-dependently and subsequently induced cell apoptosis. Western blot study indicated that upregulation of cyclin B1 and p-cdc2 was related with G2/M arrest. 6f-induced cell apoptosis was associated with both mitochondrial and death receptor pathway. In conclusion, our data showed that 6f, among the newly synthetic compounds, exhibited highest anti-proliferative activity by disrupting the microtubule polymerisation, causing G2/M arrest and subsequently inducing cell apoptosis in HT-1080 cells. Hence, 6f is a promising microtubule depolymerising agent for the treatment of various cancers

  19. Neurochemical Correlates of Accumbal Dopamine D2 and Amygdaloid 5-HT1B Receptor Densities on Observational Learning of Aggression

    PubMed Central

    Suzuki, Hideo; Lucas, Louis R.

    2015-01-01

    Social learning theory postulates that individuals learn to engage in aggressive behavior through observing an aggressive social model. Prior studies have shown that repeatedly observing aggression, also called “chronic passive exposure to aggression,” changes accumbal dopamine D2 receptor (D2R) and amygdaloid 5-HT1B receptor (5-HT1BR) densities in observers. But, the association between these outcomes remains unknown. Thus, our study used a rat paradigm to comprehensively examine the linkage between aggression, D2R density in the nucleus accumbens core (AcbC) and shell (AcbSh), and 5-HT1BR density in the medial (MeA), basomedial (BMA), and basolateral (BLA) amygdala following chronic passive exposure to aggression. Male Sprague-Dawley rats (N = 72) were passively exposed to either aggression or non-aggression acutely (1 day) or chronically (23 days). When observer rats were exposed to aggression chronically, they showed increased aggressive behavior and reduced D2R density in the bilateral AcbSh. On the other hand, exposure to aggression, regardless of exposure length, increased 5-HT1BR density in the bilateral BLA. Finally, low D2R in the AcbSh significantly interacted with high 5-HT1BR density in the BLA in predicting high levels of aggression in observer rats. Our results advance our understanding of the neurobiological mechanisms for observational learning of aggression, highlighting that dopamine-serotonin interaction, or AcbSh-BLA interaction, may contribute to a risk factor for aggression in observers who chronically witness aggressive interactions. PMID:25650085

  20. Platelet and intestinal 5-HT2A receptor mRNA in autistic spectrum disorders - results of a pilot study.

    PubMed

    Kazek, Beata; Huzarska, Małgorzata; Grzybowska-Chlebowczyk, Urszula; Kajor, Maciej; Ciupińska-Kajor, Monika; Woś, Halina; Marszał, Elzbieta

    2010-01-01

    The etiology and pathogenesis of autistic spectrum disorders (ASD) are still unknown. Platelet hyperserotonemia has been detected in 25-60% of autistic children. Higher incidence of gastrointestinal problems in people with autism is observed. The aim was compare the expression of platelet 5-HT(2A)r mRNA in autistic and non autistic groups. In a subgroup of patients with gastrointestinal problems an upper gastrointestinal tract endoscopy was performed and additionally the expression of 5-HT(2A) receptor mRNA in the duodenum was assessed. The examination was conducted in 79 children - 51 with ASD and 28 without autistic traits. Statistically significant differences between the study and control groups were proven in gastrointestinal problems. The analyses reveal a significantly higher level of 5-HT(2A)r mRNA in platelets of the study group patients, which could suggest serotonin system dysregulation.

  1. Discovery of new anti-depressants from structurally novel 5-HT3 receptor antagonists: design, synthesis and pharmacological evaluation of 3-ethoxyquinoxalin-2-carboxamides.

    PubMed

    Mahesh, Radhakrishnan; Devadoss, Thangaraj; Pandey, Dilip Kumar; Bhatt, Shvetank

    2011-02-15

    A novel series of 3-ethoxyquinoxalin-2-carboxamides were designed as per the pharmacophoric requirements of 5-HT(3) receptor antagonist using ligand-based approach. The desired carboxamides were synthesized from the key intermediate, 3-ethoxyquinoxalin-2-carboxylic acid by coupling with appropriate amines in the presence of 1-(3-dimethylaminopropyl)-3-ethylcarbodiimide hydrochloride (EDC·HCl) and 1-hydroxybenzotriazole (HOBt). The 5-HT(3) receptor antagonism was evaluated in longitudinal muscle myenteric plexus preparation from guinea pig ileum against 5-HT(3) agonist, 2-methy-5-HT, which was expressed in the form of pA(2) values. Compound 6h (3-ethoxyquinoxalin-2-yl)(4-methylpiperazin-1-yl)methanone was found to be the most active compound, which expressed a pA(2) value of 7.7. In forced swim test, the compounds with higher pA(2) value exhibited good anti-depressant-like activity and compounds with lower pA(2) value failed to show activity as compared to the vehicle-treated group. Copyright © 2010 Elsevier Ltd. All rights reserved.

  2. Detection of the index tumour and tumour volume in prostate cancer using T2-weighted and diffusion-weighted magnetic resonance imaging (MRI) alone.

    PubMed

    Rud, Erik; Klotz, Dagmar; Rennesund, Kristin; Baco, Eduard; Berge, Viktor; Lien, Diep; Svindland, Aud; Lundeby, Eskild; Berg, Rolf E; Eri, Lars M; Eggesbø, Heidi B

    2014-12-01

    To examine the performance of T2-weighted (T2W) and diffusion-weighted (DW) magnetic resonance imaging (MRI) for detecting the index tumour in patients with prostate cancer and to examine the agreement between MRI and histology when assessing tumour volume (TV) and overall tumour burden. The study included 199 consecutive patients with biopsy confirmed prostate cancer randomised to MRI before radical prostatectomy from December 2009 to July 2012. MRI-detected tumours (MRTs) were ranked from 1 to 3 according to decreasing volume and were compared with histologically detected tumours (HTs) ranked from 1 to 3, with HT 1 = index tumour. Whole-mount section histology was used as a reference standard. The TVs of true-positive MRTs (MRTVs 1-3) were compared with the TVs found by histology (HTVs 1-3). All tumours were registered on a 30-sector map and by classifying each sector as positive/negative, the rate of true-positive and -negative sectors was calculated. The detection rate for the HT 1 (index tumour) was 92%; HT 2, 45%; and HT 3, 37%. The MRTV 1-3 vs the HTV 1-3 were 2.8 mL vs 4.0 mL (index tumour, P < 0.001), 1.0 mL vs 0.9 mL (tumour 2, P = 0.413), and 0.6 mL vs 0.5 mL (tumour 3, P = 0.492). The rate of true-positive and -negative sectors was 50% and 88%, κ = 0.39. A combination of T2W and DW MRI detects the index tumour in 92% of cases, although MRI underestimates both TV and tumour burden compared with histology. © 2014 The Authors. BJU International © 2014 BJU International.

  3. UDP-Glucosyltransferases from Rice, Brachypodium, and Barley: Substrate Specificities and Synthesis of Type A and B Trichothecene-3-O-β-d-glucosides

    PubMed Central

    Malachová, Alexandra; Piątkowska, Marta; Hametner, Christian; Šofrová, Jana; Jaunecker, Günther; Häubl, Georg; Lemmens, Marc

    2018-01-01

    Trichothecene toxins are confirmed or suspected virulence factors of various plant-pathogenic Fusarium species. Plants can detoxify these to a variable extent by glucosylation, a reaction catalyzed by UDP-glucosyltransferases (UGTs). Due to the unavailability of analytical standards for many trichothecene-glucoconjugates, information on such compounds is limited. Here, the previously identified deoxynivalenol-conjugating UGTs HvUGT13248 (barley), OsUGT79 (rice) and Bradi5g03300 (Brachypodium), were expressed in E. coli, affinity purified, and characterized towards their abilities to glucosylate the most relevant type A and B trichothecenes. HvUGT13248, which prefers nivalenol over deoxynivalenol, is also able to conjugate C-4 acetylated trichothecenes (e.g., T-2 toxin) to some degree while OsUGT79 and Bradi5g03300 are completely inactive with C-4 acetylated derivatives. The type A trichothecenes HT-2 toxin and T-2 triol are the kinetically preferred substrates in the case of HvUGT13248 and Bradi5g03300. We glucosylated several trichothecenes with OsUGT79 (HT-2 toxin, T-2 triol) and HvUGT13248 (T-2 toxin, neosolaniol, 4,15-diacetoxyscirpenol, fusarenon X) in the preparative scale. NMR analysis of the purified glucosides showed that exclusively β-d-glucosides were formed regio-selectively at position C-3-OH of the trichothecenes. These synthesized standards can be used to investigate the occurrence and toxicological properties of these modified mycotoxins. PMID:29509722

  4. UDP-Glucosyltransferases from Rice, Brachypodium, and Barley: Substrate Specificities and Synthesis of Type A and B Trichothecene-3-O-β-d-glucosides.

    PubMed

    Michlmayr, Herbert; Varga, Elisabeth; Malachová, Alexandra; Fruhmann, Philipp; Piątkowska, Marta; Hametner, Christian; Šofrová, Jana; Jaunecker, Günther; Häubl, Georg; Lemmens, Marc; Berthiller, Franz; Adam, Gerhard

    2018-03-06

    Trichothecene toxins are confirmed or suspected virulence factors of various plant-pathogenic Fusarium species. Plants can detoxify these to a variable extent by glucosylation, a reaction catalyzed by UDP-glucosyltransferases (UGTs). Due to the unavailability of analytical standards for many trichothecene-glucoconjugates, information on such compounds is limited. Here, the previously identified deoxynivalenol-conjugating UGTs HvUGT13248 (barley), OsUGT79 (rice) and Bradi5g03300 ( Brachypodium ), were expressed in E. coli , affinity purified, and characterized towards their abilities to glucosylate the most relevant type A and B trichothecenes. HvUGT13248, which prefers nivalenol over deoxynivalenol, is also able to conjugate C-4 acetylated trichothecenes (e.g., T-2 toxin) to some degree while OsUGT79 and Bradi5g03300 are completely inactive with C-4 acetylated derivatives. The type A trichothecenes HT-2 toxin and T-2 triol are the kinetically preferred substrates in the case of HvUGT13248 and Bradi5g03300. We glucosylated several trichothecenes with OsUGT79 (HT-2 toxin, T-2 triol) and HvUGT13248 (T-2 toxin, neosolaniol, 4,15-diacetoxyscirpenol, fusarenon X) in the preparative scale. NMR analysis of the purified glucosides showed that exclusively β-D-glucosides were formed regio-selectively at position C-3-OH of the trichothecenes. These synthesized standards can be used to investigate the occurrence and toxicological properties of these modified mycotoxins.

  5. Structural modifications of N-(1,2,3,4-tetrahydronaphthalen-1-yl)-4-aryl-1-piperazinehexanamides: influence on lipophilicity and 5-HT7 receptor activity. Part III.

    PubMed

    Leopoldo, Marcello; Lacivita, Enza; De Giorgio, Paola; Fracasso, Claudia; Guzzetti, Sara; Caccia, Silvio; Contino, Marialessandra; Colabufo, Nicola A; Berardi, Francesco; Perrone, Roberto

    2008-09-25

    Starting from the previously reported 5-HT 7 receptor agents 4-7 with N-(1,2,3,4-tetrahydronaphthalen-1-yl)-4-aryl-1-piperazinehexanamide structure, the 1-(2-methylthiophenyl)-, 1-(2-diphenyl)-, 1-(2-isopropylphenyl)-, and 1-(2-methoxyphenyl)piperazine derivatives 8-31 were designed with the primary aim to obtain new compounds endowed with suitable physicochemical properties for rapid and extensive penetration into the brain. The affinities for 5-HT 7, 5-HT 1A, and D 2 receptors of compounds 8-31 were assessed, and several compounds displayed 5-HT 7 receptor affinities in the nanomolar range. Among these, N-(4-cyanophenylmethyl)-4-(2-diphenyl)-1-piperazinehexanamide (25) showed high 5-HT 7 receptor affinity (Ki = 0.58 nM), high selectivity over 5-HT 1A and D 2 receptors (324- and 245-fold, respectively), and agonist properties (maximal effect = 82%, EC 50 = 0.60 microM). After intraperitoneal injection in mice, 25 rapidly reached the systemic circulation and entered the brain. Its brain concentration-time profile paralleled that in plasma, indicating that 25 rapidly and freely distributes across the blood-brain barrier. Compound 25 underwent N-dealkylation to the corresponding 1-arylpiperazine metabolite.

  6. Mouse in vivo neutralization of Escherichia coli Shiga toxin 2 with monoclonal antibodies

    USDA-ARS?s Scientific Manuscript database

    Escherichia coli (E. coli) food contaminations pose serious health and food safety concerns, and have been the subject of massive food recalls. Shiga toxin 2 (Stx2)-producing E. coli has been identified as the major cause of hemorrhagic colitis and hemolytic uremic syndrome (HUS), the most severe di...

  7. Lorcaserin, a 5-HT2C Agonist, Decreases Nicotine Self-Administration in Female Rats

    PubMed Central

    Johnson, Joshua E.; Slade, Susan; Wells, Corinne; Cauley, Marty; Petro, Ann; Rose, Jed E.

    2011-01-01

    Lorcaserin, a selective 5-hydroxytryptamine2C (5-HT2C) agonist, has been shown to facilitate weight loss in obese populations. It was assessed for its efficacy in reducing nicotine self-administration in young adult female Sprague-Dawley rats. The effect of short-term doses (subcutaneous) on nicotine self-administration (0.03 mg/kg per infusion) with a fixed ratio 1 schedule was assessed in 3-h sessions. Short-term lorcaserin doses (0.3125–20 mg/kg) were administered in a counterbalanced order. Significant reduction of nicotine self-administration was achieved with all of the short-term doses in this range. Tests of lorcaserin on locomotor activity detected prominent sedative effects at doses greater than 1.25 mg/kg with more modest transient effects seen at 0.625 to 1.25 mg/kg. Long-term effects of lorcaserin on locomotor activity were tested with repeated injections with 0.625 mg/kg lorcaserin 10 times over 2 weeks. This low lorcaserin dose did not cause an overall change in locomotor activity relative to that of saline-injected controls. Long-term lorcaserin (0.625 mg/kg) significantly reduced nicotine self-administration over a 2-week period of repeated injections. Long-term lorcaserin at this same dose had no significant effects on food self-administration over the same 2-week period of repeated injections. These studies support development of the 5-HT2C agonist lorcaserin to aid tobacco smoking cessation. PMID:21636655

  8. Lorcaserin, a 5-HT2C agonist, decreases nicotine self-administration in female rats.

    PubMed

    Levin, Edward D; Johnson, Joshua E; Slade, Susan; Wells, Corinne; Cauley, Marty; Petro, Ann; Rose, Jed E

    2011-09-01

    Lorcaserin, a selective 5-hydroxytryptamine(2C) (5-HT(2C)) agonist, has been shown to facilitate weight loss in obese populations. It was assessed for its efficacy in reducing nicotine self-administration in young adult female Sprague-Dawley rats. The effect of short-term doses (subcutaneous) on nicotine self-administration (0.03 mg/kg per infusion) with a fixed ratio 1 schedule was assessed in 3-h sessions. Short-term lorcaserin doses (0.3125-20 mg/kg) were administered in a counterbalanced order. Significant reduction of nicotine self-administration was achieved with all of the short-term doses in this range. Tests of lorcaserin on locomotor activity detected prominent sedative effects at doses greater than 1.25 mg/kg with more modest transient effects seen at 0.625 to 1.25 mg/kg. Long-term effects of lorcaserin on locomotor activity were tested with repeated injections with 0.625 mg/kg lorcaserin 10 times over 2 weeks. This low lorcaserin dose did not cause an overall change in locomotor activity relative to that of saline-injected controls. Long-term lorcaserin (0.625 mg/kg) significantly reduced nicotine self-administration over a 2-week period of repeated injections. Long-term lorcaserin at this same dose had no significant effects on food self-administration over the same 2-week period of repeated injections. These studies support development of the 5-HT(2C) agonist lorcaserin to aid tobacco smoking cessation.

  9. Discovery of three toxin peptides with Kv1.3 channel and IL-2 cytokine-inhibiting activities from Non-Buthidae scorpions, Chaerilus tricostatus and Chaerilus tryznai.

    PubMed

    Ding, Li; Chen, Jing; Hao, Jinbo; Zhang, Jiahui; Huang, Xuejun; Hu, Fangfang; Wu, Zheng; Liu, Yaru; Li, Wenxin; Cao, Zhijian; Wu, Yingliang; Li, Jian; Li, Shan; Liu, Hongyan; Wu, Wenlong; Chen, Zongyun

    2017-05-01

    Non-Buthidae venomous scorpions are huge natural sources of toxin peptides; however, only a few studies have been done to understand their toxin peptides. Herein, we describe three new potential immunomodulating toxin peptides, Ctri18, Ctry68 and Ctry2908, from two non-Buthidae scorpions, Chaerilus tricostatus and Chaerilus tryznai. Sequence alignment analyses showed that Ctri18, Ctry68 and Ctry2908 are three new members of the scorpion toxin α-KTx15 subfamily. Electrophysiological experiments showed that Ctri18, Ctry68 and Ctry2908 blocked the Kv1.3 channel at micromole to nanomole levels, but had weak effects on potassium channel KCNQ1 and sodium channel Nav1.4, which indicated that Ctri18, Ctry68 and Ctry2908 might have specific inhibiting effects on the Kv1.3 channel. ELISA experiments showed that Ctri18, Ctry68 and Ctry2908 inhibited IL-2 cytokine secretions of activated T lymphocyte in human PBMCs. Excitingly, consistent with the good Kv1.3 channel inhibitory activity, Ctry2908 inhibited cytokine IL-2 secretion in nanomole level, which indicated that Ctry2908 might be a new lead drug template toward Kv1.3 channels. Together, these studies discovered three new toxin peptides, Ctri18, Ctry68 and Ctry2908, with Kv1.3 channel and IL-2 cytokine-inhibiting activities from two scorpions, C. tricostatus and C. tryznai, and highlighted that non-Buthidae venomous scorpions are new natural toxin peptide sources. Copyright © 2017 Elsevier Inc. All rights reserved.

  10. The attentional blink is not affected by backward masking of T2, T2-mask SOA, or level of T2 impoverishment.

    PubMed

    Jannati, Ali; Spalek, Thomas M; Lagroix, Hayley E P; Di Lollo, Vincent

    2012-02-01

    Identification of the second of two targets (T2) is impaired when presented shortly after the first (T1). This attentional blink (AB) is thought to arise from a delay in T2 processing during which T2 is vulnerable to masking. Conventional studies have measured T2 accuracy which is constrained by the 100% ceiling. We avoided this problem by using a dynamic threshold-tracking procedure that is inherently free from ceiling constraints. In two experiments we examined how AB magnitude is affected by three masking-related factors: (a) presence/absence of T2 mask, (b) T2-mask stimulus onset asynchrony (SOA), and (c) level of T2 impoverishment (signal-to-noise ratio [SNR]). In Experiment 1, overall accuracy decreased with T2-mask SOA. The magnitude of the AB, however, was invariant with SOA and with mask presence/absence. Experiment 2 further showed that the AB was invariant with T2 SNR. The relationship among mask presence/absence, SOA, and T2 SNR and the AB is encompassed in a qualitative model.

  11. Expression of deleted, atoxic atypical recombinant beta2 toxin in a baculovirus system and production of polyclonal and monoclonal antibodies.

    PubMed

    Serroni, Anna; Magistrali, Chiara Francesca; Pezzotti, Giovanni; Bano, Luca; Pellegrini, Martina; Severi, Giulio; Di Pancrazio, Chiara; Luciani, Mirella; Tittarelli, Manuela; Tofani, Silvia; De Giuseppe, Antonio

    2017-05-25

    Clostridium perfringens is an important animal and human pathogen that can produce more than 16 different major and minor toxins. The beta-2 minor toxin (CPB2), comprising atypical and consensus variants, appears to be involved in both human and animal enterotoxaemia syndrome. The exact role of CPB2 in pathogenesis is poorly investigated, and its mechanism of action at the molecular level is still unknown because of the lack of specific reagents such as monoclonal antibodies against the CPB2 protein and/or the availability of a highly purified antigen. Previous studies have reported that purified wild-type or recombinant CPB2 toxin, expressed in a heterologous system, presented cytotoxic effects on human intestinal cell lines. Undoubtedly, for this reason, to date, these purified proteins have not yet been used for the production of monoclonal antibodies (MAbs). Recently, monoclonal antibodies against CPB2 were generated using peptides designed on predicted antigenic epitopes of this toxin. In this paper we report, for the first time, the expression in a baculovirus system of a deleted recombinant C-terminal 6xHis-tagged atypical CPB2 toxin (rCPB2 Δ1-25 -His 6 ) lacking the 25 amino acids (aa) of the N-terminal putative signal sequence. A high level of purified recombinant rCPB2 Δ1-25 -His 6 was obtained after purification by Ni 2+ affinity chromatography. The purified product showed no in vitro and in vivo toxicity. Polyclonal antibodies and twenty hybridoma-secreting Mabs were generated using purified rCPB2 Δ1-25 -His 6 . Finally, the reactivity and specificity of the new antibodies were tested against both recombinant and wild-type CPB2 toxins. The high-throughput of purified atoxic recombinant CPB2 produced in insect cells, allowed to obtain monoclonal and polyclonal antibodies. The availability of these molecules could contribute to develop immunoenzymatic methods and/or to perform studies about the biological activity of CPB2 toxin.

  12. Neonatal stimulation of 5-HT(2) receptors reduces androgen receptor expression in the rat anteroventral periventricular nucleus and sexually dimorphic preoptic area.

    PubMed

    Dakin, C L; Wilson, C A; Kalló, I; Coen, C W; Davies, D C

    2008-05-01

    Masculinization of the brain is dependent upon a perinatal surge in testosterone. It also requires a transient decrease in hypothalamic 5-HT concentration and turnover and an increase in androgen receptor (AR) expression during the second postnatal week. We have previously shown that increasing 5-HT activity over this period in male or androgenized female rats feminizes their adult behaviour and also feminizes the size of their anteroventral periventricular nucleus (AVPV) and sexually dimorphic nucleus of the preoptic area (SDN-POA). To investigate the role of 5-HT in sexual differentiation of the brain, 5-HT activity was raised over postnatal days 8-16 in male, female and androgenized female rats by daily administration of the 5-HT(2) receptor agonist (-)[2,5 dimethoxy-4-iodophenyl]-2-amino propane hydrochloride [(-)DOI]. By postnatal day 18, the size of the AVPV and SDN-POA was sexually dimorphic; their sizes were feminized by (-)DOI treatment. In the absence of (-)DOI treatment, there were significantly more AR-immunoreactive cells in the AVPV of males, and in the SDN-POA of males and androgenized females, than in those of females on postnatal day 18. (-)DOI treatment reduced the number of AR-immunoreactive cells in the AVPV and SDN-POA of males and androgenized females, but not of females, by postnatal day 18. These results suggest that 5-HT(2) receptor activation can influence sexual differentiation of the brain by controlling AR expression.

  13. Vibrio cholerae ACE stimulates Ca(2+)-dependent Cl(-)/HCO(3)(-) secretion in T84 cells in vitro.

    PubMed

    Trucksis, M; Conn, T L; Wasserman, S S; Sears, C L

    2000-09-01

    ACE, accessory cholera enterotoxin, the third enterotoxin in Vibrio cholerae, has been reported to increase short-circuit current (I(sc)) in rabbit ileum and to cause fluid secretion in ligated rabbit ileal loops. We studied the ACE-induced change in I(sc) and potential difference (PD) in T84 monolayers mounted in modified Ussing chambers, an in vitro model of a Cl(-) secretory cell. ACE added to the apical surface alone stimulated a rapid increase in I(sc) and PD that was concentration dependent and immediately reversed when the toxin was removed. Ion replacement studies established that the current was dependent on Cl(-) and HCO(3)(-). ACE acted synergistically with the Ca(2+)-dependent acetylcholine analog, carbachol, to stimulate secretion in T84 monolayers. In contrast, the secretory response to cAMP or cGMP agonists was not enhanced by ACE. The ACE-stimulated secretion was dependent on extracellular and intracellular Ca(2+) but was not associated with an increase in intracellular cyclic nucleotides. We conclude that the mechanism of secretion by ACE involves Ca(2+) as a second messenger and that this toxin stimulates a novel Ca(2+)-dependent synergy.

  14. Translating 5-HT receptor pharmacology.

    PubMed

    Sanger, G J

    2009-12-01

    Since metoclopramide was first described (in 1964) there have been several attempts to develop compounds which retained gastrointestinal prokinetic activity (via 5-HT(4) receptor activation) but without the limiting side effects associated with dopamine D(2) receptor antagonism. Early compounds (mosapride, cisapride, renzapride, tegaserod) were identified before several of the 5-HT receptors were even described (including 5-HT(4) and 5-HT(2B)), whereas prucalopride came later. Several compounds were hampered by non-selectivity, introducing cardiac liability (cisapride: activity at human Ether-a-go-go Related Gene) or potentially, a reduced intestinal prokinetic activity caused by activity at a second 5-HT receptor (renzapride: antagonism at the 5-HT(3) receptor; tegaserod: antagonism at the 5-HT(2B) receptor). Poor intrinsic activity at gastrointestinal 5-HT(4) receptors has also been an issue (mosapride, tegaserod). Perhaps prucalopride has now achieved the profile of good selectivity of action and high intrinsic activity at intestinal 5-HT(4) receptors, without clinically-meaningful actions on 5-HT(4) receptors in the heart. The progress of this compound for treatment of chronic constipation, as well as competitor molecules such as ATI-7505 and TD-5108, will now be followed with interest as each attempts to differentiate themselves from each other. Perhaps at last, 5-HT(4) receptor agonists are being given the chance to show what they can do.

  15. A novel 5HT3 antagonist 4i (N-(3-chloro-2-methylphenyl)quinoxalin-2-carboxamide) prevents diabetes-induced depressive phenotypes in mice: Modulation of serotonergic system.

    PubMed

    Gupta, Deepali; Thangaraj, Devadoss; Radhakrishnan, Mahesh

    2016-01-15

    Despite the presence of a multitudinous pharmacotherapy, diabetes-induced depressive disorder remains undertreated. Evidence suggests that brain serotonergic deficits are associated with depressive-like behavior in diabetes and that 5HT3 receptor (5HT3R) antagonists have serotonergic facilitatory effects. This study examined the effects of a novel 5HT3R antagonist, 4i (N-(3-chloro-2-methylphenyl)quinoxalin-2-carboxamide), in diabetes-induced depressive phenotypes. Experimentally, (1) to evaluate the effects of 4i, mice with 8-weeks of diabetes (induced by streptozotocin, 200mg/kg, i.p.) were treated with vehicle, 4i (0.5 and 1mg/kg/day, i.p.), fluoxetine (10mg/kg/day, i.p.) for 4-weeks and subjected to neurobehavioral assays, followed by biochemical estimation of serotonin levels in midbrain, prefrontal-cortex and cerebellum. (2) To evaluate the role of 5HT3R in the postulated effect of 4i, diabetic mice were given 4i (1mg/kg/day, i.p.) after 1h of 1-(m-chlorophenyl)-biguanide (mCPBG, a 5HT3R agonist, 10mg/kg/day, i.p.) treatment and subjected to the same protocol. The results showed that diabetic mice exhibited a significant behavioral deficit, including depression-like behavior in forced swim test, anxiety-like in open field test and sociability deficits in social interaction test, along with a significant decrease in serotonin level in these brain regions. 4i (1mg/kg), similar to fluoxetine, prevented these behavioral abnormalities and normalized brain serotonin levels. 4i (0.5mg/kg) ameliorated only diabetes-induced depressive-like behavior and serotonin deficits, but not anxiety-like effects. mCPBG blunted 4i-mediated behavioral response and increase in brain serotonin levels. Altogether, this study suggests that 4i prevents diabetes-induced depressive phenotypes in mice, which may involve antagonism of 5HT3Rs and increase in serotonin levels in discrete brain regions. Copyright © 2015 Elsevier B.V. All rights reserved.

  16. Decreased social behaviour following 3,4-methylenedioxymethamphetamine (MDMA) is accompanied by changes in 5-HT2A receptor responsivity.

    PubMed

    Bull, Eleanor J; Hutson, Peter H; Fone, Kevin C F

    2004-02-01

    This study examined the involvement of the 5-HT(2A) receptor in the long-term anxiogenic effect of a brief exposure of young rats to 3,4-methylenedioxymethamphetamine (MDMA) using the social interaction and elevated plus-maze paradigms. Wistar rats (post-natal day (PND) 28) received either MDMA (5 mg/kg i.p.) or saline (1 ml/kg i.p.) hourly for 4 h on 2 consecutive days. Locomotor activity was measured for 60 min after the first injection and core body temperature was recorded at regular intervals over 4 h. On PND 84, without further drug administration, social interaction was assessed between treatment-matched rat pairs derived from separate litters. On PND 86, rats received either the 5-HT(2A/2C) receptor agonist, 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI, 1 mg/kg i.p.) or saline and locomotor activity, wet-dog shakes and back muscle contractions were monitored. The change in elevated plus-maze behaviour was assessed following the same injection on PND 87. Acutely, MDMA produced a significant hyperlocomotion and hyperthermia (p<0.01). Following 55 days of abstinence, social interaction was reduced by 27% in MDMA pre-treated rats compared with that in controls (p<0.01). On the elevated plus-maze, pre-treatment with MDMA prevented the anxiogenic effect of DOI. On PND 92, hippocampal, frontal cortical and striatal 5-hydroxytryptamine (5-HT) was significantly reduced in MDMA pre-treated rats by between 16% and 22%, without any accompanying change in [(3)H]paroxetine binding in cortical homogenates. In conclusion, exposure of young rats to repeated MDMA caused serotonin depletion and induced 'anxiety-like' behaviour in the social interaction test accompanied by a long-lasting reduction in specific 5-HT(2A) receptor mediated behaviour.

  17. Expression, crystallization and preliminary X-ray diffraction studies of recombinant Clostridium perfringens β2-toxin

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Gurjar, Abhijit A.; Yennawar, Neela H.; Yennawar, Hemant P.

    2007-06-01

    The cloning, expression, purification and crystallization of recombinant Clostridium perfringens β2-toxin is described. The crystals diffracted to 2.9 Å resolution. Clostridium perfringens is a Gram-positive sporulating anaerobic bacterium that is responsible for a wide spectrum of diseases in animals, birds and humans. The virulence of C. perfringens is associated with the production of several enterotoxins and exotoxins. β2-toxin is a 28 kDa exotoxin produced by C. perfringens. It is implicated in necrotic enteritis in animals and humans, a disease characterized by a sudden acute onset with lethal hemorrhagic mucosal ulceration. The recombinant expression, purification and crystallization of β2-toxin using themore » batch-under-oil technique are reported here. Native X-ray diffraction data were obtained to 2.9 Å resolution on a synchrotron beamline at the F2 station at Cornell High Energy Synchrotron Source (CHESS) using an ADSC Quantum-210 CCD detector. The crystals belong to space group R3, with a dimer in the asymmetric unit; the unit-cell parameters are a = b = 103.71, c = 193.48 Å, α = β = 90, γ = 120° using the hexagonal axis setting. A self-rotation function shows that the two molecules are related by a noncrystallographic twofold axis with polar angles ω = 90.0, ϕ = 210.3°.« less

  18. Comparative investigation on the effect of T-2 mycotoxin on lipid peroxidation and antioxidant status in different poultry species.

    PubMed

    Mézes, M; Barta, M; Nagy, G

    1999-02-01

    The effect of low dose T-2 toxin was investigated in chicken, duck and goose. The purpose of the present study was to investigate the effect of T-2 toxin on the lipid peroxidation and on the activity of glutathione redox system (amount of reduced and oxidised glutathione and the activity of glutathione peroxidase) in blood and liver. The treatment lasted days and two samples were taken, first at the time of lowest feed intake and second when the intake was the same as the control. It was found, that lipid per oxidation as detected by the amount of malondialdehyde increased in all of the species and tissues but the changes varied by species. The most sensitive species was goose followed by duck and chicken, and the most sensitive tissue was the liver followed by blood plasma and red blood cells.

  19. Antipsychotics differ in their ability to internalise human dopamine D2S and human serotonin 5-HT1A receptors in HEK293 cells.

    PubMed

    Heusler, Peter; Newman-Tancredi, Adrian; Loock, Timothé; Cussac, Didier

    2008-02-26

    Antipsychotic drugs act preferentially via dopamine D(2) receptor blockade, but interaction with serotonin 5-HT(1A) receptors has attracted interest as additional target for antipsychotic treatment. As receptor internalisation is considered crucial for drug action, we tested the propensity of antipsychotics to internalise human (h)D(2S) receptors and h5-HT(1A) receptors. Agonist-induced internalisation of hemaglutinin (HA)-tagged hD(2S) and HA-h5-HT(1A) receptors expressed in HEK293 cells was increased by coexpression of G-protein coupled receptor kinase 2 and beta-arrestin2. At the HA-hD(2S) receptor, dopamine, quinpirole and bromocriptine behaved as full agonists, while S(-)-3-(3-hydroxyphenyl)-N-n-propylpiperidine [(-)-3PPP] and sarizotan were partial agonists. The typical antipsychotic, haloperidol, and the atypical compounds, olanzapine, nemonapride, ziprasidone and clozapine did not internalise HA-hD(2S) receptors, whereas aripiprazole potently internalised these receptors (>50% relative efficacy). Among antipsychotics with combined D(2)/5-HT(1A) properties, bifeprunox and (3-exo)-8-benzoyl-N-[[(2S)7-chloro-2,3-dihydro-1,4-benzodioxin-1-yl]methyl]-8-azabicyclo-[3.2.1]octane-3-methanamine (SSR181507) partially internalised HA-hD(2S) receptors, piperazine, 1-(2,3-dihydro-1,4-benzodioxin-5-yl)-4-[[5-(4-fluorophenyl)-3-pyridinyl]methyl (SLV313) and N-[(2,2-dimethyl-2,3-dihydro-benzofuran-7-yloxy)ethyl]-3-(cyclopent-1-enyl)-benzylamine (F15063) were inactive. At the HA-h5-HT(1A) receptor, serotonin, (+)-8-hydroxy-2-(di-n-propylamino)tetralin [(+)-8-OH-DPAT] and sarizotan were full agonists, buspirone acted as partial agonist. (-)-Pindolol showed little activity and no internalising properties were manifested for the 5-HT(1A) receptor antagonist N-[2-[4-(2-methoxyphenyl)-1-piperazinyl]-ethyl]-N-(2-pyridinyl)cyclohexanecarboxamide (WAY100635). Most antipsychotics induced HA-h5-HT(1A) receptor internalisation, with an efficacy rank order: nemonapride>F15063>SSR181507

  20. Targeted Silencing of Anthrax Toxin Receptors Protects against Anthrax Toxins*

    PubMed Central

    Arévalo, Maria T.; Navarro, Ashley; Arico, Chenoa D.; Li, Junwei; Alkhatib, Omar; Chen, Shan; Diaz-Arévalo, Diana; Zeng, Mingtao

    2014-01-01

    Anthrax spores can be aerosolized and dispersed as a bioweapon. Current postexposure treatments are inadequate at later stages of infection, when high levels of anthrax toxins are present. Anthrax toxins enter cells via two identified anthrax toxin receptors: tumor endothelial marker 8 (TEM8) and capillary morphogenesis protein 2 (CMG2). We hypothesized that host cells would be protected from anthrax toxins if anthrax toxin receptor expression was effectively silenced using RNA interference (RNAi) technology. Thus, anthrax toxin receptors in mouse and human macrophages were silenced using targeted siRNAs or blocked with specific antibody prior to challenge with anthrax lethal toxin. Viability assays were used to assess protection in macrophages treated with specific siRNA or antibody as compared with untreated cells. Silencing CMG2 using targeted siRNAs provided almost complete protection against anthrax lethal toxin-induced cytotoxicity and death in murine and human macrophages. The same results were obtained by prebinding cells with specific antibody prior to treatment with anthrax lethal toxin. In addition, TEM8-targeted siRNAs also offered significant protection against lethal toxin in human macrophage-like cells. Furthermore, silencing CMG2, TEM8, or both receptors in combination was also protective against MEK2 cleavage by lethal toxin or adenylyl cyclase activity by edema toxin in human kidney cells. Thus, anthrax toxin receptor-targeted RNAi has the potential to be developed as a life-saving, postexposure therapy against anthrax. PMID:24742682

  1. Association between serotonin 5-HT-2C receptor gene (HTR2C) polymorphisms and obesity- and mental health-related phenotypes in a large population-based cohort.

    PubMed

    Vimaleswaran, K S; Zhao, J H; Wainwright, N W; Surtees, P G; Wareham, N J; Loos, R J F

    2010-06-01

    Studies have shown that common single-nucleotide polymorphisms (SNPs) in the serotonin 5-HT-2C receptor (HTR2C) are associated with antipsychotic agent-induced weight gain and the development of behavioural and psychological symptoms. We aimed to analyse whether variation in the HTR2C is associated with obesity- and mental health-related phenotypes in a large population-based cohort. Six tagSNPs, which capture all common genetic variation in the HTR2C gene, were genotyped in 4978 men and women from the European Prospective Investigation into Cancer (EPIC)-Norfolk study, an ongoing prospective population-based cohort study in the United Kingdom. To confirm borderline significant associations, the -759C/T SNP (rs3813929) was genotyped in the remaining 16 003 individuals from the EPIC-Norfolk study. We assessed social and psychological circumstances using the Health and Life Experiences Questionnaire. Genmod models were used to test associations between the SNPs and the outcomes. Logistic regression was performed to test for association of SNPs with obesity- and mental health- related phenotypes. Of the six HTR2C SNPs, only the T allele of the -759C/T SNP showed borderline significant associations with higher body mass index (BMI) (0.23 kg m(-2); (95% confidence interval (CI): 0.01-0.44); P=0.051) and increased risk of lifetime major depressive disorder (MDD) (Odds ratio (OR): 1.13 (95% CI: 1.01-1.22), P=0.02). The associations between the -759C/T and BMI and lifetime MDD were independent. As associations only achieved borderline significance, we aimed to validate our findings on the -759C/T SNP in the full EPIC-Norfolk cohort (n=20 981). Although the association with BMI remained borderline significant (beta=0.20 kg m(-2); 95% CI: 0.04-0.44, P=0.09), that with lifetime MDD (OR: 1.01; 95% CI: 0.94-1.09, P=0.73) was not replicated. Our findings suggest that common HTR2C gene variants are unlikely to have a major role in obesity- and mental health-related traits in the

  2. 5-HT2 receptor distribution shown by [18F] setoperone PET in high-functioning autistic adults.

    PubMed

    Beversdorf, David Q; Nordgren, Richard E; Bonab, Ali A; Fischman, Alan J; Weise, Steven B; Dougherty, Darin D; Felopulos, Gretchen J; Zhou, Feng C; Bauman, Margaret L

    2012-01-01

    The serotonergic system is implicated in disordered emotional behavior. Autism is characterized by impaired processing of emotional information. The serotonergic (5-HT) system is also critically involved in brain development, and abnormal brain synthesis of serotonin is observed in autism. Furthermore, whole blood and platelet serotonin have been reported to be elevated in autism. The authors examined the CNS serotonin system in autism in vivo. 5-HT2 receptors were visualized by PET imaging of [18F]setoperone-binding in this pilot study of 6 high-functioning autistic adults and 10 matched-control participants. Autism subjects had less thalamic [18F]setoperone binding than controls, when covaried for age, but no difference reached significance in other areas. A negative relationship between thalamic binding and history of language impairment was also observed. Further studies will be needed to gain a clearer picture of the role of the 5-HT system in autism.

  3. Translocation of the Catalytic Domain of Diphtheria Toxin across Planar Phospholipid Bilayers by Its Own T Domain

    NASA Astrophysics Data System (ADS)

    Oh, Kyoung Joon; Senzel, Lisa; Collier, R. John; Finkelstein, Alan

    1999-07-01

    The T domain of diphtheria toxin is known to participate in the pH-dependent translocation of the catalytic C domain of the toxin across the endosomal membrane, but how it does so, and whether cellular proteins are also required for this process, remain unknown. Here, we report results showing that the T domain alone is capable of translocating the entire C domain across model, planar phospholipid bilayers in the absence of other proteins. The T domain therefore contains the entire molecular machinery for mediating transfer of the catalytic domain of diphtheria toxin across membranes.

  4. Tianeptine: 5-HT uptake sites and 5-HT(1-7) receptors modulate memory formation in an autoshaping Pavlovian/instrumental task.

    PubMed

    Meneses, Alfredo

    2002-05-01

    Recent studies using invertebrate and mammal species have revealed that, endogenous serotonin (5-hydroxytryptamine, 5-HT) modulates cognitive processes, particularly learning and memory, though, at present, it is unclear the manner, where, and how long 5-HT systems are involved. Hence in this work, an attempt was made to study the effects of 5-HT endogenous on memory formation, using a 5-HT uptake facilitator (tianeptine) and, selective 5-HT(1-7) receptor antagonists to determine whether 5-HT uptake sites and which 5-HT receptors are involved, respectively. Results showed that post-training tianeptine injection enhanced memory consolidation in an autoshaping Pavlovian/instrumental learning task, which has been useful to detect changes on memory formation elicited by drugs or aging. On interaction experiments, ketanserin (5-HT(1D/2A/2C) antagonist) slightly enhanced tianeptine effects, while WAY 100635 (5-HT(1A) antagonist), SB-224289 (5-HT(1B) inverse agonist), SB-200646 (5-HT(2B/2C) antagonist), ondansetron (5-HT(3) antagonist), GR 127487 (5-HT(4) antagonist), Ro 04-6790 (5-HT(6) antagonist), DR 4004 (5-HT(7) antagonist), or fluoxetine (an inhibitor of 5-HT reuptake) blocked the facilitatory tianeptine effect. Notably, together tianeptine and Ro 04-6790 impaired learning consolidation. Moreover, 5-HT depletion completely reversed the tianeptine effect. Tianeptine also normalized an impaired memory elicited by scopolamine (an antimuscarinic) or dizocilpine (non-competitive glutamatergic antagonist), while partially reversed that induced by TFMPP (5-HT(1B/1D/2A-2C/7) agonist/antagonist). Finally, tianeptine-fluoxetine coadministration had no effect on learning consolidation; nevertheless, administration of an acetylcholinesterase inhibitor, phenserine, potentiated subeffective tianeptine or fluoxetine doses. Collectively, these data confirmed that endogenously 5-HT modulates, via uptake sites and 5-HT(1-7) receptors, memory consolidation, and are consistent with the

  5. Studies To Examine Potential Tolerability Differences between the 5-HT2C Receptor Selective Agonists Lorcaserin and CP-809101.

    PubMed

    Higgins, Guy A; Silenieks, Leonardo B; Patrick, Amy; De Lannoy, Ines A M; Fletcher, Paul J; Parker, Linda A; MacLusky, Neil J; Sullivan, Laura C; Chavera, Teresa A; Berg, Kelly A

    2017-05-17

    Lorcaserin (LOR) is a selective 5-HT 2C receptor agonist that has been FDA approved as a treatment for obesity. The most frequently reported side-effects of LOR include nausea and headache, which can be dose limiting. We have previously reported that in the rat, while LOR produced unconditioned signs characteristic of nausea/malaise, the highly selective 5-HT 2C agonist CP-809101 (CP) produced fewer equivalent signs. Because this may indicate a subclass of 5-HT 2C agonists having better tolerability, the present studies were designed to further investigate this apparent difference. In a conditioned gaping model, a rodent test of nausea, LOR produced significantly higher gapes compared to CP consistent with it having higher emetogenic properties. Subsequent studies were designed to identify features of each drug that may account for such differences. In rats trained to discriminate CP-809101 from saline, both CP and LOR produced full generalization suggesting a similar interoceptive cue. In vitro tests of functional selectivity designed to examine signaling pathways activated by both drugs in CHO (Chinese hamster ovary) cells expressing h5-HT 2C receptors failed to identify evidence for biased signaling differences between LOR and CP. Thus, both drugs showed similar profiles across PLC, PLA 2 , and ERK signaling pathways. In studies designed to examine pharmacokinetic differences between LOR and CP, while drug plasma levels correlated with increasing dose, CSF levels did not. CSF levels of LOR increased proportionally with dose; however CSF levels of CP plateaued from 6 to 12 mg/kg. Thus, the apparently improved tolerability of CP likely reflects a limit to CNS levels attained at relatively high doses.

  6. The susceptibility of the mallard duck (Anas platyrhynchos) to Clostridium botulinum C2 toxin

    USGS Publications Warehouse

    Jensen, Wayne I.; Duncan, Ruth M.

    1980-01-01

    Most strains of Clostridium botulinum type C, after having lost their capacity to produce their dominant toxin (C1) as a result of being“cured”of their prophages, continue to produce C2, a trypsin-activable toxin reported by other investigators. While of relatively low toxicity when administered perorally to the adult mallard duck (Anas platyrhynchos), it was highly toxic when given parenterally. By the intravenous route, for example, it was more than 1, 000 times as toxic as C1 toxin by the same route, when compared on the basis of mouse intraperitoneal toxicity. The cause of death in every instance was massive pulmonary edema and hemorrhage rather than the respiratory paralysis that occurs in C1 intoxication.抄録

  7. Synthesis and serotonergic activity of 3-[2-(pyrrolidin-1-yl)ethyl]indoles: potent agonists for the h5-HT1D receptor with high selectivity over the h5-HT1B receptor.

    PubMed

    Sternfeld, F; Guiblin, A R; Jelley, R A; Matassa, V G; Reeve, A J; Hunt, P A; Beer, M S; Heald, A; Stanton, J A; Sohal, B; Watt, A P; Street, L J

    1999-02-25

    The design, synthesis, and biological evaluation of a novel series of 3-[2-(pyrrolidin-1-yl)ethyl]indoles with excellent selectivity for h5-HT1D (formerly 5-HT1Dalpha) receptors over h5-HT1B (formerly 5-HT1Dbeta) receptors are described. Clinically effective antimigraine drugs such as Sumatriptan show little selectivity between h5-HT1D and h5-HT1B receptors. The differential expression of h5-HT1D and h5-HT1B receptors in neural and vascular tissue prompted an investigation of whether a compound selective for the h5-HT1D subtype would have the same clinical efficacy but with reduced side effects. The pyrrolidine 3b was initially identified as having 9-fold selectivity for h5-HT1D over h5-HT1B receptors. Substitution of the pyrrolidine ring of 3b with methylbenzylamine groups gave compounds with nanomolar affinity for the h5-HT1D receptor and 100-fold selectivity with respect to h5-HT1B receptors. Modification of the indole 5-substituent led to the oxazolidinones 24a,b with up to 163-fold selectivity for the h5-HT1D subtype and improved selectivity over other serotonin receptors. The compounds were shown to be full agonists by measurement of agonist-induced [35S]GTPgammaS binding in CHO cells expressed with h5-HT receptors. This study suggests that the h5-HT1D and h5-HT1B receptors can be differentiated by appropriate substitution of the ligand in the region which binds to the aspartate residue and reveals a large binding pocket in the h5-HT1D receptor domain which is absent for the h5-HT1B receptor. The compounds described herein will be important tools to delineate the role of h5-HT1D receptors in migraine.

  8. The Attentional Blink Is Not Affected by Backward Masking of T2, T2-Mask SOA, or Level of T2 Impoverishment

    ERIC Educational Resources Information Center

    Jannati, Ali; Spalek, Thomas M.; Lagroix, Hayley E. P.; Di Lollo, Vincent

    2012-01-01

    Identification of the second of two targets (T2) is impaired when presented shortly after the first (T1). This "attentional blink" (AB) is thought to arise from a delay in T2 processing during which T2 is vulnerable to masking. Conventional studies have measured T2 accuracy which is constrained by the 100% ceiling. We avoided this problem by using…

  9. Cannabinoid inhibition of adenylate cyclase-mediated signal transduction and interleukin 2 (IL-2) expression in the murine T-cell line, EL4.IL-2.

    PubMed

    Condie, R; Herring, A; Koh, W S; Lee, M; Kaminski, N E

    1996-05-31

    Cannabinoid receptors negatively regulate adenylate cyclase through a pertussis toxin-sensitive GTP-binding protein. In the present studies, signaling via the adenylate cyclase/cAMP pathway was investigated in the murine thymoma-derived T-cell line, EL4.IL-2. Northern analysis of EL4.IL-2 cells identified the presence of 4-kilobase CB2 but not CB1 receptor-subtype mRNA transcripts. Southern analysis of genomic DNA digests for the CB2 receptor demonstrated identical banding patterns for EL4.IL-2 cells and mouse-derived DNA, both of which were dissimilar to DNA isolated from rat. Treatment of EL4.IL-2 cells with either cannabinol or Delta9-THC disrupted the adenylate cyclase signaling cascade by inhibiting forskolin-stimulated cAMP accumulation which consequently led to a decrease in protein kinase A activity and the binding of transcription factors to a CRE consensus sequence. Likewise, an inhibition of phorbol 12-myristate 13-acetate (PMA)/ionomycin-induced interleukin 2 (IL-2) protein secretion, which correlated to decreased IL-2 gene transcription, was induced by both cannabinol and Delta9-THC. Further, cannabinoid treatment also decreased PMA/ionomycin-induced nuclear factor binding to the AP-1 proximal site of the IL-2 promoter. Conversely, forskolin enhanced PMA/ionomycin-induced AP-1 binding. These findings suggest that inhibition of signal transduction via the adenylate cyclase/cAMP pathway induces T-cell dysfunction which leads to a diminution in IL-2 gene transcription.

  10. Hybrid solar cells with outstanding short-circuit currents based on a room temperature soft-chemical strategy: the case of P3HT:Ag2S.

    PubMed

    Lei, Yan; Jia, Huimin; He, Weiwei; Zhang, Yange; Mi, Liwei; Hou, Hongwei; Zhu, Guangshan; Zheng, Zhi

    2012-10-24

    P3HT:Ag(2)S hybrid solar cells with broad absorption from the UV to NIR band were directly fabricated on ITO glass by using a room temperature, low energy consumption, and low-cost soft-chemical strategy. The resulting Ag(2)S nanosheet arrays facilitate the construction of a perfect percolation structure with organic P3HT to form ordered bulk heterojunctions (BHJ); without interface modification, the assembled P3HT:Ag(2)S device exhibits outstanding short-circuit current densities (J(sc)) around 20 mA cm(-2). At the current stage, the optimized device exhibited a power conversion efficiency of 2.04%.

  11. Polymorphisms in DNA repair genes increase the risk for type 2 diabetes mellitus and hypertension.

    PubMed

    Das, Sambuddha; Purkayastha, Sukanya; Roy, Hirakjyoti; Sinha, Anima; Choudhury, Yashmin

    2018-06-09

    We investigated the effect of polymorphisms in four DNA repair genes, viz. RAD18 Arg302Gln (G>A) (rs373572), XPD Asp312Asn (G>A) (rs1799793), APE1 Asp148Glu (T>G) (rs3136820), and OGG1 Ser326Cys (C>G) (rs1052133) on the risk for type 2 diabetes mellitus (T2DM) and hypertension (HT) in association with smoking, tobacco chewing, and alcohol consumption in a population from Northeast India. The study subjects were comprised of 70 patients suffering from both T2DM and HT and 83 healthy controls. Genotyping was performed using ARMS-PCR for XPD Asp312Asn (G>A) and PCR-CTPP for RAD18 Arg302Gln (G>A), APE1 Asp148Glu (T>G) and OGG1 Ser326Cys (C>G). The RAD18 Gln/Gln genotype was found to significantly increase the risk for T2DM and HT by 30 fold. Significant high risk was observed for individuals with XPD Asn/Asn-RAD18 Arg/Gln genotypes. Smoking was found to be the single most important independent risk factor for T2DM and HT. This study concludes that RAD18 Arg302Gln and XPD Asp312Asn polymorphisms might increase the risk for T2DM and HT in association with smoking, tobacco chewing, and/or alcohol consumption, while APE1 Asp148Glu (T>G) and OGG1 Ser326Cys (C>G) polymorphisms do not contribute to such risk.

  12. Saccharomyces boulardii Protease Inhibits the Effects of Clostridium difficile Toxins A and B in Human Colonic Mucosa

    PubMed Central

    Castagliuolo, Ignazio; Riegler, Martin F.; Valenick, Leyla; LaMont, J. Thomas; Pothoulakis, Charalabos

    1999-01-01

    Saccharomyces boulardii is a nonpathogenic yeast used in the treatment of Clostridium difficile diarrhea and colitis. We have reported that S. boulardii inhibits C. difficile toxin A enteritis in rats by releasing a 54-kDa protease which digests the toxin A molecule and its brush border membrane (BBM) receptor (I. Castagliuolo, J. T. LaMont, S. T. Nikulasson, and C. Pothoulakis, Infect. Immun. 64:5225–5232, 1996). The aim of this study was to further evaluate the role of S. boulardii protease in preventing C. difficile toxin A enteritis in rat ileum and determine whether it protects human colonic mucosa from C. difficile toxins. A polyclonal rabbit antiserum raised against purified S. boulardii serine protease inhibited by 73% the proteolytic activity present in S. boulardii conditioned medium in vitro. The anti-protease immunoglobulin G (IgG) prevented the action of S. boulardii on toxin A-induced intestinal secretion and mucosal permeability to [3H]mannitol in rat ileal loops, while control rabbit IgG had no effect. The anti-protease IgG also prevented the effects of S. boulardii protease on digestion of toxins A and B and on binding of [3H]toxin A and [3H]toxin B to purified human colonic BBM. Purified S. boulardii protease reversed toxin A- and toxin B-induced inhibition of protein synthesis in human colonic (HT-29) cells. Furthermore, toxin A- and B-induced drops in transepithelial resistance in human colonic mucosa mounted in Ussing chambers were reversed by 60 and 68%, respectively, by preexposing the toxins to S. boulardii protease. We conclude that the protective effects of S. boulardii on C. difficile-induced inflammatory diarrhea in humans are due, at least in part, to proteolytic digestion of toxin A and B molecules by a secreted protease. PMID:9864230

  13. Cardiac Iron Determines Cardiac T2*, T2, and T1 in the Gerbil Model of Iron Cardiomyopathy

    PubMed Central

    Wood, John C.; Otto-Duessel, Maya; Aguilar, Michelle; Nick, Hanspeter; Nelson, Marvin D.; Coates, Thomas D.; Pollack, Harvey; Moats, Rex

    2010-01-01

    Background Transfusional therapy for thalassemia major and sickle cell disease can lead to iron deposition and damage to the heart, liver, and endocrine organs. Iron causes the MRI parameters T1, T2, and T2* to shorten in these organs, which creates a potential mechanism for iron quantification. However, because of the danger and variability of cardiac biopsy, tissue validation of cardiac iron estimates by MRI has not been performed. In this study, we demonstrate that iron produces similar T1, T2, and T2* changes in the heart and liver using a gerbil iron-overload model. Methods and Results Twelve gerbils underwent iron dextran loading (200 mg · kg−1 · wk−1) from 2 to 14 weeks; 5 age-matched controls were studied as well. Animals had in vivo assessment of cardiac T2* and hepatic T2 and T2* and postmortem assessment of cardiac and hepatic T1 and T2. Relaxation measurements were performed in a clinical 1.5-T magnet and a 60-MHz nuclear magnetic resonance relaxometer. Cardiac and liver iron concentrations rose linearly with administered dose. Cardiac 1/T2*, 1/T2, and 1/T1 rose linearly with cardiac iron concentration. Liver 1/T2*, 1/T2, and 1/T1 also rose linearly, proportional to hepatic iron concentration. Liver and heart calibrations were similar on a dry-weight basis. Conclusions MRI measurements of cardiac T2 and T2* can be used to quantify cardiac iron. The similarity of liver and cardiac iron calibration curves in the gerbil suggests that extrapolation of human liver calibration curves to heart may be a rational approximation in humans. PMID:16027257

  14. Effect of perindopril and bisoprolol on IL-2, INF-γ, hs-CRP, and T-cell stimulation and its correlations with blood pressure in mild and moderate hypertension
.

    PubMed

    Madej, Andrzej; Dąbek, Józefa; Majewski, Michał; Szuta, Justyna

    2018-05-29

    Cardiovascular disease is the leading global cause of death. Its development is largely determined by lifestyle and cardiovascular risk factors, among which arterial hypertension (HT) plays a key role. Available data indicate a significant role of inflammation in the pathophysiology of HT. The aim of this study was to assess concentrations of mediators of inflammation in patients with mild and moderate HT and its modulation with antihypertensive treatment. 56 subjects: 17 with first- and 20 with second-grade HT, and 19 healthy subjects constituting a control group. Antihypertensive therapy: in first-grade HT, perindopril 5 mg daily, and in second-grade HT, bisoprolol (5 mg daily) additionally were ordered. Before and after a 4-week treatment, interleukine-2, interferon-γ, and high-sensitivity C-reactive protein serum concentrations were assessed. hs-CRP concentration was higher in hypertensive patients, and the difference was statistically significant in patients with second-grade HT compared to the control group (1.42 mg/L vs. 2.55 mg/L; p = 0.003). Antihypertensive treatment was associated with hs-CRP decrease. IL-2 concentration was 14.8% (p = 0.005) higher in first- and 22.2% (p = 0.002) higher in second-grade HT compared to the controls. Treatment significantly reduced concentration of IL-2. IFN-γ concentrations, compared to the control group, were higher by 19.6% (p < 0.001) and 39.9% (p < 0.001) in 1st and 2nd grade HT, respectively. Increased values of blood pressure were accompanied by higher concentrations of cytokines and hs-CRP. Reversal of adverse T-cell stimulation was observed after 4-week therapy.
.

  15. Evaluation of efficacy of resin hemoperfusion in patients with acute 2,4-dinitrophenol poisoning by dynamic monitoring of plasma toxin concentration.

    PubMed

    Zhao, Xue-hong; Jiang, Jiu-kun; Lu, Yuan-qiang

    2015-08-01

    The intoxications caused by 2,4-dinitrophenol (2,4-DNP), even death, have been frequently reported in recent years. This study aims to investigate the dynamic changes of plasma toxin concentration and explore the clinical value of resin hemoperfusion (HP) in the treatment of patients with acute 2,4-DNP poisoning. We reported 16 cases of acute 2,4-DNP poisoning through occupational exposure due to ignoring the risk of poisoning. The blood samples were collected from the 14 survivors. According to the different treatments of resin HP, the survivors were divided into routine HP (n=5) and intensive HP (n=9) groups. Ultra high performance liquid chromatography/ tandem mass spectroscopy (UPLC-MS/MS) was used to detect the 2,4-DNP concentration in plasma in this study. The 14 survivors recovered very well after treatment. The initial plasma 2,4-DNP concentrations (C1) of survivors ranged from 0.25 to 41.88 µg/ml (mean (12.56±13.93) µg/ml). A positive correlation existed between initial plasma 2,4-DNP concentration (C1) and temperature. The elimination of 2,4-DNP was slow and persistent, and the total clearance rates of plasma toxin from the 1st to 3rd day (R3), the 3rd to 7th day (R3-7), and the 1st to 7th day (R7), were only (53.03±14.04)%, (55.25±10.50)%, and (78.29±10.22)%, respectively. The plasma toxin was cleared up to 25 d after poisoning in most of the patients. The R3, R3-7, and R7 in the intensive HP group were all apparently higher than those in the routine HP group, with statistical significance (P<0.05). Simultaneously, the elimination half-life (t1/2) of 2,4-DNP in the intensive HP group was apparently shorter than that in the routine HP group, with statistical significance (P<0.05). The clinicians should be aware of this slow and persistent process in the elimination of plasma 2,4-DNP. Higher initial plasma toxin concentration resulted in a more severe fever for the patient. According to the limited data, longer and more frequent resin HP may

  16. Pertussis toxin-sensitive G-protein mediates the alpha 2-adrenergic receptor inhibition of melatonin release in photoreceptive chick pineal cell cultures

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Pratt, B.L.; Takahashi, J.S.

    The avian pineal gland is a photoreceptive organ that has been shown to contain postjunctional alpha 2-adrenoceptors that inhibit melatonin synthesis and/or release upon receptor activation. Physiological response and (32P)ADP ribosylation experiments were performed to investigate whether pertussis toxin-sensitive guanine nucleotide-binding proteins (G-proteins) were involved in the transduction of the alpha 2-adrenergic signal. For physiological response studies, the effects of pertussis toxin on melatonin release in dissociated cell cultures exposed to norepinephrine were assessed. Pertussis toxin blocked alpha 2-adrenergic receptor-mediated inhibition in a dose-dependent manner. Pertussis toxin-induced blockade appeared to be noncompetitive. One and 10 ng/ml doses of pertussis toxinmore » partially blocked and a 100 ng/ml dose completely blocked norepinephrine-induced inhibition. Pertussis toxin-catalyzed (32P)ADP ribosylation of G-proteins in chick pineal cell membranes was assessed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and autoradiography. Membranes were prepared from cells that had been pretreated with 0, 1, 10, or 100 ng/ml pertussis toxin. In the absence of pertussis toxin pretreatment, two major proteins of 40K and 41K mol wt (Mr) were labeled by (32P)NAD. Pertussis toxin pretreatment of pineal cells abolished (32P) radiolabeling of the 40K Mr G-protein in a dose-dependent manner. The norepinephrine-induced inhibition of both cAMP efflux and melatonin release, as assessed by RIA of medium samples collected before membrane preparation, was also blocked in a dose-dependent manner by pertussis toxin. Collectively, these results suggest that a pertussis toxin-sensitive 40K Mr G-protein labeled by (32P)NAD may be functionally associated with alpha 2-adrenergic signal transduction in chick pineal cells.« less

  17. Isolation and Characteristics of Shiga Toxin 2f-Producing Escherichia coli among Pigeons in Kyushu, Japan

    PubMed Central

    Murakami, Koichi; Etoh, Yoshiki; Ichihara, Sachiko; Maeda, Eriko; Takenaka, Shigeyuki; Horikawa, Kazumi; Narimatsu, Hiroshi; Kawano, Kimiko; Kawamura, Yoshiaki; Ito, Kenitiro

    2014-01-01

    An increasing number of Shiga toxin 2f-producing Escherichia coli (STEC2f) infections in humans are being reported in Europe, and pigeons have been suggested as a reservoir for the pathogen. In Japan, there is very little information regarding carriage of STEC2f by pigeons, prompting the need for further investigation. We collected 549 samples of pigeon droppings from 14 locations in Kyushu, Japan, to isolate STEC2f and to investigate characteristics of the isolates. Shiga toxin stx 2f gene fragments were detected by PCR in 16 (2.9%) of the 549 dropping samples across four of the 14 locations. We obtained 23 STEC2f-isolates from seven of the original samples and from three pigeon dropping samples collected in an additional sampling experiment (from a total of seven locations across both sampling periods). Genotypic and phenotypic characteristics were then examined for selected isolates from each of 10 samples with pulsed-field gel electrophoresis profiles. Eight of the stx 2f gene fragments sequenced in this study were homologous to others that were identified in Europe. Some isolates also contained virulence-related genes, including lpfA O26, irp 2, and fyuA, and all of the 10 selected isolates maintained the eae, astA, and cdt genes. Moreover, five of the 10 selected isolates contained sfpA, a gene that is restricted to Shiga toxin-producing E. coli O165:H2 and sorbitol-fermenting Shiga toxin-producing E. coli O157:NM. We document serotypes O152:HNM, O128:HNM, and O145:H34 as STEC2f, which agrees with previous studies on pigeons and humans. Interestingly, O119:H21 was newly described as STEC2f. O145:H34, with sequence type 722, was described in a German study in humans and was also isolated in the current study. These results revealed that Japanese zoonotic STEC2f strains harboring several virulence-related factors may be of the same clonal complexes as some European strains. These findings provide useful information for public health-related disease management

  18. Purification and characterization of Yersinia enterocolitica and Yersinia pestis LcrV-cholera toxin A(2)/B chimeras.

    PubMed

    Tinker, Juliette K; Davis, Chadwick T; Arlian, Britni M

    2010-11-01

    Yersinia pestis is a virulent human pathogen and potential biological weapon. Despite a long history of research on this organism, there is no licensed vaccine to protect against pneumonic forms of Y. pestis disease. In the present study, plasmids were constructed to express cholera toxin A(2)/B chimeric molecules containing the LcrV protective antigen from Yersinia enterocolitica and Y. pestis. These chimeras were expressed and purified to high yields from the supernatant of transformed Escherichia coli. Western and GM(1) ELISA assays were used to characterize the composition, receptor-binding and relative stability of the LcrV-CTA(2)/B chimera in comparison to cholera toxin. In addition, we investigated the ability of the Y. pestis LcrV-CTA(2)/B chimera to bind to and internalize into cultured epithelial cells and macrophages by confocal microscopy. These studies indicate that the uptake and trafficking of the LcrV antigen from the chimera is comparable to the trafficking of native toxin. Together these findings report that stable, receptor-binding, non-toxic LcrV-cholera toxin A(2)/B chimeras can be expressed at high levels in E. coli and purified from the supernatant. In addition, the internalization of antigen in vitro reported here supports the development of these molecules as novel mucosal vaccine candidates. Copyright 2010 Elsevier Inc. All rights reserved.

  19. Purification and characterization of Yersinia enterocolitica and Yersinia pestis LcrV–cholera toxin A2/B chimeras

    PubMed Central

    Davis, Chadwick T.; Arlian, Britni M.

    2010-01-01

    Yersinia pestis is a virulent human pathogen and potential biological weapon. Despite a long history of research on this organism, there is no licensed vaccine to protect against pneumonic forms of Y. pestis disease. In the present study, plasmids were constructed to express cholera toxin A2/B chimeric molecules containing the LcrV protective antigen from Y. enterocolitica and Y. pestis. These chimeras were expressed and purified to high yields from the supernatant of transformed E. coli. Western and GM1 ELISA assays were used to characterize the composition, receptor-binding and relative stability of the LcrV-CTA2/B chimera in comparison to cholera toxin. In addition, we investigated the ability of the Y. pestis LcrV-CTA2/B chimera to bind to and internalize into cultured epithelial cells and macrophages by confocal microscopy. These studies indicate that the uptake and trafficking of the LcrV antigen from the chimera is comparable to the trafficking of native toxin. Together these findings report that stable, receptor-binding, non-toxic LcrV-cholera toxin A2/B chimeras can be expressed at high levels in E. coli and purified from the supernatant. In addition, the internalization of antigen in vitro reported here supports the development of these molecules as novel mucosal vaccine candidates. PMID:20438844

  20. Native serotonin 5-HT3 receptors expressed in Xenopus oocytes differ from homopentameric 5-HT3 receptors.

    PubMed

    van Hooft, J A; Kreikamp, A P; Vijverberg, H P

    1997-09-01

    Efficacies of the 5-hydroxytryptamine (serotonin) 5-HT3 receptor (5-HT3R) agonists 2-methyl-5-HT, dopamine, and m-chlorophenylbiguanide on 5-HT3R native to N1E-115 cells and on homopentameric 5-HT3R expressed in Xenopus oocytes were determined relative to that of 5-HT. Efficacies of 2-methyl-5-HT and dopamine on 5-HT3R native to differentiated N1E-115 cells are high (54 and 36%) as compared with their efficacies on homopentameric 5-HT3R-A(L) and 5-HT3R-A(S) receptors expressed in oocytes (4-8%). m-Chlorophenylbiguanide does not distinguish between 5-HT3R in N1E-115 cells and in oocytes. The distinct pharmacological profile of 5-HT3R native to differentiated N1E-115 cells is conserved when poly(A)+ mRNA from these cells is expressed in oocytes. The results indicate that, apart from the known 5-HT3R subunits, N1E-115 cells express additional proteins involved in 5-HT3R function.

  1. Susceptibility of Skeletal Muscle to Coxsackie A2 Virus Infection: Effects of Botulinum Toxin and Denervation

    NASA Astrophysics Data System (ADS)

    Andrew, Clifford G.; Drachman, Daniel B.; Pestronk, Alan; Narayan, Opendra

    1984-02-01

    Coxsackie A viruses can infect denervated but not innervated mature skeletal muscles. The role of synaptic transmission in preventing susceptibility to Coxsackievirus infection was studied by surgically denervating leg muscles of mice or injecting the muscles with botulinum toxin to block quantal release of acetylcholine. Control muscles were injected with heat-inactivated toxin. Subsequent injection of Coxsackie A2 virus resulted in extensive virus replication and tissue destruction in the denervated and botulinum toxin-treated muscles, while the control muscles showed only minimal changes. This suggests that the susceptibility of skeletal muscle to Coxsackievirus infection is regulated by synaptic transmission.

  2. The central action of the 5-HT2 receptor agonist 1-(2,5-dimethoxy-4-iodophenyl)-2-aminopropane (DOI) on cardiac inotropy and vascular resistance in the anaesthetized cat

    PubMed Central

    Ramage, Andrew G; de Burgh Daly, M

    1998-01-01

    Experiments were carried out to determine the effects of the application of the selective 5-HT2 receptor agonist DOI intravenously (in the presence of the peripherally acting 5-HT2 receptor antagonist, BW501C67, 1 mg kg−1, i.v.) or to the `glycine sensitive area' of the ventral surface (30 μg each side) on the left ventricular inotropic (left ventricular dP/dt max) and vascularly isolated hindlimb responses in anaesthetized cats. For the ventral surface experiments, NMDA (10 μg each side) was applied to act as a positive control. In all experiments heart rate and mean arterial blood pressure were held constant to exclude any secondary effects caused by changes in these variables.DOI (n=6) i.v or on the ventral surface had no effect on left ventricular dP/dt max but caused a significant increase in hindlimb perfusion pressure of 40±9 and 50±14 mmHg, respectively. Respiration was unaffected. NMDA (n=6), applied to the ventral surface, caused significant increases in both left ventricular dP/dt max and hindlimb perfusion pressure of 1950±349 mmHg s−1 and 69±17 mmHg respectively, with no associated change in left ventricular end-diastolic pressure. The amplitude of respiratory movements increased.It is concluded that activation of 5-HT2 receptors at the level of the rostral ventrolateral medulla (RVLM) excites sympathetic premotor neurons and/or their antecedents controlling hindlimb vascular resistance but not those controlling the inotropic effects on the left ventricle. PMID:9863644

  3. Prevalence and factors associated with type 2 diabetes mellitus and hypertension among the hill tribe elderly populations in northern Thailand.

    PubMed

    Apidechkul, Tawatchai

    2018-06-05

    Type 2 diabetes mellitus (T2DM) and hypertension (HT) are major noncommunicable health problems in both developing and developed countries, including Thailand. Each year, a large amount of money is budgeted for treatment and care. Hill tribe people are a marginalized population in Thailand, and members of its elderly population are vulnerable to health problems due to language barriers, lifestyles, and daily dietary intake. An analytic cross-sectional study was conducted to estimate the prevalence of T2DM and HT and to assess the factors associated with T2DM and HT. The study populations were hill tribe elderly adults aged ≥  60 years living in Chiang Rai Province, Thailand. A simple random method was used to select the targeted hill tribe villages and participants into the study. A validated questionnaire, physical examination form, and 5-mL blood specimen were used as research instruments. Fasting plasma glucose and blood pressure were examined and used as outcome measurements. Chi-square tests and logistic regression were used for detecting the associations between variables at the significance level alpha=0.05. In total, 793 participants participated in the study; 49.6% were male, and 51.7% were aged 60-69 years. A total of 71.5% were Buddhist, 93.8% were uneducated, 62.9% were unemployed, and 89 % earned an income of < 5,000 baht/month. The overall prevalence of T2DM and HT was 16.8% and 45.5%, respectively. Approximately 9.0% individuals had comorbidity of T2DM and HT. Members of the Lahu, Yao, Karen, and Lisu tribes had a greater odds of developing T2DM than did those of the Akha tribe. Being overweight, having a parental history of T2DM, and having high cholesterol were associated with T2DM development. In contrast, those who engaged in highly physical activities and exercise had lower odds of developing T2DM than did those who did not. Regarding HT, being female, having a high dietary salt intake, being overweight, and having a parental history of HT

  4. Sedative effect of Clozapine is a function of 5-HT2A and environmental novelty.

    PubMed

    Joshi, Radhika S; Quadros, Rolen; Drumm, Michael; Ain, Rupasri; Panicker, Mitradas M

    2017-01-01

    Antipsychotic drugs are the mainstay in the treatment of schizophrenia and bipolar disorder. However, antipsychotics often exhibit sedation or activity suppression among many other side effects, and the factors that influence them remain poorly understood. We now show, using a 5-HT 2A knockout (Htr2a -/- ) mouse, that environmental circumstances can affect suppression of activity induced by the atypical antipsychotic- Clozapine. We observed that Htr2a -/- mice were more resistant to Clozapine-induced suppression of activity (CISA) and this behaviour was dependent on the environment being 'novel'. In their 'home' environment, at identical doses the mice exhibited CISA. Interestingly, the effect of genotype and environmental novelty on CISA could not be extended to the other antipsychotics that were tested, i.e. Haloperidol and Risperidone. Haloperidol-induced activity suppression was independent of context and genotype. Whereas context affected Risperidone-induced activity suppression only in the Htr2a +/+ mice. Furthermore, we observed that caffeine, a stimulant, elicited resistance to CISA similar to that seen in the 'novel' context. Our study establishes a previously unknown interaction between the environmental context, 5-HT 2A and CISA and emphasises the role of non-pharmacological factors such as environment on the effects of the drug, which seem antipsychotic-specific. Our findings should advance the understanding of the side effects of individual antipsychotics and the role of environment to overcome side effects such as sedation. Copyright © 2016 Elsevier B.V. and ECNP. All rights reserved.

  5. Differential effects of 5-HT2C receptor activation by WAY 161503 on nicotine-induced place conditioning and locomotor activity in rats.

    PubMed

    Hayes, Dave J; Mosher, Tera M; Greenshaw, Andrew J

    2009-02-11

    Numerous studies indicate a role for both the serotonin 2C receptor (5-HT(2C)) and the nicotinic acetylcholine receptor in locomotion, reinforcement and motivated behaviours. Nicotine, a potent nicotinic acetylcholine receptor agonist, interacts with the dopaminergic and serotonergic systems and is known to positively affect reward-related behaviours. The current study examined the effects of 5-HT(2C) receptor activation on nicotine-induced (0.6 mg/kg) place conditioning and spontaneous locomotion. Using Sprague-Dawley rats, the effects of the selective 5-HT(2C) receptor agonist WAY 161503 (0-1.0 mg/kg) and the selective 5-HT(2C) receptor antagonist SB 242084 (1.0 mg/kg) alone, in combination, and on nicotine-induced (0.6 mg/kg) spontaneous locomotor activity were assessed. The effects of WAY 161503 (1.0, 3.0 mg/kg) were also investigated in nicotine-induced place conditioning using a two-compartment biased design; amphetamine (1.0 mg/kg) served as a positive control. As differential effects were observed between place conditioning and locomotor activity, the subjects used in the place conditioning experiments were also tested for effects on locomotor activity. WAY 161503 decreased baseline and nicotine-induced locomotor activity at the highest dose tested (1.0mg/kg) and these effects were attenuated by SB 242084. Amphetamine and nicotine both induced robust place preferences and WAY 161503 did not have any effects in the context of place conditioning. In contrast, WAY 161503 (1.0 mg/kg) blocked nicotine-induced locomotor activity. These results suggest that 5-HT(2C) receptors may play an inhibitory role in nicotine-induced locomotor activity, but do not appear to influence place conditioning under the current conditions.

  6. Tailoring the interface using thiophene small molecules in TiO2/P3HT hybrid solar cells.

    PubMed

    Freitas, Flavio S; Clifford, John N; Palomares, Emilio; Nogueira, Ana F

    2012-09-14

    In this paper we focus on the effect of carboxylated thiophene small molecules as interface modifiers in TiO(2)/P3HT hybrid solar cells. Our results show that small differences in the chemical structure of these molecules, for example, the presence of the -CH(2)- group in the 2-thiopheneacetic acid (TAA), can greatly increase the TiO(2) surface wettability, improving the TiO(2)/polymer contact. This effect is important to enhance exciton splitting and charge separation.

  7. The Staphylococcus aureus Alpha-Toxin Perturbs the Barrier Function in Caco-2 Epithelial Cell Monolayers by Altering Junctional Integrity

    PubMed Central

    Vikström, Elena; Magnusson, Karl-Eric; Vécsey-Semjén, Beatrix; Colque-Navarro, Patricia; Möllby, Roland

    2012-01-01

    Increased microvascular permeability is a hallmark of sepsis and septic shock. Intestinal mucosal dysfunction may allow translocation of bacteria and their products, thereby promoting sepsis and inflammation. Although Staphylococcus aureus alpha-toxin significantly contributes to sepsis and perturbs the endothelial barrier function, little is known about possible effects of S. aureus alpha-toxin on human epithelial barrier functions. We hypothesize that S. aureus alpha-toxin in the blood can impair the intestinal epithelial barrier and thereby facilitate the translocation of luminal bacteria into the blood, which may in turn aggravate a septic condition. Here, we showed that staphylococcal alpha-toxin disrupts the barrier integrity of human intestinal epithelial Caco-2 cells as evidenced by decreased transepithelial electrical resistance (TER) and reduced cellular levels of junctional proteins, such as ZO-1, ZO-3, and E-cadherin. The Caco-2 cells also responded to alpha-toxin with an elevated cytosolic calcium ion concentration ([Ca2+]i), elicited primarily by calcium influx from the extracellular environment, as well as with a significant reduction in TER, which was modulated by intracellular calcium chelation. Moreover, a significantly larger reduction in TER and amounts of the junctional proteins, viz., ZO-3 and occludin, was achieved by basolateral than by apical application of the alpha-toxin. These experimental findings thus support the hypothesis that free staphylococcal alpha-toxin in the bloodstream may cause intestinal epithelial barrier dysfunction and further aggravate the septic condition by promoting the release of intestinal bacteria into the underlying tissues and the blood. PMID:22354024

  8. Isolation, purification, and characterization of antimicrobial compound 6-[1,2-dimethyl-6-(2-methyl-allyloxy)-hexyl]-3-(2-methoxy-phenyl)-chromen-4-one from Penicillium sp. HT-28.

    PubMed

    Kaur, Harpreet; Arora, Daljit Singh; Sharma, Vishal

    2014-08-01

    A fungal culture (Penicillium sp., HT-28), isolated from soil has been evaluated for its bioactivity, which showed broad spectrum antimicrobial activity and was effective against methicillin-resistant Staphylococcus aureus (MRSA) also. Statistical optimization of the medium by response surface methodology (RSM) enhanced the antimicrobial activity up to 1.8-fold. Column chromatography was used to isolate the active compound (A), which was characterized to be 6-[1,2-dimethyl-6-(2-methyl-allyloxy)-hexyl]-3-(2-methoxy-phenyl)-chromen-4-one by various spectroscopic techniques such as infrared (IR), (1)H and (13)C nuclear magnetic resonance (NMR) spectra, and mass spectroscopy. Minimum inhibitory concentration (MIC) of the active compound (A) ranged from 0.5 to 15 μg/mL. Viable cell count studies of the active compound (A) showed S. aureus, Escherichia coli, Staphylococcus epidermidis, and Salmonella typhimurium 1 to be the most sensitive. The compound retained its bioactivity after treating it at 100 °C for 1 h. Furthermore, the compound (A) when tested for its biosafety was found neither to be cytotoxic nor mutagenic. The study demonstrated that an apparently novel compound isolated from Penicillium sp. (HT-28) seems to be a stable and potent antimicrobial.

  9. Influence of T-2 Mycotoxin on Host Resistance to Candida albicans Infections in Mice

    DTIC Science & Technology

    1987-04-20

    body (18, 21). In candidiasis , a3 in most other infectious diseases, the major host defense mechanism is cell-mediated immunity. Our studies denons rate...ate M819, obtained from a human case of candidiasis (Dr. Howard W. Larsh, Mis ri State Chest Hospital, Mt. Vernon, Mo) was used in all studies. The org...these studies present evidence that multiple exposure to T-2 toxin has a significant synergistic effect on the pathogenesis of the opportunistic

  10. Cholera Toxin Inhibits the T-Cell Antigen Receptor-Mediated Increases in Inositol Trisphosphate and Cytoplasmic Free Calcium

    NASA Astrophysics Data System (ADS)

    Imboden, John B.; Shoback, Dolores M.; Pattison, Gregory; Stobo, John D.

    1986-08-01

    The addition of monoclonal antibodies to the antigen receptor complex on the malignant human T-cell line Jurkat generates increases in inositol trisphosphate and in the concentration of cytoplasmic free calcium. Exposure of Jurkat cells to cholera toxin for 3 hr inhibited these receptor-mediated events and led to a selective, partial loss of the antigen receptor complex from the cellular surface. None of the effects of cholera toxin on the antigen receptor complex were mimicked by the B subunit of cholera toxin or by increasing intracellular cAMP levels with either forskolin or 8-bromo cAMP. These results suggest that a cholera toxin substrate can regulate signal transduction by the T-cell antigen receptor.

  11. Actions of novel antipsychotic agents on apomorphine-induced PPI disruption: influence of combined serotonin 5-HT1A receptor activation and dopamine D2 receptor blockade.

    PubMed

    Auclair, Agnès L; Kleven, Mark S; Besnard, Joël; Depoortère, Ronan; Newman-Tancredi, Adrian

    2006-09-01

    The dopamine D1/D2 agonist apomorphine (0.63 mg/kg) disrupted prepulse inhibition (PPI) of acoustic startle in rats, a model of sensorimotor gating deficits observed in schizophrenia. All current antipsychotics, which antagonize D2 receptors, prevent this apomorphine-induced deficit. A novel class of antipsychotics possesses, in addition to D2 antagonist property, various levels of 5-HT1A agonist activity. Considering that the latter itself produces PPI deficits, it appeared necessary to assess the potential of this novel class of antipsychotics to reverse apomorphine-PPI deficits. Potent D2 antagonists, like haloperidol (0.63-2.5 mg/kg), risperidone (0.63-10 mg/kg), and olanzapine (0.63-40 mg/kg) prevented apomorphine PPI disruption. The atypical antipsychotics, clozapine (40 mg/kg), nemonapride (0.01-2.5 mg/kg), ziprasidone (10 mg/kg), and aripiprazole (0.01 and 10 mg/kg), which all exhibit 5-HT1A agonist properties, reversed PPI deficits at some doses only, whereas the anti-dyskinetic agent sarizotan (0.16-10 mg/kg), an efficacious 5-HT1A agonist, did not. New generation antipsychotics with marked 5-HT1A agonist properties, such as SLV313 and SSR181507 (0.0025-10 mg/kg and 0.16-10 mg/kg, respectively) did not reverse these deficits whereas bifeprunox (0.04-2.5 mg/kg) did. To reveal the contribution of 5-HT1A agonist properties in the lack of effects of SLV313 and SSR181507, we pretreated rats with the 5-HT1A antagonist WAY100635 (0.63 mg/kg). Under these conditions, significant reversal of PPI deficit was observed, indicating that D2 antagonist properties of SLV313 and SSR181507 are now sufficient to overcome the disruptive effects of apomorphine. To summarize, antipsychotics possessing agonist efficacy at 5-HT1A receptors exhibit diverse profiles against apomorphine-induced PPI deficits, depending on the balance between D2 and 5-HT1A activities, suggesting that they may display distinct activity on some aspects of gating deficits in schizophrenic patients.

  12. Lorcaserin (APD356), a selective 5-HT(2C) agonist, reduces body weight in obese men and women.

    PubMed

    Smith, Steven R; Prosser, Warren A; Donahue, David J; Morgan, Michael E; Anderson, Christen M; Shanahan, William R

    2009-03-01

    Lorcaserin (APD356) is a potent, selective 5-HT(2C) agonist with ~15-fold and 100-fold selectivity vs. 5-HT(2A) and 5-HT(2B) receptors, respectively. This study evaluated the safety and efficacy of lorcaserin for weight reduction in obese patients during a 12-week period. The randomized, double-blind, placebo-controlled, parallel-arm study enrolled 469 men and women between ages 18 and 65 and with BMI 30-45 kg/m(2). Patients received placebo, lorcaserin 10 mg q.d., lorcaserin 15 mg q.d., or lorcaserin 10 mg b.i.d. for 12 weeks, and were counseled to maintain their usual diet and activity. The primary end point was change in weight from baseline to day 85 by completer analysis. Safety analyses included echocardiograms at Screening and day 85/study exit. Lorcaserin was associated with progressive weight loss of 1.8 kg, 2.6 kg, and 3.6 kg at 10 mg q.d., 15 mg q.d., and 10 mg b.i.d., respectively, compared to placebo weight loss of 0.3 kg (P < 0.001 for each group). Similar results were seen by intent-to-treat last observation-carried forward (ITT-LOCF) analysis. The proportions of completers achieving > or =5% of initial body weight were 12.8, 19.5, 31.2, and 2.3% in the 10 mg q.d., 15 mg q.d., 10 mg b.i.d., and placebo groups, respectively. The most frequent adverse events (AEs) were transient headache, nausea, and dizziness. Echocardiograms showed no apparent drug-related effects on heart valves or pulmonary artery pressure (PAP). Lorcaserin was well tolerated and efficacious for weight reduction in this 12-week study. Longer-term trials employing behavior modification will be needed to more fully assess its safety and efficacy.

  13. Constitutively active 5-HT2/α1 receptors facilitate muscle spasms after human spinal cord injury

    PubMed Central

    D'Amico, Jessica M.; Murray, Katherine C.; Li, Yaqing; Chan, K. Ming; Finlay, Mark G.; Bennett, David J.

    2013-01-01

    In animals, the recovery of motoneuron excitability in the months following a complete spinal cord injury is mediated, in part, by increases in constitutive serotonin (5-HT2) and norepinephrine (α1) receptor activity, which facilitates the reactivation of calcium-mediated persistent inward currents (CaPICs) without the ligands serotonin and norepinephrine below the injury. In this study we sought evidence for a similar role of constitutive monoamine receptor activity in the development of spasticity in human spinal cord injury. In chronically injured participants with partially preserved sensory and motor function, the serotonin reuptake inhibitor citalopram facilitated long-lasting reflex responses (spasms) previously shown to be mediated by CaPICs, suggesting that in incomplete spinal cord injury, functional descending sources of monoamines are present to activate monoamine receptors below the lesion. However, in participants with motor or motor/sensory complete injuries, the inverse agonist cyproheptadine, which blocks both ligand and constitutive 5-HT2/α1 receptor activity, decreased long-lasting reflexes, whereas the neutral antagonist chlorpromazine, which only blocks ligand activation of these receptors, had no effect. When tested in noninjured control participants having functional descending sources of monoamines, chlorpromazine was effective in reducing CaPIC-mediated motor unit activity. On the basis of these combined results, it appears that in severe spinal cord injury, facilitation of persistent inward currents and muscle spasms is mainly mediated by the activation of constitutive 5-HT2 and α1 receptor activity. Drugs that more selectively block these constitutively active monoamine receptors may provide better oral control of spasticity, especially in motor complete spinal cord injury where reducing motoneuron excitability is the primary goal. PMID:23221402

  14. Effects of autoshaping procedures on 3H-8-OH-DPAT-labeled 5-HT1a binding and 125I-LSD-labeled 5-HT2a binding in rat brain.

    PubMed

    Tomie, Arthur; Di Poce, Jason; Aguado, Allison; Janes, Amy; Benjamin, Daniel; Pohorecky, Larissa

    2003-06-13

    Effects of experience with Pavlovian autoshaping procedures on lever-press autoshaping conditioned response (CR) performance and 3H-8-OH-DPAT-labeled binding of 5-HT(1a) receptors as well as 125I-LSD-labeled binding of 5-HT(2a) receptors were evaluated in four groups of male Long-Evans hooded rats. Two groups of rats (Group Paired High CR and Group Paired Low CR) received Pavlovian autoshaping procedures wherein the presentation of a lever (conditioned stimulus, CS) was followed by the response-independent presentation of food (unconditioned stimulus, US). Rats in Group Paired High CR (n=12) showed more rapid CR acquisition and higher asymptotic levels of lever-press autoshaping CR performance relative to rats in Group Low CR (n=12). Group Omission (n=9) received autoshaping with an omission contingency, such that performing the lever-press autoshaping CR resulted in the cancellation the food US, while Group Random (n=9) received presentations of lever CS and food US randomly with respect to one another. Though Groups Omission and Random did not differ in lever-press autoshaping CR performance, Group Omission showed significantly lower levels of 3H-8-OH-DPAT-labeled 5-HT(1a) binding in post-synaptic areas (frontal cortex, septum, caudate putamen), as well as significantly higher plasma corticosterone levels than Group Random. In addition, Group Random showed higher levels of 3H-8-OH-DPAT-labeled 5-HT(1a) binding in pre-synaptic somatodendritic autoreceptors on dorsal raphe nucleus relative to each of the other three groups. Autoradiographic analysis of 125I-LSD-labeled 5-HT(2a) receptor binding revealed no significant differences between Groups Paired High CR and Paired Low CR or between Groups Omission and Random in any brain regions.

  15. Cannabidiol restores intestinal barrier dysfunction and inhibits the apoptotic process induced by Clostridium difficile toxin A in Caco-2 cells.

    PubMed

    Gigli, Stefano; Seguella, Luisa; Pesce, Marcella; Bruzzese, Eugenia; D'Alessandro, Alessandra; Cuomo, Rosario; Steardo, Luca; Sarnelli, Giovanni; Esposito, Giuseppe

    2017-12-01

    Clostridium difficile toxin A is responsible for colonic damage observed in infected patients. Drugs able to restore Clostridium difficile toxin A-induced toxicity have the potential to improve the recovery of infected patients. Cannabidiol is a non-psychotropic component of Cannabis sativa, which has been demonstrated to protect enterocytes against chemical and/or inflammatory damage and to restore intestinal mucosa integrity. The purpose of this study was to evaluate (a) the anti-apoptotic effect and (b) the mechanisms by which cannabidiol protects mucosal integrity in Caco-2 cells exposed to Clostridium difficile toxin A. Caco-2 cells were exposed to Clostridium difficile toxin A (30 ng/ml), with or without cannabidiol (10 -7 -10 -9  M), in the presence of the specific antagonist AM251 (10 -7  M). Cytotoxicity assay, transepithelial electrical resistence measurements, immunofluorescence analysis and immunoblot analysis were performed in the different experimental conditions. Clostridium difficile toxin A significantly decreased Caco-2 cells' viability and reduced transepithelial electrical resistence values and RhoA guanosine triphosphate (GTP), bax, zonula occludens-1 and occludin protein expression, respectively. All these effects were significantly and concentration-dependently inhibited by cannabidiol, whose effects were completely abolished in the presence of the cannabinoid receptor type 1 (CB1) antagonist, AM251. Cannabidiol improved Clostridium difficile toxin A-induced damage in Caco-2 cells, by inhibiting the apoptotic process and restoring the intestinal barrier integrity, through the involvement of the CB1 receptor.

  16. Both exogenous 5-HT and endogenous 5-HT, released by fluoxetine, enhance distension evoked propulsion in guinea-pig ileum in vitro.

    PubMed

    Gwynne, Rachel M; Clarke, Amanda J; Furness, John B; Bornstein, Joel C

    2014-01-01

    The roles of 5-HT3 and 5-HT4 receptors in the modulation of intestinal propulsion by luminal application of 5-HT and augmentation of endogenous 5-HT effects were studied in segments of guinea-pig ileum in vitro. Persistent propulsive contractions evoked by saline distension were examined using a modified Trendelenburg method. When 5-HT (30 nM), fluoxetine (selective serotonin reuptake inhibitor; 1 nM), 2-methyl-5-HT (5-HT3 receptor agonist; 1 mM), or RS 67506 (5-HT4 receptor agonist, 1 μM) was infused into the lumen, the pressure needed to initiate persistent propulsive activity fell significantly. A specific 5-HT4 receptor antagonist, SB 207266 (10 nM in lumen), abolished the effects of 5-HT, fluoxetine, and RS 67506, but not those of 2-methyl-5-HT. Granisetron (5-HT3 receptor antagonist; 1 μM in lumen) abolished the effect of 5-HT, fluoxetine, RS 67506, and 2-methyl-5-HT. The NK3 receptor antagonist SR 142801 (100 nM in lumen) blocked the effects of 5-HT, fluoxetine, and 2-methyl-5-HT. SB 207266, granisetron, and SR 142801 had no effect by themselves. Higher concentrations of fluoxetine (100 and 300 nM) and RS 67506 (3 and 10 μM) had no effect on the distension threshold for propulsive contractions. These results indicate that luminal application of exogenous 5-HT, or increased release of endogenous mucosal 5-HT above basal levels, acts to lower the threshold for propulsive contractions in the guinea-pig ileum via activation of 5-HT3 and 5-HT4 receptors and the release of tachykinins. The results further indicate that basal release of 5-HT is insufficient to alter the threshold for propulsive motor activity.

  17. miR-29a-3p/T-bet Regulatory Circuit Is Altered in T Cells of Patients With Hashimoto's Thyroiditis.

    PubMed

    Tokić, Stana; Štefanić, Mario; Glavaš-Obrovac, Ljubica; Kishore, Amit; Navratilova, Zdenka; Petrek, Martin

    2018-01-01

    Hashimoto's thyroiditis (HT) is a common autoimmune thyroid disorder that frequently evolves from asymptomatic, T-cell mediated chronic inflammation toward overt hypothyroidism. Previously, we have demonstrated a role for T-bet, a T helper 1/CD8 + T cell transcription factor (TF), and FoxP3, a regulatory T cell TF, in disease progression and severity, but the basis behind their altered mRNA expression remains unknown. In this study, we aimed to leverage the role for microRNAs, representing negative transcriptional regulators, across the spectrum of HT clinical presentations using the same, well-characterized RNA sample cohort. Ten hypothyroid, untreated patients (hypoHT), 10 hypothyroid cases rendered euthyroid by l-thyroxine therapy (substHT), 11 spontaneously euthyroid HT subjects (euHT), and 10 healthy controls (ctrl) were probed for three candidate immunoregulatory miRNA (miR-9-5p, miR-29a-3p, and miR-210-3p) using quantitative real-time PCR measurements. Data were normalized to U6snRNA and fold difference in expression calculated by the efficiency corrected 2 -ΔΔCt model. Compared to healthy controls, peripheral blood (PB) T cells of HT patients exhibited significantly diminished miR-29a-3p expression levels [median expression levels (IQR), HT vs CTRL, 0.62 (0.44-1.01) vs 1.373 (0.63-2.7), P  = 0.046], and a similar, but not significant decline in miR-210-3p abundance [HT vs CTRL, 0.64 (0.39-1.31) vs 1.2 (0.5-2.56), P  = 0.24, Wilcoxon test]. A significant inverse correlation was observed between the two differentially expressed transcripts, T-bet mRNA and miR-29a-3p. Moreover, altered miR-29a-3p/T-bet expression in T cells of untreated HT patients was related to low serum FT4, high serum thyrotropin, and decreased thyroid volumes. Of note, miR-210-3p expression was positively correlated to HIF1α, and inversely to FoxP3 mRNA levels, but no evidence of differential expression for any of these miRNA-mRNA pairs was observed. Finally, miR-9-5p

  18. The cytolethal distending toxin from the chancroid bacterium Haemophilus ducreyi induces cell-cycle arrest in the G2 phase.

    PubMed

    Cortes-Bratti, X; Chaves-Olarte, E; Lagergård, T; Thelestam, M

    1999-01-01

    The potent cytolethal distending toxin produced by Haemophilus ducreyi is a putative virulence factor in the pathogenesis of chancroid. We studied its action on eukaryotic cells, with the long-term goal of understanding the pathophysiology of the disease. Intoxication of cultured human epithelial-like cells, human keratinocytes, and hamster fibroblasts was irreversible, and appeared as a gradual distention of three- to fivefold the size of control cells. Organized actin assemblies appeared concomitantly with cell enlargement, promoted by a mechanism that probably does not involve small GTPases of the Rho protein family. Intoxicated cells did not proliferate. Similar to cells treated with other cytolethal distending toxins, these cells accumulated in the G2 phase of the cell cycle, demonstrating an increased level of the tyrosine phosphorylated (inactive) form of the cyclin-dependent kinase p34(cdc2). DNA synthesis was not affected until several hours after this increase, suggesting that the toxin acts directly on some kinase/phosphatase in the signaling network controlling the p34(cdc2) activity. We propose that this toxin has an important role both in the generation of chancroid ulcers and in their slow healing. The toxin may also be an interesting new tool for molecular studies of the eukaryotic cell- cycle machinery.

  19. The cytolethal distending toxin from the chancroid bacterium Haemophilus ducreyi induces cell-cycle arrest in the G2 phase

    PubMed Central

    Cortes-Bratti, Ximena; Chaves-Olarte, Esteban; Lagergård, Teresa; Thelestam, Monica

    1999-01-01

    The potent cytolethal distending toxin produced by Haemophilus ducreyi is a putative virulence factor in the pathogenesis of chancroid. We studied its action on eukaryotic cells, with the long-term goal of understanding the pathophysiology of the disease. Intoxication of cultured human epithelial-like cells, human keratinocytes, and hamster fibroblasts was irreversible, and appeared as a gradual distention of three- to fivefold the size of control cells. Organized actin assemblies appeared concomitantly with cell enlargement, promoted by a mechanism that probably does not involve small GTPases of the Rho protein family. Intoxicated cells did not proliferate. Similar to cells treated with other cytolethal distending toxins, these cells accumulated in the G2 phase of the cell cycle, demonstrating an increased level of the tyrosine phosphorylated (inactive) form of the cyclin-dependent kinase p34cdc2. DNA synthesis was not affected until several hours after this increase, suggesting that the toxin acts directly on some kinase/phosphatase in the signaling network controlling the p34cdc2 activity. We propose that this toxin has an important role both in the generation of chancroid ulcers and in their slow healing. The toxin may also be an interesting new tool for molecular studies of the eukaryotic cell- cycle machinery. PMID:9884340

  20. Ca2+ signaling and emesis: Recent progress and new perspectives.

    PubMed

    Zhong, Weixia; Picca, Andrew J; Lee, Albert S; Darmani, Nissar A

    2017-01-01

    Cisplatin-like chemotherapeutics cause vomiting via calcium (Ca 2+ )-dependent release of multiple neurotransmitters (dopamine, serotonin, substance P, etc.) from the gastrointestinal enterochromaffin cells and/or the brainstem. Intracellular Ca 2+ signaling is triggered by activation of diverse emetic receptors (including tachykininergic NK 1 , serotonergic 5-HT 3 , dopaminergic D 2 , cholinergic M 1 , or histaminergic H 1 ) , whose activation in vomit-competent species can evoke emesis. Other emetogens such as cisplatin, rotavirus NSP4 protein and bacterial toxins can also induce intracellular Ca 2+ elevation. Netupitant is a highly selective neurokinin NK 1 receptor (NK 1 R) antagonist and palonosetron is a selective second-generation serotonin 5-HT 3 receptor (5-HT 3 R) antagonist with a distinct pharmacological profile. An oral fixed combination of netupitant/palonosetron (NEPA; Akynzeo(®)) with >85% antiemetic efficacy is available for use in the prevention of acute and delayed chemotherapy-induced nausea and vomiting (CINV). Cannabinoid CB 1 receptor agonists possess broad-spectrum antiemetic activity since they prevent vomiting caused by a variety of emetic stimuli including the chemotherapeutic agent cisplatin, 5-HT 3 R agonists, and D 2 R agonists. Our findings demonstrate that application of the L-type Ca 2+ channel (LTCC) agonist FPL 64176 and the intracellular Ca 2+ mobilizing agent thapsigargin (a sarco/endoplasmic reticulum Ca 2+ -ATPase inhibitor) cause vomiting in the least shrew. On the other hand, blockade of LTCCs by corresponding antagonists (nifedipine or amlodipine) not only provide broad-spectrum antiemetic efficacy against diverse agents that specifically activate emetogenic receptors such as 5-HT 3 , NK 1 , D 2 , and M 1 receptors, but can also potentiate the antiemetic efficacy of palonosetron against the non-specific emetogen, cisplatin. In this review, we will provide an overview of Ca 2+ involvement in the emetic process; discuss the

  1. Enhanced performance of P3HT/(PCBM:ZnO:TiO{sub 2}) blend based hybrid organic solar cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Ikram, M., E-mail: mianraj.1981@gmail.com; Murray, R.; Imran, M.

    Highlights: • We fabricated hybrid bulk heterojunction organic solar cells. • TiO{sub 2} and ZnO nanoparticles replace PCBM with fixed amount of P3HT in active layer • PCE was significantly improved by the introduction of TiO{sub 2} and ZnO. • A possible route toward low-cost OPV. • To the best of my knowledge, this work is the first time going to report. - Abstract: Quaternary blend hybrid organic solar cells enjoy both an increased light absorption range and an easy method to fabricate because of the simple structure. In this study effects of mixing inorganic metal oxides (ZnO and TiO{submore » 2}) nanoparticles to the active layer of organic photovoltaics devices were investigated. The active layer primarily consists of various ratios of electron donor poly (3-hexylthiophene) (P3HT) and an electron acceptor [6,6]-phenyl-C61-butyric acid methyl ester (PCBM) together with nanostructured ZnO and TiO{sub 2} dispersed in chlorobenzene (CB) and 1,2-dichlorobenzene (DCB). The ratio of PCBM to nanoparticles was varied keeping the ratio of P3HT to acceptor material constant. Mixing of nanoparticle plays a significant role in the resulting power conversion efficiency (PCE) of the devices. An increased PCE for ZnO/TiO{sub 2} doped devices can be attributed to increased absorption in the visible region and enhanced charge collection due to the percolation networks formed by metal oxides nanoparticles.« less

  2. T2-Weighted Liver MRI Using the MultiVane Technique at 3T: Comparison with Conventional T2-Weighted MRI

    PubMed Central

    Kang, Kyung A; Kim, EunJu; Jeong, Woo Kyoung; Choi, Dongil; Lee, Won Jae; Jung, Sin-Ho; Baek, Sun-Young

    2015-01-01

    Objective To assess the value of applying MultiVane to liver T2-weighted imaging (T2WI) compared with conventional T2WIs with emphasis on detection of focal liver lesions. Materials and Methods Seventy-eight patients (43 men and 35 women) with 86 hepatic lesions and 20 pancreatico-biliary diseases underwent MRI including T2WIs acquired using breath-hold (BH), respiratory-triggered (RT), and MultiVane technique at 3T. Two reviewers evaluated each T2WI with respect to artefacts, organ sharpness, and conspicuity of intrahepatic vessels, hilar duct, and main lesion using five-point scales, and made pairwise comparisons between T2WI sequences for these categories. Diagnostic accuracy (Az) and sensitivity for hepatic lesion detection were evaluated using alternative free-response receiver operating characteristic analysis. Results MultiVane T2WI was significantly better than BH-T2WI or RT-T2WI for organ sharpness and conspicuity of intrahepatic vessels and main lesion in both separate reviews and pairwise comparisons (p < 0.001). With regard to motion artefacts, MultiVane T2WI or BH-T2WI was better than RT-T2WI (p < 0.001). Conspicuity of hilar duct was better with BH-T2WI than with MultiVane T2WI (p = 0.030) or RT-T2WI (p < 0.001). For detection of 86 hepatic lesions, sensitivity (mean, 97.7%) of MultiVane T2WI was significantly higher than that of BH-T2WI (mean, 89.5%) (p = 0.008) or RT-T2WI (mean, 84.9%) (p = 0.001). Conclusion Applying the MultiVane technique to T2WI of the liver is a promising approach to improving image quality that results in increased detection of focal liver lesions compared with conventional T2WI. PMID:26357498

  3. Botulinum toxin as a therapeutic agent to prevent relapse in deep bite patients.

    PubMed

    Mücke, Thomas; Löffel, Anja; Kanatas, Anastasios; Karnezi, Sandy; Rana, Majeed; Fichter, Andreas; Haarmann, Stephan; Wolff, Klaus-Dietrich; Loeffelbein, Denys John

    2016-05-01

    The etiology of deep bite is multifactorial. One of the causes is increased muscular activity. This makes the treatment of deep bite malocclusions difficult and often results in relapse in many cases. In this work we compared patients with surgical orthognathic treatment only and surgical orthognathic treatment with additional injections of botulinum toxin after mandibular advancement for class II division 2 malocclusion. This is a prospective study. Adult patients were assessed pretreatment (T1), posttreatment (T2), and long-term after 1 year (T3). In total, 32 patients (mean age, 30.7 years; 23 women and 9 men) reached the study end point (T3); 24 patients were treated without botulinum toxin and 8 patients received preoperative injections of botulinum toxin. Significant differences between both groups were observed, with a more stable result for the experimental group treated with botulinum toxin. In a selective group of adult patients with a class II division II incisor relationship and with a class II skeletal base, botulinum toxin injections can effectively prevent relapse. This may present an alternative to a conventional myotomy. Copyright © 2016 European Association for Cranio-Maxillo-Facial Surgery. Published by Elsevier Ltd. All rights reserved.

  4. Ocular Hypotensive Response in Nonhuman Primates of (8R)-1-[(2S)-2-Aminopropyl]-8,9-dihydro-7H-pyrano[2,3-g]indazol-8-ol a Selective 5-HT2 Receptor Agonist.

    PubMed

    May, Jesse A; Sharif, Najam A; McLaughlin, Marsha A; Chen, Hwang-Hsing; Severns, Bryon S; Kelly, Curtis R; Holt, William F; Young, Richard; Glennon, Richard A; Hellberg, Mark R; Dean, Thomas R

    2015-11-25

    Recently, it has been reported that 5-HT2 receptor agonists effectively reduce intraocular pressure (IOP) in a nonhuman primate model of glaucoma. Although 1-[(2S)-2-aminopropyl]indazol-6-ol (AL-34662) was shown to have good efficacy in this nonhuman primate model of ocular hypertension as well as a desirable physicochemical and permeability profile, subsequently identified cardiovascular side effects in multiple species precluded further clinical evaluation of this compound. Herein, we report selected structural modifications that resulted in the identification of (8R)-1-[(2S)-2-aminopropyl]-8,9-dihydro-7H-pyrano[2,3-g]indazol-8-ol (13), which displayed an acceptable profile to support advancement for further preclinical evaluation as a candidate for proof-of-concept studies in humans.

  5. The 5-HT2C receptor gene Cys23Ser polymorphism influences the intravaginal ejaculation latency time in Dutch Caucasian men with lifelong premature ejaculation

    PubMed Central

    Janssen, Paddy KC; van Schaik, Ron; Olivier, Berend; Waldinger, Marcel D

    2014-01-01

    It has been postulated that the persistent short intravaginal ejaculation latency time (IELT) of men with lifelong premature ejaculation (LPE) is related to 5-hydroxytryptamine (HT)2C receptor functioning. The aim of this study was to investigate the relationship of Cys23Ser 5-HT2C receptor gene polymorphism and the duration of IELT in men with LPE. Therefore, a prospective study was conducted in 64 Dutch Caucasian men with LPE. Baseline IELT during coitus was assessed by stopwatch over a 1-month period. All men were genotyped for Cys23Ser 5-HT2C receptor gene polymorphism. Allele frequencies and genotypes of Cys and Ser variants of 5-HT2C receptor gene polymorphism were determined. Association between Cys/Cys and Ser/Ser genotypes and the natural logarithm of the IELT in men with LPE were investigated. As a result, the geometric mean, median and natural mean IELT were 25.2, 27.0, 33.9 s, respectively. Of all men, 20.0%, 10.8%, 23.1% and 41.5% ejaculated within 10, 10–20, 20–30 and 30–60 s after vaginal penetration. Of the 64 men, the Cys/Cys and Ser/Ser genotype frequency for the Cys23Ser polymorphism of the 5-HT2C receptor gene was 81% and 19%, respectively. The geometric mean IELT of the wildtypes (Cys/Cys) is significantly lower (22.6 s; 95% CI 18.3–27.8 s) than in male homozygous mutants (Ser/Ser) (40.4 s; 95% CI 20.3–80.4 s) (P = 0.03). It is concluded that Cys23Ser 5-HT2C receptor gene polymorphism is associated with the IELT in men with LPE. Men with Cys/Cys genotype have shorter IELTs than men with Ser/Ser genotypes. PMID:24799636

  6. The 5-HT2C receptor gene Cys23Ser polymorphism influences the intravaginal ejaculation latency time in Dutch Caucasian men with lifelong premature ejaculation.

    PubMed

    Janssen, Paddy Kc; Schaik, Ron van; Olivier, Berend; Waldinger, Marcel D

    2014-01-01

    It has been postulated that the persistent short intravaginal ejaculation latency time (IELT) of men with lifelong premature ejaculation (LPE) is related to 5-hydroxytryptamine (HT)2C receptor functioning. The aim of this study was to investigate the relationship of Cys23Ser 5-HT2C receptor gene polymorphism and the duration of IELT in men with LPE. Therefore, a prospective study was conducted in 64 Dutch Caucasian men with LPE. Baseline IELT during coitus was assessed by stopwatch over a 1-month period. All men were genotyped for Cys23Ser 5-HT2C receptor gene polymorphism. Allele frequencies and genotypes of Cys and Ser variants of 5-HT2C receptor gene polymorphism were determined. Association between Cys/Cys and Ser/Ser genotypes and the natural logarithm of the IELT in men with LPE were investigated. As a result, the geometric mean, median and natural mean IELT were 25.2, 27.0, 33.9 s, respectively. Of all men, 20.0%, 10.8%, 23.1% and 41.5% ejaculated within 10, 10-20, 20-30 and 30-60 s after vaginal penetration. Of the 64 men, the Cys/Cys and Ser/Ser genotype frequency for the Cys23Ser polymorphism of the 5-HT2C receptor gene was 81% and 19%, respectively. The geometric mean IELT of the wildtypes (Cys/Cys) is significantly lower (22.6 s; 95% CI 18.3-27.8 s) than in male homozygous mutants (Ser/Ser) (40.4 s; 95% CI 20.3-80.4 s) (P = 0.03). It is concluded that Cys23Ser 5-HT2C receptor gene polymorphism is associated with the IELT in men with LPE. Men with Cys/Cys genotype have shorter IELTs than men with Ser/Ser genotypes.

  7. Effects of 7-day repeated treatment with the 5-HT2A inverse agonist/antagonist pimavanserin on methamphetamine vs. food choice in male rhesus monkeys.

    PubMed

    Banks, Matthew L

    2016-08-01

    Preclinical drug vs. food choice is an emerging group of drug self-administration procedures that have shown predictive validity to clinical drug addiction. Emerging data suggest that serotonin (5-HT)2A receptors modulate mesolimbic dopamine function, such that 5-HT2A antagonists blunt the abuse-related neurochemical effects of monoamine transporter substrates, such as amphetamine or methamphetamine. Whether subchronic 5-HT2A antagonist treatment attenuates methamphetamine reinforcement in any preclinical drug self-administration procedure is unknown. The study aim was therefore to determine 7-day treatment effects with the 5-HT2A inverse agonist/antagonist pimavanserin on methamphetamine vs. food choice in monkeys. Behavior was maintained under a concurrent schedule of food delivery (1g pellets, fixed-ratio 100 schedule) and intravenous methamphetamine injections (0-0.32 mg/kg/injection, fixed-ratio 10 schedule) in male rhesus monkeys (n=3). Methamphetamine choice dose-effect functions were determined daily before and during 7-day repeated pimavanserin (1.0-10mg/kg/day, intramuscular) treatment periods. Under control conditions, increasing methamphetamine doses resulted in a corresponding increase in methamphetamine vs. food choice. Repeated pimavanserin administration failed to attenuate methamphetamine choice and produce a reciprocal increase in food choice in any monkey up to doses (3.2-10mg/kg) that suppressed rates of operant responding primarily during components where behavior was maintained by food pellets. Repeated 5-HT2A receptor inverse agonist/antagonist treatment did not attenuate methamphetamine reinforcement under a concurrent schedule of intravenous methamphetamine and food presentation in nonhuman primates. Overall, these results do not support the therapeutic potential of 5-HT2A inverse agonists/antagonists as candidate medications for methamphetamine addiction. Copyright © 2016 The Author(s). Published by Elsevier Ireland Ltd.. All rights

  8. Abrogated Freud-1/CC2D1A repression of 5-HT1A autoreceptors induces fluoxetine-resistant anxiety/depression-like behavior

    PubMed Central

    Vahid-Ansari, Faranak; Daigle, Mireille; Manzini, M. Chiara; Tanaka, Kenji F.; Hen, René; Geddes, Sean D.; Béïque, Jean-Claude; James, Jonathan; Merali, Zul; Albert, Paul R.

    2017-01-01

    Freud-1/CC2D1A represses the gene transcription of serotonin-1A (5-HT1A) autoreceptors, which negatively regulate 5-HT tone. To test the role of Freud-1 in vivo, we generated mice with adulthood conditional knockout of Freud-1 in 5-HT neurons (cF1ko). In cF1ko mice, 5-HT1A autoreceptor protein, binding and hypothermia response were increased, with reduced 5-HT content and neuronal activity in the dorsal raphe. The cF1ko mice displayed increased anxiety- and depression-like behavior that was resistant to chronic antidepressant (fluoxetine) treatment. Using conditional Freud-1/5-HT1A double knockout (cF1/1A dko) to disrupt both Freud-1 and 5-HT1A genes in 5-HT neurons, no increase in anxiety- or depression-like behaviour was seen upon knockout of Freud-1 on the 5-HT1A autoreceptor-negative background, rather a reduction in depression-like behaviour emerged. These studies implicate transcriptional dys-regulation of 5-HT1A autoreceptors by the repressor Freud-1 in anxiety and depression and provide a clinically relevant genetic model of antidepressant resistance. Targeting specific transcription factors like Freud-1 to restore transcriptional balance may augment response to antidepressant treatment. PMID:29101244

  9. MDMA-induced loss of parvalbumin interneurons within the dentate gyrus is mediated by 5HT2A and NMDA receptors

    PubMed Central

    Collins, Stuart A.; Gudelsky, Gary A.; Yamamoto, Bryan K.

    2015-01-01

    MDMA is a widely abused psychostimulant which causes a rapid and robust release of the monoaminergic neurotransmitters dopamine and serotonin. Recently, it was shown that MDMA increases extracellular glutamate concentrations in the dorsal hippocampus, which is dependent on serotonin release and 5HT2A/2C receptor activation. The increased extracellular glutamate concentration coincides with a loss of parvalbumin-immunoreactive (PV-IR) interneurons of the dentate gyrus region. Given the known susceptibility of PV interneurons to excitotoxicity, we examined whether MDMA-induced increases in extracellular glutamate in the dentate gyrus are necessary for the loss of PV cells in rats. Extracellular glutamate concentrations increased in the dentate gyrus during systemic and local administration of MDMA. Administration of the NMDA receptor antagonist, MK-801, during systemic injections of MDMA, prevented the loss of PV-IR interneurons seen 10 days after MDMA exposure. Local administration of MDL100907, a selective 5HT2A receptor antagonist, prevented the increases in glutamate caused by reverse dialysis of MDMA directly into the dentate gyrus and prevented the reduction of PV-IR. These findings provide evidence that MDMA causes decreases in PV within the dentate gyrus through a 5HT2A receptor-mediated increase in glutamate and subsequent NMDA receptor activation. PMID:25936514

  10. Synthesis and characterization of two layered aluminophosphates, ( T) 2HAl 2P 3O 12 ( T=2-BuNH 3+) and ( T)H 2Al 2P 3O 12 ( T=pyH +)

    NASA Astrophysics Data System (ADS)

    Chippindale, Ann M.; Powell, Anthony V.; Bull, Lucy M.; Jones, Richard H.; Cheetham, Anthony K.; Thomas, John M.; Xu, Ruren

    1992-01-01

    Two new aluminophosphates, ( T) 2HAl 2P 3O 12 ( T=2-BuNH 3+) ( I) and ( T)H 2Al 2P 3O 12 ( T=pyH +) ( II) with the same framework stoichiometry but different layer structures have been prepared under nonaqueous conditions and the structures determined by single-crystal X-ray diffraction. Compound ( I) crystallizes in the monoclinic space group P2 1/ c ( Z=4), with lattice parameters a=9.261(1) b=8.365(6), c=27.119(4) Å, β=91.50(1)δ, and V=2100.1 Å 3 ( R=0.072 and R w=0.090). The structure consists of Al-and P-centered tetrahedra linked to form layers. Protonated 2-butylamine molecules are located in the interlayer spaces and hydrogen bonded to the layers through NH 3+ groups. Weak hydrophobic van der Waals' interactions between alkyl groups of the 2-BuNH 3+ cations hold the layers together. Compound ( II) crystallizes in the triclinic space group P-1 ( Z=2), with a=8.574(2), b=8.631(3), c=10.371(2) Å, α=81.84(3), β=87.53(2), γ=69.07(2)δ, and V=709.49Å 3 ( R=0.039 and R w=0.052). The structure contains tetrahedrally coordinated P atoms and both tetrahedral and trigonal pyramidal Al atoms linked to form layers which are held together through hydrogen bonding, creating cavities in which pyH + cations reside.

  11. Nonstandard neutrino interactions at DUNE, T2HK and T2HKK

    DOE PAGES

    Liao, Jiajun; Marfatia, Danny; Whisnant, Kerry

    2017-01-17

    Here, we study the matter effect caused by nonstandard neutrino interactions (NSI) in the next generation long-baseline neutrino experiments, DUNE, T2HK and T2HKK. If multiple NSI parameters are nonzero, the potential of these experiments to detect CP violation, determine the mass hierarchy and constrain NSI is severely impaired by degeneracies between the NSI parameters and by the generalized mass hierarchy degeneracy. In particular, a cancellation between leading order terms in the appearance channels when ϵ eτ= cot θ 23ϵ eμ, strongly affects the sensitivities to these two NSI parameters at T2HK and T2HKK. We also study the dependence of themore » sensitivities on the true CP phase and the true mass hierarchy, and find that overall DUNE has the best sensitivity to the magnitude of the NSI parameters, while T2HKK has the best sensitivity to CP violation whether or not there are NSI. Furthermore, for T2HKK a smaller off-axis angle for the Korean detector is better overall. We find that due to the structure of the leading order terms in the appearance channel probabilities, the NSI sensitivities in a given experiment are similar for both mass hierarchies, modulo the phase change δ→δ + 180°.« less

  12. Nonstandard neutrino interactions at DUNE, T2HK and T2HKK

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Liao, Jiajun; Marfatia, Danny; Whisnant, Kerry

    Here, we study the matter effect caused by nonstandard neutrino interactions (NSI) in the next generation long-baseline neutrino experiments, DUNE, T2HK and T2HKK. If multiple NSI parameters are nonzero, the potential of these experiments to detect CP violation, determine the mass hierarchy and constrain NSI is severely impaired by degeneracies between the NSI parameters and by the generalized mass hierarchy degeneracy. In particular, a cancellation between leading order terms in the appearance channels when ϵ eτ= cot θ 23ϵ eμ, strongly affects the sensitivities to these two NSI parameters at T2HK and T2HKK. We also study the dependence of themore » sensitivities on the true CP phase and the true mass hierarchy, and find that overall DUNE has the best sensitivity to the magnitude of the NSI parameters, while T2HKK has the best sensitivity to CP violation whether or not there are NSI. Furthermore, for T2HKK a smaller off-axis angle for the Korean detector is better overall. We find that due to the structure of the leading order terms in the appearance channel probabilities, the NSI sensitivities in a given experiment are similar for both mass hierarchies, modulo the phase change δ→δ + 180°.« less

  13. Impacts of elevated CO2 on exogenous Bacillus thuringiensis toxins and transgene expression in transgenic rice under different levels of nitrogen.

    PubMed

    Jiang, Shoulin; Lu, Yongqing; Dai, Yang; Qian, Lei; Muhammad, Adnan Bodlah; Li, Teng; Wan, Guijun; Parajulee, Megha N; Chen, Fajun

    2017-11-07

    Recent studies have highlighted great challenges of transgene silencing for transgenic plants facing climate change. In order to understand the impacts of elevated CO 2 on exogenous Bacillus thuringiensis (Bt) toxins and transgene expression in transgenic rice under different levels of N-fertilizer supply, we investigated the biomass, exogenous Bt toxins, Bt-transgene expression and methylation status in Bt rice exposed to two levels of CO 2 concentrations and nitrogen (N) supply (1/8, 1/4, 1/2, 1 and 2 N). It is elucidated that the increased levels of global atmospheric CO 2 concentration will trigger up-regulation of Bt toxin expression in transgenic rice, especially with appropriate increase of N fertilizer supply, while, to some extent, the exogenous Bt-transgene expression is reduced at sub-N levels (1/4 and 1/2N), even though the total protein of plant tissues is reduced and the plant growth is restricted. The unpredictable and stochastic occurrence of transgene silencing and epigenetic alternations remains unresolved for most transgenic plants. It is expected that N fertilization supply may promote the expression of transgenic Bt toxin in transgenic Bt rice, particularly under elevated CO 2 .

  14. In vivo binding of /sup 125/I-LSD to serotonin 5-HT/sub 2/ receptors in mouse brain

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Hartig, P.R.; Scheffel, U., Frost, J.J.; Wagner, H.N. Jr.

    The binding of /sup 125/I-LSD (2-(/sup 125/I)-lysergic acid diethylamide) was studied in various mouse brain regions following intravenous injection of the radioligand. The high specific activity of /sup 125/I-LSD enabled the injection of low mass doses (14ng/kg), which are well below the threshold for induction of any known physiological effect of the probe. The highest levels of /sup 125/I-LSD binding were found in the frontal cortex, olfactory tubercles, extra-frontal cortex and striatum while the lowest level was found in the cerebellum. Binding was saturable in the frontal cortex but increased linearly in the cerebellum with increasing doses of /sup 125/I-LSD.more » Serotonergic compounds potently inhibited /sup 125/I-LSD binding in cortical regions, olfactory tubercles, and hypothalamus but had no effect in the cerebellum. Dopaminergic compounds caused partial inhibition of binding in the striatum while adrenergic compounds were inactive. From these studies the authors conclude that /sup 125/I-LSD labels serotonin 5-HT/sub 2/ receptor sites in cortical regions with no indication that other receptor sites are labeled. In the olfactory tubercles and hypothalamus, /sup 125/I-LSD labeling occurs predominantly or entirely at serotonic 5-HT/sub 2/ sites. In the striatum, /sup 125/I-LSD labels approximately equal proportions of serotonergic and dopaminergic sites. These data indicate that /sup 125/I-LSD labels serotonin receptors in vivo and suggests that appropriate derivatives of 2I-LSD may prove useful for tomographic imaging of serotonin 5-HT/sub 2/ receptors in the mammalian cortex.« less

  15. Enhanced scratching evoked by PAR-2 agonist and 5-HT but not histamine in a mouse model of chronic dry skin itch.

    PubMed

    Akiyama, T; Carstens, M Iodi; Carstens, E

    2010-11-01

    Chronic itch is a symptom of many skin conditions and systemic disease, and it has been hypothesized that the chronic itch may result from sensitization of itch-signaling pathways. We induced experimental chronic dry skin on the rostral back of mice, and observed a significant increase in spontaneous hindlimb scratches directed to the dry skin. Spontaneous scratching was significantly attenuated by a PAR-2 antibody and 5-HT2A receptor antagonist, indicating activation of these receptors by endogenous mediators released under dry skin conditions. We also observed a significant increase in the number of scratch bouts evoked by acute intradermal injections of a protease-activated receptor (PAR)-2 agonist and serotonin (5-HT), but not histamine. We additionally investigated if pruritogen-evoked activity of dorsal root ganglion (DRG) neurons is enhanced in this model. DRG cells from dry skin mice exhibited significantly larger responses to the PAR-2 agonist and 5-HT, but not histamine. Spontaneous scratching may reflect ongoing itch, and enhanced pruritogen-evoked scratching may represent hyperknesis (enhanced itch), both potentially due to sensitization of itch-signaling neurons. The correspondence between enhanced behavioral scratching and DRG cell responses suggest that peripheral pruriceptors that respond to proteases and 5-HT, but not histamine, may be sensitized in dry skin itch. Copyright © 2010 International Association for the Study of Pain. Published by Elsevier B.V. All rights reserved.

  16. Efficacy of Genetically Modified Bt Toxins Alone and in Combinations Against Pink Bollworm Resistant to Cry1Ac and Cry2Ab

    PubMed Central

    Tabashnik, Bruce E.; Fabrick, Jeffrey A.; Unnithan, Gopalan C.; Yelich, Alex J.; Masson, Luke; Zhang, Jie; Bravo, Alejandra; Soberón, Mario

    2013-01-01

    Evolution of resistance in pests threatens the long-term efficacy of insecticidal proteins from Bacillus thuringiensis (Bt) used in sprays and transgenic crops. Previous work showed that genetically modified Bt toxins Cry1AbMod and Cry1AcMod effectively countered resistance to native Bt toxins Cry1Ab and Cry1Ac in some pests, including pink bollworm (Pectinophora gossypiella). Here we report that Cry1AbMod and Cry1AcMod were also effective against a laboratory-selected strain of pink bollworm resistant to Cry2Ab as well as to Cry1Ab and Cry1Ac. Resistance ratios based on the concentration of toxin killing 50% of larvae for the resistant strain relative to a susceptible strain were 210 for Cry2Ab, 270 for Cry1Ab, and 310 for Cry1Ac, but only 1.6 for Cry1AbMod and 2.1 for Cry1AcMod. To evaluate the interactions among toxins, we tested combinations of Cry1AbMod, Cry1Ac, and Cry2Ab. For both the resistant and susceptible strains, the net results across all concentrations tested showed slight but significant synergism between Cry1AbMod and Cry2Ab, whereas the other combinations of toxins did not show consistent synergism or antagonism. The results suggest that the modified toxins might be useful for controlling populations of pink bollworm resistant to Cry1Ac, Cry2Ab, or both. PMID:24244692

  17. Evaluation in Monkey of Two Candidate PET Radioligands, [11C]RX-1 and [18F]RX-2, for Imaging Brain 5-HT4 Receptors

    PubMed Central

    LOHITH, TALAKAD G.; XU, RONG; TSUJIKAWA, TETSUYA; MORSE, CHERYL L.; ANDERSON, KACEY B.; GLADDING, ROBERT L.; ZOGHBI, SAMI S.; FUJITA, MASAHIRO; INNIS, ROBERT B.; PIKE, VICTOR W.

    2014-01-01

    The serotonin subtype-4 (5-HT4) receptor, which is known to be involved physiologically in learning and memory, and pathologically in Alzheimer’s disease, anxiety and other neuropsychiatric disorders – has few radioligands readily available for imaging in vivo. We have previously reported two novel 5-HT4 receptor radioligands, namely [methoxy-11C](1-butylpiperidin-4-yl)methyl 4-amino-3-methoxybenzoate; [11C]RX-1) and the [18F]3-fluoromethoxy analog ([18F]RX-2), and in this study we evaluated them by PET in rhesus monkey. Brain scans were performed at baseline, receptor preblock or displacement conditions using SB 207710, a 5-HT4 receptor antagonist, on the same day for [11C]RX-1 and on different days for [18F]RX-2. Specific-to-nondisplaceable ratio (BPND) was measured with the simplified reference tissue model from all baseline scans. To determine specific binding, total distribution volume (VT) was also measured in some monkeys by radiometabolite-corrected arterial input function after ex vivo inhibition of esterases from baseline and blocked scans. Both radioligands showed moderate to high peak brain uptake of radioactivity (2–6 SUV). Regional BPND values were in the rank order of known 5-HT4 receptor distribution with a trend for higher BPND values from [18F]RX-2. One-tissue compartmental model provided good fits with well identified VT values for both radioligands. In the highest 5-HT4 receptor density region, striatum, 50–60% of total binding was specific. The VT in receptor-poor cerebellum reached stable values by about 60 min for both radioligands indicating little influence of radiometabolites on brain signal. In conclusion, both [11C]RX-1 and [18F]RX-2 showed positive attributes for PET imaging of brain 5-HT4 receptors, validating the radioligand design strategy. PMID:25088028

  18. AtaT blocks translation initiation by N-acetylation of the initiator tRNAfMet.

    PubMed

    Jurėnas, Dukas; Chatterjee, Sneha; Konijnenberg, Albert; Sobott, Frank; Droogmans, Louis; Garcia-Pino, Abel; Van Melderen, Laurence

    2017-06-01

    Toxin-antitoxin (TA) loci are prevalent in bacterial genomes. They are suggested to play a central role in dormancy and persister states. Under normal growth conditions, TA toxins are neutralized by their cognate antitoxins, and under stress conditions, toxins are freed and inhibit essential cellular processes using a variety of mechanisms. Here we characterize ataR-ataT, a novel TA system, from enterohemorrhagic Escherichia coli. We show that the toxin AtaT is a GNAT family enzyme that transfers an acetyl group from acetyl coenzyme A to the amine group of the methionyl aminoacyl moiety of initiator tRNA. AtaT specifically modifies Met-tRNA fMet , but no other aminoacyl-tRNAs, including the elongator Met-tRNA Met . We demonstrate that once acetylated, AcMet-tRNA fMet fails to interact with initiation factor-2 (IF2), resulting in disruption of the translation initiation complex. This work reveals a new mechanism of translation inhibition and confirms Met-tRNA fMet as a prime target to efficiently block cell growth.

  19. Ethanol induced adaptations in 5-HT2c receptor signaling in the bed nucleus of stria terminalis: Implications for anxiety during ethanol withdrawal

    PubMed Central

    Marcinkiewcz, Catherine A.; Dorrier, Cayce E.; Lopez, Alberto J.; Kash, Thomas L.

    2015-01-01

    One of the hallmarks of alcohol dependence is the presence of a withdrawal syndrome during abstinence, which manifests as physical craving for alcohol accompanied by subjective feelings of anxiety. Using a model of chronic intermittent ethanol (CIE) vapor in mice, we investigated the role of serotonin2c signaling in the BNST as a neural substrate underlying ethanol-induced anxiety during withdrawal. Mice were subjected to a 5-day CIE regimen of 16 hours of ethanol vapor exposure followed by an 8 hour “withdrawal” period between exposures. After the 5th and final exposure, mice were withdrawn for 24 hours or 1 week before experiments began. Anxiety-like behavior was assessed in the social approach, light dark, and open field test with mice showing deficits in social, but not general anxiety-like behavior that was alleviated by pretreatment with the 5HT2c-R antagonist SB 242,084 (3 mg/kg, i.p.) 24 hours and 1 week post-CIE. Using immunohistochemistry and whole cell patch clamp electrophysiology, we also found that CIE increased FOS-IR and enhanced neuronal excitability in the ventral BNST (vBNST) 24 hrs into withdrawal in a 5HT2c-R dependent manner. This enhanced excitability persisted for 1 week post-CIE. We also found that mCPP, a 5HT2c/b agonist, induced a more robust depolarization in cells of the vBNST in CIE mice, confirming that 5HT2c-R signaling is upregulated in the vBNST following CIE. Taken together, these results suggest that CIE upregulates 5HT2c-R signaling in the vBNST, leading to increased excitability. This enhanced excitability of the vBNST may drive increased anxiety-like behavior during ethanol withdrawal. PMID:25229718

  20. Reinforced spatial alternation as an animal model of obsessive-compulsive disorder (OCD): investigation of 5-HT2C and 5-HT1D receptor involvement in OCD pathophysiology.

    PubMed

    Tsaltas, Eleftheria; Kontis, Dimitris; Chrysikakou, Sofia; Giannou, Haralambos; Biba, Angeliki; Pallidi, Stella; Christodoulou, Angeliki; Maillis, Antonis; Rabavilas, Andreas

    2005-05-15

    This study introduces a laboratory model of compulsive behavior based on persistence in the context of rewarded spatial alternation. Rats were screened for spontaneous persistence during T-maze reinforced alternation. Experiment 1: One high and one low spontaneous persistence group (n = 8) received 20 injections of fluoxetine, a matched pair saline, both followed by 4 days of meta-chlorophenylpiperazine (mCPP) challenge. Experiment 2: Five matched groups of rats (n = 9) received pretreatment (20 injections) with fluoxetine, mCPP, desipramine, diazepam or saline, followed by 4 days of mCPP challenge (fluoxetine in mCPP group). After washout, animals received 2 days of naratriptan, followed by another 2-day mCPP challenge. In both experiments mCPP significantly increased persistence in saline controls. Fluoxetine also acutely increased persistence scores: after a gradual return to baseline, these scores showed tolerance to mCPP. Experiment 1: This pattern was significant in high but not low initial persistence groups. Experiment 2: Fluoxetine and mCPP showed cross-tolerance. Neither desipramine nor diazepam protected against mCPP challenge. Persistence scores returned to baseline during washout and naratriptan and were thereafter increased by another mCPP challenge in all but the fluoxetine and mCPP groups, suggesting 5-HT2C receptor mediation. This model is based on spontaneous persistence behavior showing pharmacological responses concordant with those of compulsive symptomatology.

  1. Expression and mutagenesis of the sea anemone toxin Av2 reveals key amino acid residues important for activity on voltage-gated sodium channels.

    PubMed

    Moran, Yehu; Cohen, Lior; Kahn, Roy; Karbat, Izhar; Gordon, Dalia; Gurevitz, Michael

    2006-07-25

    Type I sea anemone toxins are highly potent modulators of voltage-gated Na-channels (Na(v)s) and compete with the structurally dissimilar scorpion alpha-toxins on binding to receptor site-3. Although these features provide two structurally different probes for studying receptor site-3 and channel fast inactivation, the bioactive surface of sea anemone toxins has not been fully resolved. We established an efficient expression system for Av2 (known as ATX II), a highly insecticidal sea anemone toxin from Anemonia viridis (previously named A. sulcata), and mutagenized it throughout. Each toxin mutant was analyzed in toxicity and binding assays as well as by circular dichroism spectroscopy to discern the effects derived from structural perturbation from those related to bioactivity. Six residues were found to constitute the anti-insect bioactive surface of Av2 (Val-2, Leu-5, Asn-16, Leu-18, and Ile-41). Further analysis of nine Av2 mutants on the human heart channel Na(v)1.5 expressed in Xenopus oocytes indicated that the bioactive surfaces toward insects and mammals practically coincide but differ from the bioactive surface of a structurally similar sea anemone toxin, Anthopleurin B, from Anthopleura xanthogrammica. Hence, our results not only demonstrate clear differences in the bioactive surfaces of Av2 and scorpion alpha-toxins but also indicate that despite the general conservation in structure and importance of the Arg-14 loop and its flanking residues Gly-10 and Gly-20 for function, the surface of interaction between different sea anemone toxins and Na(v)s varies.

  2. Gaining insight into the T _2^*-T2 relationship in surface NMR free-induction decay measurements

    NASA Astrophysics Data System (ADS)

    Grombacher, Denys; Auken, Esben

    2018-05-01

    One of the primary shortcomings of the surface nuclear magnetic resonance (NMR) free-induction decay (FID) measurement is the uncertainty surrounding which mechanism controls the signal's time dependence. Ideally, the FID-estimated relaxation time T_2^* that describes the signal's decay carries an intimate link to the geometry of the pore space. In this limit the parameter T_2^* is closely linked to a related parameter T2, which is more closely linked to pore-geometry. If T_2^* ˜eq {T_2} the FID can provide valuable insight into relative pore-size and can be used to make quantitative permeability estimates. However, given only FID measurements it is difficult to determine whether T_2^* is linked to pore geometry or whether it has been strongly influenced by background magnetic field inhomogeneity. If the link between an observed T_2^* and the underlying T2 could be further constrained the utility of the standard surface NMR FID measurement would be greatly improved. We hypothesize that an approach employing an updated surface NMR forward model that solves the full Bloch equations with appropriately weighted relaxation terms can be used to help constrain the T_2^*-T2 relationship. Weighting the relaxation terms requires estimating the poorly constrained parameters T2 and T1; to deal with this uncertainty we propose to conduct a parameter search involving multiple inversions that employ a suite of forward models each describing a distinct but plausible T_2^*-T2 relationship. We hypothesize that forward models given poor T2 estimates will produce poor data fits when using the complex-inversion, while forward models given reliable T2 estimates will produce satisfactory data fits. By examining the data fits produced by the suite of plausible forward models, the likely T_2^*-T2 can be constrained by identifying the range of T2 estimates that produce reliable data fits. Synthetic and field results are presented to investigate the feasibility of the proposed technique.

  3. 1,2,3,4-bis(p-methylbenzylidene sorbitol) accelerates crystallization and improves hole mobility of poly(3-hexylthiophene)

    NASA Astrophysics Data System (ADS)

    Yuan, Nana; Huo, Hong

    2016-02-01

    The addition of 1,2,3,4-bis(p-methylbenzylidene sorbitol) (MDBS) does not change the nucleation mechanism or the crystal form of poly(3-hexylthiophene) (P3HT), but its presence increases the crystallization temperature (T c) of P3HT, decreases the crystallization half-time (t 1/2) and accelerates P3HT crystallization, which indicates that MDBS is an effective nucleating agent for P3HT. An acceleration of P3HT crystallization by the addition of MDBS decreases the crystalline size and crystallinity of P3HT, and enhances the connectivity between ordered regions of P3HT, leading to the hole mobility rising from 1.99 × 10-6 to 7.57 × 10-5 cm2 V-1s-1 in P3HT:PCBM blend based hole-only devices with sandwich configurations. Our results suggest that accelerating P3HT crystallization by adding a nucleating agent might be an important factor to improve the hole mobility and balance the electron and hole mobility in a photovoltaic blend.

  4. Cryo-electron microscopy study of bacteriophage T4 displaying anthrax toxin proteins

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Fokine, Andrei; Bowman, Valorie D.; Battisti, Anthony J.

    2007-10-25

    The bacteriophage T4 capsid contains two accessory surface proteins, the small outer capsid protein (Soc, 870 copies) and the highly antigenic outer capsid protein (Hoc, 155 copies). As these are dispensable for capsid formation, they can be used for displaying proteins and macromolecular complexes on the T4 capsid surface. Anthrax toxin components were attached to the T4 capsid as a fusion protein of the N-terminal domain of the anthrax lethal factor (LFn) with Soc. The LFn-Soc fusion protein was complexed in vitro with Hoc{sup -}Soc{sup -}T4 phage. Subsequently, cleaved anthrax protective antigen heptamers (PA63){sub 7} were attached to the exposedmore » LFn domains. A cryo-electron microscopy study of the decorated T4 particles shows the complex of PA63 heptamers with LFn-Soc on the phage surface. Although the cryo-electron microscopy reconstruction is unable to differentiate on its own between different proposed models of the anthrax toxin, the density is consistent with a model that had predicted the orientation and position of three LFn molecules bound to one PA63 heptamer.« less

  5. The serotonin transporter (5-HTTLPR) but not serotonin receptor (5-HT2C Cys23Ser) variant is associated with bipolar I disorder in Kurdish population from Western Iran.

    PubMed

    Mohammadi, Sahar; Khazaie, Habibolah; Rahimi, Ziba; Vaisi-Raygani, Asad; Zargooshi, Newsha; Rahimi, Zohreh

    2015-03-17

    The role of 5-HTTLPR and 5-HT2C Cys23Ser polymorphisms in the psychopathology of mood disorders and suicide behavior is controversial. The aim of present study was to investigate the association between 5-HTTLPR and 5-HT2C Cys23Ser variants and susceptibility to bipolar I disorder (BID). The 5-HT2C genotypes were studied in 152 patients with BID and 173 gender- and age-matched healthy individuals with Kurds ethnic background from Western Iran using PCR and PCR-RFLP methods. In recessive model (SS vs. LL+LS) the SS genotype was associated with 1.79-fold increased risk of BID (p=0.018). Also, the presence of S allele increased the risk of adult-onset BID by 1.76-fold (p=0.027). No association was detected between 5-HTTLPR genotypes and alleles with suicide attempt. The frequency of 5-HT2C Ser allele in patients and controls were 12.3 and 12.5%, respectively. Mutant allele of HT2C Ser had higher frequency in female (14.7%) than male (10.5%, p=0.27) patients. The frequency of HT2C Ser allele in patients with a family history of BID tended to be higher (15.7%) than those without a family history of the disease (11.8%). The frequency of HT2C Ser allele in suicide attempter women was higher (16.7%) than those without a suicide attempt (14.3%). Our findings demonstrate 5-HTTLPR polymorphism might be a risk factor for BID and adult-onset BID in Kurds population. However, we found the lack of an association between 5-HT2C Cys/Ser variants and the risk of BID. Copyright © 2015 Elsevier Ireland Ltd. All rights reserved.

  6. T2L2/Jason-2, first year of processing activities

    NASA Astrophysics Data System (ADS)

    Exertier, P.

    2009-11-01

    The T2L2 (Time Transfer by Laser Link) project, developed by CNES and OCA will permit the synchronization of remote ultra stable clocks and the determination of their performances over intercontinental distances. The principle is derived from laser telemetry technology with a dedicated space equipment designed to record arrival time of laser pulses at the satellite. T2L2 was accepted in 2005 to be on board the Jason-2 altimetry satellite. It has been successfully launched from Vandenberg (CA, US) in June 2008. T2L2 acquired the first laser pulses a few days after the launch. First analysis permitted to validate some important characteristics of the instrument such as sensitivity, noise, dynamic, event timer precision and ground to space time stability.

  7. Toxicity induced by F. poae-contaminated feed and the protective effect of Montmorillonite supplementation in broilers.

    PubMed

    Yang, Lingchen; Zhao, Zhiyong; Deng, Yifeng; Zhou, Zhenlei; Hou, Jiafa

    2014-12-01

    The T-2 and HT-2 toxins, the main metabolites of Fusarium poae, induce toxicity in broilers and accumulate in tissues. Consequently, during the breeding process of broilers, diets are frequently supplemented with physical adsorbents to protect birds against the toxicity induced by mycotoxins. In the present research, T-2 and HT-2 were produced in maize inoculated with F. poae. Mont, the strongest adsorbent based on in vitro adsorption ratios, was added to the contaminated diet. One-day-old chickens were randomly and equally divided into the following four groups: control diet group, Mont supplemented diet group, contaminated diet group and detoxification diet group. The experiment lasted for 42 days. Compared to the control group, the contaminated group showed significant decrease in body weight, feed intake and TP (P < 0.05), and marked increase in FCR, ALP, AST and ALT activity, T-2/HT-2 residues in the tissues and the relative expressions of apoptosis-related mRNAs (P < 0.05). Mont supplementation provided protection for the treated broilers in terms of performance, blood biochemistry, hepatic function, T-2/HT-2 residue of tissues and apoptosis. Therefore, Mont may be suitable as a detoxification agent for T-2/HT-2 in feed for broilers. Copyright © 2014 Elsevier Ltd. All rights reserved.

  8. PKCδ Knockout Mice Are Protected from Dextromethorphan-Induced Serotonergic Behaviors in Mice: Involvements of Downregulation of 5-HT1A Receptor and Upregulation of Nrf2-Dependent GSH Synthesis.

    PubMed

    Tran, Hai-Quyen; Lee, Youngho; Shin, Eun-Joo; Jang, Choon-Gon; Jeong, Ji Hoon; Mouri, Akihiro; Saito, Kuniaki; Nabeshima, Toshitaka; Kim, Hyoung-Chun

    2018-02-22

    We investigated whether a specific serotonin (5-HT) receptor-mediated mechanism was involved in dextromethorphan (DM)-induced serotonergic behaviors. We firstly observed that the activation of 5-HT 1A receptor, but not 5-HT 2A receptor, contributed to DM-induced serotonergic behaviors in mice. We aimed to determine whether the upregulation of 5-HT 1A receptor induced by DM facilitates the specific induction of certain PKC isoform, because previous reports suggested that 5-HT 1A receptor activates protein kinase C (PKC). A high dose of DM (80 mg/kg, i.p.) induced a selective induction of PKCδ out of PKCα, PKCβI, PKCβII, PKCξ, and PKCδ in the hypothalamus of wild-type (WT) mice. More importantly, 5-HT 1A receptor co-immunoprecipitated PKCδ in the presence of DM. Consistently, rottlerin, a pharmacological inhibitor of PKCδ, or PKCδ knockout significantly protected against increases in 5-HT 1A receptor gene expression, 5-HT turnover rate, and serotonergic behaviors induced by DM. Treatment with DM resulted in an initial increase in nuclear factor erythroid-2-related factor 2 (Nrf2) nuclear translocation and DNA-binding activity, γ-glutamylcysteine (GCL) mRNA expression, and glutathione (GSH) level. This compensative induction was further potentiated by rottlerin or PKCδ knockout. However, GCL mRNA and GSH/GSSG levels were decreased 6 and 12 h post-DM. These decreases were attenuated by PKCδ inhibition. Our results suggest that interaction between 5-HT 1A receptor and PKCδ is critical for inducing DM-induced serotonergic behaviors and that inhibition of PKCδ attenuates the serotonergic behaviors via downregulation of 5-HT 1A receptor and upregulation of Nrf2-dependent GSH synthesis.

  9. Efficient hybrid solar cell with P3HT:PCBM and Cu2ZnSnS4 nanocrystals

    NASA Astrophysics Data System (ADS)

    Jang, Se-Jung; Thuy Ho, Nhu; Lee, Min Hyung; Kim, Yong Soo

    2017-06-01

    Recently, Cu2ZnSnS4 (CZTS) with band gap about 1.50 eV is predicted to become an ideal light absorption material due to the abundant component elements in the crust being nontoxic and environmentally friendly. However, CZTS solar cells made by high temperature and vacuum-processed are at a perceived cost disadvantage in compared with solution-processed systems such as organic and hybrid solar cells. In this study, we propose a hybrid solar configurations with solution-processed CZTS nanocrystals and P3HT:PCBM bulk heterojunction. The forming double heterojunction, as charge can be separated at both the P3HT:PCBM and CZTS:PCBM interface is attributed to enhance the light harvesting efficiency. As a result, organic solar cells with CZTS nanocrystals show the higher efficiency 3.32 % compare to 2.65 % of reference organic solar cells. A 25 % improvement of power conversion efficiency is obtained by the increasing in short-circuit current and fill factor.

  10. Molecular characterization of Calocedrus rupestris Aver., H.T. Nguyen & L.K. Phan, 2008 (Cupressaceae) based on ITS1 partial sequence.

    PubMed

    Long, P K; Trang, N T P; Averyanov, L V; Loc, P K

    2011-11-21

    Calocedrus rupestris Aver., H.T. Nguyen & L.K. Phan was described in 2008 based on some morphological characters that were not sufficiently significant to discriminate it as a species distinct from C. macrolepis Kurz. We applied a new approach to resolve these conflicting views by using sequence data from DNA (ITS) to elucidate phylogenetic relationships between the two species. Analyses of a partial ITS1 sequence in 5 individuals of 2 subpopulations of C. macrolepis and 18 individuals of 8 subpopulations of C. rupestris collected in Vietnam were done. Molecular characterization of the two species showed its low divergence with the lack of autapomorphic characters. In addition, the ITS1 partial sequences of some C. rupestris individuals were identical with C. macrolepis. Due to the less distinctive morphology between C. rupestris and C. macrolepis, the divergence between them does not exceed the interspecific levels, and therefore, C. rupestris could not be regarded as an independent species in relation to C. macrolepis but only as one of its varieties, C. macrolepis var. rupestris (Aver., H.T. Nguyen & L.K. Phan) L.K. Phan, Long K. Phan & Aver.

  11. A single-step purification and molecular characterization of functional Shiga toxin 2 variants from pathogenic Escherichia coli

    USDA-ARS?s Scientific Manuscript database

    Shiga toxin (Stx) 2 variants, Stx2a, Stx2c, Stx2d and Stx2g were purified to homogeneity from bacterial culture supernatants by a one-step monoclonal anti-Stx affinity chromatography method. The method was based on the binding affinity of these Stxs for a monoclonal antibody against the Stx2 A-subun...

  12. Preoperative detection of malignant liver tumors: Comparison of 3D-T2-weighted sequences with T2-weighted turbo spin-echo and single shot T2 at 1.5 T.

    PubMed

    Barat, Maxime; Soyer, Philippe; Dautry, Raphael; Pocard, Marc; Lo-Dico, Rea; Najah, Haythem; Eveno, Clarisse; Cassinotto, Christophe; Dohan, Anthony

    2018-03-01

    To assess the performances of three-dimensional (3D)-T2-weighted sequences compared to standard T2-weighted turbo spin echo (T2-TSE), T2-half-Fourier acquisition single-shot turbo spin-echo (T2-HASTE), diffusion weighted imaging (DWI) and 3D-T1-weighted VIBE sequences in the preoperative detection of malignant liver tumors. From 2012 to 2015, all patients of our institution undergoing magnetic resonance imaging (MRI) examination for suspected malignant liver tumors were prospectively included. Patients had contrast-enhanced 3D-T1-weighted, DWI, 3D-T2-SPACE, T2-HASTE and T2-TSE sequences. Imaging findings were compared with those obtained at follow-up, surgery and histopathological analysis. Sensitivities for the detection of malignant liver tumors were compared for each sequence using McNemar test. A subgroup analysis was conducted for HCCs. Image artifacts were analyzed and compared using Wilcoxon paired signed rank-test. Thirty-three patients were included: 13 patients had 40 hepatocellular carcinomas (HCC) and 20 had 54 liver metastases. 3D-T2-weighted sequences had a higher sensitivity than T2-weighted TSE sequences for the detection of malignant liver tumors (79.8% versus 68.1%; P < 0.001). The difference did not reach significance for HCC. T1-weighted VIBE and DWI had a higher sensitivity than T2-weighted sequences. 3D-T2-weighted-SPACE sequences showed significantly less artifacts than T2-weitghted TSE. 3D-T2-weighted sequences show very promising performances for the detection of liver malignant tumors compared to T2-weighted TSE sequences. Copyright © 2018 Elsevier B.V. All rights reserved.

  13. Effects of Escherichia coli subtilase cytotoxin and Shiga toxin 2 on primary cultures of human renal tubular epithelial cells.

    PubMed

    Márquez, Laura B; Velázquez, Natalia; Repetto, Horacio A; Paton, Adrienne W; Paton, James C; Ibarra, Cristina; Silberstein, Claudia

    2014-01-01

    Shiga toxin (Stx)-producing Escherichia coli (STEC) cause post-diarrhea Hemolytic Uremic Syndrome (HUS), which is the most common cause of acute renal failure in children in many parts of the world. Several non-O157 STEC strains also produce Subtilase cytotoxin (SubAB) that may contribute to HUS pathogenesis. The aim of the present work was to examine the cytotoxic effects of SubAB on primary cultures of human cortical renal tubular epithelial cells (HRTEC) and compare its effects with those produced by Shiga toxin type 2 (Stx2), in order to evaluate their contribution to renal injury in HUS. For this purpose, cell viability, proliferation rate, and apoptosis were assayed on HRTEC incubated with SubAB and/or Stx2 toxins. SubAB significantly reduced cell viability and cell proliferation rate, as well as stimulating cell apoptosis in HRTEC cultures in a time dependent manner. However, HRTEC cultures were significantly more sensitive to the cytotoxic effects of Stx2 than those produced by SubAB. No synergism was observed when HRTEC were co-incubated with both SubAB and Stx2. When HRTEC were incubated with the inactive SubAA272B toxin, results were similar to those in untreated control cells. Similar stimulation of apoptosis was observed in Vero cells incubated with SubAB or/and Stx2, compared to HRTEC. In conclusion, primary cultures of HRTEC are significantly sensitive to the cytotoxic effects of SubAB, although, in a lesser extent compared to Stx2.

  14. Promoter Sequence of Shiga Toxin 2 (Stx2) Is Recognized In Vivo, Leading to Production of Biologically Active Stx2

    PubMed Central

    Bentancor, Leticia V.; Mejías, Maria P.; Pinto, Alípio; Bilen, Marcos F.; Meiss, Roberto; Rodriguez-Galán, Maria C.; Baez, Natalia; Pedrotti, Luciano P.; Goldstein, Jorge; Ghiringhelli, Pablo D.; Palermo, Marina S.

    2013-01-01

    ABSTRACT Shiga toxins (Stx) are the main agent responsible for the development of hemolytic-uremic syndrome (HUS), the most severe and life-threatening systemic complication of infection with enterohemorrhagic Escherichia coli (EHEC) strains. We previously described Stx2 expression by eukaryotic cells after they were transfected in vitro with the stx2 gene cloned into a prokaryotic plasmid (pStx2). The aim of this study was to evaluate whether mammalian cells were also able to express Stx2 in vivo after pStx2 injection. Mice were inoculated by hydrodynamics-based transfection (HBT) with pStx2. We studied the survival, percentage of polymorphonuclear leukocytes in plasma, plasma urea levels, and histology of the kidneys and the brains of mice. Mice displayed a lethal dose-related response to pStx2. Stx2 mRNA was recovered from the liver, and Stx2 cytotoxic activity was observed in plasma of mice injected with pStx2. Stx2 was detected by immunofluorescence in the brains of mice inoculated with pStx2, and markers of central nervous system (CNS) damage were observed, including increased expression of glial fibrillary acidic protein (GFAP) and fragmentation of NeuN in neurons. Moreover, anti-Stx2B-immunized mice were protected against pStx2 inoculation. Our results show that Stx2 is expressed in vivo from the wild stx2 gene, reproducing pathogenic damage induced by purified Stx2 or secondary to EHEC infection. PMID:24085779

  15. MDMA-induced loss of parvalbumin interneurons within the dentate gyrus is mediated by 5HT2A and NMDA receptors.

    PubMed

    Collins, Stuart A; Gudelsky, Gary A; Yamamoto, Bryan K

    2015-08-15

    MDMA is a widely abused psychostimulant which causes a rapid and robust release of the monoaminergic neurotransmitters dopamine and serotonin. Recently, it was shown that MDMA increases extracellular glutamate concentrations in the dorsal hippocampus, which is dependent on serotonin release and 5HT2A/2C receptor activation. The increased extracellular glutamate concentration coincides with a loss of parvalbumin-immunoreactive (PV-IR) interneurons of the dentate gyrus region. Given the known susceptibility of PV interneurons to excitotoxicity, we examined whether MDMA-induced increases in extracellular glutamate in the dentate gyrus are necessary for the loss of PV cells in rats. Extracellular glutamate concentrations increased in the dentate gyrus during systemic and local administration of MDMA. Administration of the NMDA receptor antagonist, MK-801, during systemic injections of MDMA, prevented the loss of PV-IR interneurons seen 10 days after MDMA exposure. Local administration of MDL100907, a selective 5HT2A receptor antagonist, prevented the increases in glutamate caused by reverse dialysis of MDMA directly into the dentate gyrus and prevented the reduction of PV-IR. These findings provide evidence that MDMA causes decreases in PV within the dentate gyrus through a 5HT2A receptor-mediated increase in glutamate and subsequent NMDA receptor activation. Copyright © 2015 Elsevier B.V. All rights reserved.

  16. Regulation of Oligomeric Organization of the Serotonin 5-Hydroxytryptamine 2C (5-HT2C) Receptor Observed by Spatial Intensity Distribution Analysis*

    PubMed Central

    Ward, Richard J.; Pediani, John D.; Godin, Antoine G.; Milligan, Graeme

    2015-01-01

    The questions of whether G protein-coupled receptors exist as monomers, dimers, and/or oligomers and if these species interconvert in a ligand-dependent manner are among the most contentious current issues in biology. When employing spatial intensity distribution analysis to laser scanning confocal microscope images of cells stably expressing either a plasma membrane-associated form of monomeric enhanced green fluorescent protein (eGFP) or a tandem version of this fluorophore, the eGFP tandem was identified as a dimer. Similar studies on cells stably expressing an eGFP-tagged form of the epidermal growth factor receptor demonstrated that, although largely a monomer in the basal state, this receptor rapidly became predominantly dimeric upon the addition of its ligand epidermal growth factor. In cells induced to express an eGFP-tagged form of the serotonin 5-hydroxytryptamine 2C (5-HT2C) receptor, global analysis of construct quantal brightness was consistent with the predominant form of the receptor being dimeric. However, detailed spatial intensity distribution analysis demonstrated the presence of multiple forms ranging from monomers to higher-order oligomers. Furthermore, treatment with chemically distinct 5-HT2C receptor antagonists resulted in a time-dependent change in the quaternary organization to one in which there was a preponderance of receptor monomers. This antagonist-mediated effect was reversible, because washout of the ligand resulted in the regeneration of many of the oligomeric forms of the receptor. PMID:25825490

  17. Oleamide administered into the nucleus accumbens shell regulates feeding behaviour via CB1 and 5-HT2C receptors.

    PubMed

    Soria-Gómez, Edgar; Márquez-Diosdado, Marianela I; Montes-Rodríguez, Corinne J; Estrada-González, Vicente; Prospéro-García, Oscar

    2010-10-01

    The central nervous system control of food intake has been extensively studied, hence, several neurotransmitter systems regulating this function are now clearly identified, for example, the endocannabinoid and serotoninergic systems. The former stimulates feeding while the latter inhibits it. Oleamide (Ole) is a cannabimimetic molecule affecting both systems. In this work, we tested the orexigenic and anorectic potential of Ole when administered into the nucleus accumbens shell (NAcS), a brain region that has been related to the orexigenic effects of cannabinoids. Additionally, we tested if Ole administered into this nucleus affects the activity of the hypothalamic nuclei involved in feeding behaviour, just as other cannabinoids do. We found a hyperphagic effect of Ole that is mediated through CB1 activation. The combination of Ole and the CB1 antagonist, AM251, produced a hypophagia that was fully blocked by SB212084, a 5-HT2C receptor antagonist. We also show that blockade of 5-HT2C and 5-HT2A receptors in the NAcS stimulates food intake. Finally, the combination of Ole and AM251 activates hypothalamic nuclei, an effect also blocked by SB242084. In conclusion, we show, for the first time, that Ole administered into the NAcS has a dual effect on feeding behaviour, acting through cannabinoid and serotonin receptors. These effects probably result from a downstream interaction with the hypothalamus.

  18. Transglutaminase 2 overexpression induces depressive-like behavior and impaired TrkB signaling in mice

    PubMed Central

    Pandya, Chirayu D; Hoda, Nasrul; Crider, Amanda; Peter, Diya; Kutiyanawalla, Ammar; Kumar, Sanjiv; Ahmed, Anthony O; Turecki, Gustavo; Hernandez, Caterina M; Terry, Alvin V

    2016-01-01

    Serotonin (5-HT) and brain derived neurotrophic factor (BDNF) are two signaling molecules that play important regulatory roles in the development and plasticity of neural circuits that are known to be altered in depression. However, the mechanism by which 5-HT regulates BDNF signaling is unknown. In the present study, we found that 5-HT treatment increases BDNF receptor, TrkB (tropomyosin related kinase B) levels in mouse primary cortical neurons via a Rac1 (RAS-related C3 botulinum toxin substrate 1)-dependent mechanism. Significant increases in the levels of transglutaminase 2 (TG2, which is implicated in transamidation of 5-HT to Rac1) are observed in the mouse prefrontal cortex (PFC) following chronic exposure to stress. We also found that TG2 levels are increased in the postmortem PFC of depressed suicide subjects relative to matched controls. Moreover, in mice, neuronal overexpression of TG2 resulted in the atrophy of neurons and reduced levels of TrkB in the PFC as well as a depressive-like phenotype. Overexpression of TG2 in mouse cortical neurons reduced TrkB levels as a result of impaired endocytosis of TrkB. TG2 inhibition by either a viral particle or pharmacological approach attenuated behavioral deficits caused by chronic unpredictable stress. Moreover, the overexpression of TrkB in the mouse PFC ameliorated the depressive-like phenotype of TG2 overexpressed mice. Taken together, these postmortem and preclinical findings identify TG2 as a critical mediator of the altered TrkB expression and depressive-like behaviors associated with chronic exposure to stress and suggest that TG2 may represent a novel therapeutic target in depression. PMID:27620841

  19. Stochasticity and bifurcations in a reduced model with interlinked positive and negative feedback loops of CREB1 and CREB2 stimulated by 5-HT.

    PubMed

    Hao, Lijie; Yang, Zhuoqin; Bi, Yuanhong

    2016-04-01

    The cyclic AMP (cAMP)-response element-binding protein (CREB) family of transcription factors is crucial in regulating gene expression required for long-term memory (LTM) formation. Upon exposure of sensory neurons to the neurotransmitter serotonin (5-HT), CREB1 is activated via activation of the protein kinase A (PKA) intracellular signaling pathways, and CREB2 as a transcriptional repressor is relieved possibly via phosphorylation of CREB2 by mitogen-activated protein kinase (MAPK). Song et al. [18] proposed a minimal model with only interlinked positive and negative feedback loops of transcriptional regulation by the activator CREB1 and the repressor CREB2. Without considering feedbacks between the CREB proteins, Pettigrew et al. [8] developed a computational model characterizing complex dynamics of biochemical pathways downstream of 5-HT receptors. In this work, to describe more simply the biochemical pathways and gene regulation underlying 5-HT-induced LTM, we add the important extracellular sensitizing stimulus 5-HT as well as the product Ap-uch into the Song's minimal model. We also strive to examine dynamical properties of the gene regulatory network under the changing concentration of the stimulus, [5-HT], cooperating with the varying positive feedback strength in inducing a high state of CREB1 for the establishment of long-term memory. Different dynamics including monostability, bistability and multistability due to coexistence of stable steady states and oscillations is investigated by means of codimension-2 bifurcation analysis. At the different positive feedback strengths, comparative analysis of deterministic and stochastic dynamics reveals that codimension-1 bifurcation with respect to [5-HT] as the parameter can predict diverse stochastic behaviors resulted from the finite number of molecules, and the number of CREB1 molecules more and more preferentially resides near the high steady state with increasing [5-HT], which contributes to long

  20. Absolute Integral Cross Sections for the State-selected Ion-Molecule Reaction N2+(X2Σg+ v+ = 0-2) + C2H2 in the Collision Energy Range of 0.03-10.00 eV

    NASA Astrophysics Data System (ADS)

    Xu, Yuntao; Xiong, Bo; Chung Chang, Yih; Ng, C. Y.

    2016-08-01

    Using the vacuum ultraviolet laser pulsed field ionization-photoion source, together with the double-quadrupole-double-octopole mass spectrometer developed in our laboratory, we have investigated the state-selected ion-molecule reaction {{{{N}}}2}+({X}2{{{{Σ }}}{{g}}}+; v + = 0-2, N+ = 0-9) + C2H2, achieving high internal-state selectivity and high kinetic energy resolution for reactant {{{{N}}}2}+ ions. The charge transfer (CT) and hydrogen-atom transfer (HT) channels, which lead to the respective formation of product {{{C}}}2{{{{H}}}2}+ and N2H+ ions, are observed. The vibrationally selected absolute integral cross sections for the CT [σ CT(v +)] and HT [[σ HT(v +)] channels obtained in the center-of-mass collision energy (E cm) range of 0.03-10.00 eV reveal opposite E cm dependences. The σ CT(v +) is found to increase as E cm is decreased, and is consistent with the long-range exothermic CT mechanism, whereas the E cm enhancement observed for the σ HT(v +) suggests effective coupling of kinetic energy to internal energy, enhancing the formation of N2H+. The σ HT(v +) curve exhibits a step at E cm = 0.70-1.00 eV, suggesting the involvement of the excited {{{C}}}2{{{{H}}}2}+({A}2{{{{Σ }}}{{g}}}+) state in the HT reaction. Contrary to the strong E cm dependences for σ CT(v +) and σ HT(v +), the effect of vibrational excitation of {{{{N}}}2}+ on both the CT and HT channels is marginal. The branching ratios and cross sections for the CT and HT channels determined in the present study are useful for modeling the atmospheric compositions of Saturn's largest moon, Titan. These cross sections and branching ratios are also valuable for benchmarking theoretical calculations on chemical dynamics of the titled reaction.

  1. Microalbuminuria and early renal response to lethal dose Shiga toxin type 2 in rats.

    PubMed

    Ochoa, Federico; Oltra, Gisela; Gerhardt, Elizabeth; Hermes, Ricardo; Cohen, Lilian; Damiano, Alicia E; Ibarra, Cristina; Lago, Nestor R; Zotta, Elsa

    2012-01-01

    In Argentina, hemolytic uremic syndrome (HUS) constitutes the most frequent cause of acute renal failure in children. Approximately 2%-4% of patients die during the acute phase, and one-third of the 96% who survive are at risk of chronic renal sequelae. Little information is available about the direct effect of Shiga toxin type 2 (Stx2) on the onset of proteinuria and the evolution of toxin-mediated glomerular or tubular injury. In this work, rats were injected intraperitoneally with recombinant Escherichia coli culture supernatant containing Stx2 (sStx2; 20 μg/kg body weight) to induce HUS. Functional, immunoblotting, and immunohistochemistry studies were carried out to determine alterations in slit diaphragm proteins and the proximal tubule endocytic system at 48 hours post-inoculation. We detected a significant increase in microalbuminuria, without changes in the proteinuria values compared to the control rats. In immunoperoxidase studies, the renal tubules and glomerular mesangium showed an increased expression of transforming growth factor β(1)(TGF-β(1)). The expression of megalin was decreased by immunoperoxidase and the cytoplasm showed a granular pattern of megalin expression by immunofluorescence techniques. Western blot analysis performed in the renal cortex from sStx2-treated and control rats using anti-nephrin and anti-podocalyxin antibodies showed a decreased expression of these proteins. We suggest that the alterations in slit diaphragm proteins and megalin expression could be related to the development of microalbuminuria in response to lethal doses of Stx2.

  2. Abrogated Freud-1/Cc2d1a Repression of 5-HT1A Autoreceptors Induces Fluoxetine-Resistant Anxiety/Depression-Like Behavior.

    PubMed

    Vahid-Ansari, Faranak; Daigle, Mireille; Manzini, M Chiara; Tanaka, Kenji F; Hen, René; Geddes, Sean D; Béïque, Jean-Claude; James, Jonathan; Merali, Zul; Albert, Paul R

    2017-12-06

    Freud-1/Cc2d1a represses the gene transcription of serotonin-1A (5-HT1A) autoreceptors, which negatively regulate 5-HT tone. To test the role of Freud-1 in vivo , we generated mice with adulthood conditional knock-out of Freud-1 in 5-HT neurons ( cF1ko ). In cF1ko mice, 5-HT1A autoreceptor protein, binding and hypothermia response were increased, with reduced 5-HT content and neuronal activity in the dorsal raphe. The cF1ko mice displayed increased anxiety- and depression-like behavior that was resistant to chronic antidepressant (fluoxetine) treatment. Using conditional Freud-1/5-HT1A double knock-out ( cF1/1A dko ) to disrupt both Freud-1 and 5-HT1A genes in 5-HT neurons, no increase in anxiety- or depression-like behavior was seen upon knock-out of Freud-1 on the 5-HT1A autoreceptor-negative background; rather, a reduction in depression-like behavior emerged. These studies implicate transcriptional dysregulation of 5-HT1A autoreceptors by the repressor Freud-1 in anxiety and depression and provide a clinically relevant genetic model of antidepressant resistance. Targeting specific transcription factors, such as Freud-1, to restore transcriptional balance may augment response to antidepressant treatment. SIGNIFICANCE STATEMENT Altered regulation of the 5-HT1A autoreceptor has been implicated in human anxiety, major depression, suicide, and resistance to antidepressants. This study uniquely identifies a single transcription factor, Freud-1, as crucial for 5-HT1A autoreceptor expression in vivo Disruption of Freud-1 in serotonin neurons in mice links upregulation of 5-HT1A autoreceptors to anxiety/depression-like behavior and provides a new model of antidepressant resistance. Treatment strategies to reestablish transcriptional regulation of 5-HT1A autoreceptors could provide a more robust and sustained antidepressant response. Copyright © 2017 the authors 0270-6474/17/3711967-12$15.00/0.

  3. Density fluctuation in HT-6M tokamak by CO2 laser scattering

    NASA Astrophysics Data System (ADS)

    Zeng, Lei; Cao, Jinxiang; Zhu, Guoliang; Ding, Weixing; Yu, Chang-Xuan; Zhang, Daqing; Li, Youyi

    1993-09-01

    The small scale density fluctuations in the interior of HT-6M Ohmic plasma have been studied by CO2 laser collective scattering system in deuterium discharges covering a wide range of nqa (chord-average density times safety factor at the limiter) and energy confinement time. The relative density fluctuation level in the interior is inversely proportional to the toroidal magnetic field and average density, and the energy confinement time (tau) E decreases with the fluctuation level increasing in the region where (tau) E linearly increases with nq0.5a and satisfies the Goldston scaling law. It is suggested that the microturbulence in the interior zone is responsible for anomalous transport in tokamaks.

  4. Study of polymorphic variants of the serotonin 2A receptor gene (5-HT2A) and its possible effects on smoking habits of a population from northeastern Brazil.

    PubMed

    Ramos Neto, E S; Mágulas, J O; Sousa, J J S; Moura, A C M; Pinto, G R; Yoshioka, F K N; Canalle, R; Motta, F J N

    2014-10-20

    Previous studies have revealed a genetic component, including genetic polymorphisms in the serotonergic pathway, particularly in the serotonin receptor gene (5-HT2A). The aim of this study was to investigate associations of the T102C (rs6313) and A-1438G (rs6311) polymorphisms with tobacco use in a population from northeastern Brazil. We evaluated these polymorphisms in 135 nonsmokers and 135 smokers using polymerase chain reaction-restricted fragment length polymorphism. The distribution of allele and genotype frequencies and associations of polymorphisms with smoking were assessed with the chi-squared (χ(2)) test, the Fisher exact test, and odds ratio (OR) with a 95% confidence interval (CI). There were no differences in the distribution of genotype and allele frequencies between nonsmokers and smokers for A-1438G (P = 0.80) and T102C (P = 0.35). However, these polymorphisms were significantly associated with habit frequency (A/G: P = 0.02, OR = 6.87, 95%CI = 1.23-38.31, P = 0.04; A/G+G/G: P = 0.04, OR = 3.67, 95%CI = 1.06-12.75, P = 0.07), age of onset (C/C: P = 0.02, OR = 3.26, 95%CI = 1.17-9.07, P = 0.03, and nicotine dependence level (A/G: P = 0.02, OR = 3.28, 95%CI = 1.17-9.18, P = 0.04; A/G+G/G: P = 0.04, OR = 2.81, 95%CI = 1.13-6.99, P = 0.04; T/C: P = 0.03, OR = 3.12, 95%CI = 1.13-8.57, P = 0.04; T/C+C/C: P = 0.02, OR = 3.06, 95%CI = 1.22-7.70, P = 0.02). Therefore, these polymorphisms may not contribute significantly to smoking initiation, they do appear to be associated with habit maintenance.

  5. Characterization of Cyt2Bc Toxin from Bacillus thuringiensis subsp. medellin

    PubMed Central

    Juárez-Pérez, Victor; Guerchicoff, Alejandra; Rubinstein, Clara; Delécluse, Armelle

    2002-01-01

    We cloned and sequenced a new cytolysin gene from Bacillus thuringiensis subsp. medellin. Three IS240-like insertion sequence elements and the previously cloned cyt1Ab and p21 genes were found in the vicinity of the cytolysin gene. The cytolysin gene encodes a protein 29.7 kDa in size that is 91.5% identical to Cyt2Ba from Bacillus thuringiensis subsp. israelensis and has been designated Cyt2Bc. Inclusions containing Cyt2Bc were purified from the crystal-negative strain SPL407 of B. thuringiensis. Cyt2Bc reacted weakly with antibodies directed against Cyt2Ba and was not recognized by an antiserum directed against the reference cytolysin Cyt1Aa. Cyt2Bc was hemolytic only upon activation with trypsin and had only one-third to one-fifth of the activity of Cyt2Ba, depending on the activation time. Cyt2Bc was also mosquitocidal against Aedes aegypti, Anopheles stephensi, and Culex quinquefasciatus, including strains resistant to the Bacillus sphaericus binary toxin. Its toxicity was half of that of Cyt2Ba on all mosquito species except resistant C. quinquefasciatus. PMID:11872472

  6. Inhibition by tetanus toxin of sodium-dependent, high-affinity [3H]5-hydroxytryptamine uptake in rat synaptosomes.

    PubMed

    Inserte, J; Najib, A; Pelliccioni, P; Gil, C; Aguilera, J

    1999-01-01

    Tetanus toxin (TeTx) is a powerful clostridial neurotoxin that inhibits Ca2+-dependent neurotransmitter secretion as do the botulinum neurotoxins (BoNTs). We found that TeTx (but not BoNT/A) produced a specific time- and dose-dependent inhibition of Na+-dependent [3H]5-hydroxytryptamine (serotonin, 5-HT) uptake in rat CNS synaptosomes. This effect was found in all CNS tryptaminergic areas, being maximal in the hippocampus and occipital cortex. TeTx produced the maximum reduction in [3H]5-HT uptake after 30 min of preincubation, being significant also at lower doses (10(-12) M) or shorter incubation times (10 min). Serotonin transport inhibitors such as fenfluramine (IC50, 11.0 +/- 0.9 microM), paroxetine (IC50, 33.5 +/- 0.1 microM), and imipramine (IC50, 89.9 +/- 5.7 microM) were 3 or 4 orders of magnitude less potent than TeTx (IC50, 8.7 +/- 1.0 nM). Of the two fragments of TeTx, (the C-terminal portion of the neurotoxin heavy chain, which is responsible for the binding to the nerve tissue) was consistently more effective than the L-H(N) fragment (the light neurotoxin chain disulfide linked to the N-terminal portion of the heavy chain, which is responsible for the toxic metalloprotease action) as inhibitor of [3H]5-HT uptake in synaptosomal preparations (56 +/- 5% and 95 +/- 3% with respect to control, respectively). Antagonism of the toxin-induced [3H]5-HT uptake blockade could not be reversed by zinc chelators but did have the ability to antagonize the TeTx inhibition of basal and K+-evoked [3H]5-HT release in rat synaptosomes. The reduction in serotonin accumulation induced by TeTx could be responsible for some tetanic symptoms that have been related to the serotonergic system.

  7. Behavioral Effects of Systemic, Infralimbic and Prelimbic Injections of a Serotonin 5-HT2A Antagonist in Carioca High- and Low-Conditioned Freezing Rats

    PubMed Central

    León, Laura A.; Castro-Gomes, Vitor; Zárate-Guerrero, Santiago; Corredor, Karen; Mello Cruz, Antonio P.; Brandão, Marcus L.; Cardenas, Fernando P.; Landeira-Fernandez, J.

    2017-01-01

    The role of serotonin (5-hydroxytryptamine [5-HT]) and 5-HT2A receptors in anxiety has been extensively studied, mostly without considering individual differences in trait anxiety. Our laboratory developed two lines of animals that are bred for high and low freezing responses to contextual cues that are previously associated with footshock (Carioca High-conditioned Freezing [CHF] and Carioca Low-conditioned Freezing [CLF]). The present study investigated whether ketanserin, a preferential 5-HT2A receptor blocker, exerts distinct anxiety-like profiles in these two lines of animals. In the first experiment, the animals received a systemic injection of ketanserin and were exposed to the elevated plus maze (EPM). In the second experiment, these two lines of animals received microinjections of ketanserin in the infralimbic (IL) and prelimbic (PL) cortices and were exposed to either the EPM or a contextual fear conditioning paradigm. The two rat lines exhibited bidirectional effects on anxiety-like behavior in the EPM and opposite responses to ketanserin. Both systemic and intra-IL cortex injections of ketanserin exerted anxiolytic-like effects in CHF rats but anxiogenic-like effects in CLF rats. Microinjections of ketanserin in the PL cortex also exerted anxiolytic-like effects in CHF rats but had no effect in CLF rats. These results suggest that the behavioral effects of 5-HT2A receptor antagonism might depend on genetic variability associated with baseline reactions to threatening situations and 5-HT2A receptor expression in the IL and PL cortices. Highlights -CHF and CLF rats are two bidirectional lines that are based on contextual fear conditioning.-CHF rats have a more “anxious” phenotype than CLF rats in the EPM.-The 5-HT2A receptor antagonist ketanserin had opposite behavioral effects in CHF and CLF rats.-Systemic and IL injections either decreased (CHF) or increased (CLF) anxiety-like behavior.-PL injections either decreased (CHF) anxiety-like behavior or

  8. Evidence for a role of 5-HT2C receptors in the motor aspects of performance, but not the efficacy of food reinforcers, in a progressive ratio schedule.

    PubMed

    Bezzina, G; Body, S; Cheung, T H C; Hampson, C L; Bradshaw, C M; Glennon, J C; Szabadi, E

    2015-02-01

    5-Hydroxytryptamine2C (5-HT2C) receptor agonists reduce the breakpoint in progressive ratio schedules of reinforcement, an effect that has been attributed to a decrease of the efficacy of positive reinforcers. However, a reduction of the breakpoint may also reflect motor impairment. Mathematical models can help to differentiate between these processes. The effects of the 5-HT2C receptor agonist Ro-600175 ((αS)-6-chloro-5-fluoro-α-methyl-1H-indole-1-ethanamine) and the non-selective 5-HT receptor agonist 1-(m-chlorophenyl)piperazine (mCPP) on rats' performance on a progressive ratio schedule maintained by food pellet reinforcers were assessed using a model derived from Killeen's Behav Brain Sci 17:105-172, 1994 general theory of schedule-controlled behaviour, 'mathematical principles of reinforcement'. Rats were trained under the progressive ratio schedule, and running and overall response rates in successive ratios were analysed using the model. The effects of the agonists on estimates of the model's parameters, and the sensitivity of these effects to selective antagonists, were examined. Ro-600175 and mCPP reduced the breakpoint. Neither agonist significantly affected a (the parameter expressing incentive value), but both agonists increased δ (the parameter expressing minimum response time). The effects of both agonists could be attenuated by the selective 5-HT2C receptor antagonist SB-242084 (6-chloro-5-methyl-N-{6-[(2-methylpyridin-3-yl)oxy]pyridin-3-yl}indoline-1-carboxamide). The effect of mCPP was not altered by isamoltane, a selective 5-HT1B receptor antagonist, or MDL-100907 ((±)2,3-dimethoxyphenyl-1-(2-(4-piperidine)methanol)), a selective 5-HT2A receptor antagonist. The results are consistent with the hypothesis that the effect of the 5-HT2C receptor agonists on progressive ratio schedule performance is mediated by an impairment of motor capacity rather than by a reduction of the incentive value of the food reinforcer.

  9. Putative antipsychotics with pronounced agonism at serotonin 5-HT1A and partial agonist activity at dopamine D2 receptors disrupt basal PPI of the startle reflex in rats.

    PubMed

    Auclair, Agnès L; Galinier, Alexandra; Besnard, Joël; Newman-Tancredi, Adrian; Depoortère, Ronan

    2007-07-01

    Prepulse inhibition (PPI) of the startle reflex has been extensively studied because it is disrupted in several psychiatric diseases, most notably schizophrenia. In rats, and to a lesser extent, in humans, PPI can be diminished by dopamine (DA) D(2)/D(3) and serotonin 5-HT(1A) receptor agonists. A novel class of potential antipsychotics (SSR181507, bifeprunox, and SLV313) possess partial agonist/antagonist properties at D(2) receptors and various levels of 5-HT(1A) activation. It thus appeared warranted to assess, in Sprague-Dawley rats, the effects of these antipsychotics on basal PPI. SSR181507, sarizotan, and bifeprunox decreased PPI, with a near-complete abolition at 2.5-10 mg/kg; SLV313 had a significant effect at 0.16 mg/kg only. Co-treatment with the 5-HT(1A) receptor antagonist WAY100,635 (0.63 mg/kg) showed that the 5-HT(1A) agonist activity of SSR181507 was responsible for its effect. By contrast, antipsychotics with low affinity and/or efficacy at 5-HT(1A) receptors, such as aripiprazole (another DA D(2)/D(3) and 5-HT(1A) ligand), and established typical and atypical antipsychotics (haloperidol, clozapine, risperidone, olanzapine, quetiapine, and ziprasidone) had no effect on basal PPI (0.01-2.5 to 2.5-40 mg/kg). The present data demonstrate that some putative antipsychotics with pronounced 5-HT(1A) agonist activity, coupled with partial agonist activity at DA D(2) receptors, markedly diminish PPI of the startle reflex in rats. These data raise the issue of the influence of such compounds on sensorimotor gating in humans.

  10. Role of IL-1 beta and 5-HT2 receptors in midbrain periaqueductal gray (PAG) in potentiating defensive rage behavior in cat.

    PubMed

    Bhatt, Suresh; Bhatt, Rekha; Zalcman, Steven S; Siegel, Allan

    2008-02-01

    Feline defensive rage, a form of aggressive behavior that occurs in response to a threat can be elicited by electrical stimulation of the medial hypothalamus or midbrain periaqueductal gray (PAG). Our laboratory has recently begun a systematic examination of the role of cytokines in the regulation of rage and aggressive behavior. It was shown that the cytokine, interleukin-2 (IL-2), differentially modulates defensive rage when microinjected into the medial hypothalamus and PAG by acting through separate neurotransmitter systems. The present study sought to determine whether a similar relationship exists with respect to interleukin 1-beta (IL-1 beta), whose receptor activation in the medial hypothalamus potentiates defensive rage. Thus, the present study identified the effects of administration of IL-1 beta into the PAG upon defensive rage elicited from the medial hypothalamus. Microinjections of IL-1 beta into the dorsal PAG significantly facilitated defensive rage behavior elicited from the medial hypothalamus in a dose and time dependent manner. In addition, the facilitative effects of IL-1 beta were blocked by pre-treatment with anti-IL-1 beta receptor antibody, while IL-1 beta administration into the PAG had no effect upon predatory attack elicited from the lateral hypothalamus. The findings further demonstrated that IL-1 beta's effects were mediated through 5-HT(2) receptors since pretreatment with a 5-HT(2C) receptors antagonist blocked the facilitating effects of IL-1 beta. An extensive pattern of labeling of IL-1 beta and 5-HT(2C) receptors in the dorsal PAG supported these findings. The present study demonstrates that IL-beta in the dorsal PAG, similar to the medial hypothalamus, potentiates defensive rage behavior and is mediated through a 5-HT(2C) receptor mechanism.

  11. (+)Lysergic acid diethylamide, but not its nonhallucinogenic congeners, is a potent serotonin 5HT1C receptor agonist

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Burris, K.D.; Breeding, M.; Sanders-Bush, E.

    Activation of central serotonin 5HT2 receptors is believed to be the primary mechanism whereby lysergic acid diethylamide (LSD) and other hallucinogens induce psychoactive effects. This hypothesis is based on extensive radioligand binding and electrophysiological and behavioral studies in laboratory animals. However, the pharmacological profiles of 5HT2 and 5HT1C receptors are similar, making it difficult to distinguish between effects due to activation of one or the other receptor. For this reason, it was of interest to investigate the interaction of LSD with 5HT1C receptors. Agonist-stimulated phosphoinositide hydrolysis in rat choroid plexus was used as a direct measure of 5HT1C receptor activation.more » (+)LSD potently stimulated phosphoinositide hydrolysis in intact choroid plexus and in cultures of choroid plexus epithelial cells, with EC50 values of 9 and 26 nM, respectively. The effect of (+)LSD in both systems was blocked by 5HT receptor antagonists with an order of activity consistent with interaction at 5HT1C receptors. Neither (+)-2-bromo-LSD nor lisuride, two nonhallucinogenic congeners of LSD, were able to stimulate 5HT1C receptors in cultured cells or intact choroid plexus. In contrast, lisuride, like (+)LSD, is a partial agonist at 5HT2 receptors in cerebral cortex slices and in NIH 3T3 cells transfected with 5HT2 receptor cDNA. The present finding that (+)LSD, but not its nonhallucinogenic congeners, is a 5HT1C receptor agonist suggests a possible role for these receptors in mediating the psychoactive effects of LSD.« less

  12. Simultaneous measurement of the HT and DT fusion burn histories in inertial fusion implosions

    DOE PAGES

    Zylstra, Alex B.; Herrmann, Hans W.; Kim, Yong Ho; ...

    2017-05-23

    Measuring the thermonuclear burn history is an important way to diagnose inertial fusion implosions. Here, using the gas Cherenkov detectors at the OMEGA laser facility, we measure the HT fusion burn in a H 2+T 2 gas-fueled implosion for the first time. Then, using multiple detectors with varied Cherenkov thresholds, we demonstrate a technique for simultaneously measuring both the HT and DT burn histories from an implosion where the total reaction yields are comparable. This new technique will be used to study material mixing and kinetic phenomena in implosions.

  13. Simultaneous measurement of the HT and DT fusion burn histories in inertial fusion implosions

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Zylstra, Alex B.; Herrmann, Hans W.; Kim, Yong Ho

    Measuring the thermonuclear burn history is an important way to diagnose inertial fusion implosions. Here, using the gas Cherenkov detectors at the OMEGA laser facility, we measure the HT fusion burn in a H 2+T 2 gas-fueled implosion for the first time. Then, using multiple detectors with varied Cherenkov thresholds, we demonstrate a technique for simultaneously measuring both the HT and DT burn histories from an implosion where the total reaction yields are comparable. This new technique will be used to study material mixing and kinetic phenomena in implosions.

  14. Biliatresone, a Reactive Natural Toxin from Dysphania glomulifera and D. littoralis: Discovery of the Toxic Moiety 1,2-Diaryl-2-Propenone.

    PubMed

    Koo, Kyung A; Lorent, Kristin; Gong, Weilong; Windsor, Peter; Whittaker, Stephen J; Pack, Michael; Wells, Rebecca G; Porter, John R

    2015-08-17

    We identified a reactive natural toxin, biliatresone, from Dysphania glomulifera and D. littoralis collected in Australia that produces extrahepatic biliary atresia in a zebrafish model. Three additional isoflavonoids, including the known isoflavone betavulgarin, were also isolated. Biliatresone is in the very rare 1,2-diaryl-2-propenone class of isoflavonoids. The α-methylene of the 1,2-diaryl-2-propenone of biliatresone spontaneously reacts via Michael addition in the formation of water and methanol adducts. The lethal dose of biliatresone in a zebrafish assay was 1 μg/mL, while the lethal dose of synthetic 1,2-diaryl-2-propen-1-one was 5 μg/mL, suggesting 1,2-diaryl-2-propenone as the toxic Michael acceptor.

  15. Biliatresone, a Reactive Natural Toxin from Dysphania glomulifera and D. littoralis: Discovery of the Toxic Moiety 1,2-Diaryl-2-Propenone

    PubMed Central

    Koo, Kyung A.; Lorent, Kristin; Gong, Weilong; Windsor, Peter; Whittaker, Stephen J.; Pack, Michael; Wells, Rebecca G.; Porter, John R.

    2016-01-01

    We identified a reactive natural toxin, biliatresone, from Dysphania glomulifera and D. littoralis collected in Australia that produces extrahepatic biliary atresia in a zebrafish model. Three additional isoflavonoids, including the known isoflavone betavulgarin, were also isolated. Biliatresone is in the very rare 1,2-diaryl-2-propenone class of isoflavonoids. The α-methylene of the 1,2-diaryl-2-propenone of biliatresone spontaneously reacts via Michael addition in the formation of water and methanol adducts. The lethal dose of biliatresone in a zebrafish assay was 1 μg/mL, while the lethal dose of synthetic 1,2-diaryl-2-propen-1-one was 5 μg/mL, suggesting 1,2-diaryl-2-propenone as the toxic Michael acceptor. PMID:26175131

  16. 42 CFR 73.3 - HHS select agents and toxins.

    Code of Federal Regulations, 2011 CFR

    2011-10-01

    ... Register and will be listed on the CDC Web site at http://www.cdc.gov/. (2) If an excluded attenuated...) Staphylococcal enterotoxins T-2 toxin Tetrodotoxin Tick-borne encephalitis complex (flavi) viruses (Central... listed in paragraph (b) of this section. (2) Recombinant nucleic acids that encode for the functional...

  17. The relaxant 5-HT receptor in the dog coronary artery smooth muscle: pharmacological resemblance to the cloned 5-ht7 receptor subtype.

    PubMed Central

    Terrón, J. A.

    1996-01-01

    1. The relaxant effect of 5-hydroxytryptamine (5-HT) in the dog isolated coronary artery deprived of endothelium is mediated by a receptor unrelated to the 5-HT1, 5-HT2, 5-HT3 or 5-HT4 types. Based upon the pharmacological characteristics of this relaxant 5-HT receptor and those reported for the new members of the 5-HT receptor family, the present study explored the possibility that the relaxant 5-HT receptor referred to above, corresponds to the cloned 5-ht7 subtype. Thus, the relaxing and/or blocking effects of several 5-HT receptor drugs as well as some typical and atypical antipsychotic drugs with high affinity for the cloned 5-ht7 receptor in precontracted ring segments were analyzed. 2. 5-HT, 5-carboxamidotryptamine (5-CT) and 5-methoxytryptamine, but not 8-OH-DPAT or sumatriptan, produced concentration-dependent relaxations in endothelium-denuded canine coronary artery rings precontracted with prostaglandin F2a (2 microM). Clozapine (1 microM) produced in some cases a small relaxing effect and antagonized 5-HT- and 5-CT-induced relaxation suggesting a partial agonist effect. In the presence of the 5-HT1D receptor antagonist, GR127935 (100 nM), the rank order of agonist potency was 5-CT > 5-HT > clozapine > or = 5-methoxytryptamine. 8-OH-DPAT and sumatriptan remained inactive as agonists. 3. In GR127935-treated preparations, methiothepin (3 nM) and mianserin (1 microM), as well as the antipsychotics, clozapine (1 microM), pimozide (300 nM), risperidone (3 nM) and spiperone (1 microM), failed to induce a significant relaxation in prostaglandin F2x-precontracted vessels, but produced significant rightward displacements of the concentration-response curves to 5-HT and 5-CT without significantly reducing the Emax. In a final set of experiments with 5-CT, metergoline (100 nM) and mesulergine (300 nM) behaved as competitive antagonists. In contrast, lisuride (3 nM) noncompetitively antagonized 5-CT-induced relaxation. The estimated affinity (apparent pKa values) of

  18. Forced swimming test and fluoxetine treatment: in vivo evidence that peripheral 5-HT in rat platelet-rich plasma mirrors cerebral extracellular 5-HT levels, whilst 5-HT in isolated platelets mirrors neuronal 5-HT changes.

    PubMed

    Bianchi, M; Moser, C; Lazzarini, C; Vecchiato, E; Crespi, F

    2002-03-01

    Low levels of central serotonin (5-HT) have been related to the state of depression, and 5-HT is the major target of the newer antidepressant drugs such as selective serotonin reuptake inhibitors (SSRIs). Neurons and platelets display structural and functional similarities, so that the latter have been proposed as a peripheral model of central functions. In particular, in blood more than 99% of 5-HT is contained in platelets, so that one could consider changes in 5-HT levels in platelets as a mirror of changes in central 5-HT. Here, this hypothesis has been studied via the analysis of the influence of: (1) the forced swimming test (FST, which has been proved to be of utility to predict the clinical efficacy of antidepressants in rodents) and (2) treatment with the SSRI fluoxetine upon 5-HT levels monitored in brain regions and in peripheral platelets by means of electrochemical in vivo and ex vivo measurements. The results obtained confirm that the FST increases immobility; furthermore they show a parallel and significant decrease in cerebral (brain homogenate) and peripheral (in platelet-rich plasma, PRP) voltammetric 5-HT levels following the FST in naive rats. In addition, subchronic treatment with fluoxetine was followed by a significant increase in 5-HT levels in PRP, while the same SSRI treatment performed within the FST resulted in a decrease in the 5-HT levels in PRP. However, this decrease was inferior to that observed without SSRI treatment. These data suggest that there is an inverse relationship between immobility and the levels of 5-HT in PRP and that these peripheral 5-HT levels are sensitive to: (1) the FST, (2) the treatment with fluoxetine and (3) the combination of both treatments, i.e. SSRI + FST. It has been reported that SSRI treatment at first inhibits the 5-HT transporter in brain, resulting in increased extracellular 5-HT, while following sustained SSRI treatments decreased intracellular levels of central 5-HT were observed. Accordingly, the

  19. Retracted: Association of ACE I/D gene polymorphism with T2DN susceptibility and the risk of T2DM developing into T2DN in a Caucasian population.

    PubMed

    Liu, Guohui; Zhou, Tian-Biao; Jiang, Zongpei; Zheng, Dongwen

    2015-03-01

    The association of the angiotensin-converting enzyme (ACE) insertion/deletion (I/D) gene polymorphism with type-2 diabetic nephropathy (T2DN) susceptibility and the risk of type-2 diabetes mellitus (T2DM) developing into T2DN in Caucasian populations is still controversial. A meta-analysis was performed to evaluate the association of ACE I/D gene polymorphism with T2DN susceptibility and the risk of T2DM developing into T2DN in Caucasian populations. A predefined literature search and selection of eligible relevant studies were performed to collect data from electronic databases. Sixteen articles were identified for the analysis of the association of ACE I/D gene polymorphism with T2DN susceptibility and the risk of T2DM developing into T2DN in Caucasian populations. ACE I/D gene polymorphism was not associated with T2DN susceptibility and the risk of patients with T2DM developing T2DN in Caucasian populations. Sensitivity analysis according to sample size of case (<100 vs. ≥100) was also performed, and the results were similar to the non-sensitivity analysis. ACE I/D gene polymorphism was not associated with T2DN susceptibility and the risk of patients with T2DM developing T2DN in Caucasian populations. However, more studies should be performed in the future. © The Author(s) 2014.

  20. Electronic properties of in-plane phase engineered 1T'/2H/1T' MoS2

    NASA Astrophysics Data System (ADS)

    Thakur, Rajesh; Sharma, Munish; Ahluwalia, P. K.; Sharma, Raman

    2018-04-01

    We present the first principles studies of semi-infinite phase engineered MoS2 along zigzag direction. The semiconducting (2H) and semi-metallic (1T') phases are known to be stable in thin-film MoS2. We described the electronic and structural properties of the infinite array of 1T'/2H/1T'. It has been found that 1T'phase induced semi-metallic character in 2H phase beyond interface but, only Mo atoms in 2H phase domain contribute to the semi-metallic nature and S atoms towards semiconducting state. 1T'/2H/1T' system can act as a typical n-p-n structure. Also high holes concentration at the interface of Mo layer provides further positive potential barriers.