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Sample records for haemonchus contortus acetylcholine

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

    NARCIS (Netherlands)

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

    1997-01-01

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

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

    NARCIS (Netherlands)

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

    1997-01-01

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

  3. Carbohydrate epitopes on Haemonchus contortus antigens

    NARCIS (Netherlands)

    Schallig, H. D.; van Leeuwen, M. A.

    1996-01-01

    Extracts of infective larvae and adults of the trichostrongylid Haemonchus contortus were studied for the presence of carbohydrate moieties. Several different lectin-binding sites were demonstrated in both stages using a panel of nine lectins. The carbohydrate specificity of the lectins used

  4. Early detection of Haemonchus contortus infection in sheep using ...

    African Journals Online (AJOL)

    Haemonchus contortus is a blood-sucking parasite causing the presence of faecal occult blood (FOB). The objective was to study three different FOB tests in order to have a new indicator of H. contortus infection in sheep that could be included in the genetic evaluation system as an alternative selection criterion to faecal ...

  5. Diagnosis, Treatment and Management of Haemonchus contortus in Small Ruminants.

    Science.gov (United States)

    Besier, R B; Kahn, L P; Sargison, N D; Van Wyk, J A

    2016-01-01

    Haemonchus contortus is a highly pathogenic, blood-feeding nematode of small ruminants, and a significant cause of mortalities worldwide. Haemonchosis is a particularly significant threat in tropical, subtropical and warm temperate regions, where warm and moist conditions favour the free-living stages, but periodic outbreaks occur more widely during periods of transient environmental favourability. The clinical diagnosis of haemonchosis is based mostly on the detection of anaemia in association with a characteristic epidemiological picture, and confirmed at postmortem by the finding of large numbers of H. contortus in the abomasum. The detection of impending haemonchosis relies chiefly on periodic monitoring for anaemia, including through the 'FAMACHA' conjunctival-colour index, or through faecal worm egg counts and other laboratory procedures. A range of anthelmintics for use against H. contortus is available, but in most endemic situations anthelmintic resistance significantly limits the available treatment options. Effective preventative programmes vary depending on environments and enterprise types, and according to the scale of the haemonchosis risk and the local epidemiology of infections, but should aim to prevent disease outbreaks while maintaining anthelmintic efficacy. Appropriate strategies include animal management programmes to avoid excessive H. contortus challenge, genetic and nutritional approaches to enhance resistance and resilience to infection, and the monitoring of H. contortus infection on an individual animal or flock basis. Specific strategies to manage anthelmintic resistance centre on the appropriate use of effective anthelmintics, and refugia-based treatment schedules. Alternative approaches, such as biological control, may also prove useful, and vaccination against H. contortus appears to have significant potential in control programmes. Copyright © 2016 Elsevier Ltd. All rights reserved.

  6. Anthelmintic activity of Cymbopogon citratus against Haemonchus contortus

    Directory of Open Access Journals (Sweden)

    Iara Tersia Freitas Macedo

    Full Text Available Parasitic nematodes are of major economic importance in livestock. An alternative for the control of parasites is phytotherapy. This study evaluated the efficacy of Cymbopogon citratus decoction (CcD, C. citratus essential oil (CcEo and citral against Haemonchus contortus using in vitro egg hatch test (EHT and larval development test (LDT and an in vivo test using a Meriones unguiculatus (gerbil model. The effect of 800 mg/kg CcEo was evaluated in gerbils that had been artificially infected with 5,000 third-stage H. contortus larvae. The effective concentrations required to inhibit 50% (EC50 of egg hatching were 0.46, 0.14 and 0.13 mg/mL for CcD, CcEo and citral, respectively. The EC50 values in the LDT were 5.04, 1.92 and 1.37 mg/mL for CcD, CcEo and citral, respectively. H. contortus population in the group treated with C. citratus essential oil was reduced by 38.5% (P< 0.05 in comparison to the control group. These results suggest that it may be possible to use C. citratusessential oil to control of H. contortus parasite of small ruminant.

  7. Characterization of the development of Haemonchus contortus ZJ strain from gerbils

    Directory of Open Access Journals (Sweden)

    Yi Yang

    2017-10-01

    Full Text Available Abstract Background Haemonchus contortus is a serious parasitic nematode in domestic ruminants around the world, including China. Haemonchus contortus has developed extensive resistance to commercial anthelmintics, which has produced a demand for new control methods, such as more effective drugs. Gerbils infected with H. contortus have previously been used as a model for anthelmintics selections, and the growth of H. contortus had been briefly examined. To enhance the model, this study provides an additional description of the development of H. contortus ZJ strain in gerbils. Results Gerbils were infected with H. contortus ZJ strain at a dose of 2000 exsheathed infective larvae (xL3s and sacrificed at 4, 7 and 18 days post-infection (dpi. Only fourth-stage larvae were found in the stomachs. About 2% of the inoculums were obtained at each of the three sampling time points. Larvae grew more slowly in gerbils than in sheep, but presented almost the same morphology. Rod-like crystalline inclusions were present in the intestinal cells of larvae, indicating that the metabolic rate of larvae was probably greatly reduced. Histological examination of stomach sections showed that larvae are located in the lumens or at the mucosal surfaces, with few inflammatory changes evident. Conclusions The development and features of H. contortus ZJ strain in gerbils were described. Our results provide supplementary information of H. contortus growth in gerbils, especially the presence of rod-like crystalline inclusions, and may contribute to improve the anthelmintic selection system.

  8. Haemonchus contortus: Characterization of the baculovirus expressed form of aminopeptidase H11

    NARCIS (Netherlands)

    Reszka, N.; Rijsewijk, F.A.M.; Zelnik, V.; Moskwa, B.; Bienkowska-Szewczyk, K.

    2007-01-01

    Recombinant form of Haemonchus contortus aminopeptidase H11, an intestinal membrane glycoprotein considered to be in its native form the most promising vaccine candidate, was produced in insect cells, characterised and tested in pilot vaccination-challenge trial on sheep. The sequence of the cloned

  9. Selection for high levamisole resistance in Haemonchus contortus monitored with an egg-hatch assay

    NARCIS (Netherlands)

    Hoekstra, R.; Borgsteede, F.H.M.; Boersema, J.H.; Roos, M.H.

    1997-01-01

    To investigate the characteristics of selection for levamisole resistance in Haemonchus contortus, the consecutive nematode generations of an in vivo selection were monitored with a newly developed egg-hatch assay. The in vivo selection was started with a population not previously exposed to any

  10. Larvicidal activity of Bacillus thuringiensis var. israelensis Cry11Aa toxin against Haemonchus contortus.

    Science.gov (United States)

    DE Lara, Ana Paula DE Souza Stori; Lorenzon, Lucas Bigolin; Vianna, Ana Muñoz; Santos, Francisco Denis Souza; Pinto, Luciano Silva; Aires Berne, Maria Elisabeth; Leite, Fábio Pereira Leivas

    2016-10-01

    Effective control of gastrointestinal parasites is necessary in sheep production. The development of anthelmintics resistance is causing the available chemically based anthelmintics to become less effective. Biological control strategies present an alternative to this problem. In the current study, we tested the larvicidal effects of Bacillus thuringiensis var. israelensis Cry11Aa toxin against Haemonchus contortus larvae. Bacterial suspensions [2 × 108 colony-forming units (CFU) g-1 of the feces] of B. thuringiensis var. israelensis and recombinant Escherichia coli expressing Cry11Aa toxin were added to naturally H. contortus egg-contaminated feces. The larvae were quantified, and significant reductions of 62 and 81% (P var. israelensis and recombinant E. coli expressing Cry11Aa toxin were then orally administered to lambs naturally infected with H. contortus. Twelve hours after administration, feces were collected and submitted to coprocultures. Significant larvae reductions (P var. israelensis is a promising new class of biological anthelmintics for treating sheep against H. contortus.

  11. Anthelmintic activity of acetone extracts from South African plants used on egg hatching of Haemonchus contortus

    CSIR Research Space (South Africa)

    Sakong, BM

    2016-01-01

    Full Text Available , C.M.L., Moraes, S.M. & Vieira, L.S., 1999, ‘In vitro ovicidal and larvicidal effect of the plants Spigelia anthelmia and Momordica charantia against the nematode Haemonchus contortus’, Ciência Animal 9, 67–73. Bizimenyera, E.S., Githiori, J... ruminants of semi-arid Brazilian northeast’, Ciência Animal 10, 294–300. Mendoza de Gives, P., Crespo, J.F., Rodriguez, D.H., Prats, V.V., Hernandez, E.L. & Fernandez, G.E.O., 1998, ‘Biological control of Haemonchus contortus infective larvae in ovine...

  12. FACTORS AFFECTING VARIABILITY OF RESISTANCE IN GAROLE SHEEP NATURALLY INFECTED WITH HAEMONCHUS CONTORTUS

    Directory of Open Access Journals (Sweden)

    Manoranjan Roy

    2013-06-01

    Full Text Available Resistance status against natural infection to Haemonchus contortus as well as influence of season, sex, body weight, and haemoglobin type on resistance levels were evaluated in 309 numbers of Garole sheep. In adult Garole sheep, egg per gram(EPG of faeces for Haemonchus contortus was varied from 300 to 1600, but overall EPG in Garole have been recorded as 829.96 ± 20.60. The effects of season, sex, and body weight and haemoglobin type on EPG were all found to be highly significant (P < 0.01. EPG count was highest during monsoon (986.27 ± 28.26, followed by summer (832.88 ± 28.26 and lowest during winter (670.74 ± 28.26 which indicated the existence of a seasonal variation of EPG. Rams had higher EPG (954.32 ± 57.93 than ewes (705.60 ± 45.79 which reflected that males appeared to be more susceptible to Haemonchus contortus infection compared to females. Animals with lower body weight (upto 10 kg showed higher EPG (1017.20 ± 54.82, then the infection level decreased as body weight increased (886.79 ± 56.23 for 10 kg to 12 kg and 737.18 ± 50.29 for 12 kg to 14 kg and lowest EPG was recorded in animals with above 14 kg body weight (678.68 ± 54.49. This study reveals Hb-BB type animals had higher EPG count (983.81 ± 18.22 in comparison to Hb-AB type animals (676.12 ± 33.96 indicating that Haemoglobin-A locus has some relation with resistance. From our study it can be concluded that resistant level of Garole sheep against Haemonchus contortus is influenced by some intrinsic factors like sex, body weight, and haemoglobin type of the sheep and also by extrinsic factor like season.

  13. Ivermectin resistant and susceptible third-stage larvae of Haemonchus contortus: cholinesterase and phosphatase activities

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    Consuelo Giménez-Pardo

    2004-03-01

    Full Text Available Cholinesterase and acid phosphatase (AP, but not alkaline phosphatase activities, were detected in cytosolic and membrane-bound fractions of ivermectin resistant and susceptible Haemonchus contortus infective-stage larvae. Some differences in acetylcholinesterase activity of cytosolic fractions and in the AP activity of these fractions as well as in the response to AP inhibitors by membrane-bound fractions were detected. Data are discussed.

  14. Antigenic Cross-reactivity among Haemonchus contortus, Oesophagostomum columbianum and Trichuris ovis of Goat

    OpenAIRE

    JAS, Ruma; GHOSH, Joydeb; DAS, Kinsuk

    2016-01-01

    Background: Cross antigenicity is the major problem in developing a reliable tool for immunodiagnosis and immunoprophylaxis of parasitic diseases. Mixed infection due to different types of gastrointestinal parasites is more common than single species infection under field condition.Methods: The present study was undertaken to detect antigenic cross-reactivity among Haemonchus contortus, Oesophagostomum columbianum and Trichuris ovis of goats by SDS-PAGE and western blot analysis using hyperim...

  15. In vitro effects of Musa x paradisiaca extracts on four developmental stages of Haemonchus contortus.

    Science.gov (United States)

    Marie-Magdeleine, C; Udino, L; Philibert, L; Bocage, B; Archimede, H

    2014-02-01

    This study was carried out to evaluate the in vitro effect of Musa x paradisiaca stem and leaf against the parasitic nematode of small ruminants Haemonchus contortus. Three extracts (aqueous, methanolic and/or dichloromethane) of Musa x paradisiaca stem and leaf were tested in vitro on four developmental stages of H. contortus using egg hatch assay (EHA), larval development assay (LDA), L3 migration inhibition assay (LMI) and adult worm motility assay (AWM). The highly significant (P67% for each extract) and the negative effect of the dichloromethane extract of leaf on adult worm motility (43% of inhibition of motility after 24h of incubation) compared to the negative controls, suggest anthelmintic properties of Musa x paradisiaca stem and leaf against H. contortus. The active principles responsible for the activity could be secondary metabolites such as terpenoid and flavonoid compounds present in the leaf and stem of the plant. Copyright © 2013 Elsevier Ltd. All rights reserved.

  16. Effect of Vaccination with Irradiated Third Stage Larvae of Haemonchus Contortus

    International Nuclear Information System (INIS)

    Beriajaya; Sukardji, P.

    2000-01-01

    The objective of this study was to determine the effect of vaccination with irradiated third stage larvae of Haemonchus contortus on immune responses in sheep. A number of 15 young male thin-tail sheep freed of worms were divided into 3 groups of 5. The first group was vaccinated with 50.000 irradiated third larvae of H. contortus . The second group was vaccinated as group 1 but without challenged. The third group was not vaccinated but challenged as group 1. Observations were carried out on egg counts, worn counts, total serum protein and antibody titer against H. contortus. The results showed there was no significant differences (P>0.05) on egg counts, worn counts and antibody titer, but a significant difference was seen on value of serum protein between vaccinated group and non vaccinated group. The results showed no protective immunity which is showed in worn counts of vaccinated and non vaccinated groups. Key word: Haemonchus contortus, irradiated larvae, sheep, vaccine

  17. The 'Toolbox' of strategies for managing Haemonchus contortus in goats: What's in and what's out.

    Science.gov (United States)

    Kearney, P E; Murray, P J; Hoy, J M; Hohenhaus, M; Kotze, A

    2016-04-15

    A dynamic and innovative approach to managing the blood-consuming nematode Haemonchus contortus in goats is critical to crack dependence on veterinary anthelmintics. H. contortus management strategies have been the subject of intense research for decades, and must be selected to create a tailored, individualized program for goat farms. Through the selection and combination of strategies from the Toolbox, an effective management program for H. contortus can be designed according to the unique conditions of each particular farm. This Toolbox investigates strategies including vaccines, bioactive forages, pasture/grazing management, behavioural management, natural immunity, FAMACHA, Refugia and strategic drenching, mineral/vitamin supplementation, copper Oxide Wire Particles (COWPs), breeding and selection/selecting resistant and resilient individuals, biological control and anthelmintic drugs. Barbervax(®), the ground-breaking Haemonchus vaccine developed and currently commercially available on a pilot scale for sheep, is prime for trialling in goats and would be an invaluable inclusion to this Toolbox. The specialised behaviours of goats, specifically their preferences to browse a variety of plants and accompanying physiological adaptations to the consumption of secondary compounds contained in browse, have long been unappreciated and thus overlooked as a valuable, sustainable strategy for Haemonchus management. These strategies are discussed in this review as to their value for inclusion into the 'Toolbox' currently, and the future implications of ongoing research for goat producers. Combining and manipulating strategies such as browsing behaviour, pasture management, bioactive forages and identifying and treating individual animals for haemonchosis, in addition to continuous evaluation of strategy effectiveness, is conducted using a model farm scenario. Selecting strategies from the Toolbox, with regard to their current availability, feasibility, economical cost

  18. Proteomic identification of galectin-11 and 14 ligands from Haemonchus contortus

    Directory of Open Access Journals (Sweden)

    Dhanasekaran Sakthivel

    2018-03-01

    Full Text Available Haemonchus contortus is the most pathogenic nematode of small ruminants. Infection in sheep and goats results in anaemia that decreases animal productivity and can ultimately cause death. The involvement of ruminant-specific galectin-11 (LGALS-11 and galectin-14 (LGALS-14 has been postulated to play important roles in protective immune responses against parasitic infection; however, their ligands are unknown. In the current study, LGALS-11 and LGALS-14 ligands in H. contortus were identified from larval (L4 and adult parasitic stages extracts using immobilised LGALS-11 and LGALS-14 affinity column chromatography and mass spectrometry. Both LGALS-11 and LGALS-14 bound more putative protein targets in the adult stage of H. contortus (43 proteins when compared to the larval stage (two proteins. Of the 43 proteins identified in the adult stage, 34 and 35 proteins were bound by LGALS-11 and LGALS-14, respectively, with 26 proteins binding to both galectins. Interestingly, hematophagous stage-specific sperm-coating protein and zinc metalloprotease (M13, which are known vaccine candidates, were identified as putative ligands of both LGALS-11 and LGALS-14. The identification of glycoproteins of H. contortus by LGALS-11 and LGALS-14 provide new insights into host-parasite interactions and the potential for developing new interventions.

  19. Effectiveness of Ivermectin and Albendazole against Haemonchus contortus in Sheep in West Java, Indonesia

    Science.gov (United States)

    Puspitasari, Silvia; Farajallah, Achmad; Sulistiawati, Erni; Muladno

    2016-01-01

    Administering a half dose of an anthelmintic is a simple method for detecting resistance in parasites infesting small ruminants. When a single anthelmintic fails in native sheep from Indonesia, a combination of anthelmintics from different chemical classes with different modes of action are administered as an alternative parasite-control strategy. This study compared the anthelmintic efficacy of ivermectin (IVM) and albendazole (ABZ) given either separately as a single dose or half dose or co-administered to sheep naturally infected with Haemonchus contortus. Twelve sheep from Bogor, West Java, Indonesia were divided into the following six treatment groups: half-dose IVM, full-dose IVM, half-dose ABZ, full-dose ABZ, combined IVM + ABZ, and control. The treatment efficacy was determined using the faecal egg count reduction test (FECRT) at day 0 (pre-treatment) and post-treatment at days 7, 14, 21, 28, 35, and 42. The efficacies of half-dose IVM, full-dose IVM, half-dose ABZ, full-dose ABZ, and the combination treatment ranged from −1900% to 100%, 99% to 100%, −167% to 100%, −467% to 89%, and −200% to 100%, respectively. The FECRT for the half-dose IVM, half-dose ABZ, full-dose ABZ showed that H. contortus is resistant to half-dose IVM and ABZ. Full-dose IVM was effective against H. contortus. The combined treatment was more effective against H. contortus than ABZ alone. PMID:27019686

  20. Effectiveness of Ivermectin and Albendazole against Haemonchus contortus in Sheep in West Java, Indonesia.

    Science.gov (United States)

    Puspitasari, Silvia; Farajallah, Achmad; Sulistiawati, Erni; Muladno

    2016-02-01

    Administering a half dose of an anthelmintic is a simple method for detecting resistance in parasites infesting small ruminants. When a single anthelmintic fails in native sheep from Indonesia, a combination of anthelmintics from different chemical classes with different modes of action are administered as an alternative parasite-control strategy. This study compared the anthelmintic efficacy of ivermectin (IVM) and albendazole (ABZ) given either separately as a single dose or half dose or co-administered to sheep naturally infected with Haemonchus contortus. Twelve sheep from Bogor, West Java, Indonesia were divided into the following six treatment groups: half-dose IVM, full-dose IVM, half-dose ABZ, full-dose ABZ, combined IVM + ABZ, and control. The treatment efficacy was determined using the faecal egg count reduction test (FECRT) at day 0 (pre-treatment) and post-treatment at days 7, 14, 21, 28, 35, and 42. The efficacies of half-dose IVM, full-dose IVM, half-dose ABZ, full-dose ABZ, and the combination treatment ranged from -1900% to 100%, 99% to 100%, -167% to 100%, -467% to 89%, and -200% to 100%, respectively. The FECRT for the half-dose IVM, half-dose ABZ, full-dose ABZ showed that H. contortus is resistant to half-dose IVM and ABZ. Full-dose IVM was effective against H. contortus. The combined treatment was more effective against H. contortus than ABZ alone.

  1. E-ADA activity in serum of lambs experimentally infected with Haemonchus contortus.

    Science.gov (United States)

    Da Silva, Aleksandro S; Fausto, Guilherme C; Grando, Thirssa H; Cadore, Carlos A; Pimentel, Victor C; Jaques, Jeandre A; Schetinger, Maria R C; Monteiro, Silvia G; Leal, Marta L R

    2013-08-01

    The aim of this study was to evaluate adenosine deaminase (E-ADA) activity in sera of lambs experimentally infected with Haemonchus contortus. We used 12 lambs divided into 2 groups; Group A had 5 healthy, non-infected animals (control) and Group B had 7 healthy animals infected with H. contortus . Lambs were infected orally with 500 larvae (L3) per animal every 2 days, for a period of 20 days, and later the infection was confirmed by examination of feces (eggs per gram [EPG] via fecal egg count). Blood collection was performed at days 0, 20, 40, 60, and 80 post-infection (PI) for analysis of E-ADA activity. Animals in Group A showed negative EPG throughout the experiment unlike those from Group B that had elevated EPG counts. E-ADA activity was reduced in the serum of animals infected with H. contortus when compared to non-infected controls at days 20, 40, 60, and 80 PI. Therefore, it is concluded that infection with H. contortus influences the E-ADA activity in lambs.

  2. Interactions Between Nutrition and Infections With Haemonchus contortus and Related Gastrointestinal Nematodes in Small Ruminants.

    Science.gov (United States)

    Hoste, H; Torres-Acosta, J F J; Quijada, J; Chan-Perez, I; Dakheel, M M; Kommuru, D S; Mueller-Harvey, I; Terrill, T H

    2016-01-01

    Interactions between host nutrition and feeding behaviour are central to understanding the pathophysiological consequences of infections of the digestive tract with parasitic nematodes. The manipulation of host nutrition provides useful options to control gastrointestinal nematodes as a component of an integrated strategy. Focussed mainly on the Haemonchus contortus infection model in small ruminants, this chapter (1) illustrates the relationship between quantitative (macro- and micro-nutrients) and qualitative (plant secondary metabolites) aspects of host nutrition and nematode infection, and (2) shows how basic studies aimed at addressing some generic questions can help to provide solutions, despite the considerable diversity of epidemiological situations and breeding systems. Copyright © 2016 Elsevier Ltd. All rights reserved.

  3. Pathology of Haemonchus contortus in New World camelids in the southeastern United States: a retrospective review.

    Science.gov (United States)

    Edwards, Erin E; Garner, Bridget C; Williamson, Lisa H; Storey, Bob E; Sakamoto, Kaori

    2016-03-01

    Most small ruminant farms in tropical climates are plagued by Haemonchus contortus, a hematophagous, abomasal parasite. Heavy burdens of this parasite can cause anemia, hypoproteinemia, weight loss, and mortality in susceptible animals. Haemonchus contortus is becoming a major health concern in New World camelids as well, namely llamas (Llama glama) and alpacas (Vicugna pacos), yet little research has been conducted regarding its prevalence or pathology in these species. Herein, we present a retrospective review of llamas and alpacas that were admitted to The University of Georgia Veterinary Teaching Hospital and Athens Diagnostic Laboratory between the years 2002 and 2013. Antemortem fecal egg count (FEC) estimates performed on 30 alpacas were negatively correlated with hematocrit, hemoglobin, and red blood cell count. Total protein was not significantly correlated with FEC. On postmortem examination, 55 of 198 camelids, including 2 from the aforementioned antemortem review, were infected with H. contortus, with llamas (42.6%) having a significantly higher infection rate than alpacas (22.2%). In 15.7% of the total cases, the parasite was the major cause of death. Common gross lesions included peritoneal, thoracic, and pericardial effusions, visceral pallor, subcutaneous edema, and serous atrophy of fat. Histologic lesions included centrilobular hepatic necrosis, hepatic atrophy, lymphoplasmacytic inflammation of the mucosa of the third gastric compartment (C3), extramedullary hematopoiesis in both the liver and spleen, and the presence of nematodes in C3. Our study emphasizes the importance of H. contortus diagnosis and herd monitoring in New World camelids, particularly llamas. © 2016 The Author(s).

  4. Effect of sericea lespedeza leaf meal pellets on adult female Haemonchus contortus in goats.

    Science.gov (United States)

    Kommuru, D S; Whitley, N C; Miller, J E; Mosjidis, J A; Burke, J M; Gujja, S; Mechineni, A; Terrill, T H

    2015-01-15

    Sericea lespedeza (SL; Lespedeza cuneata) is a perennial warm-season forage rich in condensed tannins (CT) that has been reported to have anthelmintic activity against small ruminant gastrointestinal nematodes (GIN), particularly Haemonchus contortus, a highly pathogenic blood-feeder, but the mechanism of action of CT against H. contortus is not clearly understood. An experiment with young goats was designed to study the effect of SL leaf meal pellets on (1) a mature H. contortus infection, and (2) the surface appearance of adult H. contortus female worms. Thirty-six female and castrated male Boer crossbred goats artificially infected with H. contortus larvae were fed 75% SL leaf meal pellets or alfalfa pellets (18 goats/treatment group) in a 28-day confinement feeding trial. Fecal and blood samples were collected weekly for fecal egg count (FEC) and packed cell volume (PCV) determination, respectively, and all goats were slaughtered at the end of the trial for adult GIN recovery and counting. Five adult female H. contortus were recovered from the abomasum of two goats from each treatment group and from a prior study in which 75% and 95% SL leaf meal pellets or a commercial feed pellet were group-fed to grazing goats (270 days old, Spanish males, 10/treatment group) at 0.91 kg/head/d for 11 weeks. Adult GIN collected were fixed and examined for evidence of surface damage using scanning electron microscopy. Feeding 75% SL pellets to young goats in confinement reduced (P<0.05) FEC compared with control animals, while total worm numbers and PCV were not influenced by treatment. Three out of the 5 adult H. contortus recovered from SL treatment goats in the confinement feeding trial had cuticular surface damage, while no damage was observed on worms from the control group. All five worms observed from both SL treatments in the grazing study showed a shrunken, disheveled cuticular surface, whereas this was not observed on worms from control animals. Overall, this work

  5. Study on the pathophysiology of Haemonchus contortus (Rudolph, 1803) in calves with the use of radioisotopes

    International Nuclear Information System (INIS)

    Gennari, S.M.

    1989-01-01

    This study was conducted to investigate the pathophysiology of Haemonchus contortus infection in friesian calves. The animals were divided in two groups of four calves each. One group was infected, orally, with 500 Haemonchus contortus larvae per kg body weight and the other one as the worm-free control. Five weeks after infection they were all housed in metabolic crates and injected with 125 I-bovine albumin, 51 Cr-red cell and 59 Fe-transferrin to study the albumin metabolism, the erythokinetic and ferrokinetic. The results showed that there was a significant reduction in the mean haematocrit values and an evidence of lower weight gains over the period of experiment in the infected calves compared with the controls. There was also a change in the distribution of albumin from the extravascular to the intravascular pool and a significant increase in the plasma and blood volumes of infected calves. The study of ferrokinetic showed an increase in the iron turnover and in the rate of iron utilization in the infected group. The blood and albumin loss by the gastrointestinal tract were similar in both groups. (author)

  6. Cross-infection between tropical goats and heifers with Haemonchus contortus.

    Science.gov (United States)

    d'Alexis, S; Mahieu, M; Jackson, F; Boval, M

    2012-03-23

    Developing effective biological control without the systematic use of anthelmintics is necessary to reduce the impact of gastrointestinal nematodes on small ruminants. Therefore, grazing management systems that use different host species to dilute nematodes in pasture appear to be promising for worm control. A trial was carried out to investigate the specificity of Haemonchus contortus for goats and cattle and to evaluate cross-infection between ruminant species. The effect of an experimental infection of 12 heifers by the free-living stages of H. contortus collected from goats (500 larvae per kg liveweight) was evaluated and compared to uninfected controls. After 28 and 35 days, egg excretion was measured. The experimental infection of heifers by H. contortus was not significant, with no egg excretion. These results, i.e., the lack of cross-infection of GIN between goats and cattle, suggest that integrated grazing using such animals could be employed for pasture dilution and decontamination. Copyright © 2011 Elsevier B.V. All rights reserved.

  7. Use of medicinal plants to control Haemonchus contortus infection in small ruminants

    Directory of Open Access Journals (Sweden)

    Sawleha Qadir1

    Full Text Available Haemonchus contortus is singly the most important of all the gastrointestinal nematodes that constrain the survival and productivity of sheep and goats owned by rural poor farmers in the developing world. This haematophagus parasite is infamous throughout the humid tropics/subtropics, being responsible for acute disease outbreaks with high levels of mortalities, particularly in young animals. Costs associated with control of this parasite in India, have been estimated to be US$ 103 million. H. contortus is also prominent amongst the reports of anthelmintic resistance that has emerged in all countries of the world that produce small ruminants. This emergence of multiple anthelmintic resistances has provided a spur for research on alternative forms of control. Recent surveys in developing countries have identified many plants that are intended and have the potential to be used as anthelmintics. This paper reviews the use of some medicinal plants as anthelmintics against H. contortus infection in small ruminants. [Veterinary World 2010; 3(11.000: 515-518

  8. In vitro anthelmintic effects of Spigelia anthelmia protein fractions against Haemonchus contortus.

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    Sandra Alves Araújo

    Full Text Available Gastrointestinal nematodes are a significant concern for animal health and well-being, and anthelmintic treatment is mainly performed through the use of chemical products. However, bioactive compounds produced by plants have shown promise for development as novel anthelmintics. The aim of this study is to assess the anthelmintic activity of protein fractions from Spigelia anthelmia on the gastrointestinal nematode Haemonchus contortus. Plant parts were separated into leaves, stems and roots, washed with distilled water, freeze-dried and ground into a fine powder. Protein extraction was performed with sodium phosphate buffer (75 mM, pH 7.0. The extract was fractionated using ammonium sulfate (0-90% and extensively dialyzed. The resulting fractions were named LPF (leaf protein fraction, SPF (stem protein fraction and RPF (root protein fraction, and the protein contents and activities of the fractions were analyzed. H. contortus egg hatching (EHA, larval exsheathment inhibition (LEIA and larval migration inhibition (LMIA assays were performed. Proteomic analysis was conducted, and high-performance liquid chromatography (HPLC chromatographic profiles of the fractions were established to identify proteins and possible secondary metabolites. S. anthelmia fractions inhibited H. contortus egg hatching, with LPF having the most potent effects (EC50 0.17 mg mL-1. During LEIA, SPF presented greater efficiency than the other fractions (EC50 0.25 mg mL-1. According to LMIA, the fractions from roots, stems and leaves also reduced the number of larvae, with EC50 values of 0.11, 0.14 and 0.21 mg mL-1, respectively. Protein analysis indicated the presence of plant defense proteins in the S. anthelmia fractions, including protease, protease inhibitor, chitinase and others. Conversely, secondary metabolites were absent in the S. anthemia fractions. These results suggest that S. anthelmia proteins are promising for the control of the gastrointestinal nematode H

  9. Antigenic Cross-reactivity among Haemonchus contortus, Oesophagostomum columbianum and Trichuris ovis of Goat

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    Ruma JAS

    2016-12-01

    Full Text Available Background: Cross antigenicity is the major problem in developing a reliable tool for immunodiagnosis and immunoprophylaxis of parasitic diseases. Mixed infection due to different types of gastrointestinal parasites is more common than single species infection under field condition.Methods: The present study was undertaken to detect antigenic cross-reactivity among Haemonchus contortus, Oesophagostomum columbianum and Trichuris ovis of goats by SDS-PAGE and western blot analysis using hyperimmune sera (HIS rose in rabbit separately against the antigens of the three nematode species.Results: Thirteen, 16 and 14 polypeptides in crude somatic antigen (CSAg of H. contortus (CSAg-Hc, O. columbianum (CSAg-Oc and T. ovis (CSAg-To, respectively, were resolved in SDS PAGE analyses. It was revealed that 54 kDa peptide was shared by H.contortus and O. columbianum, whereas 47 kDa peptide was shared by O. columbianum and T. ovis. Western blot analyses revealed that three immunogenic polypeptides (MW 54, 49 and 42 kDa in CSAg-Hc, five in CSAg-Oc (54, 47, 44, 38 and 35.5 kDa and CSAg-To and five polypeptides (90, 51, 47, 39.5 and 31 kDa in CSAg-To cross-reacted with the heterologous HIS. Four species-specific immunoreactive polypeptides (92, 85, 65 and 39 kDa of H. contortus and two (72 & 26 kDa in O. columbianum were also identified in the study. Conclusion: The shared polypeptides and species-specific polypeptides might be evaluated as protective antigen and subsequently exploitation for developing immunodiagnostic and for immunoprophylactic tools of for these common nematode species. 

  10. Antigenic Cross-reactivity among Haemonchus contortus, Oesophagostomum columbianum and Trichuris ovis of Goat.

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    Jas, Ruma; Ghosh, Joydeb; DAS, Kinsuk

    2016-01-01

    Cross antigenicity is the major problem in developing a reliable tool for immunodiagnosis and immunoprophylaxis of parasitic diseases. Mixed infection due to different types of gastrointestinal parasites is more common than single species infection under field condition. The present study was undertaken to detect antigenic cross-reactivity among Haemonchus contortus, Oesophagostomum columbianum and Trichuris ovis of goats by SDS-PAGE and western blot analysis using hyperimmune sera (HIS) rose in rabbit separately against the antigens of the three nematode species. Thirteen, 16 and 14 polypeptides in crude somatic antigen (CSAg) of H. contortus (CSAg-Hc), O. columbianum (CSAg-Oc) and T. ovis (CSAg-To), respectively, were resolved in SDS PAGE analyses. It was revealed that 54 kDa peptide was shared by H.contortus and O. columbianum , whereas 47 kDa peptide was shared by O. columbianum and T. ovis . Western blot analyses revealed that three immunogenic polypeptides (MW 54, 49 and 42 kDa) in CSAg-Hc, five in CSAg-Oc (54, 47, 44, 38 and 35.5 kDa) and CSAg-To and five polypeptides (90, 51, 47, 39.5 and 31 kDa) in CSAg-To cross-reacted with the heterologous HIS. Four species-specific immunoreactive polypeptides (92, 85, 65 and 39 kDa) of H. contortus and two (72 & 26 kDa) in O. columbianum were also identified in the study. The shared polypeptides and species-specific polypeptides might be evaluated as protective antigen and subsequently exploitation for developing immunodiagnostic and for immunoprophylactic tools of for these common nematode species.

  11. The potential to control Haemonchus contortus in indigenous South African goats with copper oxide wire particles.

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    Vatta, A F; Waller, P J; Githiori, J B; Medley, G F

    2009-06-10

    The high prevalence of resistance of Haemonchus contortus to all major anthelmintic groups has prompted investigations into alternative control methods in South Africa, including the use of copper oxide wire particle (COWP) boluses. To assess the efficacy of COWP against H. contortus in indigenous South African goats, 18 male faecal egg-count-negative goats were each given ca.1200 infective larvae of H. contortus three times per week during weeks 1 and 2 of the experiment. These animals made up an "established" infection group (ESTGRP). At the start of week 7, six goats were each given a 2-g COWP bolus orally; six goats received a 4-g COWP bolus each and six animals were not treated. A further 20 goats constituted a "developing" infection group (DEVGRP). At the beginning of week 1, seven of the DEVGRP goats were given a 2-g COWP bolus each; seven goats were treated with a 4-g COWP bolus each and no bolus was given to a further six animals. During weeks 1-6, each of these DEVGRP goats was given ca. 400 H. contortus larvae three times per week. All 38 goats were euthanized for worm recovery from the abomasa and small intestines in week 11. In the ESTGRP, the 2-g and 4-g COWP boluses reduced the worm burdens by 95% and 93%, respectively compared to controls (mean burden+/-standard deviation, SD: 23+/-33, 30+/-56 and 442+/-518 worms, P=0.02). However, in the DEVGRP goats, both the 2-g and 4-g COWP treatments were ineffective in reducing the worm burdens relative to the controls (mean burdens+/-SD: 1102+/-841, 649+/-855, 1051+/-661 worms, P=0.16). Mean liver copper levels did not differ between the ESTGRP goats treated with 2-g COWP, 4-g COWP or no COWP (mean+/-standard error of the mean, SEM, in ppm: 93.7+/-8.3; 101.5+/-8.3; 71.8+/-8.3, P=0.07) nor did they differ between the DEVGRP goats (mean+/-SEM, in ppm: 74.1+/-9.1; 75.4+/-9.1; 74.9+/-10.0, P>0.99). The copper values were considered adequate, but not high, for goats. The COWP boluses have the potential to be used

  12. Safety and serologic response to a Haemonchus contortus vaccine in alpacas.

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    VanHoy, Grace; Carman, Michelle; Habing, Greg; Lakritz, Jeffrey; Hinds, C Austin; Niehaus, Andrew; Kaplan, Ray M; Marsh, Antoinette E

    2018-03-15

    Haemonchosis in camelids remains a challenging disease to treat, and prevention has become increasingly problematic due to widespread anthelmintic resistance. Barbervax ® is an adjuvanted vaccine containing natural H-11, H-gal-GP antigens obtained from Haemonchus contortus adults via a proprietary process and solubilized in Quil A. This vaccine is approved for use in Australia, after demonstrating its safety and efficacy in sheep and goats. There are no published studies evaluating Barbervax in other ruminants/pseudoruminants such as camelids which can be parasitized with H. contortus. The vaccine utilizes a mixture of the parasite gut mucosal membrane enzymes including H-gal-GP and H11, involved in digesting a blood meal from the host. This study monitored the safety profile of the Barbervax ® vaccine in a group of adolescent alpacas. Although designed into the original study of vaccine efficacy, the experimental infection with viable H. contortus third stage larvae could not be completed due to lack of detectable significant variation of infection following experimental challenge. Twelve alpacas (158 + 15 days) were randomized to vaccination with Barbervax ® or no treatment. Three doses of Barbervax ® were administered at 3 week intervals and investigators involved in animal monitoring and sample collection were blinded to the groupings. Clinical pathologic parameters were evaluated 7 days before vaccination, and 1 and 2 months post-vaccination. Daily clinical observations were made and specific observations regarding the injection site and rectal temperatures were monitored in each alpaca twice daily for 1 week following vaccination. Fecal egg counts, packed cell volume, and total protein were monitored following challenge with 1500 H. contortus larvae on days 42, 46, and 50. An increase in rectal temperature for a duration of 2 days (range 2-4 days) was observed post-vaccination. Vaccinated alpacas were lethargic for 2-3 days following vaccination

  13. Anthelmintic activity of acetone extracts from South African plants used on egg hatching of Haemonchus contortus

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    Gerda Fouche

    2016-07-01

    Full Text Available The nematode, Haemonchus contortus, is responsible for major economic losses in the livestock industry. The management of parasites such as H. contortus has been through the use of synthetic parasiticides. This has resulted in the presence of residues in meat and milk, which affects food safety. The development of resistance to available anthelmintics coupled with their high cost has further complicated matters. This has led to the investigation of alternative methods to manage nematodes, including the use of plants and plant extracts as a potential source of novel anthelmintics. Acetone extracts were prepared from 15 South African plant species and their anthelmintic activity determined using the egg hatch assay (EHA. The leaf extract of Cleome gynandra had the best inhibitory activity (68% ± 3% at a concentration of 2.5 mg/mL, followed by the stem extract of Maerua angolensis (65% ± 5%. The extracts had a relatively low toxicity on Vero cells determined by the MTT (3-(4,5-dimethylthiazol-2-yl-2,5- diphenyltetrazolium bromide cellular assay.

  14. Early detection of Haemonchus contortus infection in sheep using three different faecal occult blood tests

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    A.V. Rodríguez

    2015-07-01

    Full Text Available Haemonchus contortus is a blood-sucking parasite causing the presence of faecal occult blood (FOB. The objective was to study three different FOB tests in order to have a new indicator of H. contortus infection in sheep that could be included in the genetic evaluation system as an alternative selection criterion to faecal worm egg count (FEC. A total of 29 Corriedale lambs were experimentally infected with 10.000 larvae of H. contortus. Stool samples were recorded for FEC and FOB tests (Hexagon, Hematest® and Multistix®, blood for packed cell volume (PCV, haemoglobin, white and red blood cell count (RBC, and FAMACHA© for scoring anaemia. At the end of the experiment lambs were slaughtered to worm burden count. Field infection was achieved in 309 Merino lambs under natural parasite challenge. FEC data were normalized through logarithmic transformation (LnFEC. Pearson correlation was estimated to examine the relationship between all traits. The three tests were able to detect the presence of FOB at day 11. FEC, PCV and RBC decreased to sub-normal values from day 18. FAMACHA© score 3 was considered to be indicative of anaemia. Most of the correlations were of high magnitude, with the exception of Multistix® test that was moderately correlated with haematological parameters, LnFEC and FEC. In field infection, most samples were negative to FOB tests and the correlations were lower than those calculated under experimental infection. In conclusion, FOB tests were able to detect haemonchosis earlier than FEC under high experimental parasite challenge. However, they were not able to detect FOB under natural mixed parasite challenge. FAMACHA© and PCV demonstrated to be good indicators of Haemonchosis, having moderate to high correlations with FEC.

  15. An in silico analysis of the glutamate dehydrogenases of Teladorsagia circumcincta and Haemonchus contortus

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    SIMON BROWN

    2014-04-01

    Full Text Available Nematode glutamate dehydrogenase (GDH amino acid sequences are very highly conserved (68-99% identity and are also very similar to those of the bovine and human enzymes (54-60% identity. The residues involved in binding nucleotides or substrates are completely conserved and tend to be located in highly conserved regions of the sequence. Based on the strong homology between the bovine, Teladorsagia circumcincta and Haemonchus contortus GDH sequences, models of the structure of the T. circumcincta and H. contortus monomers were constructed. The structure of the T. circumcincta monomer obtained using SWISS-MODEL was very similar to that of the bovine enzyme monomer and the backbone of the polypetide deviated very little from that of the bovine enzyme monomer. Despite the sequence differences between the bovine and T. circumcincta enzymes, the relative positions and orientations of the residues involved in ligand binding were very similar. The reported Km for NADP+ of T. circumcincta is about 35 and times that of the bovine enzyme, whereas the Kms of the two enzymes for glutamate, -ketoglutarate and NAD(PH are much more similar. The residue corresponding to S267 of the bovine enzyme is involved in binding the 2′-phosphate of NADP+ and is replaced in the T. circumcincta and H. contortus sequences by a tryptophan. The partial occlusion of the NAD(P-binding site by the tryptophan sidechain and the loss of at least one potential H-bond provided by the serine may explain the lower affinity of the T. circumcincta for NADP+.

  16. PREVALENCE OF HAEMONCHUS CONTORTUS IN NATURALLY INFECTED SMALL RUMINANTS GRAZING IN THE POTOHAR AREA OF PAKISTAN

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    F. R. CHAUDARY, M. F. U. KHAN1 AND M. QAYYUM

    2007-04-01

    Full Text Available The present study was carried out to investigate the prevalence and seasonal trend of the Haemonchus contortus in sheep and goats in the Potohar areas of northern Punjab, Pakistan from December 2004 to January 2006. Faecal samples collected from 968 sheep and 961 goats of different breeds were examined by the modified McMaster technique using saturated solution of sodium chloride. Results revealed that the infection was significantly (P<0.05 higher in sheep compared to goats. The peak infection level was recorded during rainy season (July-October. On the other hand, low infection level was noted from December upto May. In sheep, highest log faecal egg counts (LFECs were recorded in Islamabad, followed by Attock, Jhelum and Chakwal. However, in goats the LFECs trend was highest in Islamabad, followed by Jhelum, Attock and Chakwal districts. A significant (P<0.05 variability in LFECs was noted between sheep and goat breeds from site-site, while no significant difference was observed between breeds at the same site. Hairy (Jattal goats and Salt-Range (Latti sheep breeds exhibited significantly reduced LFECs level along with higher packed cell volume (PCV and haemoglobin (Hb levels compared to other breeds. Moreover, FAMACHA© chart scoring in relation with worm infection (FECs was more valid in sheep than goats. High prevalence of H. contortus in Potohar areas was due to favourable agro-climatic conditions that favour the development and survival of the free-living stages of H. contortus. The findings are discussed with regard to their relevance for strategic control of haemonchosis in small ruminants.

  17. Climate-driven changes to the spatio-temporal distribution of the parasitic nematode, Haemonchus contortus, in sheep in Europe.

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    Rose, Hannah; Caminade, Cyril; Bolajoko, Muhammad Bashir; Phelan, Paul; van Dijk, Jan; Baylis, Matthew; Williams, Diana; Morgan, Eric R

    2016-03-01

    Recent climate change has resulted in changes to the phenology and distribution of invertebrates worldwide. Where invertebrates are associated with disease, climate variability and changes in climate may also affect the spatio-temporal dynamics of disease. Due to its significant impact on sheep production and welfare, the recent increase in diagnoses of ovine haemonchosis caused by the nematode Haemonchus contortus in some temperate regions is particularly concerning. This study is the first to evaluate the impact of climate change on H. contortus at a continental scale. A model of the basic reproductive quotient of macroparasites, Q0 , adapted to H. contortus and extended to incorporate environmental stochasticity and parasite behaviour, was used to simulate Pan-European spatio-temporal changes in H. contortus infection pressure under scenarios of climate change. Baseline Q0 simulations, using historic climate observations, reflected the current distribution of H. contortus in Europe. In northern Europe, the distribution of H. contortus is currently limited by temperatures falling below the development threshold during the winter months and within-host arrested development is necessary for population persistence over winter. In southern Europe, H. contortus infection pressure is limited during the summer months by increased temperature and decreased moisture. Compared with this baseline, Q0 simulations driven by a climate model ensemble predicted an increase in H. contortus infection pressure by the 2080s. In northern Europe, a temporal range expansion was predicted as the mean period of transmission increased by 2-3 months. A bimodal seasonal pattern of infection pressure, similar to that currently observed in southern Europe, emerges in northern Europe due to increasing summer temperatures and decreasing moisture. The predicted patterns of change could alter the epidemiology of H. contortus in Europe, affect the future sustainability of contemporary

  18. Morphological variation between isolates of the nematode Haemonchus contortus from sheep and goat populations in Malaysia and Yemen.

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    Gharamah, A A; Rahman, W A; Siti Azizah, M N

    2014-03-01

    Haemonchus contortus is a highly pathogenic nematode parasite of sheep and goats. This work was conducted to investigate the population and host variations of the parasitic nematode H. contortus of sheep and goats from Malaysia and Yemen. Eight morphological characters were investigated, namely the total body length, cervical papillae, right spicule, left spicule, right barb, left barb, gubernaculum and cuticular ridge (synlophe) pattern. Statistical analysis showed the presence of morphological variation between populations of H. contortus from Malaysia and Yemen, with minor variation in the synlophe pattern of these isolates. Isolates from each country were grouped together in the scatterplots with no host isolation. Body, cervical papillae and spicule lengths were the most important characters that distinguished between populations of the two countries. This variation between Malaysia and Yemen may be attributed to geographical isolation and the possible presence of a different isolate of this worm in each country.

  19. The nutritional status affects the complete blood count of goats experimentally infected with Haemonchus contortus.

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    Cériac, S; Jayles, C; Arquet, R; Feuillet, D; Félicité, Y; Archimède, H; Bambou, J-C

    2017-11-09

    Gastrointestinal nematode (GIN) remains the most important pathogenic constraint of small ruminant production worldwide. The improvement of the host immune response against GIN though breeding for improved animal resistance, vaccination and nutritional supplementation appear as very promising methods. The objective of this study was to investigate the effect of four nutritional status differing in protein and energy levels (Hay: 5.1 MJ/Kg of dry matter (DM) and 7.6% of crude protein (CP), Ban: 8.3 MJ/Kg of DM and 7.5% of CP, Soy: 7.6 MJ/Kg of DM and 17.3% of CP, BS: 12.7 MJ/Kg of DM and 7.4% of CP) on the haematological disturbances due to Haemonchus contortus infection in Creole kid goats. No significant effect of the nutritional status was observed for faecal egg count (FEC) but the experimental infection induced haematological disturbances whose intensity and lengthening were dependent on the nutritional status. A transient marked regenerative macrocytic hypochromic anaemia as revealed by a decrease of packed cell volume (PCV), red blood cells (RBC) and hemoglobin and an increase of reticulocytes was observed in all infected groups except Hay. In this latter, the anaemia settled until the end of the experiment. Furthermore, H. contortus induced a thrombocytopenia significantly more pronounced in the group under the lowest nutritional status in term of protein (Hay and Ban). A principal component analysis revealed that the variables that discriminated the nutritional status were the average daily gain (ADG) and the PCV, considered as measures of the level of resilience to H. contortus infection. Moreover, the variables that discriminated infected and non-infected animals were mostly related to the biology of RBC (i.e. size and hemoglobin content) and they were correlated with FEC. The severity and the lengthening of the regenerative anaemia and the thrombocytopenia induced by H. contortus have been affected by the nutritional status. The protein enriched

  20. Effect of dietary supplementation on resistance to experimental infection with Haemonchus contortus in Creole kids.

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    Bambou, J C; Archimède, H; Arquet, R; Mahieu, M; Alexandre, G; González-Garcia, E; Mandonnet, N

    2011-06-10

    The aim of the present study was to test the effect of dietary supplementation on resistance to experimental infection with Haemonchus contortus in Creole kids. One trial with three replicates involved a total of 154 female kids that were chosen from three successive cohorts of the Creole flock of INRA-Gardel in 2007. The kids were placed into four treatments according to the amount of concentrate they received: G0 (no concentrate and a quality Dichantium spp. hay ad libitum, HAY), G1 (HAY+100g commercial concentrate d(-1)), G2 (HAY+200 g commercial concentrate d(-1)), G3 (HAY+300 g commercial concentrate d(-1)). The G0-G3 groups were infected with a single dose of 10,000 H. contortus third stage larvae (L(3)) at Day 0 (D0). Each infected group was comprised of one half resistant and one half susceptible genetically indexed kids. The average breeding values on egg excretion at 11 months of age were distant of 0.70, 0.65, 0.61 and 0.61 genetic standard deviations in G0, G1, G2 and G3, respectively. The faecal egg count (FEC), packed cell volume (PCV), eosinophilia (EOSI) and dry matter intake (DMI) indices were monitored weekly until 42 days post-infection. Enzyme-linked immunosorbent assay was carried out on serum samples to determine the level of IgA anti-H. contortus L(3) crude extracts and adult excretion/secretion products (ESP). The 10,000 L(3) dose received by the kids induced a severe infection: 8000 eggs per gram at the FEC peak, a PCV less than 15% and mortality. Interestingly, the supplemented animals in G3 showed a higher level of EOSI but a lower level of IgA anti-L3 and IgA anti-ESP than non-supplemented animals (G0). Resistant and susceptible kids had significantly different FEC variations within the groups. Susceptible kids had a 1.6 times higher egg output than resistant kids in G0. This difference was not found in the supplemented groups. The results of this study showed that supplementary feeding improved resistance of Creole kids to H. contortus

  1. Genetic diversity of Haemonchus contortus isolated from sympatric wild blue sheep (Pseudois nayaur) and sheep in Helan Mountains, China.

    Science.gov (United States)

    Shen, Dong-Dong; Wang, Ji-Fei; Zhang, Dan-Yu; Peng, Zhi-Wei; Yang, Tian-Yun; Wang, Zhao-Ding; Bowman, Dwight D; Hou, Zhi-Jun; Liu, Zhen-Sheng

    2017-09-19

    Haemonchus contortus is known among parasitic nematodes as one of the major veterinary pathogens of small ruminants and results in great economic losses worldwide. Human activities, such as the sympatric grazing of wild with domestic animals, may place susceptible wildlife hosts at risk of increased prevalence and infection intensity with this common small ruminant parasite. Studies on phylogenetic factors of H. contortus should assist in defining the amount of the impact of anthropogenic factors on the extent of sharing of agents such as this nematode between domestic animals and wildlife. H. contortus specimens (n = 57) were isolated from wild blue sheep (Pseudois nayaur) inhabiting Helan Mountains (HM), China and additional H. contortus specimens (n = 20) were isolated from domestic sheep that were grazed near the natural habitat of the blue sheep. Complete ITS2 (second internal transcribed spacer) sequences and partial sequences of the nad4 (nicotinamide dehydrogenase subunit 4 gene) gene were amplified to determine the sequence variations and population genetic diversities between these two populations. Also, 142 nad4 haplotype sequences of H. contortus from seven other geographical regions of China were retrieved from database to further examine the H. contortus population structure. Sequence analysis revealed 10 genotypes (ITS2) and 73 haplotypes (nad4) among the 77 specimens, with nucleotide diversities of 0.007 and 0.021, respectively, similar to previous studies in other countries, such as Pakistan, Malaysia and Yemen. Phylogenetic analyses (BI, MP, NJ) of nad4 sequences showed that there were no noticeable boundaries among H. contortus populations from different geographical origin and population genetic analyses revealed that most of the variation (94.21%) occurred within H. contortus populations. All phylogenetic analyses indicated that there was little genetic differentiation but a high degree of gene flow among the H. contortus populations among

  2. In vitro and in vivo Nematocidal Activity of Allium sativum and Tagetes erecta Extracts Against Haemonchus contortus.

    Science.gov (United States)

    Palacio- Landín, Josefina; Mendoza-de Gives, Pedro; Salinas-Sánchez, David Osvaldo; López-Arellano, María Eugenia; Liébano-Hernández, Enrique; Hernández-Velázquez, Victor Manuel; Valladares-Cisneros, María Guadalupe

    2015-12-01

    In the Mexican ethno-medicine, a number of plants have shown a successful anthelmintic activity. This fact could be crucial to identify possible green anti-parasitic strategies against nematodes affecting animal production. This research evaluated the in vitro and in vivo nematocidal effects of two single and combined plant extracts: bulbs of Allium sativum (n-hexane) and flowers of Tagetes erecta (acetone). The in vivo assay evaluated the administration of extracts either individually or combined against Haemonchus contortus in experimentally infected gerbils. The in vitro larvicidal activity percentage (LAP) of A. sativum and T. erecta extracts against H. contortus (L3) was determined by means of individual and combined usage of the extracts. Similarly, the extracts were evaluated in terms of reduction in the parasitic population in gerbils infected with H. contortus by individual and combined usage. The LAP at 40 mg/mL was 68% with A. sativum and 36.6% with T. erecta. The combination caused 83.3% mortality of parasites. The oral administration of A. sativum and T. erecta extracts at 40 mg/mL, caused 68.7% and 53.9% reduction of the parasitic burden, respectively. Meanwhile, the combined effect of both extracts shown 87.5% reduction. This study showed evidence about the effect of A. sativum and T. erecta plant extracts by means of individual and combined usage against H. contortus in in vitro and in vivo bioassays in artificially H. contortus-infected gerbils as a model.

  3. Ovicidal and larvicidal activity of the crude extracts from Phytolacca icosandra against Haemonchus contortus.

    Science.gov (United States)

    Hernández-Villegas, M M; Borges-Argáez, R; Rodriguez-Vivas, R I; Torres-Acosta, J F J; Méndez-Gonzalez, M; Cáceres-Farfan, M

    2011-06-30

    The development of anthelmintic resistance has impacted on the success of conventional anthelmintics (AH) for the control of gastrointestinal nematodes in grazing/browsing sheep and goats. Medicinal plants from the traditional herbolary in Mexico may provide new candidates that can be explored as alternative sources of AHs for ruminants. This study evaluated the leaf extracts derived from Phytolacca icosandra against infective L(3) larvae and eggs from Haemonchus contortus collected from sheep. Three extracts of different polarities were obtained from the leaf plants using ethanol, n-hexane and dichloromethane as the solvents. The effectiveness of the in vitro AH activity of the plant extracts was evaluated using larval migration inhibition (LMI) and egg hatch (EHA) assays. For the LMI assays, the ethanolic extract of P. icosandra showed 55.4% inhibition of larval migration at 2mg/mL (p<0.05). The dichloromethane extract of P. icosandra showed 67.1% inhibition of migration at 3mg/mL (p<0.05) and a dose-dependent response with an LD(50) of 0.90 mg/mL. The n-hexane extract failed to show inhibition of larval migration at any concentration explored. In the EHA for the ethanol extract, the lowest concentration tested (0.15 mg/mL) resulted in inhibition of egg hatching greater than 72.6%. Therefore, the LD(50) could not be calculated for this extract. The LD(50) of the dichloromethane extract of P. icosandra was 0. 28 mg/mL. An egg hatch inhibition greater than 90% was observed with both the ethanolic and dichloromethane extracts when using a concentration of 0.90 mg/mL or higher. The n-hexane extract failed to show egg hatch inhibition at any concentration tested. The AH activity reported for P. icosandra could be attributable to the flavonoids, steroids, terpenoids, coumarins and/or saponins that were present in the ethanolic and dichloromethane extracts. A combination of more than one component may also explain the observed AH activity against the H. contortus life

  4. [Ovicidal and larvicidal activity in vitro of Eucalyptus globulus essential oils on Haemonchus contortus].

    Science.gov (United States)

    Macedo, Iara T F; Bevilaqua, Claudia M L; de Oliveira, Lorena M B; Camurça-Vasconcelos, Ana L F; Vieira, Luiz da S; Oliveira, Fabrício R; Queiroz-Junior, Eudson M; Portela, Bruno G; Barros, Renata S; Chagas, Ana C S

    2009-01-01

    The objective of this work was to evaluate ovicidal and larvicidal effects of Eucalyptus globulus essential oil (EGEO) on Haemonchus contortus. The chemical composition determination of EGEO was through gas chromatography and mass spectrometry. Egg hatch test (EHT) was performed in concentrations 21.75; 17.4; 8.7; 5.43 e 2.71 mg x mL(-1). In larval development test (LDT) were used the concentrations 43.5; 21.75; 10.87; 5.43 e 2.71 mg x mL(-1). Each trial was conducted by negative control with Tween 80 (3%) and positive control, 0.02 mg x mL(-1) of thiabendazole in EHT and 0.008 mg x mL(-1) of ivermectin in LDT. The maximum effectiveness of EGEO on eggs was 99.3% in concentration of 21.75 mg x mL(-1) and on larvae was 98.7% in concentration 43.5 mg x mL(-1). The concentration of EGEO that inhibits 50% of the eggs and larvae was 8.3 and 6.92 mg x mL(-1), respectively. The oil chemical analysis identified as main component the monoterpen 1,8-cineol. EGEO presented ovicidal and larvicidal activities in vitro, revealing a good potential for use in the control of sheep and goat gastrointestinal nematodes.

  5. Piper aduncum against Haemonchus contortus isolates: cross resistance and the research of natural bioactive compounds

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    Yousmel Alemán Gaínza

    Full Text Available Abstract The anthelminthic activity of the essential oil (EO of Piper aduncum L. was tested in vitro on eggs and larvae of resistant (Embrapa2010 and susceptible (McMaster isolates of Haemonchus contortus. The EO was obtained by steam distillation and its components identified by chromatography. EO concentrations of 12.5 to 0.02 mg/mL were used in the egg hatch test (EHT and concentrations of 3.12 to 0.01 mg/mL in the larval development test (LDT. Inhibition concentrations (IC were determined by the SAS Probit procedure, and significant differences assessed by ANOVA followed by Tukey’s test. In the EHT, the IC50 for the susceptible isolate was 5.72 mg/mL. In the LDT, the IC50 and IC90 were, respectively, 0.10 mg/mL and 0.34 mg/mL for the susceptible isolate, and 0.22 mg/mL and 0.51 mg/mL for the resistant isolate. The EO (dillapiole 76.2% was highly efficacious on phase L1. Due to the higher ICs obtained for the resistant isolate, it was raised the hypothesis that dillapiole may have a mechanism of action that resembles those of other anthelmintic compounds. We further review and discuss studies, especially those conducted in Brazil, that quantified the major constituents of P. aduncum-derived EO.

  6. Piper aduncum against Haemonchus contortus isolates: cross resistance and the research of natural bioactive compounds.

    Science.gov (United States)

    Gaínza, Yousmel Alemán; Fantatto, Rafaela Regina; Chaves, Francisco Celio Maia; Bizzo, Humberto Ribeiro; Esteves, Sérgio Novita; Chagas, Ana Carolina de Souza

    2016-01-01

    The anthelminthic activity of the essential oil (EO) of Piper aduncum L. was tested in vitro on eggs and larvae of resistant (Embrapa2010) and susceptible (McMaster) isolates of Haemonchus contortus. The EO was obtained by steam distillation and its components identified by chromatography. EO concentrations of 12.5 to 0.02 mg/mL were used in the egg hatch test (EHT) and concentrations of 3.12 to 0.01 mg/mL in the larval development test (LDT). Inhibition concentrations (IC) were determined by the SAS Probit procedure, and significant differences assessed by ANOVA followed by Tukey's test. In the EHT, the IC50 for the susceptible isolate was 5.72 mg/mL. In the LDT, the IC50 and IC90 were, respectively, 0.10 mg/mL and 0.34 mg/mL for the susceptible isolate, and 0.22 mg/mL and 0.51 mg/mL for the resistant isolate. The EO (dillapiole 76.2%) was highly efficacious on phase L1. Due to the higher ICs obtained for the resistant isolate, it was raised the hypothesis that dillapiole may have a mechanism of action that resembles those of other anthelmintic compounds. We further review and discuss studies, especially those conducted in Brazil, that quantified the major constituents of P. aduncum-derived EO.

  7. Variability of resistance in Black Bengal goats naturally infected with Haemonchus contortus.

    Science.gov (United States)

    Kumar, Ratnesh; Ranjan, Sanjeev; Vishnu, P Guru; Negi, Mamta; Senapati, P K; Charita, V Gnani

    2015-03-01

    A total 290 Black Bengal goats (6 buck, 109 doe and 175 kids born from 11 sires) were studied to evaluate the variability of resistance in Black Bengal goats naturally infected with Haemonchus contortus. The variability of resistance in Black Bengal goat was studied for both genetic and non-genetic factors like village, sex, age dam, sire, dam resistance group and offspring resistance group. Male kids have slightly higher resistance than female kids although it was not significant. Resistance of kids was increased as age increases and kid population showed significantly different resistance status among the offspring resistant groups. The doe population showed significantly different LEPG as per the resistance group in all the collections. The present study found that the resistance of kids under sire were varied significantly and observed that the kids under sire 1, 6-8 were significantly more resistant than the kids of the sire 2, 5 and 11 in 3rd collection and it is also noticed that maternal genetic effect has a very little impact on resistance of kids. Males (buck) were most resistant and the kids were least resistant and the resistance of dam was in between the male and kids population.

  8. Effect of copper oxide wire particles dosage and feed supplement level on Haemonchus contortus infection in lambs.

    Science.gov (United States)

    Burke, J M; Miller, J E; Olcott, D D; Olcott, B M; Terrill, T H

    2004-09-02

    The objective of the experiment was to determine the optimal dose of copper oxide wire particles (COWPs) to reduce infection of Haemonchus contortus in male lambs. Five to six-month-old hair breed lambs were housed on concrete and fed 450 (L; n = 25) or 675 g (H; n = 25) corn/soybean meal supplement and bermudagrass hay. In July, lambs were inoculated with 10,000 L(3) larvae (97% H. contortus; Day 0). Lambs were administered 0, 2, 4, or 6 g COWP on Day 28. Concentrations of copper in the liver were determined. There were no effects of supplement level on concentrations of copper in the liver and a linear relationship existed between COWP treatment and concentrations of copper in liver (P copper in the liver of the COWP treatment groups. PCV values were more favorable for lambs fed the higher level of supplement, especially when FEC were greater than 8000 epg.

  9. Dried, ground banana plant leaves (Musa spp.) for the control of Haemonchus contortus and Trichostrongylus colubriformis infections in sheep.

    Science.gov (United States)

    Gregory, L; Yoshihara, E; Ribeiro, B L M; Silva, L K F; Marques, E C; Meira, E B S; Rossi, R S; Sampaio, P H; Louvandini, H; Hasegawa, M Y

    2015-12-01

    To evaluate the anthelmintic effect of Musa spp. leaves, 12 animals were artificially infected with Haemonchus contortus, and another 12 animals were infected with Trichostrongylus colubriformis. Then, both treatment groups were offered 400 g of dried ground banana plant leaves, and the control animals were offered only 1000 g of coast cross hay. During the trials, the animals received weekly physical examinations. The methods used to evaluate the efficiency of this treatment were packed cell volume, total plasma protein and faecal egg counts, and egg hatchability tests were performed on days -2, +3, +6, +9, +13 and +15. Coproculture tests were performed on day -2 to confirm monospecific infections. In the FEC and EHT, a statistically significant difference (0.04, 0.005; p  0.05) for Haemochus contortus group in all tests. Our results confirmed previous findings suggesting that dried ground banana plant leaves possess anthelmintic activity.

  10. Effectiveness of copper oxide wire particles for Haemonchus contortus control in sheep.

    Science.gov (United States)

    Knox, M R

    2002-04-01

    To assess the efficacy of copper oxide wire particles (COWP) for the control of H contortus infections in grazing sheep. In experiment 1, 40 worm-free Merino hoggets (11 to 12 months of age) were divided into four equal groups and allocated to separate 0.8 ha pasture plots. Two groups then received 2.5 g COWP whereas the other two groups were untreated. From 1 week after COWP treatment all lambs received a weekly infection of 2000 H contortus larvae. At week 8, six sheep from the untreated group were then allocated to two groups and treated with either 2.5 or 5.0 g of COWP to establish therapeutic efficacy of treatment. Experiment 2 followed a similar protocol but was conducted with 40 worm-free Merino lambs (3 to 4 months of age) and no assessment of therapeutic efficacy was made. In experiment 1 no significant difference in faecal worm egg counts was observed between treatments and faecal worm egg counts remained less than 3000 epg in all animals. Total worm counts were reduced by 37% by COWP treatment (P = 0.055). Both 2.5 g and 5.0 g doses of COWP at 8 weeks of infection reduced faecal worm egg counts by > 85% with the higher dose giving an earlier response to treatment. In experiment 2, faecal worm egg counts at 4 and 6 weeks were reduced by more than 90% in the COWP treated lambs and worm numbers were 54% lower after 6 weeks when all remaining untreated lambs had to be treated for haemonchosis. Mean faecal worm egg counts in the COWP lambs remained below 3500 epg and clinical disease did not develop in the majority of lambs before the end of the experiment at 10 weeks. Treatment with COWPs appears to have the potential to reduce establishment and worm fecundity of Haemonchus spp for an extended period and may offer livestock producers a supplementary means of reducing larval contamination of pasture particularly in areas where anthelmintic resistance is a problem and copper supplementation is likely to be beneficial.

  11. Persistence of the efficacy of copper oxide wire particles against Haemonchus contortus in sheep.

    Science.gov (United States)

    Galindo-Barboza, A J; Torres-Acosta, J F J; Cámara-Sarmiento, R; Sandoval-Castro, C A; Aguilar-Caballero, A J; Ojeda-Robertos, N F; Reyes-Ramírez, R; España-España, E

    2011-03-10

    The aim was to determine the persistent efficacy of copper oxide wire particles (COWP) against Haemonchus contortus in sheep, using the harmonization guidelines protocol. Thirty-six male lambs (2 months old) reared free of gastrointestinal nematodes were used (average body weight of 10.8±3.8kg). Before and for the duration of the study, lambs were kept in raised cages with slatted floors and were offered ad libitum a complete mixed diet. Animals were divided into six groups (n=6): one non-treated control group (G0) and five groups treated with one COWP capsule (1.7g of copper oxide; Copinox(®)). Animals in each group were treated on pre-defined dates before the artificial infection was applied: days -35 (G1), -28 (G2), -21 (G3), -14 (G4) and -7 (G5). On day 0 animals were infected with 3700 H. contortus infective larvae per animal. Animals were humanely slaughtered between days 22 and 23 post-infection. The abomasums were individually washed to obtain the contents. These organs were subjected to separate artificial digestions. Adult parasites were counted from the abomasum contents and the larvae from the digested material. Worm burden geometric means were calculated for each group. A significant worm burden reduction in either of the treated groups (G1, G2, G3, G4, and G5) compared to the control (G0) was considered as persistence of the anthelmintic effect. Copper levels were determined from individual liver samples of each animal. The geometric mean worm burden of the control group (G0) was 1959. Compared to the control, worm burdens geometric means were significantly reduced in groups G1 (1108), G4 (528) and G5 (1063) (P<0.03). Efficacies in G1, G4 and G5 were 43.4%, 73.0% and 45.7% respectively. No significant reduction was found for G2 (1342) and G3 (1430). A larger quantity of Cu was found in the livers of treated animals compared to the control group (P<0.05) except for G3 (P=0.06). A negative association between Cu liver content and worm burdens was

  12. Study on the irradiated vaccine of the haemonchus contortus worm in lamb

    Energy Technology Data Exchange (ETDEWEB)

    Soekardji; Arifin, M.; Murnihati, I. (National Atomic Energy Agency, Jakarta (Indonesia). Pasar Djumat Research Centre)

    1983-12-01

    The effect of /sup 60/Co gamma rays on the third larvae of haemonchus contortus has been studied in lambs. Inoculation of 1.000 irradiated L/sub 3/ from BPPH (Balai Penelitian Penyakit Hewan) Bogor strain (400, 500, 600 and 700 Gy) was done. The production of eggs were examined, also the effect of radiation on anaemia, and live weight gain of the lambs. The eggs production in every lamb was: control group (K)=116.0 +- 94.83; R/sub 1/ (radiation group at 400 Gy)=93.5 +- 64.05; R/sub 2/(radiation group at 500 Gy)=29.63 +- 19.67; R/sub 3/(radiation group at 600 Gy)=173.3 +- 181.34; and R/sub 4/(radiation group at 700 Gy)=125.2 +- 114.22. The specific clinical symptoms were anaemia; the data obtained were K=31.38 +- 0.49%; R/sub 1/=30.76 +- 1.21%; R/sub 2/=39.50 +- 2.27%; R/sub 3/=28.65 +- 0.92% and R/sub 4/=28.60 +- 1.44%. Average weight gain of the surviving animals was K=0.96 +- 0.49kg; R/sub 1/=3.82 +- 1.56kg; R/sub 2/=3.38 +- 0.84kg; R/sub 3/=1.57 +- 0.53kg; and R/sub 4/=1.99 +- 0.90kg. The experimental analysis of these data showed that the groups inoculated with irradiated L/sub 3/ compared with control were highly significant different, while among the 5 group were respectively highly significant.

  13. Introgression of ivermectin resistance genes into a susceptible Haemonchus contortus strain by multiple backcrossing.

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    Elizabeth Redman

    2012-02-01

    Full Text Available Anthelmintic drug resistance in livestock parasites is already widespread and in recent years there has been an increasing level of anthelmintic drug selection pressure applied to parasitic nematode populations in humans leading to concerns regarding the emergence of resistance. However, most parasitic nematodes, particularly those of humans, are difficult experimental subjects making mechanistic studies of drug resistance extremely difficult. The small ruminant parasitic nematode Haemonchus contortus is a more amenable model system to study many aspects of parasite biology and investigate the basic mechanisms and genetics of anthelmintic drug resistance. Here we report the successful introgression of ivermectin resistance genes from two independent ivermectin resistant strains, MHco4(WRS and MHco10(CAVR, into the susceptible genome reference strain MHco3(ISE using a backcrossing approach. A panel of microsatellite markers were used to monitor the procedure. We demonstrated that after four rounds of backcrossing, worms that were phenotypically resistant to ivermectin had a similar genetic background to the susceptible reference strain based on the bulk genotyping with 18 microsatellite loci and individual genotyping with a sub-panel of 9 microsatellite loci. In addition, a single marker, Hcms8a20, showed evidence of genetic linkage to an ivermectin resistance-conferring locus providing a starting point for more detailed studies of this genomic region to identify the causal mutation(s. This work presents a novel genetic approach to study anthelmintic resistance and provides a "proof-of-concept" of the use of forward genetics in an important model strongylid parasite of relevance to human hookworms. The resulting strains provide valuable resources for candidate gene studies, whole genome approaches and for further genetic analysis to identify ivermectin resistance loci.

  14. Gastrointestinal parasites of captive European bison Bison bonasus (L.) with a sign of reduced efficacy of Haemonchus contortus to fenbendazole.

    Science.gov (United States)

    Pyziel, Anna M; Björck, Sven; Wiklund, Rikard; Skarin, Moa; Demiaszkiewicz, Aleksander W; Höglund, Johan

    2018-01-01

    The history of European bison Bison bonasus Linnaeus, 1758 has been stormy since its extinction in the wild after the First World War. Due to the fact that the species was restored from just 12 founders, further expansion has suffered from low genetic variability, rendering the bison vulnerable to various pathogens due to inbreeding depression. Parasites are recognised as a key biological threat to bison population. Thus, parasitological examination including monitoring of the level of anthelmintic resistance in a herd should be a routine procedure involved in management and protection of European bison. This study was conducted in a group of 27 bison kept in a European bison breeding centre in Sweden. In April 2015, a faecal egg count reduction test (FECRT) was performed in animals with ≥ 100 gastrointestinal nematode (GIN) eggs per gram faeces, to determine effectiveness of fenbendazole (FBZ) treatment. Additionally, the third stage larvae were cultured for molecular examination by a conventional PCR as well as by real-time quantitative PCR (q-PCR) for detection of the blood-sucking nematode Haemonchus contortus. Faecal sampling was conducted 1 day before and 8 days after deworming each animal. Anthelmintic treatment turned to be entirely efficient toward intestinal nematodes of genera Nematodirus and Trichuris, whereas shedding of strongylid eggs from the subfamily Ostertagiinae was reduced from 81 to 30%. Polymerase chain reaction (PCR) on cultured third-stage larvae (L3) before treatment was positive for H. contortus, Ostertagia ostertagi and Cooperia oncophora, whereas post-treatment examination revealed exclusively the DNA of H. contortus. Thus, only H. contortus was involved in post-treatment faecal egg count (FEC). FECRT showed that the reduction in strongylid FEC to FBZ in the examined bison herd was 87% (95%-confidence intervals [95% CI] = 76-93), suggesting reduced efficacy of FBZ to strongylid GIN including mainly H. contortus.

  15. Survival and infectivity studies of in-vitro cultivated larvae of Haemonchus contortus in Sheep and Goats in Nigeria

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    A O Sonibare

    Full Text Available The survival and infectivity of indigenous isolates of Haemonchus contortus and in vitro cultivated infective larvae (L3 were studied. Active motile gravid females, adult and immature parasites were morphologically and morphometrically characterized and isolated from Haemonchus infected goats. These were subsequently inoculated into the abomasum of parasite free lambs through left sided laparatomy for production of fertile eggs to be used for coproculture. Harvested faeces from these lambs were cultured in a nutritive medium and kept in the incubator at 320C and room temperature 24-280C to obtain in vitro cultivated L of H. contortus. The infectivity and survival of in vitro cultivated L3 stored in refrigerator at 4-80C and under room temperature 24-280C were evaluated. Incubation period of 3 coproculture was observed to be shorter in medium at 320C than under room temperature 24-280C. The storage of in vitro cultivated L under different temperature range showed that the quantity of active larvae in a given volume 3 decreased at p<0.05 from the 5th day under 24-280C while at 4-80C became significantly lower from 20th day. Infectivity of isolate kept at 4-80C was sustained for 18-20 days while at 24-280C it was maintained for 10-12 days. Laboratory invitro cultivated L of H. contortus had limited infectivity and larvae mortality increase with time under different 3 temperatures. [Vet. World 2011; 4(12.000: 533-536

  16. Effects of commonly used chemical fertilizers on development of free-living stages of Haemonchus contortus in experimentally infected pasture

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    Tapas Kumar Roul

    2017-07-01

    Full Text Available Aim: The effects of N-P-K fertilizers in the form of urea, single super phosphate and muriate of potash on development of free-living stages of Haemonchus contortus were studied. Materials and Methods: Five parasite free experimental plots of 1 mx1 m area, each of paddy leaves (15-day-old and an equal number of Cynodon dactylon grass were infested with about 10x104 eggs/ml phosphate buffer saline along with the application of the calculated amount of fertilizers solution. On the 10th day of posttreatment, the pasture was cut, processed, larvae recovered by Baermann method and counted, which was expressed as number of L3 per kg dry matter (DM of pasture. Results: The average recovered population of L3 of H. contortus per kg DM varied significantly (p0.05. Conclusion: This study shown that when N-P-K fertilizers administered at recommended level, significantly reduced larval translation of H. contortus minimizing pasture infectivity for the free range grazing animals.

  17. Comparative in vitro efficacy of kefir produced from camel, goat, ewe and cow milk on Haemonchus contortus.

    Science.gov (United States)

    Alimi, D; Rekik, M; Akkari, H

    2018-04-18

    One of the great challenges of veterinary parasitology is the search for alternative methods for controlling gastrointestinal parasites in small ruminants. Milk kefir is a traditional source of probiotic, with great therapeutic potential. The objective of this study was to investigate the anthelmintic effects of kefir on the abomasal nematode Haemonchus contortus from sheep. The study used camel, goat, ewe and cow milk as a starting material, to produce camel, goat, cow and ewe milk kefir. All kefirs showed a significant concentration-dependent effect on H. contortus egg hatching at all tested concentrations. The highest inhibition (100%) of eggs was observed with camel milk kefir at a concentration 0.125 mg/ml. In relation to the effect of kefirs on the survival of adult parasites, all kefirs induced concentration-dependent mortality in adults, with variable results. The complete mortality (100%) of adults of H. contortus occurred at concentrations in the range 0.25-2 mg/ml. The highest inhibition of motility (100%) of worms was observed after 8 h post exposure with camel milk kefir at 0.25 mg/ml. These findings indicate that kefir can be considered a potential tool to control haemonchosis in sheep. Further investigations are needed to assess the active molecules in kefir responsible for its anthelmintic properties and to investigate similar in vivo effects.

  18. Biotransformation of albendazole and activities of selected detoxification enzymes in Haemonchus contortus strains susceptible and resistant to anthelmintics.

    Science.gov (United States)

    Vokřál, Ivan; Jirásko, Robert; Stuchlíková, Lucie; Bártíková, Hana; Szotáková, Barbora; Lamka, Jiří; Várady, Marián; Skálová, Lenka

    2013-09-23

    The increased activity of drug-metabolizing enzymes can protect helminths against the toxic effect of anthelmintics. The aim of this study was to compare the metabolism of the anthelmintic drug albendazole (ABZ) and the activities of selected biotransformation and antioxidant enzymes in three different strains of Haemonchus contortus: the ISE strain (susceptible to common anthelmintics), the BR strain (resistant to benzimidazole anthelmintics) and the WR strain (multi-resistant). H. contortus adults were collected from the abomasum of experimentally infected lambs. In vitro (subcellular fractions of H. contortus homogenate) as well as ex vivo (living nematodes cultivated in flasks with medium) experiments were performed. HPLC with spectrofluorimetric and mass-spectrometric detection was used in the analysis of ABZ metabolites. The in vitro activities of oxidation/antioxidation and conjugation enzymes toward model substrates were also assayed. The in vitro data showed significant differences between the susceptible (ISE) and resistant (BR, WR) strains regarding the activities of peroxidases, catalase and UDP-glucosyltransferases. S-oxidation of ABZ was significantly lower in BR than in the ISE strain. Ex vivo, four ABZ metabolites were identified: ABZ sulphoxide and three ABZ glucosides. In the resistant strains BR and WR, the ex vivo formation of all ABZ glucosides was significantly higher than in the susceptible ISE strain. The altered activities of certain detoxifying enzymes might partly protect the parasites against the toxic effect of the drugs as well as contribute to drug-resistance in these parasites. Copyright © 2013 Elsevier B.V. All rights reserved.

  19. Comparison of two in vitro methods for the detection of ivermectin resistance in Haemonchus contortus in sheep

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    Urda Dolinská M.

    2016-06-01

    Full Text Available Gastrointestinal parasitic nematodes in sheep cause severe economic losses. Anthelmintics are the most commonly used drugs for prophylaxis and therapy against parasitic helminths. The problem of drug resistance has developed for all commercially available anthelmintics in several genera and classes of helminths. In vitro and in vivo tests are used to detect anthelmintic resistance. Two in vitro methods (larval migration inhibition test and micromotility test for the detection of ivermectin (IVM resistance were compared using IVM-resistant and IVM-susceptible isolates of Haemonchus contortus. The degree of resistance for each test was expressed as a resistance factor (RF. The micromotility test was more sensitive for quantitatively measuring the degree of resistance between susceptible and resistant isolates. The RFs for this test for IVM and eprinomectin ranged from 1.00 to 108.05 and from 3.87 to 32.32, respectively.

  20. The efficacy of a slow-release albendazole capsule against Haemonchus contortus with known resistance to albendazole

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    Alan D. Fisher

    2013-12-01

    Full Text Available Controlled-release albendazole capsules (CRCs are currently registered for use in Australia and New Zealand as anthelmintic treatment in sheep. However, reports on the efficacy of such products on resistant parasite populations are sometimes controversial. This is the first study to report on the efficacy of such products under South African field conditions in sheep harbouring a population of Haemonchus contortus with known multiple anthelmintic resistance, including to albendazole. Treatment groups were comprised of CRC-treated and single dose albendazole-treated sheep, as well as negative controls. Groups were compared by using faecal egg count reduction tests, FAMACHA© anaemia scoring, conception rates and comparative weight gains over three and a half months. Based on a comparison of faecal egg counts, no advantage could be found using CRCs. Moreover, the use of the product actually decreased weight gain when compared with the control group animals.

  1. Feed resource selection of Criollo goats artificially infected with Haemonchus contortus: nutritional wisdom and prophylactic self-medication.

    Science.gov (United States)

    Ventura-Cordero, J; González-Pech, P G; Jaimez-Rodriguez, P R; Ortiz-Ocampo, G I; Sandoval-Castro, C A; Torres-Acosta, J F J

    2018-06-01

    Previous cafeteria studies suggested that a moderate natural gastrointestinal nematode (GIN) infection did not modify the resource selection of adult Criollo goats towards tannin-rich plants compared with worm-free goats. A higher infection with Haemonchus contortus could trigger a change in the resource selection behaviour towards tannin-rich foliage. Alternatively, goats might select plant species solely to meet their nutritional requirements. A cafeteria study investigated the effect of a high artificial infection with H. contortus on the feed resource selection of goats. Adult Criollo goats (37.5±4.8 kg BW) with browsing experience were distributed in two groups: the infected group (IG) with six animals artificially infected with H. contortus (6000 L3/animal); and the non-infected group (NIG) with six animals maintained worm-free. The experiment included two 5-day periods with additional 5-day adaptation period. In the first period, animals were offered foliage of five plant species with a decreasing gradient of condensed tannins (CT) (Mimosa bahamensis, Gymnopodium floribundum, Havardia albicans, Acacia pennatula, Lysiloma latisiliqum), and three plant species with negligible CT content (Leucaena leucocephala, Piscidia piscipula and Brosimum alicastrum). In the second period the foliage of B. alicastrum was withdrawn. A grain-based concentrate feed was offered daily at 1% BW in DM basis. Dry matter and nutrient intake was determined. Foliage selection of each experimental group was determined using the Chesson selection index. The H. contortus egg count per gram of faeces (EPG) was determined for infected goats twice daily. Chesson index showed a similar pattern of foliage selection on periods 1 and 2. Mean EPG of goats in IG was 2028±259 EPG during period 1 and 1 293±198 EPG during period 2 (P>0.05). During period 1, the selection pattern was highest for B. alicastrum (tannin-free), followed by a tannin-rich plant (M. bahamensis). These two plants remained

  2. In vitro anthelmintic activity of five tropical legumes on the exsheathment and motility of Haemonchus contortus infective larvae.

    Science.gov (United States)

    von Son-de Fernex, Elke; Alonso-Díaz, Miguel Angel; Valles-de la Mora, Braulio; Capetillo-Leal, Concepción M

    2012-08-01

    This study investigated the in vitro anthelmintic (AH) activity of five tropical legume plants [Arachis pintoi CIAT 22160 (A.p. 22160), Gliricidia sepium, Cratylia argentea (C.a. Yacapani), C. argentea CIAT 22386 (C.a. 22386), C. argentea Veranera (C.a. Veranera)] against Haemonchus contortus infective larvae and the role of tannins/polyphenolic compounds in the AH effect. Lyophilized leaf extracts of each plant were evaluated using the Larval Exsheathment Inhibition Assay (LEIA) and the larval migration inhibition assay (LMIA). The role of tannins/polyphenolic compounds in the AH effect was evaluated in both assays using polyethylene glycol (PEG) to remove tannins from the solutions. At the highest concentration (1200μg of extract/ml), A. pintoi 22160, C.a. Yacapani, C.a. Veranera and C.a. 22386 completely inhibited the exsheathment process of H. contortus (P<0.01). At the same concentration (1200μg of extract/ml), the inhibition of larval migration for C.a. 22386, C.a. Veranera and G. sepium was 66.0%, 35.9% and 39.2% (relative to the PBS control), respectively. In both bioassays (LEIA and LMIA), the AH effect shown by each plant was blocked after the addition of polyethylene glycol (PEG), corroborating the role of tannins/polyphenolic compounds. Copyright © 2012 Elsevier Inc. All rights reserved.

  3. Duddingtonia flagrans chlamydospores in nutritional pellets: effect of storage time and conditions on the trapping ability against Haemonchus contortus larvae.

    Science.gov (United States)

    Fitz-Aranda, J A; Mendoza-de-Gives, P; Torres-Acosta, J F J; Liébano-Hernández, E; López-Arellano, M E; Sandoval-Castro, C A; Quiroz-Romero, H

    2015-01-01

    The study evaluated the effect of storage time and conditions of nutritional pellets (NP) containing Duddingtonia flagrans chlamydospores on its in vitro trapping ability against Haemonchus contortus L3 larvae. The treated batch (200 NP) contained 4 × 106 chlamydospores of the FTH0-8 strain, whereas the control batch (200 NP) was produced without spores. Both NP batches were exposed to four experimental storage conditions: (T1) shelves (indoors); (T2) refrigeration (4°C); (T3) outdoors under a roof; and (T4) 100% outdoors. Each group comprised 48 NP with spores and 48 NP without spores (control). The ability of D. flagrans spores to trap H. contortus L3 larvae was evaluated for 8 weeks for each storage condition. For that purpose, six randomly selected NP with spores were compared to their respective control NP. Each NP was individually crushed. The crushed material (1 g) was placed on the surface of a 2% water agar plate with 200 H. contortus L3 larvae. Plates were sealed and were incubated at room temperature for 8 days. The whole content of every plate was transferred to a Baermann apparatus to recover the remaining larvae. There was a clear larval reduction in the NP with spores, compared to the respective control NP in the four storage conditions (PT2), 77 ± 6.1 (T1), 81.5 ± 3.8 (T4) and 82.1 ± 2.5 (T3). Larval reductions were similar at all times and were not affected by storage conditions or storage time (R 20.05). The long-term shelf-life of the chlamydospores in the NP suggests that this spore dosage technology is a viable option.

  4. Impact of the post-weaning parasitism history on an experimental Haemonchus contortus infection in Creole goat kids.

    Science.gov (United States)

    Ceï, W; Mahieu, M; Philibert, L; Arquet, R; Alexandre, G; Mandonnet, N; Bambou, J C

    2015-01-15

    Gastrointestinal nematode (GIN) infections have an important negative impact on small ruminant production. The selection of genotypes resistant to these parasitic infections is a promising alternative control strategy. Thus, resistance against GIN is an important component of small ruminant breeding schemes, based on phenotypic measurements of resistance in immune mature infected animals. In this study we evaluated both the impact of the post-weaning parasitism history on the response to an experimental Haemonchus contortus infection of resistant and susceptible Creole kids chosen on the basis of their estimated breeding value, and the interaction with the kid's genetic status. During the post-weaning period (from 3 months until 7 months of age) Creole kids were reared at pasture according to four different levels of a mixed rotational stocking system with Creole cattle: 100% (control), 75% (GG75), 50% (GG50), and 25% (GG25) of the total stocking rate of the pasture. The level of infection of the kids decreased significantly at 50% and 25% of the total stocking rate. After the post-weaning period at pasture, at 11 months of age kids were experimentally infected with H. contortus. The faecal egg counts (FEC) were significantly lower in the groups showing the highest FEC at pasture. This result suggests that a degree of protection against an experimental H. contortus infection occurred during the post-weaning period and was dependant on the level of parasitism. Interestingly, no interaction was observed between this level of protection and the genetic status. In conclusion, the level of post-weaning natural parasitism history at pasture would not influence the genetic status evaluation. More generally our results suggest that it would be better to expose kids to a high level of gastrointestinal parasitism during the post-weaning period in order to increase the basal level of resistance thereafter. Copyright © 2014 Elsevier B.V. All rights reserved.

  5. E-ADA activity in erythrocytes of lambs experimentally infected with Haemonchus contortus and its possible functional correlations with anemia.

    Science.gov (United States)

    Da Silva, Aleksandro S; Schafer, Andressa S; Aires, Adelina R; Tonin, Alexandre A; Pimentel, Victor C; Oliveira, Camila B; Zanini, Daniela; Schetinger, Maria R C; Lopes, Sonia T A; Leal, Marta L R

    2013-12-01

    The aim of this study was to evaluate the ecto-adenosine deaminase (E-ADA) activity in erythrocytes of lambs experimentally infected with Haemonchus contortus, correlating it with the degrees of anemia of the experimental animals. A total of 14 healthy lambs, with negative fecal exam for parasites, were to carry out the present study. They were divided into two groups, composed by seven animals: Group A represented the healthy animals (uninfected), while in Group B the animals were infected with 15,000 larvae of H. contortus. Blood was drawn on the days 15, 45 and 75 post-infection (PI) in order to perform the hematological analysis, as well as the mensuration of E-ADA activity in erythrocytes. Parasitological stool exam were performed on the same days mentioned above to follow up the evolution of the infection, as well to determine the number of eggs per gram of feces (EPG). On day 15PI, the animals presented negative EPG and there was not significant (P>0.05) difference between groups in relation to E-ADA activity and hematologic parameters. Animals in Group B had positive EPG for helminths on days 45 and 75 PI, accompanied by varying degrees of anemia, when compared to Group A. At the same periods E-ADA activity was significantly (PADA in erythrocytes and hematocrit on days 45 (r = -0.76) and 75 (r = -0.85)PI. Based on these results and in the scientific literature, it is possible to conclude that the E-ADA may participate on mechanisms related with the pathogenesis and host response against anemia caused by H. contortus. Copyright © 2013 Elsevier Ltd. All rights reserved.

  6. In vitro anthelmintic activity of Combretum molle (R. Br. ex G. Don) (Combretaceae) against Haemonchus contortus ova and larvae.

    Science.gov (United States)

    Ademola, I O; Eloff, J N

    2010-04-19

    Parasitic nematodes, especially Haemonchus contortus (Rudolphi), are among the most common and economically important causes of disease in sheep and goats owned by pastoralists and small holder farmers in Africa. The control of these infections relies mainly on the use of anthelmintic drugs. However, herbal preparations are widely used by pastoralists and small holder farmers for the treatment of their livestock against helminth parasites. The anthelmintic effect of acetone leaf extract and fractions of Combretum molle was investigated to determine the relative efficacy of the components against gastrointestinal sheep nematodes. The fractions were obtained by solvent:solvent extraction from the acetone extract. These were evaluated for nematocidal activity by means of an egg hatch (EHA) and larval a development and viability assay (LDVA) in vitro. The effect of the test extracts on the hatchability of eggs and development of first to third stage larvae and the survival rate of the third stage larvae. H. contortus, were used to determine the relative bioactivities. Best-fit LC(50) values were computed using global model of nonlinear regression curve-fitting. The extracts inhibited egg hatching and development of the larvae of H. contortus in a concentration-dependent manner. Best-fit LC(50) values for the egg hatch test were 0.866, 0.333, 0.833, 0.747, and 0.065mg/mL for acetone extract, n-butanol, hexane, chloroform, and 35% water in methanol fractions, respectively. The best-fit LC(50) values for the LDVA were 0.604, 0.362, 1.077, 0.131 and 0.318mg/mL for the acetone extract, butanol, hexane, chloroform, and 35% water in methanol fractions, respectively. In the EHA the 35% water in methanol fraction was significantly more active than all the other fractions (pmolle leaf could find application in anthelmintic therapy in veterinary practice.

  7. In vitro screening of six anthelmintic plant products against larval Haemonchus contortus with a modified methyl-thiazolyl-tetrazolium reduction assay.

    Science.gov (United States)

    Hördegen, P; Cabaret, J; Hertzberg, H; Langhans, W; Maurer, V

    2006-11-03

    Because of the increasing anthelmintic resistance and the impact of conventional anthelmintics on the environment, it is important to look for alternative strategies against gastrointestinal nematodes. Phytotherapy could be one of the major options to control these pathologies. Extracts or ingredients of six different plant species were tested against exsheathed infective larvae of Haemonchus contortus using a modified methyl-thiazolyl-tetrazolium (MTT) reduction assay. Pyrantel tartrate was used as reference anthelmintic. Bromelain, the enzyme complex of the stem of Ananas comosus (Bromeliaceae), the ethanolic extracts of seeds of Azadirachta indica (Meliaceae), Caesalpinia crista (Caesalpiniaceae) and Vernonia anthelmintica (Asteraceae), and the ethanolic extracts of the whole plant of Fumaria parviflora (Papaveraceae) and of the fruit of Embelia ribes (Myrsinaceae) showed an anthelmintic efficacy of up to 93%, relative to pyrantel tartrate. Based on these results obtained with larval Haemonchus contortus, the modified MTT reduction assay could be a possible method for testing plant products with anthelmintic properties.

  8. In vitro screening of six anthelmintic plant products against larval Haemonchus contortus with a modified methyl-thiazolyl-tetrazolium reduction assay

    OpenAIRE

    Hördegen, P.; Cabaret, J.; Hertzberg, H.; Langhans, W.; Maurer, V.

    2006-01-01

    Because of the increasing anthelmintic resistance and the impact of conventional anthelmintics on the environment, it is important to look for alternative strategies against gastrointestinal nematodes. Phytotherapy could be one of the major options to control these pathologies. Extracts or ingredients of six different plant species were tested against exsheathed infective larvae of Haemonchus contortus using a modified methyl-thiazolyltetrazolium (MTT) reduction assay. Pyrantel tartrate was u...

  9. Differences in transcription between free-living and CO2-activated third-stage larvae of Haemonchus contortus

    Directory of Open Access Journals (Sweden)

    Zhong Weiwei

    2010-04-01

    Full Text Available Abstract Background The disease caused by Haemonchus contortus, a blood-feeding nematode of small ruminants, is of major economic importance worldwide. The infective third-stage larva (L3 of this gastric nematode is enclosed in a cuticle (sheath and, once ingested with herbage by the host, undergoes an exsheathment process that marks the transition from the free-living (L3 to the parasitic (xL3 stage. This study explored changes in gene transcription associated with this transition and predicted, based on comparative analysis, functional roles for key transcripts in the metabolic pathways linked to larval development. Results Totals of 101,305 (L3 and 105,553 (xL3 expressed sequence tags (ESTs were determined using 454 sequencing technology, and then assembled and annotated; the most abundant transcripts encoded transthyretin-like, calcium-binding EF-hand, NAD(P-binding and nucleotide-binding proteins as well as homologues of Ancylostoma-secreted proteins (ASPs. Using an in silico-subtractive analysis, 560 and 685 sequences were shown to be uniquely represented in the L3 and xL3 stages, respectively; the transcripts encoded ribosomal proteins, collagens and elongation factors (in L3, and mainly peptidases and other enzymes of amino acid catabolism (in xL3. Caenorhabditis elegans orthologues of transcripts that were uniquely transcribed in each L3 and xL3 were predicted to interact with a total of 535 other genes, all of which were involved in embryonic development. Conclusion The present study indicated that some key transcriptional alterations taking place during the transition from the L3 to the xL3 stage of H. contortus involve genes predicted to be linked to the development of neuronal tissue (L3 and xL3, formation of the cuticle (L3 and digestion of host haemoglobin (xL3. Future efforts using next-generation sequencing and bioinformatic technologies should provide the efficiency and depth of coverage required for the determination of the

  10. A report on the metabolism of iron in goats artificially infected with Haemonchus contortus

    International Nuclear Information System (INIS)

    Perpuse, W.G.; Yumul, B.Y.; Anden, A.

    1976-03-01

    The determination of iron metabolism in goats artificially infected with N. contortus using tracer method has been conducted. Radioferric chloride ( 59 Fe) was given orally and intravenously and the distribution and utilization in goats were determined. Results showed that anemic goats have a higher absorption rate compared to non-anemic goats. Of all the organs examined the bone marrow showed the highest activity

  11. Anthelmintic efficacy of five tropical native Australian plants against Haemonchus contortus and Trichostrongylus colubriformis in experimentally infected goats (Capra hircus).

    Science.gov (United States)

    Moreno, F C; Gordon, I J; Knox, M R; Summer, P M; Skerrat, L F; Benvenutti, M A; Saumell, C A

    2012-06-08

    The study of the anthelmintic properties of plants rich in plant secondary metabolites can provide ecologically sound methods for the treatment of parasites on grazing animals. The purpose of the present study was to evaluate the anthelmintic effect of five tropical native Australian plant species rich in plant secondary metabolites on adult Haemonchus contortus and Trichostrongylus colubriformis in experimentally infected goats. Thirty young, nematode-free goats were infected with 2500 H. contortus and 5000 T. colubriformis infective larvae thrice weekly for a week (day 1-7 of the experiment). On day 27 after first infection, the goats were allocated into six groups of five animals per group. From day 28 to day 35, fresh leaves from Acacia salicina, Acacia nilotica, Eucalyptus corymbia, Casuarina cunninghamiana and Eucalyptus drepanophylla were included in the goats diet. Five groups were offered leaves from one of these plant species and one group, the untreated control, received only the basal diet formulated with 20% Medicago sativa and 80% Avena sativa. Following plant material administration, the goats were monitored daily until day 40 and then slaughtered on day 41. Total faecal worm egg output, total production of larvae recovered from faecal cultures, total post-mortem worm burdens and the per capita fecundity of female worms were estimated. The toxicity of the plant species for the goats was measured by histopathological analyses of liver and kidney samples. Results showed that goats feeding on the plant material rich in plant secondary metabolites had significantly lower egg output compared to the control goats (P0.05), the per capita fecundity was significantly reduced by E. corymbia, A. nilotica and A. salicina (Pgoats can benefit from the short-term ingestion of plant secondary metabolites, which reduce the total faecal egg output and thus decrease the potential for re-infection from the pasture. Crown Copyright © 2012. Published by Elsevier B.V. All

  12. Persistence of the efficacy of copper oxide wire particles against Haemonchus contortus in grazing South African goats.

    Science.gov (United States)

    Vatta, A F; Waller, P J; Githiori, J B; Medley, G F

    2012-11-23

    A study was conducted to examine the duration of anthelmintic effect of copper oxide wire particles (COWP) in grazing goats, as data for the persistence of efficacy of COWP in this host species is limited. Forty-eight indigenous male goats were infected naturally by grazing them on Haemonchus contortus-infected pasture. When the faecal egg count (FEC) in the goats was 3179 ± 540 eggs per gram of faeces (mean ± standard error), half the animals were treated with 4 g COWP (day 0; mean live weight=25.5 ± 0.8 kg). Eight treated (COWP) and eight non-treated (CONTROL) goats were removed from the pasture on each of days 7, 28 and 56, maintained for 27 or 29 days in concrete pens and then humanely slaughtered for nematode recovery. Mean liver copper levels were in the high range in the goats removed from pasture at day 7 (treated: 191 ± 19.7 ppm; untreated: 120 ± 19.7 ppm; P=0.022), but had dropped to normal levels at days 28 and 56. The mean H. contortus burdens of the treated versus the non-treated goats were, respectively, 184 ± 48 and 645 ± 152 for the goats removed from pasture at day 7 (71% reduction; P=0.004), 207 ± 42 and 331 ± 156 at day 28 (37% reduction; P=0.945) and 336 ± 89 and 225 ± 53 at day 56 (-49% reduction; P=0.665). Weekly monitoring of FECs after treatment until slaughter indicated that the COWP-treated goats had lower FECs than the controls, the treatment main effect being significant at days 7, 28 and 56 (Pcopper levels return to normal two to three months after COWP treatment. Copyright © 2012 Elsevier B.V. All rights reserved.

  13. Phenobarbital induction and chemical synergism demonstrate the role of UDP-glucuronosyltransferases in detoxification of naphthalophos by Haemonchus contortus larvae.

    Science.gov (United States)

    Kotze, Andrew C; Ruffell, Angela P; Ingham, Aaron B

    2014-12-01

    We used an enzyme induction approach to study the role of detoxification enzymes in the interaction of the anthelmintic compound naphthalophos with Haemonchus contortus larvae. Larvae were treated with the barbiturate phenobarbital, which is known to induce the activity of a number of detoxification enzymes in mammals and insects, including cytochromes P450 (CYPs), UDP-glucuronosyltransferases (UDPGTs), and glutathione (GSH) S-transferases (GSTs). Cotreatment of larvae with phenobarbital and naphthalophos resulted in a significant increase in the naphthalophos 50% inhibitory concentration (IC50) compared to treatment of larvae with the anthelmintic alone (up to a 28-fold increase). The phenobarbital-induced drug tolerance was reversed by cotreatment with the UDPGT inhibitors 5-nitrouracil, 4,6-dihydroxy-5-nitropyrimidine, probenecid, and sulfinpyrazone. Isobologram analysis of the interaction of 5-nitrouracil with naphthalophos in phenobarbital-treated larvae clearly showed the presence of strong synergism. The UDPGT inhibitors 5-nitrouracil, 4,6-dihydroxy-5-nitropyrimidine, and probenecid also showed synergistic effects with non-phenobarbital-treated worms (synergism ratio up to 3.2-fold). This study indicates that H. contortus larvae possess one or more UDPGT enzymes able to detoxify naphthalophos. In highlighting the protective role of this enzyme group, this study reveals the potential for UDPGT enzymes to act as a resistance mechanism that may develop under drug selection pressure in field isolates of this species. In addition, the data indicate the potential for a chemotherapeutic approach utilizing inhibitors of UDPGT enzymes as synergists to increase the activity of naphthalophos against parasitic worms and to combat detoxification-mediated drug resistance if it arises in the field. Copyright © 2014, American Society for Microbiology. All Rights Reserved.

  14. Evaluation of Copper Supplementation to Control Haemonchus contortus Infections of Sheep in Sweden

    Directory of Open Access Journals (Sweden)

    Rydzik A

    2004-09-01

    Full Text Available A pen study was conducted to assess the effect of providing daily copper mineral supplement, or copper wire particle (COWP capsules, on established or incoming mixed nematode infections in young sheep. For lambs with established (6 week old infections, COWP resulted in 97% and 56% reduction of the adult and early L4 stages of H. contortus, respectively, compared with controls (p Teladorsagia circumcincta infections in the COWP lambs compared with controls (p H. contortus infections, but lack of parasites during the grazing season prevented an adequate assessment from being made. These results indicate that there is little, if any, benefit from a parasite control standpoint in recommending copper therapy, specifically to control parasites in Swedish sheep flocks.

  15. Single vs. double dose of copper oxide wire particles (COWP) for treatment of anthelmintic resistant Haemonchus contortus in weanling lambs.

    Science.gov (United States)

    Schweizer, Nikki M; Foster, Derek M; Knox, William B; Sylvester, Hannah J; Anderson, Kevin L

    2016-10-15

    Haemonchus contortus parasitism is a major disease of sheep, with these parasites frequently demonstrating multi-drug class anthelmintic resistance. Copper oxide wire particles (COWP) have shown potential as adjuncts or alternatives to anthelmintics in resistant flocks. The purpose of this study was to compare the efficacy of two different COWP treatment regimens or placebo in the control of H. contortus in weaned lambs within a flock historically shown to have multi-drug resistant H. contortus using the DrenchRite ® assay. Data from 43 lambs within 3 treatment groups in a double blind study were included in the experiment. Treatments were administered as a total of 2 boluses, each given on separate occasions (day 0 and day 42), so that each lamb received either 2 placebos, a single dose of 2g COWP followed by placebo, or two doses of 1g COWP. Strongyle-type fecal egg counts (FEC) were performed at initial treatment (day 0), on day 10, at second treatment (day 42), on day 52, and at study end (day 84). At the start of the trial, mean±standard deviation FEC were 1634.4±825.2, 2241.7±1496.8, and 2013.3±1194.2epg for the 2g, 1g×2, and control groups, respectively. At the end of the trial, FEC were 757.1±825.3, 483.4±557.2, and 1660.0±1345.3epg for the 2g, 1g×2, and control groups, respectively. Lambs given a 2g single dose of COWP or a 1g dose of COWP twice had reductions in strongyle-type FEC (p≤0.01) from trial start to trial end, whereas lambs given placebo did not. Average daily gains did not differ significantly among groups. Although copper is potentially toxic to sheep, no signs of toxicity were observed during this trial, which was consistent with similar studies at this treatment dose. The study indicated that administering COWP to lambs at weaning reduced FEC. Copyright © 2016 Elsevier B.V. All rights reserved.

  16. Atividade ovicida e larvicida in vitro do óleo essencial de Eucalyptus globulus sobre Haemonchus contortus Ovicidal and larvicidal activity in vitro of Eucalyptus globulus essential oils on Haemonchus contortus

    Directory of Open Access Journals (Sweden)

    Iara T. F. Macedo

    2009-09-01

    Full Text Available O objetivo deste trabalho foi avaliar o efeito do óleo essencial de Eucalyptus globulus (OEEG sobre a eclosão e desenvolvimento de larvas de Haemonchus contortus. A determinação da composição química do OEEG foi feita por meio de cromatografia gasosa acoplada à espectrometria de massa. No teste de eclosão de ovos (TEO, utilizaram-se as concentrações 21,75; 17,4; 8,7; 5,43 e 2,71 mg.mL-1; e no de desenvolvimento larvar (TDL as concentrações foram 43,5; 21,75; 10,87; 5,43 e 2,71 mg.mL-1. Cada ensaio foi acompanhado por um controle positivo, 0,02 mg.mL-1 tiabendazol, no TEO e 0,008 mg.mL-1 ivermectina no TDL, e por um controle negativo com o diluente Tween 80 (3%. A eficácia máxima obtida pelo OEEG sobre ovos foi de 99,3% na concentração de 21,75 mg.mL-1 e, sobre larvas, 98,7% na concentração de 43,5 mg.mL-1. A concentração do OEEG, que inviabilizou 50% dos ovos e das larvas do parasito, foi de 8,3 e 6,92 mg.mL-1, respectivamente. A análise química do óleo identificou como principal componente o monoterpeno 1,8-cineol. O OEEG apresentou atividade ovicida e larvicida in vitro sobre H. contortus, determinando um bom potencial para utilização no controle de nematóides gastrintestinais de ovinos e caprinos.The objective of this work was to evaluate ovicidal and larvicidal effects of Eucalyptus globulus essential oil (EGEO on Haemonchus contortus. The chemical composition determination of EGEO was through gas chromatography and mass spectrometry. Egg hatch test (EHT was performed in concentrations 21.75; 17.4; 8.7; 5.43 e 2.71 mg.mL-1. In larval development test (LDT were used the concentrations 43.5; 21.75; 10.87; 5.43 e 2.71 mg.mL-1. Each trial was conducted by negative control with Tween 80 (3% and positive control, 0.02 mg.mL -1 of thiabendazole in EHT and 0.008 mg.mL-1 of ivermectin in LDT. The maximum effectiveness of EGEO on eggs was 99.3% in concentration of 21.75 mg.mL-1 and on larvae was 98.7% in concentration 43

  17. Predatory activity of Butlerius nematodes and nematophagous fungi against Haemonchus contortus infective larvae

    Directory of Open Access Journals (Sweden)

    Manoel Eduardo da Silva

    Full Text Available Abstract The purpose of this study was to evaluate the predatory activity of the nematode Butlerius spp. and fungal isolates of Duddingtonia flagrans, Clonostachys rosea, Arthrobotrys musiformis and Trichoderma esau against H. contortus infective larvae (L3 in grass pots. Forty-eight plastic gardening pots containing 140 g of sterile soil were used. Panicum spp. grass seeds (200 mg were sown into each pot and individually watered with 10 mL of tap water. Twelve days after seeding, the pots were randomly divided into 6 groups (n=8. Two thousand H. contortus infective larvae (L3 were added to each group. Additionally, the following treatments were established: Group 1 – 2000 Butlerius spp. larvae; group 2 – A. musiformis (1x107 conidia; group 3 – T. esau (1x107 conidia; group 4 – C. rosea (1x107 conidia, group 5 – D. flagrans (1x107conidia and Group 6 – no biological controller (control group. The larval population of H. contortus exposed to Butlerius spp. was reduced by 61.9%. Population reductions of 90.4, 66.7, 61.9 and 85.7% were recorded in the pots containing A. musiformis, T. esau, C. rosea and D. flagrans, respectively. The results of this study indicate that the predatory nematode Butlerius spp. and the assessed fungi display an important predatory activity can be considered suitable potential biological control agents.

  18. Annotation of two large contiguous regions from the Haemonchus contortus genome using RNA-seq and comparative analysis with Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Roz Laing

    Full Text Available The genomes of numerous parasitic nematodes are currently being sequenced, but their complexity and size, together with high levels of intra-specific sequence variation and a lack of reference genomes, makes their assembly and annotation a challenging task. Haemonchus contortus is an economically significant parasite of livestock that is widely used for basic research as well as for vaccine development and drug discovery. It is one of many medically and economically important parasites within the strongylid nematode group. This group of parasites has the closest phylogenetic relationship with the model organism Caenorhabditis elegans, making comparative analysis a potentially powerful tool for genome annotation and functional studies. To investigate this hypothesis, we sequenced two contiguous fragments from the H. contortus genome and undertook detailed annotation and comparative analysis with C. elegans. The adult H. contortus transcriptome was sequenced using an Illumina platform and RNA-seq was used to annotate a 409 kb overlapping BAC tiling path relating to the X chromosome and a 181 kb BAC insert relating to chromosome I. In total, 40 genes and 12 putative transposable elements were identified. 97.5% of the annotated genes had detectable homologues in C. elegans of which 60% had putative orthologues, significantly higher than previous analyses based on EST analysis. Gene density appears to be less in H. contortus than in C. elegans, with annotated H. contortus genes being an average of two-to-three times larger than their putative C. elegans orthologues due to a greater intron number and size. Synteny appears high but gene order is generally poorly conserved, although areas of conserved microsynteny are apparent. C. elegans operons appear to be partially conserved in H. contortus. Our findings suggest that a combination of RNA-seq and comparative analysis with C. elegans is a powerful approach for the annotation and analysis of strongylid

  19. Metabolism of albendazole, ricobendazole and flubendazole in Haemonchus contortus adults: Sex differences, resistance-related differences and the identification of new metabolites

    Directory of Open Access Journals (Sweden)

    Lucie Raisová Stuchlíková

    2018-04-01

    Full Text Available Haemonchus contortus (family Trichostrongylidae, Nematoda, a hematophagous gastrointestinal parasite found in small ruminants, has a great ability to develop resistance to anthelmintic drugs. We studied the biotransformation of the three benzimidazole anthelmintics: albendazole (ABZ, ricobendazole (albendazole S-oxide; RCB and flubendazole (FLU in females and males of H. contortus in both a susceptible ISE strain and resistant IRE strain. The ex vivo cultivation of living nematodes in culture medium with or without the anthelmintics was used. Ultrasensitive UHPLC/MS/MS analysis revealed 9, 7 and 12 metabolites of ABZ, RCB and FLU, respectively, with most of these metabolites now described in the present study for the first time in H. contortus. The structure of certain metabolites shows the presence of biotransformation reactions not previously reported in nematodes. There were significant qualitative and semi-quantitative differences in the metabolites formed by male and female worms. In most cases, females metabolized drugs more extensively than males. Adults of the IRE strain were able to form many more metabolites of all the drugs than adults of the ISE strain. Some metabolites were even found only in adults of the IRE strain. These findings suggest that increased drug metabolism may play a role in resistance to benzimidazole drugs in H. contortus. Keywords: Drug resistance, Drug metabolism, Anthelmintics, Benzimidazole, Nematode

  20. Persistence of the efficacy of copper oxide wire particles against Haemonchus contortus in grazing South African goats

    Science.gov (United States)

    Vatta, A.F.; Waller, P.J.; Githiori, J.B.; Medley, G.F.

    2012-01-01

    A study was conducted to examine the duration of anthelmintic effect of copper oxide wire particles (COWP) in grazing goats, as data for the persistence of efficacy of COWP in this host species is limited. Forty-eight indigenous male goats were infected naturally by grazing them on Haemonchus contortus-infected pasture. When the faecal egg count (FEC) in the goats was 3179 ± 540 eggs per gram of faeces (mean ± standard error), half the animals were treated with 4 g COWP (day 0; mean live weight = 25.5 ± 0.8 kg). Eight treated (COWP) and eight non-treated (CONTROL) goats were removed from the pasture on each of days 7, 28 and 56, maintained for 27 or 29 days in concrete pens and then humanely slaughtered for nematode recovery. Mean liver copper levels were in the high range in the goats removed from pasture at day 7 (treated: 191 ± 19.7 ppm; untreated: 120 ± 19.7 ppm; P = 0.022), but had dropped to normal levels at days 28 and 56. The mean H. contortus burdens of the treated versus the non-treated goats were, respectively, 184 ± 48 and 645 ± 152 for the goats removed from pasture at day 7 (71% reduction; P = 0.004), 207 ± 42 and 331 ± 156 at day 28 (37% reduction; P = 0.945) and 336 ± 89 and 225 ± 53 at day 56 (−49% reduction; P = 0.665). Weekly monitoring of FECs after treatment until slaughter indicated that the COWP-treated goats had lower FECs than the controls, the treatment main effect being significant at days 7, 28 and 56 (P goats removed from pasture at day 28 (P ≤ 0.001). Packed cell volumes increased during the course of the experiment (day, P goats removed from pasture at day 28 (CONTROL 28 d, 28.65 ± 0.52% goats, extending at most to 28 days after treatment. However, repeated COWP administration at three-month intervals may be safe, given that liver copper levels return to normal two to three months after COWP treatment. PMID:22789299

  1. Structural and functional characterization of a novel gene, Hc-daf-22, from the strongylid nematode Haemonchus contortus.

    Science.gov (United States)

    Guo, Xiaolu; Zhang, Hongli; Zheng, Xiuping; Zhou, Qianjin; Yang, Yi; Chen, Xueqiu; Du, Aifang

    2016-07-29

    The strongylid nematode Haemonchus contortus is a parasite of major concern for modern livestock husbandry because hostile environmental conditions may induce diapause in the early fourth-stage larvae. A new gene Hc-daf-22 was identified which is the homologue of Ce-daf-22 and human SCPx. Genome walking and RACE were performed to obtain the whole cDNA and genomic sequence of this gene. Using qRT-PCR with all developmental stages as templates to explore the transcription level and micro-injection was applied to confirm the promoter activity of the 5'-flanking region. Overexpression, rescue and RNA interference experiments were performed in N2, daf-22 mutant (ok 693) strains of C. elegans to study the gene function of Hc-daf-22. The full length gene of Hc-daf-22 (6,939 bp) contained 16 exons separated by 15 introns, and encoded a cDNA of 1,602 bp (533 amino acids, estimated at about 59.3 kDa) with a peak in L3 and L4 in transcriptional level. The Hc-DAF-22 protein was consisted of a 3-oxoacyl-CoA thiolase domain and a SCP2 domain and evolutionarily conserved. The 1,548 bp fragment upstream of the 5'-flanking region was confirmed to have promoter activity compared with 5'-flanking region of Ce-daf-22. The rescue experiment by micro-injection of daf-22 (ok693) mutant strain showed significant increase in body size and brood size in the rescued worms with significantly reduced or completely absent fat granules confirmed by Oil red O staining, indicating that Hc-daf-22 could partially rescue the function of Ce-daf-22. Furthermore, RNAi with Hc-daf-22 could partially silence the endogenous Ce-daf-22 in N2 worms and mimic the phenotype of daf-22 (ok693) mutants. The gene Hc-daf-22 was isolated and its function identified using C. elegans as a model organism. Our results indicate that Hc-daf-22 shared similar characteristics and function with Ce-daf-22 and may play an important role in peroxisomal β-oxidation and the development in H. contortus.

  2. Evaluation of multiple low doses of copper oxide wire particles compared with levamisole for control of Haemonchus contortus in lambs.

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    Burke, J M; Miller, J E

    2006-06-30

    High levels of anthelmintic resistance in gastrointestinal nematodes (GIN) of small ruminants have created the need for alternative approaches to parasite control. Copper oxide wire particles (COWP; 2g) have proven effective in decreasing GIN infection in lambs. However, the risk of copper toxicity has limited the usefulness of this approach. Recently, smaller doses (0.5 and 1g) have proven effective in GIN control, reducing the risk of toxicity. The objective of this study was to examine the effectiveness and risk of toxicity using multiple small doses of COWP for GIN control in lambs between weaning and market weight. Dorper crossbred ram lambs were orally administered levamisole (Levasol, 8.0mg/kg; n=8), 0.5g (n=9), or 1g COWP (n=9) at weaning (Day 0; 118+/-2 days of age; late May 2005) and again at 6-week intervals for a total of four treatments. A pooled fecal culture determined that Haemonchus contortus was the predominant gastrointestinal parasite at weaning. Lambs grazed bermudagrass pastures and were supplemented with up to 500g corn/soybean meal and free choice trace mineralized salt. Fecal egg counts (FEC), packed cell volume (PCV), and plasma aspartate aminotransferase (AST) activity were determined every 14 days and lambs weighed every 28 days. GIN infection reached a peak at Day 42 (high FEC, low PCV). COWP effectively reduced FEC on Days 0 and 42 compared with the previous week, but did not reduce FEC on Days 84 and 126 (treatment by time interaction, Pcopper in the liver on Day 155 were greater in COWP-treated lambs (Pcopper toxicity.

  3. Experiences with integrated concepts for the control of Haemonchus contortus in sheep and goats in the United States.

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    Terrill, Thomas H; Miller, James E; Burke, Joan M; Mosjidis, Jorge A; Kaplan, Ray M

    2012-05-04

    The generally warm, moist environmental conditions in the southern United States (U.S.) are ideal for survival and growth of the egg and larval stages of Haemonchus contortus and other gastrointestinal nematodes (GIN) of sheep and goats. Consequently, infection with GIN is the greatest threat to economic small ruminant production in this region. With anthelmintic resistance now reaching epidemic proportions in small ruminants in the U.S., non-chemical control alternatives are critically needed. The Southern Consortium for Small Ruminant Parasite Control (SCSRPC) was formed in response to this crisis and over the last decade has successfully validated the use of several novel control technologies, including FAMACHA(©) for the implementation of targeted selective treatments (TST), copper oxide wire particles (COWP), nematode-trapping fungi, and grazing or feeding hay of the high-tannin perennial legume sericea lespedeza [Lespedeza cuneata (Dum.-Cours. G. Don)]. Producer attitudes toward GIN control in the U.S. have been shifting away from exclusive dependence upon anthelmintics toward more sustainable, integrated systems of parasite control. Some novel control technologies have been readily adopted by producers in combination with appropriate diagnostic tools, such as FAMACHA(©). Others techniques are still being developed, and will be available for producer use as they are validated. Although new drugs will likely be available to U.S. goat and sheep producers in the future, these will also be subject to development of anthelmintic resistance. Therefore, the adoption and implementation of sustainable GIN control principles will remain important. With emerging markets for grass-fed or organic livestock, there will continue to be a critical need for research and outreach on development and on-farm application of integrated GIN control systems for small ruminants in the U.S. for the foreseeable future. Copyright © 2011 Elsevier B.V. All rights reserved.

  4. Eco-epidemiologia de Haemonchus contortus bahienses, ecotipo presente en ovinos de zonas aridas de Venezuela Eco-epidemiologia de Haemonchus contortus bahiensis, ecotipo presente em ovinos de zonas áridas da Venezuela Ecoepidemiology of Haemonchus contortus bahiensis: ecotype present in sheep of Venezuelan arid zone

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    Gustavo Morales

    1987-09-01

    Full Text Available Haemonchus contortus bahiensis Grisi, 1974 parasita a los ovinos de zonas áridas de Venezuela. En las hembras de dicho ecotipo la distribución de frecuencias por nosotros observada fue de un 15,32% para el morfotipo con lengüeta supravulvar, de un 51,61% para el botonado y de un 33,07% para el liso. Correspondiéndole a dicho ecotipo un índice de diversidad de Shannon-Weaver de 1,44 bits. Las medidas corporales, el tamaño de los huevos y el symlophe son similares en los tres morfotipos. La disposición espacial tanto de los machos como de las hembras en el seno de la población de hospedadores es de tipo contagiosa siendo bien descrita por el parámetro K de la distibución binominal negativa. Se evidenció la existencia de una compleja interacción entre la abundancia, la agregación y la prevalencia de dicho ecotipo y se discute su importancia sobre el equilibrio hospedador-parásito.Nas fêmesas do dito ecotipo a distribuição de freqüência observada foi de 15,32% para o morfotipo com lingueta supra-vulvar, de 51,61% para o tipo com botão vulvare de 33,07% para o tipo liso. A esse ecotipo corresponde um índice de diversidade de Shannon-Weaver igual a 1,44 bits. As medidas corporais, o tamanho dos ovos e o número de estrias cuticulares longitudinais são semelhantes nos três morfotipos. A disposição espacial, tanto dos machos quanto das fêmeas na população de hospedeiros é do tipo contagiosa, sendo bem descrita pelo parâmetro K da distribuição binomial negativa. Se verificou a existência de uma complexa interação entre a abundância, a agregação e a prevalência do citado ecotipo, discutindo-se sua importância no equilíbrio hospedeiro-parasito.The frequency distribution of female Haemonchus contortus bahiensis Grisi, 1974 in sheep from Venezuelan arid zones is 15.32% for the type with vulvar flap, 51.6% for the vulvar-knob and 33.07% for smooth type. A Shannon-Weaver diversity index corresponding to 1.44 bits was

  5. Evaluation of resistance in a selected field strain of Haemonchus contortus to ivermectin and moxidectin using the Larval Migration on Agar Test

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    Fernanda S. Fortes

    2013-02-01

    Full Text Available Haemonchus contortus is one of the most common and economically significant causes of disease in small ruminants worldwide, and the control programs of parasitic nematodes - including H. contortus - rely mostly on the use of anthelmintic drugs. The consequence of the use of this, as the sole sanitary strategy to avoid parasite infections, was the reduction of the efficacy of all chemotherapeutic products with a heavy selection for resistance. The widespread of anthelmintic resistance and the difficulty of its early diagnosis has been a major concern for the sustainable parasite management on farms. The objective of this research was to determine and compare the ivermectin (IVM and moxidectin (MOX effect in a selected field strain of H. contortus with a known resistance status, using the in vitro larval migration on agar test (LMAT. Third stage larvae of the selected isolate were obtained from faecal cultures of experimentally infected sheep and incubated in eleven increasing diluted concentrations of IVM and MOX (6, 12, 24, 48, 96, 192, 384, 768, 1536, 3072 and 6144µg/mL. The dose-response sigmoidal curves were obtained using the R² value of >0.90 and the lethal concentration (LC50 dose for the tested anthelmintic drugs using a four-parameter logistic model. The LC50 value for MOX was significantly lower than IVM (1.253µg/mL and 91.06µg/mL, identifying the H. contortus isolate as considerably less susceptible to IVM compared to MOX. Furthermore, the LMAT showed a high consistency (p<0.0001 and provided to be a useful diagnostic tool for monitoring the resistance status of IVM and MOX in H. contortus field isolate, as well as it may be used for official routine drug monitoring programs under the Ministry of Agriculture (MAPA guidance.

  6. Core alpha1-->3-fucose is a common modification of N-glycans in parasitic helminths and constitutes an important epitope for IgE from Haemonchus contortus infected sheep

    NARCIS (Netherlands)

    van Die, I.; Gomord, V.; Kooyman, F. N.; van den Berg, T. K.; Cummings, R. D.; Vervelde, L.

    1999-01-01

    Synthesis of parasite specific IgE plays a critical role in the defence against helminth infections. We report here that IgE from serum from Schistosoma mansoni infected mice and Haemonchus contortus infected sheep recognizes complex-type N-glycans from Arabidopsis thaliana, which contain

  7. Comparison of parasitological and productive traits of Criollo lambs native to the central Mexican Plateau and Suffolk lambs experimentally infected with Haemonchus contortus.

    Science.gov (United States)

    Alba-Hurtado, F; Romero-Escobedo, E; Muñoz-Guzmán, M A; Torres-Hernández, G; Becerril-Pérez, C M

    2010-09-20

    The study compares the parasitological and productive traits of Criollo lambs native to the central Mexican Plateau (CNCMP) and Suffolk (SU) lambs experimentally infected with Haemonchus contortus. CNCMP lambs (n=20) and SU lambs (n=15) were infected with L3 of H. contortus while five lambs of each genotype were kept as controls. Fecal egg count (FEC), packed cell volume (PCV), blood eosinophil number (BEN), ocular mucous membrane color (as measured by the FAMACHA index), changes in body condition score (BCS) and cumulative live weight gain (CLWG) were measured weekly during a 20-week period. On week 20, all animals were euthanized and the number of adult worms (AW) in the abomasum was counted. Infected SU lambs had higher (p<0.05) FEC and AW mean values compared to CNCMP lambs, which had a higher mean BEN count (p<0.05). Infected lambs had lower PCV values than controls, regardless of genotype, and had a negative correlation (r=-0.84, p<0.05) with the FAMACHA index. BCS tended to decline for infected SU lambs and increased slightly for infected CNCMP lambs. CLWG differed in all groups (p<0.05); infected SU lambs gained 12.1+/-1.9kg, infected CNCMP lambs gained 18.8+/-0.7kg, control SU lambs gained 34.6+/-1.6kg, and control CNCMP lambs gained 26.9+/-0.8kg. In conclusion, CNCMP lambs had a smaller worm burden, a better ability to maintain their productive traits, and were less affected by infection with Haemonchus contortus.

  8. Chemical composition and efficacy in the egg-hatching inhibition of essential oil of Piper aduncum against Haemonchus contortus from sheep

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    Gisele Lopes Oliveira

    Full Text Available Piper aduncum L., Piperaceae, has been used to treat mainly inflammatory diseases and has shown several biological activities such as insecticidal and larvicidal. The aim of this study was to analyze the chemical composition of essential oil of P. aduncum and its efficacy to egg-hatching inhibition of Haemonchus contortus from sheep. The essential oil was obtained from leaves and analysed by gas chromatography coupled to flame ionization detector and gas chromatography coupled to mass spectrometry. It was possible to characterize 22 different substances, among them monoterpenes (80.6% and sesquiterpenes (13.9%. The major compound was identified as 1,8-cineole (55.8%. Eggs of the nematode were exposed to four concentrations of the essential oil. Levamisole phosphate was used as positive control. The essential oil showed to be effective in inhibiting H. contortus hatchability and the LC90 was calculated as 8.9 mg.ml-1. These results can point out the P. aduncumessential oil and its chemical components as potential alternative to control of H. contortus

  9. In Vitro Anthelmintic Activity of Crude Extracts of Aerial Parts of Cissus quadrangularis L. and Leaves of Schinus molle L. against Haemonchus contortus

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    Selamawit Zenebe

    2017-01-01

    Full Text Available Background. Haemonchus contortus, the causative agent of Haemonchosis, is the most economically important parasite in small ruminant production. Control with chemotherapy has not been successful due to rapid emergence of drug-resistant strains. There is a continuous search for alternative leads particularly from plants. The study aimed to evaluate the anthelmintic activity of crude methanolic extracts of leaves of Schinus molle and aerial parts of Cissus quadrangularis against H. contortus. Methods. Adult motility test and egg hatching inhibition assay were employed to investigate the in vitro adulticidal and egg hatching inhibitory effects of the extracts. Results. Higher concentrations of the extracts (10 and 5 mg/ml had a significantly superior adulticidal activity (p<0.05 compared to the negative control and lower concentration levels, which was comparable to albendazole. Similarly, the relative egg hatch inhibition efficacy of S. molle and C. quadrangularis extracts indicated a maximum of 96% and 88% egg hatch inhibition, respectively, within the 48 hrs of exposure at 1 mg/ml. Conclusion. The current study evidenced that the crude methanolic extracts of the plants have promising adulticidal and egg hatching inhibitory effects against H. contortus.

  10. In Vitro Anthelmintic Activity of Crude Extracts of Aerial Parts of Cissus quadrangularis L. and Leaves of Schinus molle L. against Haemonchus contortus.

    Science.gov (United States)

    Zenebe, Selamawit; Feyera, Teka; Assefa, Solomon

    2017-01-01

    Haemonchus contortus, the causative agent of Haemonchosis, is the most economically important parasite in small ruminant production. Control with chemotherapy has not been successful due to rapid emergence of drug-resistant strains. There is a continuous search for alternative leads particularly from plants. The study aimed to evaluate the anthelmintic activity of crude methanolic extracts of leaves of Schinus molle and aerial parts of Cissus quadrangularis against H. contortus. Methods . Adult motility test and egg hatching inhibition assay were employed to investigate the in vitro adulticidal and egg hatching inhibitory effects of the extracts. Higher concentrations of the extracts (10 and 5 mg/ml) had a significantly superior adulticidal activity ( p molle and C. quadrangularis extracts indicated a maximum of 96% and 88% egg hatch inhibition, respectively, within the 48 hrs of exposure at 1 mg/ml. The current study evidenced that the crude methanolic extracts of the plants have promising adulticidal and egg hatching inhibitory effects against H. contortus .

  11. Age of Haemonchus contortus third stage infective larvae is a factor influencing the in vitro assessment of anthelmintic properties of tannin containing plant extracts.

    Science.gov (United States)

    Castañeda-Ramírez, G S; Mathieu, C; Vilarem, G; Hoste, H; Mendoza-de-Gives, P; González-Pech, P G; Torres-Acosta, J F J; Sandoval-Castro, C A

    2017-08-30

    The larval exsheathment inhibition assay (LEIA) of infective larvae (L 3 ) is an in vitro method used to evaluate the anthelmintic (AH) activity of tannin-containing plant extracts against different species of gastrointestinal nematodes, including Haemonchus contortus. Some conditions remain to be defined in order to standardize the LEIA, i.e. the optimal age of larvae produced from donor animals to use in the assays. Therefore, this study aimed at identifying the effect of age and age-related vitality of H. contortus infective larvae produced under tropical conditions, on the in vitro AH activity measured with the LEIA. The same acetone:water (70:30) extract from Acacia pennatula leaves was used to perform respective LEIA tests with H. contortus L 3 of different ages (1-7 weeks). Each week, the L 3 were tested against different concentrations of extract (1200, 600, 400, 200, 100, 40μg/mL of extract) plus a PBS control. Bioassays were performed with a benzimidazole (Bz) resistant H. contortus (Paraíso) strain. In order to identify changes in L 3 vitality on different weeks (1-7), two assays testing larval motility were included only with PBS: the larval migration assay (LMA) and the larval motility observation assay (LMOA). Mean effective concentrations causing 50% and 90% exsheathment inhibition (EC 50 , EC 90 ) were obtained for every week using respective Probit analyses. On the first week, the larvae had lowest EC 50 and EC 90 (39.4 and 65.6μg/mL) compared to older larvae (P0.05), while older larvae tended to show higher EC 50 and EC 90 (Page of larvae (r≥-0.83; P age. More stable efficacy results were found between two to five weeks of age. Copyright © 2017 Elsevier B.V. All rights reserved.

  12. Effect of six tropical tanniferous plant extracts on larval exsheathment of Haemonchus contortus Efeito de seis extratos de plantas taniníferas tropicais sobre o desembainhamento larvar de Haemochus contortus

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    Lorena Mayana Beserra de Oliveira

    2011-06-01

    Full Text Available Tanniferous plants represent a promising alternative for controlling gastrointestinal nematodes of small ruminants. This experiment evaluated the effects of extracts from the leaf and stem of Anadenanthera colubrina, Leucaena leucocephala and Mimosa tenuiflora on larval exsheathment of Haemonchus contortus in vitro and verified the role of tannins in this process. Third-stage larvae of H. contortus were incubated with extracts for 3 hours and were exposed to sodium hypochlorite solution. The extracts were tested at 300 µg.mL-1 and accompanied by controls: phosphate buffer solution (PBS and polyvinyl polypyrrolidone (PVPP. The larval exsheathment was evaluated for 60 minutes, and the results were subjected to the Kruskal-Wallis test (p 0.05, except for L. leucocephala and M. tenuiflora leaf extracts. However, pre-incubation with PVPP of these two extracts significantly changed larval exsheathment when compared to the non-treated extracts (p Plantas taniníferas representam uma promissora alternativa de controle dos nematóides gastrintestinais de pequenos ruminantes. Esse experimento avaliou in vitro os efeitos dos extratos das folhas e caules de Anadenanthera colubrina, Leucaena leucocephala e Mimosa tenuiflora sobre o desembainhamento larvar de Haemonchus contortus e verificou o papel dos taninos nesse processo. Larvas de terceiro estádio de H. contortus foram incubadas com 300 µg.mL-1 de extrato por 3 horas e expostas a uma solução de hipoclorito de sódio. O ensaio foi acompanhado por controles: solução salina tamponada com fosfato (PBS e polivinilpolipirrolidona (PVPP. O desembainhamento larvar foi avaliado durante 60 minutos e os resultados submetidos ao teste Kruskal-Wallis (p 0,05, exceto nos extratos das folhas de L. leucocephala e M. tenuiflora. Entretanto, a pré-incubação desses dois extratos com PVPP alterou significativamente o desembainhamento quando comparado com extratos não-tratados (p < 0,05. Esses resultados sugerem

  13. In vitro AND in vivo ANTHELMINTIC EFFECT OF Coffea arabica RESIDUES AGAINTS AN Haemonchus contortus ISOLATE WITH LOW SUSCEPTIBILITY TO TANNINS

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    G.I. Ortiz-Ocampo

    2016-05-01

    Full Text Available The present study evaluated in vitro and in vivo anthelmintic (AH effect of the percolated by-product of Coffea arabica against a Haemonchus contortus isolate with low susceptibility to tannins. Firstly, the susceptibility to tannins of Paraíso H. contortus isolate was evaluated with the in vitro larval exsheathment inhibition assay (LEIA using an acetone:water (70:30 extract of Acacia pennatula. Afterwards, the in vitro AH effect of acetone:water extracts of percolated C. arabica by-products (Clean and Smooth (CS and Shade Grown (SG Starbucks® were evaluated. Then, the in vivo AH effect was determined in growing sheep (4 months old and 15.2 kg bodyweight, infected with H. contortus (Paraiso isolate and fed with a diet containing the percolated by-product of C. arabica. Three experimental groups were formed (n = 6 lambs: GC Group (control diet without C. arabica, GCA Group (feed containing 10% C. arabica and, GCA+PEG (polyethilenglycol Group (feed with 10% C. arabica + PEG as a tannin inhibitor. Fecal samples were obtained daily from the rectum of lambs (days 1 to 20 of the study. The H. contortus isolate tolerated the A. pennatula extract at concentrations of 150 and 300 μg extract/ml PBS (P > 0.05, which confirmed its low susceptibility to tannins. The C. arabica CS extract reduced the exsheatment from 150 μg of extract/ml PBS and the SG variety from 1200 μg of extract/ml PBS (P0.05. In conclusion, the C. arabica percolated by-product extracts showed in vitro AH effect against exsheathment. However, the consumption of feed containing 10% of the percolated by-product did not show an in vivo AH effect in sheep infected with the Paraiso H. contortus isolate. The latter could be due to the low susceptibility towards the secondary compounds of tannin rich plants of the H. contortus isolate used in this study.

  14. Comparative Response of the West African Dwarf Goats to Experimental Infections with Red Sokoto and West African Dwarf Goat Isolates of Haemonchus contortus

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    Lucas Atehmengo Ngongeh

    2015-01-01

    Full Text Available Response of the West African Dwarf (WAD goats to two different isolates of Haemonchus contortus, the Red Sokoto (RS goat isolate (RSHc and the WAD goat isolate (WADHc (isolated from WAD goats, was studied by experimental infections of 4–6-month-old male WAD goat kids. Group 1 and Group 2 goats were each infected with 4500 infective larvae (L3 of RSHc and WADHc, respectively. Group 3 animals served as uninfected control. Prepatent period (PPP, faecal egg counts (FEC, worm burden (WB, body weight (BW, packed cell volume (PCV, and body condition score (BCS were determined. WAD goats infected with RSHc isolate and the ones infected with WADHc isolate had mean PPP of 19.63±0.26 and 19.50±0.19, respectively. Goats infected with WADHc isolate had significantly higher FEC (P=0.004 and WB (P=0.001. BW were significantly higher (P=0.004 both in the controls and in Group 2 goats infected with WADHc isolate than in Group 1 goats infected with the RSHc isolate. BCS of animals in both infected groups dropped significantly (P=0.001. There was a significant drop in PCV (P=0.004 of both infected groups in comparison. Both isolates of H. contortus were pathogenic to the host.

  15. Conservation of a microRNA cluster in parasitic nematodes and profiling of miRNAs in excretory-secretory products and microvesicles of Haemonchus contortus.

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    Henry Y Gu

    2017-11-01

    Full Text Available microRNAs are small non-coding RNAs that are important regulators of gene expression in a range of animals, including nematodes. We have analysed a cluster of four miRNAs from the pathogenic nematode species Haemonchus contortus that are closely linked in the genome. We find that the cluster is conserved only in clade V parasitic nematodes and in some ascarids, but not in other clade III species nor in clade V free-living nematodes. Members of the cluster are present in parasite excretory-secretory products and can be detected in the abomasum and draining lymph nodes of infected sheep, indicating their release in vitro and in vivo. As observed for other parasitic nematodes, H. contortus adult worms release extracellular vesicles (EV. Small RNA libraries were prepared from vesicle-enriched and vesicle-depleted supernatants from both adult worms and L4 stage larvae. Comparison of the miRNA species in the different fractions indicated that specific miRNAs are packaged within vesicles, while others are more abundant in vesicle-depleted supernatant. Hierarchical clustering analysis indicated that the gut is the likely source of vesicle-associated miRNAs in the L4 stage, but not in the adult worm. These findings add to the growing body of work demonstrating that miRNAs released from parasitic helminths may play an important role in host-parasite interactions.

  16. Comparative Response of the West African Dwarf Goats to Experimental Infections with Red Sokoto and West African Dwarf Goat Isolates of Haemonchus contortus.

    Science.gov (United States)

    Ngongeh, Lucas Atehmengo; Onyeabor, Amaechi

    2015-01-01

    Response of the West African Dwarf (WAD) goats to two different isolates of Haemonchus contortus, the Red Sokoto (RS) goat isolate (RSHc) and the WAD goat isolate (WADHc) (isolated from WAD goats), was studied by experimental infections of 4-6-month-old male WAD goat kids. Group 1 and Group 2 goats were each infected with 4500 infective larvae (L3) of RSHc and WADHc, respectively. Group 3 animals served as uninfected control. Prepatent period (PPP), faecal egg counts (FEC), worm burden (WB), body weight (BW), packed cell volume (PCV), and body condition score (BCS) were determined. WAD goats infected with RSHc isolate and the ones infected with WADHc isolate had mean PPP of 19.63 ± 0.26 and 19.50 ± 0.19, respectively. Goats infected with WADHc isolate had significantly higher FEC (P = 0.004) and WB (P = 0.001). BW were significantly higher (P = 0.004) both in the controls and in Group 2 goats infected with WADHc isolate than in Group 1 goats infected with the RSHc isolate. BCS of animals in both infected groups dropped significantly (P = 0.001). There was a significant drop in PCV (P = 0.004) of both infected groups in comparison. Both isolates of H. contortus were pathogenic to the host.

  17. Site-Directed Mutagenesis Study Revealed Three Important Residues in Hc-DAF-22, a Key Enzyme Regulating Diapause of Haemonchus contortus

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    Yan Huang

    2017-11-01

    Full Text Available Haemonchus contortus (H. contortus is one of the most important parasites of small ruminants, especially goats and sheep. The complex life cycle of this nematode is a main obstacle for the control and prevention of haemonchosis. So far, a special form of arrested development called diapause different from the dauer stage in Caenorhabditis elegans (C. elegans has been found in many parasitic nematodes. In our previous study, we have characterized a novel gene Hc-daf-22 from H. contortus sharing high homology with Ce-daf-22 and functional analysis showed this gene has similar biological function with Ce-daf-22. In this study, Hc-daf-22 mutants were constructed using site-directed mutagenesis, and carried out rescue experiments, RNA interference (RNAi experiments and in vitro enzyme activity analysis with the mutants to further explore the precise function site of Hc-DAF-22. The results showed that Hc-daf-22 mutants could be expressed in the rescued ok693 worms and the expression positions were mainly in the intestine which was identical with that of Hc-daf-22 rescued worms. Through lipid staining we found that Hc-daf-22 could rescue daf-22 mutant (ok693 from the fatty acid metabolism deficiency while Hc-daf-22 mutants failed. Brood size and body length analyses in rescue experiment along with body length and life span analyses in RNAi experiment elucidated that Hc-daf-22 resembled Ce-daf-22 in effecting the development and capacity of C. elegans and mutants impaired the function of Hc-daf-22. Together with the protease activity assay, this research revealed three important active resides 84C/299H/349H in Hc-DAF-22 by site-directed mutagenesis.

  18. Anthelmintic effect of carob pods and sainfoin hay when fed to lambs after experimental trickle infections with Haemonchus contortus and Trichostrongylus colubriformis.

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    Arroyo-Lopez, Celia; Manolaraki, Foteini; Saratsis, Anastasios; Saratsi, Katerina; Stefanakis, Alexandros; Skampardonis, Vasileios; Voutzourakis, Nikolaos; Hoste, Hervé; Sotiraki, Smaragda

    2014-01-01

    The aim of the study was to compare the in vivo anthelmintic activity of sainfoin hay (Onobrychis viciifolia) and carob pod meal (Ceratonia siliqua) against gastrointestinal nematodes. Seven days before infection, 64 naive lambs were assigned to four different groups: Group S received sainfoin hay and group CAR was fed with carob pods. The remaining lambs received lucerne hay (Medicago sativa) and were assigned to positive (non-treated, NT) and negative (treated, T) control groups (treatment with albendazole). On day 0, lambs were artificially trickle infected for 6 weeks, with a mixture of infective larvae of Haemonchus contortus and Trichostrongylus colubriformis. Parasitological and pathophysiological parameters were measured repeatedly during the 2-month study. Compared to the NT group, decreases in egg excretion were observed in the CAR and S groups with significant differences only found for sainfoin (p < 0.05). At necropsy, group S showed decreases in the total worm numbers of both nematode species with significant differences for H. contortus. In contrast, no differences were noticed for the CAR group. Compared to the NT group, lower values for fecundity of female H. contortus were found in the S and CAR groups, however differences were non-significant. No differences in body weight gains were found between groups. Consistent results were found showing significantly higher packed cell volume (PCV) values in the T and S groups compared to NT and CAR groups. Overall, these results confirm a positive effect associated with the feeding of lambs with tanniniferous resources on host resilience (PCV values) and against gastrointestinal parasitic nematodes by affecting some biological traits of worm populations (e.g. eggs per gram of faeces and worm numbers). However, the anthelmintic effects differed between the two tannin-containing resources, which might be associated with the quantity and/or quality of secondary metabolites (condensed tannins and/or other

  19. Anthelmintic effect of carob pods and sainfoin hay when fed to lambs after experimental trickle infections with Haemonchus contortus and Trichostrongylus colubriformis

    Directory of Open Access Journals (Sweden)

    Arroyo-Lopez Celia

    2014-01-01

    Full Text Available The aim of the study was to compare the in vivo anthelmintic activity of sainfoin hay (Onobrychis viciifolia and carob pod meal (Ceratonia siliqua against gastrointestinal nematodes. Seven days before infection, 64 naive lambs were assigned to four different groups: Group S received sainfoin hay and group CAR was fed with carob pods. The remaining lambs received lucerne hay (Medicago sativa and were assigned to positive (non-treated, NT and negative (treated, T control groups (treatment with albendazole. On day 0, lambs were artificially trickle infected for 6 weeks, with a mixture of infective larvae of Haemonchus contortus and Trichostrongylus colubriformis. Parasitological and pathophysiological parameters were measured repeatedly during the 2-month study. Compared to the NT group, decreases in egg excretion were observed in the CAR and S groups with significant differences only found for sainfoin (p < 0.05. At necropsy, group S showed decreases in the total worm numbers of both nematode species with significant differences for H. contortus. In contrast, no differences were noticed for the CAR group. Compared to the NT group, lower values for fecundity of female H. contortus were found in the S and CAR groups, however differences were non-significant. No differences in body weight gains were found between groups. Consistent results were found showing significantly higher packed cell volume (PCV values in the T and S groups compared to NT and CAR groups. Overall, these results confirm a positive effect associated with the feeding of lambs with tanniniferous resources on host resilience (PCV values and against gastrointestinal parasitic nematodes by affecting some biological traits of worm populations (e.g. eggs per gram of faeces and worm numbers. However, the anthelmintic effects differed between the two tannin-containing resources, which might be associated with the quantity and/or quality of secondary metabolites (condensed tannins and

  20. Site-Directed Mutagenesis Study Revealed Three Important Residues in Hc-DAF-22, a Key Enzyme Regulating Diapause of Haemonchus contortus.

    Science.gov (United States)

    Huang, Yan; Zheng, Xiuping; Zhang, Hongli; Ding, Haojie; Guo, Xiaolu; Yang, Yi; Chen, Xueqiu; Zhou, Qianjin; Du, Aifang

    2017-01-01

    Haemonchus contortus ( H. contortus ) is one of the most important parasites of small ruminants, especially goats and sheep. The complex life cycle of this nematode is a main obstacle for the control and prevention of haemonchosis. So far, a special form of arrested development called diapause different from the dauer stage in Caenorhabditis elegans ( C. elegans ) has been found in many parasitic nematodes. In our previous study, we have characterized a novel gene Hc-daf-22 from H. contortus sharing high homology with Ce-daf-22 and functional analysis showed this gene has similar biological function with Ce-daf-22 . In this study, Hc-daf-22 mutants were constructed using site-directed mutagenesis, and carried out rescue experiments, RNA interference (RNAi) experiments and in vitro enzyme activity analysis with the mutants to further explore the precise function site of Hc-DAF-22. The results showed that Hc-daf-22 mutants could be expressed in the rescued ok693 worms and the expression positions were mainly in the intestine which was identical with that of Hc-daf-22 rescued worms. Through lipid staining we found that Hc-daf-22 could rescue daf-22 mutant ( ok693 ) from the fatty acid metabolism deficiency while Hc-daf-22 mutants failed. Brood size and body length analyses in rescue experiment along with body length and life span analyses in RNAi experiment elucidated that Hc-daf-22 resembled Ce-daf-22 in effecting the development and capacity of C. elegans and mutants impaired the function of Hc-daf-22 . Together with the protease activity assay, this research revealed three important active resides 84C/299H/349H in Hc-DAF-22 by site-directed mutagenesis.

  1. Effects of single or trickle Haemonchus contortus experimental infection on digestibility and host responses of naïve Creole kids reared indoor.

    Science.gov (United States)

    Bambou, J C; Cei, W; Camous, S; Archimède, H; Decherf, A; Philibert, L; Barbier, C; Mandonnet, N; González-García, E

    2013-01-31

    The objective of this study was to compare the effects of the type of Haemonchus contortus experimental infection (trickle infection, TI versus single infection, SI) on feed intake, nutrients digestibility, parasitological and haematological measures, and plasma leptin in Creole kids. The animals were infected over 2 periods (challenge 1 and challenge 2) of 6 weeks each, corresponding respectively to the primary and the secondary infection. Periods prior infection (1 week each) were considered as controls. The primary infection was realized with 35 Creole kids (18.40±3.76 kg BW) housed in individual boxes and fed a hay-based diet. The secondary infection continued with 29 kids (21.90±3.40 kg BW) from the initial 35. A total of 6 kids and 8 kids were slaughtered for measuring nematode burden at the end of the primary and the secondary infection, respectively. Measurements of nutrients digestibility were made at 0, 3 and 5 weeks post-infection for both challenges. Faecal egg count (FEC), blood eosinophilia and packed cell volume (PCV) were monitored weekly. Feed intake (dry matter intake, DMI) and nutrients digestibility were negatively affected by H. contortus infection only during the primary infection. Plasma leptin changed significantly over time (P=0.0002) but was not affected by the infection type. Effect of infection type was observed only on crude protein digestibility during the primary infection, which was lower in the TI group (P<0.01). The overall level of blood eosinophilia was significantly higher in the TI group (P<0.0001) during both challenges. The overall FEC mean was significantly higher in the SI compared with the TI groups, during both challenges (P<0.02). These results were related to the mean female length significantly higher in the SI group compared with the TI group during challenge 1 (P=0.004), and the number of adult nematode significantly lower in the TI group compared with the SI group during the challenge 2 (P=0.05). The results

  2. Effect of the infection with the nematode Haemonchus contortus (Strongylida: Trichostrongylidae on the haematological, biochemical, clinical and reproductive traits in rams

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    Mariem Rouatbi

    2016-08-01

    Full Text Available This study aimed to investigate the effect of Haemonchus contortus infection on rams’ haematological, biochemical and clinical parameters and reproductive performances. A total number of 12 Barbarine rams (control and infected were included in the experiment. The infected group received 30 000 H. contortus third-stage larvae orally. Each ram’s ejaculate was immediately evaluated for volume, sperm cell concentration and mortality rate. At the end of the experiment (day 82 post-infection, which lasted 89 days, serial blood samples were collected in order to assess plasma testosterone and luteinising hormone (LH concentrations. There was an effect of time, infection and their interaction on haematological parameters (p < 0.001. In infected rams, haematocrit, red blood cell count and haemoglobin started to decrease from 21 days post-infection. There was an effect of time and infection for albumin. For total protein, only infection had a statistically significant effect. For glucose, only time had a statistically significant effect. Concentrations were significantly lower in infected rams compared to control animals. A significant effect of infection and time on sperm concentrations and sperm mortality was observed. The effect of infection appears in time for sperm concentrations at days 69 and 76 post-infection. Sperm mortality rate was significantly higher in infected animals at day 46 post-infection when compared to control group (p < 0.05. Finally, plasma testosterone traits (average concentration, cumulated levels during the sampling period and pulse frequency were depressed in infected rams when compared to control counterparts; none of these endocrine traits were affected for plasma LH.

  3. Interaction between the effects of evaporation rate and amount of simulated rainfall on development of the free-living stages of Haemonchus contortus.

    Science.gov (United States)

    O'Connor, Lauren J; Kahn, Lewis P; Walkden-Brown, Stephen W

    2008-08-17

    A factorial experiment (3 x 4 x 2 x 3) was conducted in programmable incubators to investigate interaction between the effects of rainfall amount, rainfall distribution and evaporation rate on development of Haemonchus contortus to L3. Sheep faeces containing H. contortus eggs were incubated on sterilised soil under variable temperatures typical of summer in the Northern Tablelands of NSW, Australia. Simulated rainfall was applied in 1 of 3 amounts (12, 24 or 32 mm) and 4 distributions (a single event on the day after deposition, or the same total amount split in 2, 3 or 4 equal events over 2, 3 or 4 days, respectively). Samples were incubated at either a Low or High rate of evaporation (Low: 2.1-3.4 mm/day and High: 3.8-6.1 mm/day), and faeces and soil were destructively sampled at 4, 7 and 14 days post-deposition. Recovery of L3 from the soil (extra-pellet L3) increased over time (up to 0.52% at day 14) and with each increment of rainfall (12 mm: evaporation rate (0.01%) compared with the Low evaporation rate (0.31%). All rainfall amounts yielded significantly different recoveries of L3 under Low evaporation rates but there was no difference between the 12 and 24 mm treatments under the High evaporation rate. The distribution of simulated rainfall did not significantly affect recovery of infective larvae. Faecal moisture content was positively associated with L3 recovery, as was the ratio of cumulative precipitation and cumulative evaporation (P/E), particularly when measured in the first 4 days post-deposition. The results show that evaporation rate plays a significant role in regulating the influence of rainfall amount on the success of L3 transmission.

  4. Hc-daf-2 encodes an insulin-like receptor kinase in the barber's pole worm, Haemonchus contortus, and restores partial dauer regulation.

    Science.gov (United States)

    Li, Facai; Lok, James B; Gasser, Robin B; Korhonen, Pasi K; Sandeman, Mark R; Shi, Deshi; Zhou, Rui; Li, Xiangrui; Zhou, Yanqin; Zhao, Junlong; Hu, Min

    2014-06-01

    Infective L3s (iL3s) of parasitic nematodes share common behavioural, morphological and developmental characteristics with the developmentally arrested (dauer) larvae of the free-living nematode Caenorhabditis elegans. It is proposed that similar molecular mechanisms regulate entry into or exit from the dauer stage in C. elegans, and the transition from free-living to parasitic forms of parasitic nematodes. In C. elegans, one of the key factors regulating the dauer transition is the insulin-like receptor (designated Ce-DAF-2) encoded by the gene Ce-daf-2. However, nothing is known about DAF-2 homologues in most parasitic nematodes. Here, using a PCR-based approach, we identified and characterised a gene (Hc-daf-2) and its inferred product (Hc-DAF-2) in Haemonchus contortus (a socioeconomically important parasitic nematode of ruminants). The sequence of Hc-DAF-2 displays significant sequence homology to insulin receptors (IR) in both vertebrates and invertebrates, and contains conserved structural domains. A sequence encoding an important proteolytic motif (RKRR) identified in the predicted peptide sequence of Hc-DAF-2 is consistent with that of the human IR, suggesting that it is involved in the formation of the IR complex. The Hc-daf-2 gene was transcribed in all life stages of H. contortus, with a significant up-regulation in the iL3 compared with other stages. To compare patterns of expression between Hc-daf-2 and Ce-daf-2, reporter constructs fusing the Ce-daf-2 or Hc-daf-2 promoter to sequence encoding GFP were microinjected into the N2 strain of C. elegans, and transgenic lines were established and examined. Both genes showed similar patterns of expression in amphidial (head) neurons, which relate to sensation and signal transduction. Further study by heterologous genetic complementation in a daf-2-deficient strain of C. elegans (CB1370) showed partial rescue of function by Hc-daf-2. Taken together, these findings provide a first insight into the roles of Hc

  5. Blood parameters of sheep with high infection of Haemonchus contortus and treated with “mushroom of the sun” (Agaricus blazei

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    Gabriela Almeida Bastos

    2016-04-01

    Full Text Available Blood and parasitological parameters of sheep experimentally infected with Haemonchus contortus and treated with “mushroom of the sun” (Agaricus blazei were assessed. Lambs infected with the nematode were divided into three groups: treated with basidiocarp powder from the fungus, wormed with trichlorfon, and a control group that did not receive anthelmintic. Split-plot design analysis was performed where the treatments were defined as plots and four periods of collection were defined as subplots. Interaction between the treatments and the collection periods for the fecal egg counts per gram of feces (FEC was observed. Treatment with “mushroom of the sun” showed anthelmintic efficacies ranging from 28.6 to 54.2%. Similar performances to weight gain were observed among the groups. A significant interaction between treatments and evaluation periods was observed for erythrocyte counts, hematocrit value, and serum concentrations of albumin and urea. The values of the mean corpuscular volume, erythrocyte distribution width, and leukocyte, neutrophil, lymphocyte, monocyte, and eosinophil counts, as well as the mean platelet volume, varied among the collection days. The administration of “mushroom of the sun” significantly reduced the FEC, did not influence on the erythrocyte count and hematocrit value, which remained within normal limits. However it increased the serum concentrations of albumin.

  6. ANTHELMINTIC EFFECTS OF DRIED GROUND BANANA PLANT LEAVES (MUSA SPP.) FED TO SHEEP ARTIFICIALLY INFECTED WITH HAEMONCHUS CONTORTUS AND TRICHOSTRONGYLUS COLUBRIFORMIS.

    Science.gov (United States)

    Gregory, Lilian; Yoshihara, Eidi; Silva, Leandro Kataoaka Fernandes; Marques, Eduardo Carvalho; Ribeiro, Bruno Leonardo Mendonça; de Souza Meira, Enoch Brandão; Rossi, Rodolfo Santos; do Amarante, Alessandro Francisco Talamini; Hasegawa, Marjorie Yumi

    2017-01-01

    Helminths is a endoparasites that cause the major losses for profitable sheep production in Brazil. The increased development of resistant strains of endoparasites have enforced the search for sustainable alternatives. The aim of this paper was to provide information about endoparasites control with banana leaves in infected sheep as alternative control strategies and see its viability. In this study, we performed two trials to investigate the anthelmintic properties of banana leaves on endoparasites in sheep. In Trial 1, twelve sheep were artificially infected with Trichostrongylus colubriformis ; in Trial 2, eleven sheep were artificially infected with Haemonchus contortus . Clinical examinations, packed cell volume, total protein, faecal egg counts (FECs) and egg hatchability tests (EHTs) were performed. At the end of the trials, the sheep were humanely slaughtered, and total worm counts were performed. In Trial 1 and 2, no significant FEC decreases were note but significant diference in EHTs were observed. Total worm counts, clinical and haematological parameters did not reveal significant changes between the treatment and control groups. These results suggest that feeding dried ground banana plant leaves to sheep may reduce the viability of Trichostrongylus colubriformis eggs, and this anthelmintic activity is potentially exploitable as part of an integrated parasite management programme. However, further investigation is needed to establish the optimal dosage, develop a convenient delivery form and confirm the economic feasibility of using banana plantation byproducts as feed for ruminant species. Abbreviations: Coproculture test (CT)., Faecal egg count (FEC)., Egg hatchability test (EHT).

  7. The anthelmintic effect of aqueous methanol extract of Combretum molle (R. Br. x. G. Don) (Combretaceae) in lambs experimentally infected with Haemonchus contortus.

    Science.gov (United States)

    Simon, M K; Ajanusi, O J; Abubakar, M S; Idris, A L; Suleiman, M M

    2012-06-08

    The aqueous methanol extract from the stem-bark of Combretum molle was evaluated for anthelmintic activity in lambs infected with Haemonchus contortus using faecal egg count (FEC) reduction assay. The extract showed a dose-dependent reduction in FEC in infected animals. At doses of 500, 1000 and 2000 mg kg(-1), the extract caused FEC reduction of 63%, 69.25% and 96.23%, respectively. Similarly, the standard anthelmintic (albendazole) at a dose of 200 mg kg(-1) produced FEC reduction of 99.24%. FEC reduction produced by the extract at doses of 500 and 1000 mg kg(-1) is below the minimum standard of 90% FEC recommended by the World Association for the Advancement of Veterinary Parasitology (WAAVP). However, there was no significant (P>0.05) difference between the means of groups treated with 1000 mg kg(-1) and 2000 mg kg(-1) compared to that of albendazole. In this study, C. molle has shown a promising anthelmintic activity against experimental haemonchosis. Nonetheless, further studies to evaluate its detailed toxicity are required for the plant extract to be developed into a useful anthelmintic drug. There is also the need to evaluate other parts of the plant (root, leaves, fruits, etc.) for the same effect. Copyright © 2012 Elsevier B.V. All rights reserved.

  8. Impact of the post-weaning nutritional history on the response to an experimental Haemonchus contortus infection in Creole goats and Black Belly sheep.

    Science.gov (United States)

    Ceï, W; Salah, N; Paut, C; Dumoulin, P-J; Arquet, R; Félicité, Y; Alexandre, G; Archimède, H; Bambou, J-C

    2016-03-15

    In small ruminants, the response against gastrointestinal nematode (GIN) infections is influenced not only by the host genotype and the physiological stage but also by environmental factors, particularly the nutritional status at the time of infection. In this study we evaluated the long-term effect and the interaction between the host species and the nutritional history on the response to GIN infection in two animal models differing in their phenotypic growth and their level of GIN resistance: Black Belly sheep and Creole goats. Lambs and kids were subjected to three distinct nutritional conditions at weaning: low dietary conditions (100% of the theoretical energy requirement for maintenance, corresponding to 548v. 484KJ/Kg BW(0.75) for lambs and kids respectively and 6% of crude protein, CP), medium dietary conditions (150% of the theoretical energy requirement for maintenance and 13% CP) and high dietary conditions (200% of the theoretical energy requirement for maintenance and 20% CP). This 3-months period was followed by a 1-month period on the medium dietary conditions for all the animals before an experimental Haemonchus contortus infection. We monitored the impact of the nutritional history (nutritional condition after weaning), on the intensity of the GIN infection by measuring individual faecal egg counts (FEC), growth rate (ADG), blood eosinophil counts and other pathophysiological parameters. The FEC, growth rate and blood eosinophil counts were significantly affected by the nutritional history in lambs but not in kids. The lowest FEC was found for lambs placed in high dietary conditions, however during the same period body weight loss was observed in this group. In low dietary conditions, kids were more resistant than lambs and the ADG was higher in lambs. However, the anaemia and the level of serum pepsinogen, marker of the abomasal mucosa integrity, were higher in kids. Our data suggest that the impact of the post-weaning nutritional history on the

  9. Benzimidazole -Resistance in Haemonchus Contortus: New PCR-RFLP Method for the Detection of Point Mutation at Codon 167 of Isotype 1 Β-Tubulin Gene

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    A Eslami

    2012-12-01

    Full Text Available Background: Due to the lack of a suitable and economic test for the analysis of the polymorphism at codon 167, we developed a new PCR-RFLP technique, based on a modified forward primer (UT-HC167 MF-primer, to identify simultaneously the SNPs at codons 167 and 200 of isotype 1 β-tubu­lin gene of Haemonchus contortus.Methods: There already are several safe and easy methods for identification of point mutations at codons 198 and 200. Due to the lack of a reliable and easy method for the detection of the single nucleo­tide polymorphism (SNP at codon 167, we developed an innovative PCR-RFLP technique based on a modified forward primer (UT-HC167 MF-primer, in which the nucleotide T at the posi­tion 443 was substituted through a nucleotide A creating a restriction site for restriction endonuc­lease SnaB I in the nucleotide sequences including codon 167. A total of 138 adult male H. contortus were collected from three different geo-climatic areas of Iran. The isolated genomic DNA of each single worm was amplified by PCR using primers flanking codon 167. The PCR product (527 bp was then amplified by semi-nested PCR using the UT-HC167 MF-primer and the reverse primer achiev­ing a PCR product of 451 bp in length. This PCR product was subsequently digested with the restriction endonucleases SnaB I and TaaI for analysis of the mutations at codons 167 and 200, respec­tively.Results: All worms had two alleles encoding for phenylalanine (BZss homozygote for both codons.Conclusion: Using the UT-HC167 MF-primer and a suitable reverse primer designed upstream from codon 200, it is possible to amplify a PCR product which can be used for analysis of the SNPs at all three mentioned codons using RFLP.

  10. Anthelmintic-resistant strains of Haemonchus contortus and Trichostrongylus colubriformis isolated from an organic sheep and goat farm in Denmark

    DEFF Research Database (Denmark)

    Pena-Espinoza, Miguel Angel; Enemark, Heidi L.; Thansborg, Stig M.

    2013-01-01

    A suspected case of anthelmintic resistance (AR) was investigated in an organic dairy sheep and goat farm. The herd was established in 2007 by purchase of animals from a number of other farms. Selection for the study was based on history of anthelmintic-treatment failure. Forty-eight lambs and 48......) or 10 mg/kg closantel (CLO). Kids were treated with 10 mg/kg FBZ, 0.3 mg/kg MOX, 14 mg/kg LEV, 0.2 mg/kg IVM or 10 mg/kg CLO. FECs were performed at day of treatment and 14 days post treatment. In a subsequent investigation, faeces from adult goats were cultured to obtain 3rd-stage nematode larvae (L3...... %. This is the first isolation of BZ-resistant H. contortus and T. colubriformis in Denmark and highlights the need for continuous surveillance of AR in conventional and organic farms....

  11. Evaluation of the phenotypic performance of a Red Maasai and Dorper double backcross resource population: indoor trickle challenge with Haemonchus contortus.

    Science.gov (United States)

    Mugambi, J M; Audho, J O; Njomo, S; Baker, R L

    2005-02-28

    Six F(1) Dorper (D) x Red Maasai (R) rams were mated to both D and R ewes to produce backcross lambs. These six double backcross resource families are being analysed to identify quantitative trait loci that may be controlling resistance to gastro-intestinal (GI) nematode parasites, mainly Haemonchus contortus. After assessing the phenotypic performance of the lambs following exposure to natural infections, the surviving lambs were drenched and moved indoors for an artificial challenge with H. contortus along with straightbred D and R lambs. A total of 1317 lambs were included in the analyses consisting of 523 3/4D, 580 3/4R, 87 D and 127 R. The D lambs were significantly and consistently heavier than R lambs and 3/4D were significantly heavier than the 3/4R lambs. The difference between the backcrosses was about half of that between the straightbreds. Resistance was assessed in terms of faecal egg counts (FEC) and total worm counts (TWC) at necropsy while packed cell volume (PCV) was used to assess resilience to weekly oral doses of 2500 infective larvae of H. contortus. No significant breed differences were observed for log transformed FEC (LFEC). A significant breed difference in PCV was recorded. The backcrosses had the higher values and while no differences were observed between the straightbreds, 3/4D had significantly higher PCV than the 3/4R. Despite the absence of breed differences in FEC the R and the 3/4R had significantly fewer worms than the D and the 3/4D. The D had significantly longer worms than the R and the 3/4D had significantly longer worms than the 3/4R. Worms recovered from D had more eggs than those recovered from R. Similarly worms from 3/4D contained more eggs than those from 3/4R. Thus, on a breed basis the breed with more worms had longer worms. In contrast, when, in a small part of the experiment two doses of larvae were used to check for any breed by dose interactions, worms from the low dose (and hence fewer worms) animals were longer

  12. In vitro and in vivo evaluation of the activity of pineapple (Ananas comosus) on Haemonchus contortus in Santa Inês sheep.

    Science.gov (United States)

    Domingues, Luciana Ferreira; Giglioti, Rodrigo; Feitosa, Karina Alves; Fantatto, Rafaela Regina; Rabelo, Márcio Dias; de Sena Oliveira, Márcia Cristina; Bechara, Gervásio Henrique; de Oliveira, Gilson Pereira; Barioni Junior, Waldomiro; de Souza Chagas, Ana Carolina

    2013-10-18

    The development of resistance to anthelmintics has prompted research into alternative methods of controlling intestinal nematodes in ruminants. This study aimed to assess the activity of Ananas comosus on Haemonchus contortus in Santa Inês sheep. The aqueous extract of pineapple skin (AEPS), bromelain from pineapple stems (B4882) and residue from pineapple processing was evaluated in in vitro and in vivo tests. The enzymatic activity of substances was analyzed by the azocasein method. The egg hatch test (EHT) and larval development test (LDT) were performed using the Embrapa2010 isolate of H. contortus. In the in vivo test, 36 sheep artificially infected with H. contortus were divided into six groups: G1: 2 g/kg BW of the aqueous extract administered for three days; G2: 2 g/kg BW of the industrial pineapple residue for 60 days; G3: 180 mg/animal of bromelain in a single dose; G4: negative control I; G5: positive control (levamisole phosphate); and G6: negative control II. The eggs per gram (EPG) in the feces were counted till 28 days after treatment. LC₅₀ and LC₉₀ were obtained by the probit procedure, while the in vivo test results were analyzed by GLM. The aqueous extract in the in vitro and in vivo test, the bromelain and industrial residue presented 0.102, 0.157, 1.864 and 0.048 enzyme units/mL, respectively. In the egg hatch test, the LC₅₀ and LC₉₀ were respectively 31 and 81 mg/mL for the aqueous extract and 0.50 and 2 mg/mL for bromelain. In the larval development test, the LC₅₀ and LC₉₀ were respectively 1.7 and 7.3 mg/mL for the aqueous extract and 0.019 and 0.086 mg/mL for bromelain. In the in vivo test, the general efficacies of the treatments in relation to the negative control were 22.6%, 42.2%, 3.65% and 89% for the aqueous extract, industrial pineapple residue, bromelain and positive control respectively. The transformed EPG values were 3.19 ± 0.59, 3.32 ± 0.25, 2.85 ± 0.66, 3.44 ± 0.50, 2.28 ± 0.93 and 2.75 ± 0.94 for

  13. Effects of in vitro exposure to ivermectin and levamisole on the expression patterns of ABC transporters in Haemonchus contortus larvae

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    Ali Raza

    2016-08-01

    Full Text Available This study investigated the interaction of ATP binding cassette (ABC transport proteins with ivermectin (IVM and levamisole (LEV in larvae of susceptible and resistant isolates of Haemonchus contortus in vitro by measuring transcription patterns following exposure to these anthelmintics. Furthermore, we studied the consequences of drug exposure by measuring the sensitivity of L3 to subsequent exposure to higher drug concentrations using larval migration assays. The most highly transcribed transporter genes in both susceptible and resistant L3 were pgp-9.3, abcf-1, mrp-5, abcf-2, pgp-3, and pgp-10. The resistant isolate showed significantly higher transcription of pgp-1, pgp-9.1 and pgp-9.2 compared to the susceptible isolate. Five P-gp genes and the haf-6 gene showed significantly higher transcription (up to 12.6-fold after 3 h exposure to IVM in the resistant isolate. Similarly, five P-gp genes, haf-6 and abcf-1 were transcribed at significantly higher levels (up to 10.3-fold following 3 h exposure to LEV in this isolate. On the other hand, there were no significant changes in transcriptional patterns of all transporter genes in the susceptible isolate following 3 and 6 h exposure to IVM or LEV. In contrast to these isolate-specific transcription changes, both isolates showed an increase in R-123 efflux following exposure to the drugs, suggesting that the drugs stimulated activity of existing transporter proteins in both isolates. Exposure of resistant larvae to IVM or LEV resulted, in some instances, in an increase in the proportion of the population able to migrate at the highest IVM concentrations in subsequent migration assays. The significant increase in transcription of some ABC transporter genes following 3 h exposure to both IVM and LEV in the resistant isolate only, suggests that an ability to rapidly upregulate protective pathways in response to drugs may be a component of the resistance displayed by this isolate.

  14. Development of a tetra-primer ARMS-PCR for detecting the E198A SNP in the isotype-1 β-tubulin gene of Haemonchus contortus populations in China.

    Science.gov (United States)

    Zongze, Zhang; Xin, Yang; Awais, Ali Ahmad; Weiqiang, Lei; Chunqun, Wang; Di, Wenda; Yanqin, Zhou; Junlong, Zhao; Rui, Fang; Min, Hu

    2018-03-15

    The tetra-primer ARMS-PCR is a rapid, simple and low cost method for single nucleotide polymorphism (SNP) genotyping and has been used to detect SNPs associated with diseases and drug resistance. E198A in the isotype-1 β-tubulin gene is one of the three SNPs associated with benzimidazole resistance in parasitic nematode Haemonchus contortus. However, up to now, only PCR-RFLP method was used to test E198A in H. contortus. In the present study, we developed a tetra-primer ARMS-PCR to detect E198A in H. contortus and the accuracy of the results was compared with that of PCR-coupled sequencing. The results showed that optimization of PCR reaction system, especially the proportion of the amount of inner and outer primers, could achieve desirable amplification effect. Three different profiles displaying three distinct genotypes could be identified clearly and intuitively on the agarose gel where the samples with amplified PCR products containing two bands of 433 bp and 200 bp in size indicated susceptible homozygous (SS), those with PCR products containing two bands of 433 bp and 284 bp in length indicated resistant homozygous (RR) and the samples with amplified PCR products containing three bands of 433 bp, 284 bp and 200 bp in size indicated heterozygous (RS). The results showed that the established method can be successfully applied to the detection of E198A in H. contortus, which has high accuracy and is easy to perform. Copyright © 2018 Elsevier B.V. All rights reserved.

  15. Evaluación del efecto de las artemisininas provenientes del extracto etanólico de Artemisia cina sobre L3 de Haemonchus contortus en una técnica de explantes abomasales

    Directory of Open Access Journals (Sweden)

    Rosa Isabel Higuera-Piedrahita

    2016-01-01

    Full Text Available El nuevo enfoque de control integrado de parásitos obliga a buscar alternativas que consideren el medioambiente, es decir, sostenibles y sustentables. El objetivo de este trabajo fue fraccionar el extracto etanólico de la planta Artemisia cina, obtener artemisininas y conocer el efecto de estas sobre la capacidad de asociación de las L3 de Haemonchus contortus en explantes abomasales. El extracto etanólico se fraccionó por medio de la metodología establecida para Artemisia japónica, y se identificaron las artemisininas por medio de cromatografía en capa fina, teniendo como referencia artemisininas comerciales. Las artemisininas se utilizaron sobre L3 desenvainada, se realizó la técnica de explantes abomasales por triplicado y se compararon así: levamisol (7.5 mg/ml, artemisinina comercial (1 mg/ml, agua y seis diferentes fracciones de A. cina a dosis de 1 mg/ml: Ac3k, Ac3b, Ac3a, Ac3h, Ac3i, Ac4b. No se obtuvieron diferencias significativas entre las fracciones de A. cina y artemisinina comercial (p>0.05. Se concluye que no existió efecto de las artemisininas obtenidas del extracto etanólico de A. cina sobre la capacidad de asociación de las L3 de H. contortus al tejido abomasal. Es importante continuar con más estudios de la artemisinina para determinar sobre qué fase del parásito afecta su viabilidad.

  16. Study on the use of sheep serum post vaccination of three phase larva (L3) Haemonchus contortus worm irradiated inoculated to rabbit

    International Nuclear Information System (INIS)

    Partodihardjo, Sukardji; Suryadi, Helmi; Purwanti, Endang; Adiwinata, Gatot; Irtisam

    1998-01-01

    Study has been done on rabbit to find out the effect of giving sheep serum post vaccination with irradiated gastric worm challenged with infective H. contortus larva. Forty eight local rabbit divided into 3 groups : K, VI and V2, each groups: consists of 4 rabbit, by using 4 times repetition. K group which only inoculated 10,000L3 H. contortus infective. V1 = group which twice serum inoculated with 21 days interval, inoculation dose were 0,3 ml serum each, and challenged with 10.000 L3 H.contortus infective. V2 = group which 3 times serum inoculation with 21 days interval, inoculation dose 0,3 ml serum each and challenged with 10.000 L3 H.contortus infective. Design model used with Completely Random Design (RAL), Duncan was used as inter treatment test. Parameter observed are total protein fraction, Larva L4 content in gaster, cumulative weight gain, and death percentage rabbits. Result of study showed that on average parameter total protein fraction on K + 2,0086 ± 0,2353 mg/dl, V1 = 3,2781 ± 0,9227 mg/dl and V2 = 5,5035 ± 1,6794 mg/d. Average parameter L4 death showed on K = 10, V1 = 8, and V2 = 4, while L4 alive on K = 3, V1 + 1 and V2 = 0. Commutative Average parameter PCV value on K = 19,91 ± 1,24%, V1= 27,31 ± 1,22%, and V2 35,71 ± 0,56 %. Average parameter weight gain per week on K = 24,46 ± 0,74 g, V1 = 26,04 ± 2.23 g, and V2 = 28,93 ± 1,67 g. Percentage average parameter death rate on K = 37,5 ± 14,46%, V1=18,75 ± 23,90% and V2 = 0± 0%. The conclusion of the study result was serum inoculation on rabbits have positive result on reducing anaemia rate, elevating weight gain, reduce death percentage, reduced L4 alive content in gaster, and able oneelevate immunity respond content that is total protein fraction on rabbits. (author)

  17. The N- and C-terminal carbohydrate recognition domains of Haemonchus contortus galectin bind to distinct receptors of goat PBMC and contribute differently to its immunomodulatory functions in host-parasite interactions.

    Science.gov (United States)

    Lu, MingMin; Tian, XiaoWei; Yang, XinChao; Yuan, Cheng; Ehsan, Muhammad; Liu, XinChao; Yan, RuoFeng; Xu, LiXin; Song, XiaoKai; Li, XiangRui

    2017-09-05

    Hco-gal-m is a tandem-repeat galectin isolated from the adult worm of Haemonchus contortus. A growing body of studies have demonstrated that Hco-gal-m could exert its immunomodulatory effects on host peripheral blood mononuclear cells (PBMC) to facilitate the immune evasion. Our previous work revealed that C-terminal and N-terminal carbohydrate recognition domains (CRD) of Hco-gal-m had different sugar binding abilities. However, whether different domains of Hco-gal-m account differently for its multiple immunomodulatory functions in the host-parasite interaction remains to be elucidated. We found that the N-terminal CRD of Hco-gal-m (MNh) and the C-terminal CRD (MCh) could bind to goat peripheral blood mononuclear cells by distinct receptors: transmembrane protein 63A (TMEM63A) was a binding receptor of MNh, while transmembrane protein 147 (TMEM147) was a binding receptor of MCh. In addition, MCh was much more potent than MNh in inhibiting cell proliferation and inducing apoptosis, while MNh was much more effective in inhibiting NO production. Moreover, MNh could suppress the transcription of interferon-γ (IFN-γ), but MCh not. Our data suggested that these two CRDs of Hco-gal-m bind to distinct receptors and contributed differently to its ability to downregulate host immune response. These results will improve our understanding of galectins from parasitic nematodes contributing to the mechanism of parasitic immune evasion and continue to illustrate the diverse range of biological activities attributable to the galectin family.

  18. Comparison of constitutive and thiabendazole-induced expression of five cytochrome P450 genes in fourth-stage larvae of Haemonchus contortus isolates with different drug susceptibility identifies one gene with high constitutive expression in a multi-resistant isolate

    Directory of Open Access Journals (Sweden)

    Esra Yilmaz

    2017-12-01

    Full Text Available Benzimidazoles (BZs remain amongst the most widely used anthelmintic drug classes against gastro-intestinal nematode infections, although their efficacy is increasingly compromised by resistance. The primary underlying mechanisms for BZ resistance are single-nucleotide polymorphisms (SNPs in the isotype 1 β-tubulin gene causing the substitutions F167Y, E198A or F200Y. However, resistance is believed to be multi-genic and previous studies have shown that isolates carrying 90–100% F200Y can vary considerably in their resistance level in the egg hatch assay (EHA. Cytochrome P450 monooxygenases (CYPs are associated with drug resistance in mammals and arthropods and have been considered as mediators of anthelmintic resistance. In Caenorhabditis elegans, several members of the CYP34/35 and CYP31 families are BZ and/or xenobiotic inducible and thiabendazole (TBZ is metabolised by CYP35D1. Here, expression of all 5 CYPs closely related to the C. elegans CYP34/35 and CYP31 families was investigated in fourth-stage larvae of two susceptible and three BZ-resistant Haemonchus contortus isolates following in vitro exposure to TBZ for 3 and 6 h using real-time RT-PCR. The resistance status of all isolates was determined using EHAs and quantification of resistance-associated β-tubulin SNPs using pyrosequencing. While none of the CYPs was TBZ inducible, constitutive expression of CYP34/35 family member HCOI100383400 was significantly 2.4–3.7-fold higher in the multi-drug resistant WR isolate with the strongest BZ resistance phenotype compared to susceptible and intermediate-level BZ-resistant isolates. Although this increase is only moderate, HCOI100383400 might still be involved in high-level BZ resistance by further decreasing susceptibility in isolates already carrying 100% of a β-tubulin SNP causing BZ resistance. Lower transcript levels were observed for all CYPs in the intermediately resistant IRE isolate in comparison to the susceptible Hc

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

    Directory of Open Access Journals (Sweden)

    Elise Courtot

    2015-12-01

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

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

    Science.gov (United States)

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

    2015-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Thomas B Duguet

    2016-07-01

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

  2. A study of the influence of breed and strain on the susceptibility of sheep to Haemonchus controtus infection

    International Nuclear Information System (INIS)

    Castelino, J.

    1976-07-01

    Breed appears to influence the susceptibility of sheep to Haemonchus contortus infection. Experiments on Masai and Merino sheep indicated that the Masai appear innately capable of reacting more vigorously to throw off H. contortus infection, this reaction being enhanced by previous experience. Following challenge, tangible resistance develops consistently in Masai, compared to a partial resistance, at best, in Merino. Resistance appears to be a short-lived phenomenon in both breeds. A field trial was carried out, to determine the influence of hemoglobin type on the susceptibility of Merino sheep. Sheep of hemoglobin types AA, AB and BB were used. A lower egg production occurred in hemoglobin type AA sheep, contrary to results obtained in the laboratory where ''massive'' experimental infection (250 Haemonchus larvae/kg liveweight) was effected. Tracer techniques involving 51 Cr- labelled erythrocytes and 125 I-tagged plasma were applied

  3. Phylogenetic Study of Haemonchus Species from Iran Based On Morpho-Molecular Characterization.

    Directory of Open Access Journals (Sweden)

    Behnam Meshgi

    2015-06-01

    Full Text Available Haemonchosis has a negative effect on the farming industry throughout the world, especially in the tropic and sub-tropic countries. The present study was carried out to differentiate Haemonchus species from its main hosts in Iran, including sheep, goat and camel.The identification took place based on the morphometrics of the spicules and molecular characters. Two hundred seventy adult male nematodes were collected from the abomasums of different ruminants (90 samples from each animal at the slaughterhouses from different localities in Iran. Samples were morphologically identified according to the spicules' morphometric measurements. In the section on molecular study, 10 samples of each Haemonchus isolates were genetically examined. A simple PCR-restriction fragment length polymorphism (PCR-RFLP assay of the second internal transcribed spacer of ribosomal DNA (ITS2-rDNA were described to confirm the PCR results.PCR-RFLP profile obtained from the restriction enzyme HPa1 in H. contortus and H. longistipes indicated 1 (278 bp and 2 (113 and 135 bp different fragments, respectively. The morphological parameters clearly distinguish H. contortus from H. longistipes. Moreover, regarding the ITS2-rDNA, sequences of 295 bp and 314 bp were obtained from H. contortus and H. longistipes, respectively.The genotypic results are in agreement with the phenotypic findings of both species.

  4. Monepantel is a non-competitive antagonist of nicotinic acetylcholine receptors from Ascaris suum and Oesophagostomum dentatum

    Directory of Open Access Journals (Sweden)

    Melanie Abongwa

    2018-04-01

    Full Text Available Zolvix® is a recently introduced anthelmintic drench containing monepantel as the active ingredient. Monepantel is a positive allosteric modulator of DEG-3/DES-2 type nicotinic acetylcholine receptors (nAChRs in several nematode species. The drug has been reported to produce hypercontraction of Caenorhabditis elegans and Haemonchus contortus somatic muscle. We investigated the effects of monepantel on nAChRs from Ascaris suum and Oesophagostomum dentatum heterologously expressed in Xenopus laevis oocytes. Using two-electrode voltage-clamp electrophysiology, we studied the effects of monepantel on a nicotine preferring homomeric nAChR subtype from A. suum comprising of ACR-16; a pyrantel/tribendimidine preferring heteromeric subtype from O. dentatum comprising UNC-29, UNC-38 and UNC-63 subunits; and a levamisole preferring subtype (O. dentatum comprising UNC-29, UNC-38, UNC-63 and ACR-8 subunits. For each subtype tested, monepantel applied in isolation produced no measurable currents thereby ruling out an agonist action. When monepantel was continuously applied, it reduced the amplitude of acetylcholine induced currents in a concentration-dependent manner. In all three subtypes, monepantel acted as a non-competitive antagonist on the expressed receptors. ACR-16 from A. suum was particularly sensitive to monepantel inhibition (IC50 values: 1.6 ± 3.1 nM and 0.2 ± 2.3 μM. We also investigated the effects of monepantel on muscle flaps isolated from adult A. suum. The drug did not significantly increase baseline tension when applied on its own. As with acetylcholine induced currents in the heterologously expressed receptors, contractions induced by acetylcholine were antagonized by monepantel. Further investigation revealed that the inhibition was a mixture of competitive and non-competitive antagonism. Our findings suggest that monepantel is active on multiple nAChR subtypes. Keywords: Monepantel, Zolvix®, Nicotinic acetylcholine

  5. Synergistic inhibition of Haemonchus contortus exsheathment by flavonoid monomers and condensed tannins

    DEFF Research Database (Denmark)

    Klongsiriwet, Chaweewan; Quijada, Jessica; Williams, Andrew Richard

    2015-01-01

    This study investigated the separate and combined anthelmintic (AH) effects of different phenolic compounds, including condensed tannins and flavonoids, all of which are known to occur in willow leaves, a potentially valuable dry season feed. A range of contrasting model tannins, which span the w...

  6. Synergistic interaction of ten essential oils against Haemonchus contortus in vitro

    Science.gov (United States)

    Anthelmintic resistance in sheep gastrointestinal nematodes is a worldwide problem. Multi-drug resistant haemonchosis is the most serious impediment for small ruminant systems, and there are no new drug candidates currently under development. Molecules from natural sources have demonstrated anthelmi...

  7. Lectin, hemolysin and protease inhibitors in seed fractions with ovicidal activity against Haemonchus contortus.

    Science.gov (United States)

    Salles, Hévila Oliveira; Braga, Ana Carolina Linhares; Nascimento, Maria Thayana dos Santos Canuto do; Sousa, Ana Márjory Paiva; Lima, Adriano Rodrigues; Vieira, Luiz da Silva; Cavalcante, Antônio Cézar Rocha; Egito, Antonio Silvio do; Andrade, Lúcia Betânia da Silva

    2014-01-01

    Bioactive molecules of plant species are promising alternatives for the chemical control of gastrointestinal nematodes in ruminants. Extracts of native and exotic seed species from Brazil's semi-arid region were tested in vitro in an egg hatch assay and the bioactivity of their proteins was investigated. Each seed species was subjected to three extractions with three types of solvents. All the seeds showed ovicidal activity, which varied according to the solvents. Higher ovicidal activity was found in the molecule fractions of low molecular weight (Albizia lebbeck, Ipomoea asarifolia, Jatropha curcas, Libidibia ferrea, Moringa oleifera and Ricinus communis (P0.05, Bonferroni test). Hemagglutinating activity was detected in the fractions of C. spectabilis and M. oleifera fractions, hemolysin activity in the A. lebbeck and M. oleifera fractions, serine protease inhibitory activity in the A. lebbeck, I. asarifolia, J. curcas, M. oleifera and R. communis fractions, cysteine protease inhibitor activity in the M. oleifera fraction, and no protein activity in the L. ferrea fraction. The results of this work reveal new plant species with a potential for use in controlling nematode parasites in goats, thus opening a new field of research involving plant protein molecules with ovicidal properties.

  8. Potential Test of Irradiated Vaccine for Haemonchus Contortus Worm and Food Supplement on Sheep

    International Nuclear Information System (INIS)

    Sukardji Partodiharjo; Arifin, M.; Endang Yuliawati; Enuh Rahardjo

    2004-01-01

    An experiment of vaccines was carried out is one of Nematoda worm present in abomasums or stomach of sheep or goat. The case in the field was high enough, may teach 60% the aim of this study is on observe the effect of gastric worm vaccination irradiated 5000 L 3 post challenge wild strain 5000 L 3 , The parameter of observe were clinic, blood twice description number of adult worms, with 3 treatments as follows; control (K), V 2 = twice vaccinations without challenge V 1 twice vaccinations with challenge. The result of the study the average of gain (gram); V 1 =97, V 2 =91 and K=31.20 (P 6 ), V 1 =6.50, V 2 =5.90 and K=6.10 (P 1 =29.30, V 2 =35.30 and K=27.50 (P 3 ), V 1 =9.30, V 2 =9.00 and K=7.40 (P 1 =8.30, V 2 =9.50 and K=6.20 (P 1 immunity respond of weight gain, erythrocyte and leucocyte to have immunity which is better than the respond of other treatments. (author)

  9. Lectin, hemolysin and protease inhibitors in seed fractions with ovicidal activity against Haemonchus contortus

    Directory of Open Access Journals (Sweden)

    Hévila Oliveira Salles

    Full Text Available Bioactive molecules of plant species are promising alternatives for the chemical control of gastrointestinal nematodes in ruminants. Extracts of native and exotic seed species from Brazil's semi-arid region were tested in vitro in an egg hatch assay and the bioactivity of their proteins was investigated. Each seed species was subjected to three extractions with three types of solvents. All the seeds showed ovicidal activity, which varied according to the solvents. Higher ovicidal activity was found in the molecule fractions of low molecular weight (0.05, Bonferroni test. Hemagglutinating activity was detected in the fractions of C. spectabilis and M. oleifera fractions, hemolysin activity in the A. lebbeck and M. oleifera fractions, serine protease inhibitory activity in the A. lebbeck, I. asarifolia, J. curcas, M. oleifera and R. communis fractions, cysteine protease inhibitor activity in the M. oleifera fraction, and no protein activity in the L. ferrea fraction. The results of this work reveal new plant species with a potential for use in controlling nematode parasites in goats, thus opening a new field of research involving plant protein molecules with ovicidal properties.

  10. Dose titration of sericea lespedeza leaf meal on Haemonchus contortus infection in lambs and kids

    Science.gov (United States)

    The objective of three experiments was to determine the impact of supplementing sericea lespedeza (Lespedeza cuneata; SL) in three concentrations in a loose or pelleted diet on gastrointestinal nematodes (GIN) in small ruminants. Experiments on lambs were conducted at the USDA, Agricultural Research...

  11. Interaction between Trypanosoma brucei and Haemonchus ...

    African Journals Online (AJOL)

    In order to investigate the immunomodulatory influence of concurrent T. brucei and H. contortus infection in West African Dwarf (WAD) goats, 28 infected and 7 uninfected (control) of 8-9 months old male WAD goats were studied. The infected goats were separated into resistant (Class 1) and susceptible (Class 2) Faecal ...

  12. Acetylcholine receptor antibody

    Science.gov (United States)

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

  13. The immunogenic effect of irradiated Hemonchus contortus larvae in goats

    International Nuclear Information System (INIS)

    de la Cruz, B.; Perfuse, W.

    1980-01-01

    The development of radiation-attenuated vaccines against parasitic disease has been reported in literature. The use of irradiated larvae as vaccines enables one to take advantage of the special immunological properties of living parasites; the radiation treatment can largely suppress the pathogenic effects of the vaccines and the reproductive capacity of the parasites involved. This report deals with our attempts to produce a radiation-attenuated vaccine against Hemonchus contortus infection in adult goats. Hemonchus contortus infection in ruminants is prevalent in the Philippines, especially among cattle and goats. The results presented indicate that a certain degree of resistance against Hemonchus contortus infection can be induced in adult goats immunized with irradiated larvae. The shelf-life of the vaccine was also determined. (author)

  14. A Biological Study of Larvae and Adult Hemonchus contortus in Goat

    Directory of Open Access Journals (Sweden)

    Yuswandi Yuswandi

    2015-11-01

    experimental animal in vivo. Before the goat necropsied, the diagnosis of H. contortus egg was done every two day post infection and started two weeks after infection. The data was analyzed descriptively. The results showed that the capability of the egg development of the worm to the L3 stadium was 0,33%, the capability of the L3 development to adult was 32,42%, and the prepaten period of H. contortus was 21 days.

  15. In vitro effects of aqueous extract from Maytenus senegalensis (Lam.) Exell stem bark on egg hatching, larval migration and adult worms of Haemonchus contortus.

    Science.gov (United States)

    Zangueu, Calvin Bogning; Olounlade, Abiodoun Pascal; Ossokomack, Marlyse; Djouatsa, Yolande Noelle Nangue; Alowanou, Goue Géorcelin; Azebaze, Anatole Guy Blaise; Llorent-Martínez, Eulogio José; de Córdova, Maria Luisa Fernández; Dongmo, Alain Bertrand; Hounzangbe-Adote, Mawulé Sylvie

    2018-05-02

    Maytenus senegalensis is a common shrub which is scattered in tropical Africa. Different parts of this plant have been reported to be useful in traditional medicine against gastrointestinal disorders and intestinal worms. This study evaluated the anthelmintic activity of the aqueous stem bark extract of M. senegalensis using egg hatch assay (EHA), larval migration inhibition assay (LMIA) and adult worms' motility inhibition assay (AMIA). On EHA, the extract concentrations tested resulted in a significant (p  50%). These in vitro results suggest the presence of some anthelmintic properties in M. senegalensis extract, which is traditionally used by small farmers in west and central Africa. These effects may be due to the flavonoids and proanthocyanidins present in the extract and need to be studied under in vivo conditions.

  16. Efficacy of closantel against ivermectin- and fenbendazole-resistant Haemonchus sp. in sheep in Ontario, Canada.

    Science.gov (United States)

    Westers, T; Jones-Bitton, A; Menzies, P; Van Leeuwen, J; Poljak, Z; Peregrine, A S

    2016-09-15

    In Ontario, Canada, widespread resistance to ivermectin and fenbendazole, the only readily available ovine anthelmintics, has been documented, primarily in Haemonchus sp. In other parts of the world, closantel has been used to control such infections; however, the drug was not currently licensed for use in Canada and the USA. A randomized controlled trial was conducted on six client-owned farms in Ontario in 2013 and 2014 to determine the efficacy of closantel (Flukiver ® 5% Oral Suspension, Elanco Animal Health, 10mg/kg bodyweight) against ivermectin- and fenbendazole-resistant Haemonchus sp. infections in periparturient ewes and grazing lambs. Three farms were randomly assigned to treat all ewes, and three farms were randomly assigned to selectively treat individual ewes at lambing, using predetermined criteria. Fecal samples were collected from a minimum of 15 randomly selected ewes and 13 lambs per group on each farm at the time of treatment and approximately 14days later. Trichostrongyle-type fecal egg counts (FEC) were performed using a modified McMaster technique with a lower detection limit of 8.3 eggs per gram of feces (epg). Haemonchus-specific FECs were determined by multiplying FECs by the proportion of Haemonchus sp. identified from coproculture for each farm; Haemonchus-specific FEC reductions were calculated for each farm. Twenty grazing lambs had FECs conducted monthly, and when mean monthly FECs surpassed 200 epg, all lambs were randomly allocated to either closantel, positive control (ivermectin, fenbendazole, or levamisole) or negative control groups. Pre-treatment Haemonchus-specific mean FECs ranged from 27 to 3359 epg in ewes and 0-5698 epg in lambs. Efficacy of closantel against Haemonchus sp. ranged from 99% (95% CI: 97%-99%) to 100% in recently lambed ewes on all farms in both years (total n=274 ewes), and from 99% (95% CI: 98%-99%) to 100% in grazing lambs in both years on all but one farm (total n=171 lambs). On the latter farm, a whole

  17. Docking to flexible nicotinic acetylcholine receptors

    DEFF Research Database (Denmark)

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

    2010-01-01

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

  18. Neuromodulation: acetylcholine and memory consolidation.

    Science.gov (United States)

    Hasselmo

    1999-09-01

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

  19. Antagonism of acetylcholine by adrenaline antagonists

    Science.gov (United States)

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

    1963-01-01

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

  20. In vitro larvicidal effects of ethanolic extract of Curcuma longa Linn. on Haemonchus larval stage

    Directory of Open Access Journals (Sweden)

    Norisal Binti Nasai

    2016-04-01

    Full Text Available Aim: Gastrointestinal helminthosis is a global problem in small ruminant production. Most parasites have developed resistance to commonly available anthelminthic compounds, and there is currently an increasing need for new compounds with more efficacies. This study evaluated the in vitro effects of ethanolic extract of Curcuma longa (EECL as a biological nematicide against third stage Haemonchus larvae (L3 isolated from sheep. Materials and Methods: Haemonchus L3 were cultured and harvested from the feces of naturally infected sheep. EECL was prepared and three concentrations; 50, 100, and 200 mg/mL were tested for their efficacies on Haemonchus L3. Levamisole at concentration 1.5 and 3 mg/mL were used as positive controls. Results: EECL showed anthelmintic activity in a dose-dependent manner with 78% worm mortality within 24 h of exposure at the highest dose rate of 200 mg/mL. There was a 100% worm mortality rate after 2 h of levamisole (3 mg/mL admisntration. However, there was a comparable larvicidal effect between when levamisole (1.5 mg/mL and EECL (200 mg were administered. Conclusion: The study shows that EECL does exhibit good anthelmintic properties at 200 mg/mL which is comparable with levamisole at 1.5 mg/mL.

  1. The α7 nicotinic acetylcholine receptor complex

    DEFF Research Database (Denmark)

    Thomsen, Morten Skøtt; Mikkelsen, Jens D

    2012-01-01

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

  2. Haemonchus resistente a lactona macrocíclica em caprinos naturalmente parasitados

    Directory of Open Access Journals (Sweden)

    Mattos Mary Jane Tweedie de

    2004-01-01

    Full Text Available O ivermectin, antiparasitário amplamente utilizado em caprinos jovens, foi avaliado sobre nematódeos gastrintestinais, em todas as fases parasitárias. Doze caprinos jovens, naturalmente parasitados foram utilizados no experimento, sendo um grupo de seis animais medicado com ivermectin, na dose de 200µg.kg-1 v.o. e o outro grupo utilizado como controle. Para a avaliação, utilizou-se o teste de redução na contagem de ovos (FECR, redução de larvas infectantes e carga parasitária. Amostras fecais de caprinos foram coletadas no dia da medicação (dia zero, aos sete e catorze dias. No décimo quarto dia pós-medicação, todos os caprinos foram sacrificados, para a coleta de helmintos e cálculo da carga parasitária total. A redução do número de ovos (FECR e a eficácia sobre adultos foi de 42,10 % e 32,62 %, respectivamente.O gênero Haemonchus representou 100 % da população do grupo medicado e 99,58 % do grupo controle. Estes resultados indicam que os caprinos já estavam parasitados por Haemonchus resistente ao ivermectin.

  3. Structural Studies of Nicotinic Acetylcholine Receptors

    DEFF Research Database (Denmark)

    Shahsavar, Azadeh; Gajhede, Michael; Kastrup, Jette

    2016-01-01

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

  4. Allosteric Modulation of Muscarinic Acetylcholine Receptors

    Czech Academy of Sciences Publication Activity Database

    Jakubík, Jan; El-Fakahany, E. E.

    2010-01-01

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

  5. Plasma concentration of acetylcholine in young women

    International Nuclear Information System (INIS)

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

    1987-01-01

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

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

    African Journals Online (AJOL)

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

  7. Haemonchosis and haemoparasites of small ruminants reared in ...

    African Journals Online (AJOL)

    The abomasa were examined by Hansen and Perry method for the presence of Haemonchus contortus while blood samples were examined using the thin blood smear and Haematocrit Centrifugation Techniques (HCT). The prevalence of Haemonchus contortus in small ruminants was 80.3% with goats and sheep having ...

  8. The effect of ketamine on intraspinal acetylcholine release

    DEFF Research Database (Denmark)

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

    2006-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Herman S. Cheung

    2016-07-01

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

  10. The α7 nicotinic acetylcholine receptor complex

    DEFF Research Database (Denmark)

    Thomsen, Morten S; Mikkelsen, Jens D

    2012-01-01

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

  11. Acetylcholine Receptor: Complex of Homologous Subunits

    Science.gov (United States)

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

    1980-06-01

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

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

    Science.gov (United States)

    Davis, Tiffany J; de Fiebre, Christopher M

    2006-01-01

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

  13. Impulsive behavior and nicotinic acetylcholine receptors.

    Science.gov (United States)

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

    2012-01-01

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

  14. Inhibition of acetylcholine synthesis in vitro

    International Nuclear Information System (INIS)

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

    1986-01-01

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

  15. Acetylcholine receptors in the human retina

    International Nuclear Information System (INIS)

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

    1985-01-01

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

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

    DEFF Research Database (Denmark)

    Larsen, Kirsten Vendelbo; Nedergaard, Ove A.

    1999-01-01

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

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

    African Journals Online (AJOL)

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

  18. Effect of aerosolized acetylcholine on bronchial blood flow.

    Science.gov (United States)

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

    1998-08-01

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

  19. The role of acetylcholine in cocaine addiction.

    Science.gov (United States)

    Williams, Mark J; Adinoff, Bryon

    2008-07-01

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

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

    International Nuclear Information System (INIS)

    Paes, Paulo Cesar de Arruda

    2002-01-01

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

  1. Acetylcholine release and inhibitory interneuron activity in hippocampal CA1

    Directory of Open Access Journals (Sweden)

    A. Rory McQuiston

    2014-09-01

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

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

    International Nuclear Information System (INIS)

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

    1988-01-01

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

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

    Science.gov (United States)

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

    2005-11-21

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

  4. Flavonoids with M1 Muscarinic Acetylcholine Receptor Binding Activity

    Directory of Open Access Journals (Sweden)

    Meyyammai Swaminathan

    2014-06-01

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

  5. Muscarinic acetylcholine receptor expression in aganglionic bowel.

    Science.gov (United States)

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

    2000-01-01

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

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

    International Nuclear Information System (INIS)

    Yang Chongjun; Tang Feiyu; Zhang Ping; Guo Yuhai

    2004-01-01

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

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

    DEFF Research Database (Denmark)

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

    2016-01-01

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

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

    African Journals Online (AJOL)

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

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

    NARCIS (Netherlands)

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

    2015-01-01

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

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

    Science.gov (United States)

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

    2012-02-01

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

  11. Effect of mixing low palatable grasses of heteropogon contortus with ipil ipil leaves on digestibility in goats

    International Nuclear Information System (INIS)

    Bashir, M.; Qamar, I.A.; Babar, R.

    2015-01-01

    The study was conducted at the National Agricultural Research Centre, Islamabad, Pakistan during 2012 to find out the effect of mixing low palatable grasses of Heteropogon contortus (HC), with tree leaves of Leucaena leucocephala (Ipil ipil, II) in the ratio of 75:25, 50:50, 25:75, along with sole species on their digestibility in small ruminants. Goats fed II/sub 100%/, HC/sub 25%/ II/sub 75%/, HC/sub 50%/ II/sub 50%/, HC/sub 75%/ II/sub 25%/ and HC/sub 100%/ had similar dry matter (DM), crude protein (CP) and crude fibre (CF) consumption among all the treatments. The digestibility percentage of dry matter intake (DMI) varied among the treatments ranging from 68.25% to 41.66%. Mixtures of low palatable grass and Ipil ipil were in general more digestible with more than 65% dry matter digestibility. The lowest digestibility of dry matter (41.66%) was observed in HC/sub 100%/. A similar trend was noted for CP digestibility. However, reverse trend was observed in digestibility of CF where highest digestibility was recorded in HC100% and lowest in II100%. It can be concluded that grass and ipil ipil leaf mixture are better regarding forage quality and nutrient digestibility and can be recommended as animal feed. (author)

  12. Branched nanotrees with immobilized acetylcholine esterase for nanobiosensor applications

    DEFF Research Database (Denmark)

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

    2010-01-01

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2003-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Chur Chin

    2010-04-01

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

  15. Branched nanotrees with immobilized acetylcholine esterase for nanobiosensor applications

    Energy Technology Data Exchange (ETDEWEB)

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

    2010-02-05

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

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

    International Nuclear Information System (INIS)

    Gibson, G.E.; Peterson, C.

    1986-01-01

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

  17. BKR.2014.003 (Sonibare) b

    African Journals Online (AJOL)

    Femi Olorunniji

    2014-03-31

    Mar 31, 2014 ... constituted 33.33% prevalence rate, while Haemonchus contortus and Oesophagostomum columbianum mixed infection ... However, in WAD, the eosinophil count ... Caprine parasitic gastrointestinal infection is worldwide in.

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

    Science.gov (United States)

    Becchetti, Andrea; Amadeo, Alida

    2016-01-01

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

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

    DEFF Research Database (Denmark)

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

    2003-01-01

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

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

    Science.gov (United States)

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

    2017-11-01

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

  1. Acetylcholine synthesis and possible functions during sea urchin development

    Directory of Open Access Journals (Sweden)

    C Angelini

    2009-06-01

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

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

    Science.gov (United States)

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

    2015-11-01

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

  3. Incidence of Haemonchus spp. and effect on haematocrit and eye colour in goats farmed under resource-poor conditions in South Africa.

    Science.gov (United States)

    Vatta, A F; Krecek, R C; Letty, B A; van der Linde, M J; Grimbeek, R J; de Villiers, J F; Motswatswe, P W; Molebiemang, G S; Boshoff, H M; Hansen, J W

    2002-01-03

    The diversity and predominance of nematode genera in goats of resource-poor farmers at Rust de Winter, Gauteng Province, Impendle, KwaZulu-Natal Province, and Kraaipan, North-West Province, South Africa, was determined by means of a longitudinal study of the nematode faecal egg counts (FECs) and differential third-stage nematode larvae. The animals were bled for haematocrit determination and scored for pallor of ocular mucous membranes using the FAMACHA( Copyright) method, an assay for clinical evaluation of anaemia caused by Haemonchus spp. Animals considered to be in danger of dying from anaemia caused by haemonchosis were selectively treated with an anthelmintic. Lower haematocrit values were registered during periods of heavier Haemonchus infection, which occurred from December/January to March for Rust de Winter; from December to March/April for Impendle; and from November/December to February or April for Kraaipan. There was agreement too between the lower haematocrits and paler mucous membranes scored according to the FAMACHA( Copyright) method. The use of this system may be recommended as part of an integrated approach to worm control in goats kept in the resource-poor areas studied.

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

    DEFF Research Database (Denmark)

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

    2011-01-01

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

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

    DEFF Research Database (Denmark)

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

    2015-01-01

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

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

    International Nuclear Information System (INIS)

    Rosenthal, L.S.

    1987-01-01

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

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

    Science.gov (United States)

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

    2011-07-01

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

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

    Institute of Scientific and Technical Information of China (English)

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

    2009-01-01

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

  9. Nucleus Accumbens Acetylcholine Receptors Modulate Dopamine and Motivation.

    Science.gov (United States)

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

    2016-11-01

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

  10. Caenorhabditis elegans nicotinic acetylcholine receptors are required for nociception

    Science.gov (United States)

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

    2014-01-01

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

  11. Between-breed variations in resistance/resilience to gastrointestinal nematodes among indigenous goat breeds in Uganda

    NARCIS (Netherlands)

    Onzima, R.B.; Mukiibi, Robert; Ampaire, A.; Benda-Beckmann, von K.; Kanis, E.

    2017-01-01

    Gastrointestinal nematodes (GINs), Haemonchus contortus, are a major health problem in goat production. Resistance to H. contortus, the most prevalent GIN in Uganda, was studied among three indigenous goat breeds to assess their differences. Twelve male goats of each breed approximately 7 months old

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

    Science.gov (United States)

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

    1973-01-01

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

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

    Science.gov (United States)

    2012-01-01

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

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

    Science.gov (United States)

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

    2001-12-01

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

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

    International Nuclear Information System (INIS)

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

    1987-01-01

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

  16. Functional interaction between Lypd6 and nicotinic acetylcholine receptors

    DEFF Research Database (Denmark)

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

    2016-01-01

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

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

    International Nuclear Information System (INIS)

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

    1986-01-01

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

  18. Influence du feu et du pâturage sur l'évolution de la phytomasse d'une savane à Heteropogon contortus de la région de Sakaraha (sud-ouest de Madagascar

    Directory of Open Access Journals (Sweden)

    Rakotoarimanana, V.

    2008-01-01

    Full Text Available Effects of Fire and Grazing on Grass Phytomass Evolution in a Heteropogon contortus Savanna (Sakaraha Region, South West Madagascar. An experimental study of the effects of fire and grazing was carried out in the Sakaraha area (S. W. Madagascar. The objectives were to evaluate the combined or separate effects of fire and grazing on the green biomass and total phytomass of a Heteropogon contortus-Poupartia caffra dominated savanna. Three burning treatments (no burning, early burning and late burning and two grazing treatments (grazed, ungrazed were compared. A three-year study showed that (1 both burning treatments led to a delayed vegetation phenophase but a higher standing phytomass at the end of the growth cycle, (2 grazing treatment led to a significant decrease of the standing phytomass and of the contribution of Poaceae.

  19. Physicochemical properties of the modeled structure of astacin metalloprotease moulting enzyme NAS-36 and mapping the druggable allosteric space of Heamonchus contortus, Brugia malayi and Ceanorhabditis elegans via molecular dynamics simulation.

    Science.gov (United States)

    Sharma, Om Prakash; Agrawal, Sonali; Kumar, M Suresh

    2013-12-01

    Nematodes represent the second largest phylum in the animal kingdom. It is the most abundant species (500,000) in the planet. It causes chronic, debilitating infections worldwide such as ascariasis, trichuriasis, hookworm, enterobiasis, strongyloidiasis, filariasis and trichinosis, among others. Molecular modeling tools can play an important role in the identification and structural investigation of molecular targets that can act as a vital candidate against filariasis. In this study, sequence analysis of NAS-36 from H. contortus (Heamonchus contortus), B. malayi (Brugia malayi) and C. elegans (Ceanorhabditis elegans) has been performed, in order to identify the conserved residues. Tertiary structure was developed for an insight into the molecular structure of the enzyme. Molecular Dynamics Simulation (MDS) studies have been carried out to analyze the stability and the physical properties of the proposed enzyme models in the H. contortus, B. malayi and C. elegans. Moreover, the drug binding sites have been mapped for inhibiting the function of NAS-36 enzyme. The molecular identity of this protease could eventually demonstrate how ex-sheathment is regulated, as well as provide a potential target of anthelmintics for the prevention of nematode infections.

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

    NARCIS (Netherlands)

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

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

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

    DEFF Research Database (Denmark)

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

    1980-01-01

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

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

    Energy Technology Data Exchange (ETDEWEB)

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

    1987-08-01

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

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

    OpenAIRE

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

    2004-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Xinli Liu

    2008-01-01

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

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

    DEFF Research Database (Denmark)

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

    2009-01-01

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

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

    Science.gov (United States)

    Acquas, E; Di Chiara, G

    1999-10-27

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

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

    Directory of Open Access Journals (Sweden)

    Abrar Ashoor

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

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

    Science.gov (United States)

    Dennis, M J; Miledi, R

    1974-03-01

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

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

    Science.gov (United States)

    Fobbs, Wambura C; Mizumori, Sheri J Y

    2014-01-01

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

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

    International Nuclear Information System (INIS)

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

    1985-01-01

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

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

    Science.gov (United States)

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

    2010-01-01

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

  12. Effects of two oxadiazolidinones on cholinesterases and acetylcholine receptors

    International Nuclear Information System (INIS)

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

    1986-01-01

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

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

    International Nuclear Information System (INIS)

    Herholz, Karl

    2008-01-01

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

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

    International Nuclear Information System (INIS)

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

    1987-01-01

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

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

    International Nuclear Information System (INIS)

    McCrea, P.D.

    1987-01-01

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

  16. Biochemical and immunological studies of the Muscarinic acetylcholine receptor

    International Nuclear Information System (INIS)

    Gainer, M.W.

    1985-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Joan Y. Holgate

    2015-06-01

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

  18. Rapid synthesis of acetylcholine receptors at neuromuscular junctions.

    Science.gov (United States)

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

    1988-10-11

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

  19. Acetylcholine receptors in dementia and mild cognitive impairment

    Energy Technology Data Exchange (ETDEWEB)

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

    2008-03-15

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

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

    Directory of Open Access Journals (Sweden)

    Peng H Benjamin

    2009-07-01

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

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

    International Nuclear Information System (INIS)

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

    2002-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Sérgio M Pinto

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

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

    DEFF Research Database (Denmark)

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

    2013-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Ritva Tikkanen

    2013-04-01

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

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

    Science.gov (United States)

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

    2015-12-01

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

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

    Science.gov (United States)

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

    2000-01-01

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

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

    International Nuclear Information System (INIS)

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

    1981-01-01

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

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

    DEFF Research Database (Denmark)

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

    2012-01-01

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

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

    Science.gov (United States)

    Kommalage, Mahinda; Höglund, A Urban

    2005-02-21

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

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

    International Nuclear Information System (INIS)

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

    1987-01-01

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

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

    International Nuclear Information System (INIS)

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

    1985-01-01

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

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

    Science.gov (United States)

    Wang, Jingyi; Lindstrom, Jon

    2018-06-01

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

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

    Science.gov (United States)

    Furuhashi, Tsubasa; Sakamoto, Kazuichi

    2016-03-25

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

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

    DEFF Research Database (Denmark)

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

    2013-01-01

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

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

    DEFF Research Database (Denmark)

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

    2015-01-01

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

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

    DEFF Research Database (Denmark)

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

    2016-01-01

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

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

    NARCIS (Netherlands)

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

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

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

    DEFF Research Database (Denmark)

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

    2012-01-01

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

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

    DEFF Research Database (Denmark)

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

    2014-01-01

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

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

    DEFF Research Database (Denmark)

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

    2009-01-01

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

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

    DEFF Research Database (Denmark)

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

    2010-01-01

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

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

    NARCIS (Netherlands)

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

    2012-01-01

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

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

    DEFF Research Database (Denmark)

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

    2010-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Ann Sluder

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

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

    NARCIS (Netherlands)

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

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

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

    Science.gov (United States)

    Uchida, Sae; Kagitani, Fusako

    2017-05-12

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2009-01-01

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

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

    NARCIS (Netherlands)

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

    2016-01-01

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

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

    NARCIS (Netherlands)

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

    2001-01-01

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

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

    NARCIS (Netherlands)

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

    1995-01-01

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

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

    NARCIS (Netherlands)

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

    2013-01-01

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2009-01-01

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

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

    Science.gov (United States)

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

    2016-01-01

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2016-01-01

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

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

    Czech Academy of Sciences Publication Activity Database

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

    2017-01-01

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

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

    NARCIS (Netherlands)

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

    2011-01-01

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

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

    NARCIS (Netherlands)

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

    2011-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Eneida R. Rabelo

    2008-01-01

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

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

    NARCIS (Netherlands)

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

    1993-01-01

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

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

    International Nuclear Information System (INIS)

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

    2006-01-01

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

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

    Science.gov (United States)

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

    2007-01-01

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

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

    Directory of Open Access Journals (Sweden)

    Elisabet Jerlhag

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

  3. Sigma receptor ligand N,N'-di-(ortho-tolyl)guanidine inhibits release of acetylcholine in the guinea pig ileum.

    Science.gov (United States)

    Cambell, B G; Keana, J F; Weber, E

    1991-11-26

    The inhibition of stimulated contractions of the guinea pig ileum longitudinal muscle/myenteric plexus preparation by sigma receptor ligands has been previously described. In this study, the stimulated release of [3H]acetylcholine from cholinergic nerve terminals in this same preparation was monitored in the presence and absence of sigma receptor ligands. N,N'-Di-(orthotolyl)guanidine (DTG) and other compounds selective for the sigma receptor inhibited stimulated [3H]acetylcholine release. These results suggest that their inhibition of stimulated contractions in this preparation was mediated by inhibition of acetylcholine release.

  4. Covalent Trapping of Methyllycaconitine at the α4-α4 Interface of the α4β2 Nicotinic Acetylcholine Receptor

    DEFF Research Database (Denmark)

    Absalom, Nathan L; Quek, Gracia; Lewis, Trevor M

    2013-01-01

    The α4β2 nicotinic acetylcholine receptors (nAChRs) are widely expressed in the brain and are implicated in a variety of physiological processes. There are two stoichiometries of the α4β2 nAChR, (α4)2(β2)3 and (α4)3(β2)2, with different sensitivities to acetylcholine (ACh), but their pharmacologi......The α4β2 nicotinic acetylcholine receptors (nAChRs) are widely expressed in the brain and are implicated in a variety of physiological processes. There are two stoichiometries of the α4β2 nAChR, (α4)2(β2)3 and (α4)3(β2)2, with different sensitivities to acetylcholine (ACh...

  5. Acetylcholine produces contraction mediated by cyclooxigenase pathway in arterial vessels in the marine fish (Isacia conceptionis

    Directory of Open Access Journals (Sweden)

    FA. Moraga

    Full Text Available Preliminary studies showed that dorsal artery contraction mediated by acetylcholine (ACh is blocked with indomethacin in intertidal fish (G. laevifrons. Our objective was to characterize the cholinergic pathway in several artery vessels of the I. conceptionis. Afferent and efferent branchial, dorsal and mesenteric arteries were dissected of 6 juvenile specimens, isometric tension studies were done using doses response curves (DRC for Ach (10–13 to 10–3 M, and cholinergic pathways were obtained by blocking with atropine or indomethacin. CRC to ACh showed a pattern of high sensitivity only in efferente branchial artery and low sensibility in all vessels. Furthermore, these contractions were blocked in the presence of atropine and indomethacin in all vessels. Our results corroborate previous results observed in intertidal species that contraction induced by acetylcholine is mediated by receptors that activate a cyclooxygenase contraction pathway.

  6. Study of the Peripheral Nerve Fibers Myelin Structure Changes during Activation of Schwann Cell Acetylcholine Receptors.

    Directory of Open Access Journals (Sweden)

    Ekaterina E Verdiyan

    Full Text Available In the present paper we consider a new type of mechanism by which neurotransmitter acetylcholine (ACh regulates the properties of peripheral nerve fibers myelin. Our data show the importance of the relationship between the changes in the number of Schwann cell (SC acetylcholine receptors (AChRs and the axon excitation (different intervals between action potentials (APs. Using Raman spectroscopy, an effect of activation of SC AChRs on the myelin membrane fluidity was investigated. It was found, that ACh stimulates an increase in lipid ordering degree of the myelin lipids, thus providing evidence for specific role of the "axon-SC" interactions at the axon excitation. It was proposed, that during the axon excitation, the SC membrane K+- depolarization and the Ca2+-influx led to phospholipase activation or exocytosis of intracellular membrane vesicles and myelin structure reorganization.

  7. Changes in acetylcholine content, release and muscarinic receptors in rat hippocampus under cold stress

    International Nuclear Information System (INIS)

    Fatranska, M.; Budai, D.; Gulya, K; Kvetnansky, R.

    1989-01-01

    The aim was to study the mechanism of the previously established decrease in acetylcholine (ACh) concentration in the rat hippocampus under cold stress. Male rats were exposed for 14 days to cold (5 degree C) or kept (controls) at room temperature (24 degree C). Acetylcholine content, release and muscarinic receptor binding were investigated in the hippocampus. Cold exposure resulted in a decrease of ACh concentration in the dorsal hippocampus. Moreover, the potassium-evoked release of ACh from hippocampal slices was increased and an increase of maximal binding capacity of [ 3 H](-) quinuclidinyl benzilate in the dorsal hippocampus of cold exposed animals was also observed. Thus the decrease of hippocampal ACh concentration under cold exposure is probably due to its increased release. On balance then, our results demonstrate that cold stress in the rat induces significant activation of the hippocampal cholinergic system

  8. Functional Characterization of CCHamide and Muscarinic Acetylcholine Receptor Signalling in Drosophila melanogaster

    DEFF Research Database (Denmark)

    Ren, Guilin Robin

    G-protein coupled receptors (GPCRs) constitute a large and ancient superfamily of membraneproteins responsible for the transduction of extracellular signals to the inside of the cells. In thisPh.D. thesis, Drosophila melanogaster (Dm) was used as a model organism to investigate a numberof topics...... is a newly discovered insect peptide hormone. The function of this novel peptide hasnot been well characterised. In this Ph.D. thesis, I identified CCHamide-2 peptides in endocrinecells of the gut and neurones of the brain of larvae and endocrine cells of the gut of adultDrosophila. Behavioural assays...... little is known about muscarinic acetylcholine receptorsignalling in insects. In this study, I found that two types of mAChRs occur in D. melanogaster, onecoupling to Gq (A-type) and the other to Gi (B-type). Both A- and B-type Dm-mAChRs can beactivated by acetylcholine (ACh), but the classical...

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

    International Nuclear Information System (INIS)

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

    1986-01-01

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

  10. Fluoxetine Alleviates Behavioral Depression while Decreasing Acetylcholine Release in the Nucleus Accumbens Shell

    OpenAIRE

    Chau, David T; Rada, Pedro V; Kim, Kay; Kosloff, Rebecca A; Hoebel, Bartley G

    2011-01-01

    Selective serotonin reuptake inhibitors, such as fluoxetine, have demonstrated the ability to alleviate behavioral depression in the forced swim test; however, the sites and mechanisms of their actions remain to be further elucidated. Previous studies have suggested that behavioral depression in the swim test is mediated in part by acetylcholine (ACh) stimulating the cholinergic M1 receptors in the nucleus accumbens (NAc) shell. The current study tested whether acute, local, and chronic, subc...

  11. Classical and atypical agonists activate M1 muscarinic acetylcholine receptors through common mechanisms

    Czech Academy of Sciences Publication Activity Database

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

    2015-01-01

    Roč. 97, Jul 2015 (2015), s. 27-39 ISSN 1043-6618 R&D Projects: GA ČR(CZ) GA305/09/0681; GA ČR(CZ) GBP304/12/G069; GA MŠk(CZ) EE2.3.30.0025 Institutional support: RVO:67985823 Keywords : muscarinic acetylcholine receptors * atypical agonists * xanomeline * activation mechanism Subject RIV: ED - Physiology Impact factor: 4.816, year: 2015

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

    International Nuclear Information System (INIS)

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

    1983-01-01

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

  13. Nicotine at clinically relevant concentrations affects atrial inward rectifier potassium current sensitive to acetylcholine.

    Science.gov (United States)

    Bébarová, Markéta; Matejovič, Peter; Švecová, Olga; Kula, Roman; Šimurdová, Milena; Šimurda, Jiří

    2017-05-01

    Nicotine abuse is associated with variety of diseases including arrhythmias, most often atrial fibrillation (AF). Altered inward rectifier potassium currents including acetylcholine-sensitive current I K(Ach) are known to be related to AF pathogenesis. Since relevant data are missing, we aimed to investigate I K(Ach) changes at clinically relevant concentrations of nicotine. Experiments were performed by the whole cell patch clamp technique at 23 ± 1 °C on isolated rat atrial myocytes. Nicotine was applied at following concentrations: 4, 40 and 400 nM; ethanol at 20 mM (∼0.09%). Nicotine at 40 and 400 nM significantly activated constitutively active component of I K(Ach) with the maximum effect at 40 nM (an increase by ∼100%); similar effect was observed at -110 and -50 mV. Changes at 4 nM nicotine were negligible on average. Coapplication of 40 nM nicotine and 20 mM ethanol (which is also known to activate this current) did not show cumulative effect. In the case of acetylcholine-induced component of I K(Ach) , a dual effect of nicotine and its correlation with the current magnitude in control were apparent: the current was increased by nicotine in the cells showing small current in control and vice versa. The effect of 40 and 400 nM nicotine on acetylcholine-induced component of I K(Ach) was significantly different at -110 and -50 mV. We conclude that nicotine at clinically relevant concentrations significantly increased constitutively active component of I K(Ach) and showed a dual effect on its acetylcholine-induced component, similarly as ethanol. Synchronous application of nicotine and ethanol did not cause additive effect.

  14. Brain nicotinic acetylcholine receptors are involved in stress-induced potentiation of nicotine reward in rats.

    Science.gov (United States)

    Javadi, Parastoo; Rezayof, Ameneh; Sardari, Maryam; Ghasemzadeh, Zahra

    2017-07-01

    The aim of the present study was to examine the possible role of nicotinic acetylcholine receptors of the dorsal hippocampus (CA1 regions), the medial prefrontal cortex or the basolateral amygdala in the effect of acute or sub-chronic stress on nicotine-induced conditioned place preference. Our results indicated that subcutaneous administration of nicotine (0.2 mg/kg) induced significant conditioned place preference. Exposure to acute or sub-chronic elevated platform stress potentiated the response of an ineffective dose of nicotine. Pre-conditioning intra-CA1 (0.5-4 µg/rat) or intra-medial prefrontal cortex (0.2-0.3 µg/rat) microinjection of mecamylamine (a non-selective nicotinic acetylcholine receptor antagonist) reversed acute stress-induced potentiation of nicotine reward as measured in the conditioned place preference paradigm. By contrast, pre-conditioning intra-basolateral amygdala microinjection of mecamylamine (4 µg/rat) potentiated the effects of acute stress on nicotine reward. Our findings also showed that intra-CA1 or intra-medial prefrontal cortex, but not intra-basolateral amygdala, microinjection of mecamylamine (4 µg/rat) prevented the effect of sub-chronic stress on nicotine reward. These findings suggest that exposure to elevated platform stress potentiates the rewarding effect of nicotine which may be associated with the involvement of nicotinic acetylcholine receptors. It seems that there is a different contribution of the basolateral amygdala, the medial prefrontal cortex or the CA1 nicotinic acetylcholine receptors in stress-induced potentiation of nicotine-induced conditioned place preference.

  15. Autoradiographic visualization of extrajunctional acetylcholine receptors in whole human biceps brachii muscle

    International Nuclear Information System (INIS)

    Askmark, H.; Gillberg, P.-G.; Aquilonius, S.-M.

    1985-01-01

    The morphological distribution of acetylcholine receptors in the whole biceps brachii muscle from a patient with amyotrophic lateral sclerosis (ALS) and from a control patient was studied by in vitro autoradiography with 3 H-alpha-bungarotoxin alphaBtx). In ALS, 3 H-alpha-Btx binding occurred over the entire muscle, while in the control case the binding was restricted to the motor end-plate region. (author)

  16. Resistência de Haemonchus placei, Cooperia punctata e Oesophagostomum radiatum à ivermectina pour-on a 500mcgkg-1 em rebanhos bovinos no Brasil

    Directory of Open Access Journals (Sweden)

    Welber Daniel Zanetti Lopes

    2014-05-01

    Full Text Available O amplo espectro de ação dos endectocidas e a praticidade de aplicação das suas formulações pour-on são fatores que têm estimulado uma maior frequência de utilização destes produtos na criação de bovinos. Neste estudo, avaliou-se a eficácia anti-helmíntica da ivermectina, administrada via pour-on, 500mcgkg-1, contra nematódeos de bovinos provenientes de diferentes rebanhos. Foram adquiridos 12 bezerros de cada um dos quatro rebanhos selecionados. Todos os 48 bezerros utilizados apresentavam médias de ovos por grama de fezes (OPG maiores que 500, considerando-se três contagens consecutivas. Na sequencia, os animais de cada rebanho foram divididos em dois grupos de seis bovinos cada, sendo um tratado com ivermectina 500mcgkg-1 via pour-on e outro mantido como controle. Quatorze dias depois do tratamento, os bovinos foram submetidos à eutanásia para contagem de endoparasitas. A ivermectina demonstrou resultados nulos de eficácia contra H. placei nos quatro rebanhos. Contra C. punctata, tal formulação teve eficácia nula nos rebanhos provenientes de Jaboticabal,SP, e de Formiga,MG, e eficácias de 75,8% e 58,4% nos rebanhos provenientes de São José do Rio Pardo,SP, e de São Sebastião do Paraíso,MG, respectivamente. Valores de eficácia de 94,2% (Jaboticabal, 0,0% (São José do Rio Pardo, 94,2% (Formiga e 39,2% (São Sebastião do Paraíso foram detectados contra O. radiatum. Com base nos resultados encontrados, pode-se concluir que as quatro populações de Haemonchus placei e de Cooperia punctata avaliadas foram resistentes à ivermectina, administrada via pour-on, 500mcgkg-1. Foram observadas cepas de Oesophagostomum radiatum resistentes à ivermectina em dois dos quatro rebanhos testados.

  17. Nerve terminal contributes to acetylcholine receptor organization at the dystrophic neuromuscular junction of mdx mice.

    Science.gov (United States)

    Marques, Maria Julia; Taniguti, Ana Paula Tiemi; Minatel, Elaine; Neto, Humberto Santo

    2007-02-01

    Changes in the distribution of acetylcholine receptors have been reported to occur at the neuromuscular junction of mdx mice and may be a consequence of muscle fiber regeneration rather than the absence of dystrophin. In the present study, we examined whether the nerve terminal determines the fate of acetylcholine receptor distribution in the dystrophic muscle fibers of mdx mice. The left sternomastoid muscle of young (1-month-old) and adult (6-month-old) mdx mice was injected with 60 microl lidocaine hydrochloride to induce muscle degeneration-regeneration. Some mice had their sternomastoid muscle denervated at the time of lidocaine injection. After 10 days of muscle denervation, nerve terminals and acetylcholine receptors were labeled with 4-Di-2-ASP and rhodamine-alpha-bungarotoxin, respectively, for confocal microscopy. In young mdx mice, 75% (n = 137 endplates) of the receptors were distributed in islands. The same was observed in 100% (n = 114 endplates) of the adult junctions. In denervated-regenerated fibers of young mice, the receptors were distributed as branches in 89% of the endplates (n = 90). In denervated-regenerated fibers of adult mice, the receptors were distributed in islands in 100% of the endplates (n = 100). These findings show that nerve-dependent mechanisms are also involved in the changes in receptor distribution in young dystrophic muscles. In older dystrophic muscles, other factors may play a role in receptor distribution.

  18. The anthelmintic levamisole is an allosteric modulator of human neuronal nicotinic acetylcholine receptors.

    Science.gov (United States)

    Levandoski, Mark M; Piket, Barbara; Chang, Jane

    2003-06-13

    L-[-]-2,3,5,6-Tetrahydro-6-phenylimidazo[2,1b]-thiazole hydrochloride (levamisole) is an anthelmintic that targets the nicotinic acetylcholine receptors of parasitic nematodes. We report here the effects of levamisole on human neuronal alpha 3 beta 2 and alpha 3 beta 4 nicotinic receptors, heterologously expressed in Xenopus oocytes and studied with the voltage clamp method. Applied alone, levamisole was a very weak partial agonist for the two subunit combinations. When co-applied with acetylcholine, micromolar concentrations of levamisole potentiated responses, while millimolar concentrations inhibited them; these effects were complex functions of both acetylcholine and levamisole concentrations. The differences in the levamisole effects on the two receptor combinations suggest that the effects are mediated by the beta subunit. Several combinations of agonist and anthelmintic gave the dual potentiation/inhibition behavior, suggesting that the modulatory effects are general. Levamisole inhibition showed macroscopic characteristics of open channel block. Several results led us to conclude that levamisole potentiation occurs through noncompetitive binding to the receptor. We propose pseudo-site binding for noncompetitive potentiation by levamisole.

  19. Monkey Adrenal Chromaffin Cells Express α6β4* Nicotinic Acetylcholine Receptors

    Science.gov (United States)

    Scadden, Mick´l; Carmona-Hidalgo, Beatriz; McIntosh, J. Michael; Albillos, Almudena

    2014-01-01

    Nicotinic acetylcholine receptors (nAChRs) that contain α6 and β4 subunits have been demonstrated functionally in human adrenal chromaffin cells, rat dorsal root ganglion neurons, and on noradrenergic terminals in the hippocampus of adolescent mice. In human adrenal chromaffin cells, α6β4* nAChRs (the asterisk denotes the possible presence of additional subunits) are the predominant subtype whereas in rodents, the predominant nAChR is the α3β4* subtype. Here we present molecular and pharmacological evidence that chromaffin cells from monkey (Macaca mulatta) also express α6β4* receptors. PCR was used to show the presence of transcripts for α6 and β4 subunits and pharmacological characterization was performed using patch-clamp electrophysiology in combination with α-conotoxins that target the α6β4* subtype. Acetylcholine-evoked currents were sensitive to inhibition by BuIA[T5A,P6O] and MII[H9A,L15A]; α-conotoxins that inhibit α6-containing nAChRs. Two additional agonists were used to probe for the expression of α7 and β2-containing nAChRs. Cells with currents evoked by acetylcholine were relatively unresponsive to the α7-selctive agonist choline but responded to the agonist 5-I-A-85380. These studies provide further insights into the properties of natively expressed α6β4* nAChRs. PMID:24727685

  20. Dopamine modulates acetylcholine release via octopamine and CREB signaling in Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Satoshi Suo

    Full Text Available Animals change their behavior and metabolism in response to external stimuli. cAMP response element binding protein (CREB is a signal-activated transcription factor that enables the coupling of extracellular signals and gene expression to induce adaptive changes. Biogenic amine neurotransmitters regulate CREB and such regulation is important for long-term changes in various nervous system functions, including learning and drug addiction. In Caenorhabditis elegans, the amine neurotransmitter octopamine activates a CREB homolog, CRH-1, in cholinergic SIA neurons, whereas dopamine suppresses CREB activation by inhibiting octopamine signaling in response to food stimuli. However, the physiological role of this activation is unknown. In this study, the effect of dopamine, octopamine, and CREB on acetylcholine signaling was analyzed using the acetylcholinesterase inhibitor aldicarb. Mutants with decreased dopamine signaling exhibited reduced acetylcholine signaling, and octopamine and CREB functioned downstream of dopamine in this regulation. This study demonstrates that the regulation of CREB by amine neurotransmitters modulates acetylcholine release from the neurons of C. elegans.

  1. Pancreatic and snake venom presynaptically active phospholipases A2 inhibit nicotinic acetylcholine receptors.

    Science.gov (United States)

    Vulfius, Catherine A; Kasheverov, Igor E; Kryukova, Elena V; Spirova, Ekaterina N; Shelukhina, Irina V; Starkov, Vladislav G; Andreeva, Tatyana V; Faure, Grazyna; Zouridakis, Marios; Tsetlin, Victor I; Utkin, Yuri N

    2017-01-01

    Phospholipases A2 (PLA2s) are enzymes found throughout the animal kingdom. They hydrolyze phospholipids in the sn-2 position producing lysophospholipids and unsaturated fatty acids, agents that can damage membranes. PLA2s from snake venoms have numerous toxic effects, not all of which can be explained by phospholipid hydrolysis, and each enzyme has a specific effect. We have earlier demonstrated the capability of several snake venom PLA2s with different enzymatic, cytotoxic, anticoagulant and antiproliferative properties, to decrease acetylcholine-induced currents in Lymnaea stagnalis neurons, and to compete with α-bungarotoxin for binding to nicotinic acetylcholine receptors (nAChRs) and acetylcholine binding protein. Since nAChRs are implicated in postsynaptic and presynaptic activities, in this work we probe those PLA2s known to have strong presynaptic effects, namely β-bungarotoxin from Bungarus multicinctus and crotoxin from Crotalus durissus terrificus. We also wished to explore whether mammalian PLA2s interact with nAChRs, and have examined non-toxic PLA2 from porcine pancreas. It was found that porcine pancreatic PLA2 and presynaptic β-bungarotoxin blocked currents mediated by nAChRs in Lymnaea neurons with IC50s of 2.5 and 4.8 μM, respectively. Crotoxin competed with radioactive α-bungarotoxin for binding to Torpedo and human α7 nAChRs and to the acetylcholine binding protein. Pancreatic PLA2 interacted similarly with these targets; moreover, it inhibited radioactive α-bungarotoxin binding to the water-soluble extracellular domain of human α9 nAChR, and blocked acetylcholine induced currents in human α9α10 nAChRs heterologously expressed in Xenopus oocytes. These and our earlier results show that all snake PLA2s, including presynaptically active crotoxin and β-bungarotoxin, as well as mammalian pancreatic PLA2, interact with nAChRs. The data obtained suggest that this interaction may be a general property of all PLA2s, which should be proved by

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

    Science.gov (United States)

    Collin, Caitlin; Hauser, Frank; Gonzalez de Valdivia, Ernesto; de Valdivia, Ernesto Gonzalez; Li, Shizhong; Reisenberger, Julia; Carlsen, Eva M M; Khan, Zaid; Hansen, Niels O; Puhm, Florian; Søndergaard, Leif; Niemiec, Justyna; Heninger, Magdalena; Ren, Guilin R; Grimmelikhuijzen, Cornelis J P

    2013-09-01

    Muscarinic acetylcholine receptors (mAChRs) play a central role in the mammalian nervous system. These receptors are G protein-coupled receptors (GPCRs), which are activated by the agonists acetylcholine and muscarine, and blocked by a variety of antagonists. Mammals have five mAChRs (m1-m5). In this study, we cloned two structurally related GPCRs from the fruit fly Drosophila melanogaster, which, after expression in Chinese hamster ovary cells, proved to be muscarinic acetylcholine receptors. One mAChR (the A-type; encoded by gene CG4356) is activated by acetylcholine (EC50, 5 × 10(-8) M) and muscarine (EC50, 6 × 10(-8) M) and blocked by the classical mAChR antagonists atropine, scopolamine, and 3-quinuclidinyl-benzilate (QNB), while the other (the B-type; encoded by gene CG7918) is also activated by acetylcholine, but has a 1,000-fold lower sensitivity to muscarine, and is not blocked by the antagonists. A- and B-type mAChRs were also cloned and functionally characterized from the red flour beetle Tribolium castaneum. Recently, Haga et al. (Nature 2012, 482: 547-551) published the crystal structure of the human m2 mAChR, revealing 14 amino acid residues forming the binding pocket for QNB. These residues are identical between the human m2 and the D. melanogaster and T. castaneum A-type mAChRs, while many of them are different between the human m2 and the B-type receptors. Using bioinformatics, one orthologue of the A-type and one of the B-type mAChRs could also be found in all other arthropods with a sequenced genome. Protostomes, such as arthropods, and deuterostomes, such as mammals and other vertebrates, belong to two evolutionarily distinct lineages of animal evolution that split about 700 million years ago. We found that animals that originated before this split, such as cnidarians (Hydra), had two A-type mAChRs. From these data we propose a model for the evolution of mAChRs.

  3. Pancreatic and snake venom presynaptically active phospholipases A2 inhibit nicotinic acetylcholine receptors.

    Directory of Open Access Journals (Sweden)

    Catherine A Vulfius

    Full Text Available Phospholipases A2 (PLA2s are enzymes found throughout the animal kingdom. They hydrolyze phospholipids in the sn-2 position producing lysophospholipids and unsaturated fatty acids, agents that can damage membranes. PLA2s from snake venoms have numerous toxic effects, not all of which can be explained by phospholipid hydrolysis, and each enzyme has a specific effect. We have earlier demonstrated the capability of several snake venom PLA2s with different enzymatic, cytotoxic, anticoagulant and antiproliferative properties, to decrease acetylcholine-induced currents in Lymnaea stagnalis neurons, and to compete with α-bungarotoxin for binding to nicotinic acetylcholine receptors (nAChRs and acetylcholine binding protein. Since nAChRs are implicated in postsynaptic and presynaptic activities, in this work we probe those PLA2s known to have strong presynaptic effects, namely β-bungarotoxin from Bungarus multicinctus and crotoxin from Crotalus durissus terrificus. We also wished to explore whether mammalian PLA2s interact with nAChRs, and have examined non-toxic PLA2 from porcine pancreas. It was found that porcine pancreatic PLA2 and presynaptic β-bungarotoxin blocked currents mediated by nAChRs in Lymnaea neurons with IC50s of 2.5 and 4.8 μM, respectively. Crotoxin competed with radioactive α-bungarotoxin for binding to Torpedo and human α7 nAChRs and to the acetylcholine binding protein. Pancreatic PLA2 interacted similarly with these targets; moreover, it inhibited radioactive α-bungarotoxin binding to the water-soluble extracellular domain of human α9 nAChR, and blocked acetylcholine induced currents in human α9α10 nAChRs heterologously expressed in Xenopus oocytes. These and our earlier results show that all snake PLA2s, including presynaptically active crotoxin and β-bungarotoxin, as well as mammalian pancreatic PLA2, interact with nAChRs. The data obtained suggest that this interaction may be a general property of all PLA2s, which

  4. Acetylcholine Attenuates Hypoxia/ Reoxygenation-Induced Mitochondrial and Cytosolic ROS Formation in H9c2 Cells via M2 Acetylcholine Receptor

    Directory of Open Access Journals (Sweden)

    Yi Miao

    2013-02-01

    Full Text Available Background: The anti-infammatory and cardioprotective effect of acetylcholine (ACh has been reported; nevertheless, whether and how ACh exhibits an antioxidant property against ischemia/reperfusion (I/R-induced oxidative stress remains obscure. Methods: In the present study, H9c2 rat cardiomyocytes were exposed to hypoxia/reoxygenation (H/R to mimic I/R injury. We estimated intracellular different sources of reactive oxygen species (ROS by measuring mitochondrial ROS (mtROS, mitochondrial DNA (mtDNA copy number, xanthine oxidase (XO and NADPH oxidase (NOX activity and expression of rac 1. Cell injury was determined by lactate dehydrogenase (LDH release and cleaved caspase-3 expression. The siRNA transfection was performed to knockdown of M2 acetylcholine receptor (M2 AChR expression. Results: 12-h hypoxia followed by 2-h reoxygenation resulted in an abrupt burst of ROS in H9c2 cells. Administration of ACh reduced the levels of ROS in a concentration-dependent manner. Compared to the H/R group, ACh decreased mtROS, recovered mtDNA copy number, diminished XO and NOX activity, rac 1 expression as well as cell injury. Co- treatment with atropine rather than hexamethonium abolished the antioxidant and cardioprotective effect of ACh. Moreover, knockdown of M2 AChR by siRNA showed the similar trends as atropine co-treatment group. Conclusions: ACh inhibits mitochondria-, XO- and NOX-derived ROS production thus protecting H9c2 cells against H/R-induced oxidative stress, and these benefcial effects are mainly mediated by M2 AChR. Our findings suggested that increasing ACh release could be a potential therapeutic strategy for treatment and prevention of I/R injury.

  5. Local application of SCH 39166 reversibly and dose-dependently decreases acetylcholine release in the rat striatum.

    Science.gov (United States)

    Acquas, E; Di Chiara, G

    1999-11-03

    The effect of local application by reverse dialysis of the dopamine D(1) receptor antagonist (-)-trans-6,7,7a,8,9, 13b-exahydro-3-chloro-2-hydroxy-N-methyl-5H-benzo-[d]-nap hto-[2, 1b]-azepine hydrochloride (SCH 39166) on acetylcholine release was studied in awake, freely moving rats implanted with concentric microdialysis probes in the dorsal striatum. In these experiments, the reversible acetylcholine esterase inhibitor, neostigmine, was added to the perfusion solution at two different concentrations, 0.01 and 0.1 microM. SCH 39166 (1, 5 and 10 microM), in the presence of 0.01 microM neostigmine, reversibly decreased striatal acetylcholine release (1 microM SCH 39166 by 8+/-4%; 5 microM SCH 39166 by 24+/-5%; 10 microM SCH 39166 by 27+/-7%, from basal). Similarly, SCH 39166, applied in the presence of a higher neostigmine concentration (0.1 microM), decreased striatal acetylcholine release by 14+/-4% at 1 microM, by 28+/-8% at 5 microM and by 30+/-5% at 10 microM, in a dose-dependent and time-dependent manner. These results are consistent with the existence of a facilitatory tone of dopamine on striatal acetylcholine transmission mediated by dopamine D(1) receptors located on striatal cholinergic interneurons.

  6. Investigation of the presence and antinociceptive function of muscarinic acetylcholine receptors in the African naked mole-rat (Heterocephalus glaber).

    Science.gov (United States)

    Jørgensen, Kristine B; Krogh-Jensen, Karen; Pickering, Darryl S; Kanui, Titus I; Abelson, Klas S P

    2016-01-01

    The present study investigated the cholinergic system in the African naked mole-rat (Heterocephalus glaber) with focus on the muscarinic acetylcholine receptor subtypes M1 and M4. The protein sequences for the subtypes m 1-5 of the naked mole-rat were compared to that of the house mouse (Mus musculus) using basic local alignment search tool (BLAST). The presence and function of M1 and M4 was investigated in vivo, using the formalin test with the muscarinic receptor agonists xanomeline and VU0152100. Spinal cord tissue from the naked mole-rat was used for receptor saturation binding studies with [(3)H]-N-methylscopolamine. The BLAST test revealed 95 % protein sequence homology showing the naked mole-rat to have the genetic potential to express all five muscarinic acetylcholine receptor subtypes. A significant reduction in pain behavior was demonstrated after administration of 8.4 mg/kg in the formalin test. Administration of 50 mg/kg VU0152100 resulted in a non-significant tendency towards antinociception. The antinociceptive effects were reversed by the muscarinic acetylcholine receptor antagonist atropine. Binding studies indicated presence of muscarinic acetylcholine receptors with a radioligand affinity comparable to that reported in mice. In conclusion, muscarinic acetylcholine receptor subtypes are present in the naked mole-rat and contribute to antinociception in the naked mole-rat.

  7. A neuronal acetylcholine receptor regulates the balance of muscle excitation and inhibition in Caenorhabditis elegans.

    Directory of Open Access Journals (Sweden)

    Maelle Jospin

    2009-12-01

    Full Text Available In the nematode Caenorhabditis elegans, cholinergic motor neurons stimulate muscle contraction as well as activate GABAergic motor neurons that inhibit contraction of the contralateral muscles. Here, we describe the composition of an ionotropic acetylcholine receptor that is required to maintain excitation of the cholinergic motor neurons. We identified a gain-of-function mutation that leads to spontaneous muscle convulsions. The mutation is in the pore domain of the ACR-2 acetylcholine receptor subunit and is identical to a hyperactivating mutation in the muscle receptor of patients with myasthenia gravis. Screens for suppressors of the convulsion phenotype led to the identification of other receptor subunits. Cell-specific rescue experiments indicate that these subunits function in the cholinergic motor neurons. Expression of these subunits in Xenopus oocytes demonstrates that the functional receptor is comprised of three alpha-subunits, UNC-38, UNC-63 and ACR-12, and two non-alpha-subunits, ACR-2 and ACR-3. Although this receptor exhibits a partially overlapping subunit composition with the C. elegans muscle acetylcholine receptor, it shows distinct pharmacology. Recordings from intact animals demonstrate that loss-of-function mutations in acr-2 reduce the excitability of the cholinergic motor neurons. By contrast, the acr-2(gf mutation leads to a hyperactivation of cholinergic motor neurons and an inactivation of downstream GABAergic motor neurons in a calcium dependent manner. Presumably, this imbalance between excitatory and inhibitory input into muscles leads to convulsions. These data indicate that the ACR-2 receptor is important for the coordinated excitation and inhibition of body muscles underlying sinusoidal movement.

  8. Prostate stem cell antigen interacts with nicotinic acetylcholine receptors and is affected in Alzheimer's disease

    DEFF Research Database (Denmark)

    Jensen, Majbrit Myrup; Mikkelsen, Jens D.; Arvaniti, Maria

    2015-01-01

    Alzheimer's disease (AD) is a neurodegenerative disorder involving impaired cholinergic neurotransmission and dysregulation of nicotinic acetylcholine receptors (nAChRs). Ly-6/neurotoxin (Lynx) proteins have been shown to modulate cognition and neural plasticity by binding to nAChR subtypes...... are present in the human brain. We further showed that PSCA forms stable complexes with the α4 nAChR subunit and decreases nicotine-induced extracellular-signal regulated kinase phosphorylation in PC12 cells. In addition, we analyzed protein levels of PSCA and Lypd6 in postmortem tissue of medial frontal...

  9. Lynx1 and Aβ1-42 bind competitively to multiple nicotinic acetylcholine receptor subtypes

    DEFF Research Database (Denmark)

    Thomsen, Morten S; Arvaniti, Maria; Jensen, Majbrit M

    2016-01-01

    Lynx1 regulates synaptic plasticity in the brain by regulating nicotinic acetylcholine receptors (nAChRs). It is not known to which extent Lynx1 can bind to endogenous nAChR subunits in the brain or how this interaction is affected by Alzheimer's disease pathology. We apply affinity purification....... Incubation with Ws-Lynx1 decreases nicotine-mediated extracellular signal-regulated kinase phosphorylation in PC12 cells and striatal neurons, indicating that binding of Ws-Lynx1 is sufficient to inhibit signaling downstream of nAChRs. The effect of nicotine in PC12 cells is independent of α7 or α4β2 n...

  10. Regional distribution of muscarinic acetylcholine receptors in the telencephalon of the pigeon (Columba livia f. domestica)

    International Nuclear Information System (INIS)

    Waechtler, K.

    1985-01-01

    The distribution of muscarinic acetylcholine receptors was studied autoradiographically in croystat sections of the pigeon telencephalon using 3 H-quinuclidinylbenzylate as a ligand. Highest receptor density was observed in the hyperstriatum ventrale, palaeostriatum augmentatum, septum, and parts of the archistriatum. In sites of known sensory input of neostriatum (field L) and ectostriatum low receptor binding was observed. Acetylcholinesterase distribution is in good agreement with the receptor picture only in the basal telencephalon. In the pallium differences in the pattern of these two components can be seen. (author)

  11. Changes in acetylcholine release from the chick retina are not associated with myopia development

    International Nuclear Information System (INIS)

    Vessey, K.A.; Cotriall, C.L.; McBrien, N.A.

    2002-01-01

    Full text: The effectiveness of muscarinic receptor antagonists in inhibiting myopia progression in animal models and humans implicates cholinergic signalling in ocular growth regulation. Therefore to determine if changes in the release of acetylcholine from the retina are involved in myopia development, the efflux of acetylcholine from the in vitro retina of normal and myopic chick eyes was investigated. Chicks were monocularly deprived (MD) of pattern vision with translucent occluders for 2 or 7 days and refractive error of MD groups and age matched normals was monitored using retinoscopy (n=6 each group). 3 H-choline-Cl (1 Ci in 7μL) was injected into the vitreous of each eye under 2.5% halothane anaesthesia. After 1hr, the eyes were enucleated, under terminal anaesthesia (sodium pentobarbital, 120 mg/kg, im). Retinas were flat-mounted on acetate filter discs and superfused with oxygenated physiological saline solution (PSS) for 30min at 0.4mL/min. Five baseline fractions were collected (B1-B5), then three stimulated fractions were collected in the presence of PSS containing 50mM KCl (K1-K3) at 2min intervals. 3 H-acetylcholine ( 3 H-ACh) in each fraction was quantified by liquid scintillation counting. Significant amounts of myopia were induced in MD eyes after 2 (-5.1±0.8D) and 7 days (-18.8±2.4D) relative to control eyes (paired t-test p 3 H-ACh release was 146±15% above basal levels (K2/B1%) from retinas of normal animals. After 2 days MD, there was no significant difference between KCl-evoked release of 3 H-ACh from deprived eyes (147 39%) compared to control eyes (198±61%, paired t-test, p=0.27) or the eyes of normal animals (ANOVA, p>0.5). Similar results were obtained following 7 days MD. The results demonstrate that evoked acetylcholine release from the chick retina of myopic eyes is unaltered relative to control or normal eyes using an in vitro approach. Copyright (2002) Australian Neuroscience Society

  12. Temperature effect on proximal to distal gradient of quantal release of acetylcholine at frog endplate

    Czech Academy of Sciences Publication Activity Database

    Samigullin, D.; Bukharaeva, E.; Nikolsky, E.; Vyskočil, František

    2003-01-01

    Roč. 28, 3-4 (2003), s. 507-514 ISSN 0364-3190 R&D Projects: GA AV ČR IAA7011902; GA ČR GA305/02/1333; GA ČR GA202/02/1213 Grant - others:RFBR(RU) 02/04/48901 Institutional research plan: CEZ:AV0Z5011922; CEZ:MSM 113100003 Keywords : neuromuscular junction * acetylcholine release * temperature Subject RIV: ED - Physiology Impact factor: 1.511, year: 2003

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

    OpenAIRE

    Du, Xiaoyi; Schoemaker, Regien; Bos, Egbert; Saxena, Pramod Ranjan

    1995-01-01

    textabstractIn the human isolated myocardium, acetylcholine (10−9 to 10−3 M) elicited a biphasic inotropic effect (a decrease in the lower and an increase in the higher concentration range) in atrial and a positive inotropic effect in ventricular trabeculae. However, under conditions of raised contractility achieved by exposure to noradrenaline (10−5 M), only negative inotropic effects were observed in both atria and ventricles. Atropine (10−6 M), but not propranolol (10−6 M), antagonized bot...

  14. [[sup 3]H]imidacloprid: synthesis of a candidate radioligand for the nicotinic acetylcholine receptor

    Energy Technology Data Exchange (ETDEWEB)

    Latli, B.; Casida, J.E. (California Univ., Berkeley, CA (United States). Dept. of Entomological Sciences)

    1992-08-01

    Imidacloprid is an exceptionally potent insecticide known from physiological studies to act at the nicotinic acetylcholine receptor. To prepare [[sup 3]H]imidacloprid as a candidate radioligand, 6-chloronicotinoyl chloride was reduced with NaB[sup 2]H[sub 4] (in model studies) or NaB[sup 3]H[sub 4] in absolute ethanol to 2-chloro-5-pyridinylmethanol which was transformed to 2-chloro-5-chloromethylpyridine on refluxing with thionyl chloride. Coupling with 4,5-dihydro-N-nitro-1H-imidazol-2-amine then gave [[sup 2]H[sub 2

  15. INFLUENCE OF ANTIBIOTICS ON THE MECHANICAL RESPONSES OF GUINEA-PIG ILEUM TO ACETYLCHOLINE AND HISTAMINE

    Directory of Open Access Journals (Sweden)

    Petroianu Andy

    1998-01-01

    Full Text Available The side effects of antibiotics have been extensively described during the last decades, however, their role on digestive motility must be better investigated. Following a line of research, the influence of penicillin, chloranfenicol tetracycline and gentamicine on longitudinal smooth muscle responses to acetylcholine and histamine were studied on guinea-pig ileum. There were no differences between the responses before and after the addition of each antibiotic. Further investigations must be performed in order to find a possible influence of antibiotics on digestive motility.

  16. Drugs interfering with Muscarinic Acetylcholine Receptors and Their effects on Place Navigation

    Czech Academy of Sciences Publication Activity Database

    Svoboda, Jan; Popelíková, Anna; Stuchlík, Aleš

    2017-01-01

    Roč. 8, Nov 9 (2017), č. článku 215. ISSN 1664-0640 R&D Projects: GA ČR(CZ) GA17-04047S; GA ČR NV17-30833A; GA MŠk(CZ) LM2015062 Grant - others:AV ČR(CZ) PAN-17-07 Program:Bilaterální spolupráce Institutional support: RVO:67985823 Keywords : scopolamine * biperiden * acetylcholine * receptor * behavior * learning Subject RIV: FH - Neurology OBOR OECD: Neurosciences (including psychophysiology Impact factor: 3.532, year: 2016

  17. Muscarinic Acetylcholine Receptor M3 Mutation Causes Urinary Bladder Disease and a Prune-Belly-like Syndrome.

    Science.gov (United States)

    Weber, Stefanie; Thiele, Holger; Mir, Sevgi; Toliat, Mohammad Reza; Sozeri, Betül; Reutter, Heiko; Draaken, Markus; Ludwig, Michael; Altmüller, Janine; Frommolt, Peter; Stuart, Helen M; Ranjzad, Parisa; Hanley, Neil A; Jennings, Rachel; Newman, William G; Wilcox, Duncan T; Thiel, Uwe; Schlingmann, Karl Peter; Beetz, Rolf; Hoyer, Peter F; Konrad, Martin; Schaefer, Franz; Nürnberg, Peter; Woolf, Adrian S

    2011-11-11

    Urinary bladder malformations associated with bladder outlet obstruction are a frequent cause of progressive renal failure in children. We here describe a muscarinic acetylcholine receptor M3 (CHRM3) (1q41-q44) homozygous frameshift mutation in familial congenital bladder malformation associated with a prune-belly-like syndrome, defining an isolated gene defect underlying this sometimes devastating disease. CHRM3 encodes the M3 muscarinic acetylcholine receptor, which we show is present in developing renal epithelia and bladder muscle. These observations may imply that M3 has a role beyond its known contribution to detrusor contractions. This Mendelian disease caused by a muscarinic acetylcholine receptor mutation strikingly phenocopies Chrm3 null mutant mice. Copyright © 2011 The American Society of Human Genetics. Published by Elsevier Inc. All rights reserved.

  18. Autonomous control of phosphatidylinositol turnover by histamine and acetylcholine receptors in the NIE-115 neuron-like cell line

    International Nuclear Information System (INIS)

    Large, T.H.; Lambert, M.P.; Cohen, N.M.; Klein, W.L.

    1986-01-01

    Histamine was found to stimulate the turnover of phosphatidylinositol (PI) in cultures of neuron-like NE-115 cells. Turnover was measured by increased production of ( 3 H)inositol phosphates (breakdown) and by accelerated incorporation of 32 P into PI (resynthesis). Data were consistent with hydrolysis of polyphosphoinositides being the initial event in receptor-stimulated PI turnover. This response to histamine desensitized within 10 min. Receptor systems for histamine and acetylcholine were tested for possible interactions: PI turnover in response to dual stimulation was approximately equal to the sum of the individual responses while prior desensitization of the acetylcholine receptor system had no effect on subsequent stimulation of the histamine receptor system. These results are consistent with the hypothesis that components of acetylcholine and histamine receptor systems responsible for PI turnover are autonomously organised and regulated. (author)

  19. An extract of lionfish (Pterois volitans) spine tissue contains acetylcholine and a toxin that affects neuromuscular transmission.

    Science.gov (United States)

    Cohen, A S; Olek, A J

    1989-01-01

    A soluble toxic extract derived from spine tissue of the lionfish (Pterois volitans) decreased heart rate and force of contraction in isolated clam and frog hearts. These actions were due to the presence of micromolar concentrations of acetylcholine in the extract. Toxicity was retained after hydrolysis of acetylcholine by exogenous acetylcholinesterase, but heart function was no longer affected. Toxin treated in this way induced muscle fibrillation in an isolated nerve-muscle preparation, followed by blockade of neuromuscular transmission. Bursts of transient depolarizations were recorded at the muscle endplate shortly after toxin addition that correlated in time with the duration of toxin-induced muscle fibrillation. These effects are thought to be due to the increased release and then depletion of acetylcholine from the nerve terminal.

  20. Analogues of neuroactive polyamine wasp toxins that lack inner basic sites exhibit enhanced antagonism toward a muscle-type mammalian nicotinic acetylcholine receptor

    DEFF Research Database (Denmark)

    Stromgaard, K; Brierley, M J; Andersen, K

    1999-01-01

    noncompetitively antagonized the nicotinic acetylcholine receptor (nAChR) in a concentration-, time-, and voltage-dependent manner. The amplitudes of acetylcholine-induced currents were compared at their peaks and at the end of a 1 s application in the presence or absence of the analogues. Most of the analogues...

  1. Human Secreted Ly-6/uPAR Related Protein-1 (SLURP-1) Is a Selective Allosteric Antagonist of α7 Nicotinic Acetylcholine Receptor

    DEFF Research Database (Denmark)

    Lyukmanova, Ekaterina N; Shulepko, Mikhail A; Kudryavtsev, Denis

    2016-01-01

    of nicotinic acetylcholine receptors (nAChRs) and muscarinic acetylcholine receptors, respectively, and anti-α7-nAChRs antibodies revealed α7 type nAChRs as an rSLURP-1 target in keratinocytes. Using affinity purification from human cortical extracts, we confirmed that rSLURP-1 binds selectively to the α7-n...

  2. The A- and B-type muscarinic acetylcholine receptors from Drosophila melanogaster couple to different second messenger pathways

    DEFF Research Database (Denmark)

    Ren, Guilin Robin; Folke, Jonas; Hauser, Frank

    2015-01-01

    Muscarinic acetylcholine receptors (mAChRs) are G protein-coupled receptors (GPCRs) that are activated by the agonists acetylcholine and muscarine and blocked by several antagonists, among them atropine. In mammals five mAChRs (m1-m5) exist of which m1, m3, and m5 are coupled to members of the Gq...

  3. Effects of BMS-902483, an α7 nicotinic acetylcholine receptor partial agonist, on cognition and sensory gating in relation to receptor occupancy in rodents.

    Science.gov (United States)

    Pieschl, Rick L; Miller, Regina; Jones, Kelli M; Post-Munson, Debra J; Chen, Ping; Newberry, Kimberly; Benitex, Yulia; Molski, Thaddeus; Morgan, Daniel; McDonald, Ivar M; Macor, John E; Olson, Richard E; Asaka, Yukiko; Digavalli, Siva; Easton, Amy; Herrington, James; Westphal, Ryan S; Lodge, Nicholas J; Zaczek, Robert; Bristow, Linda J; Li, Yu-Wen

    2017-07-15

    The α7 nicotinic acetylcholine receptor is thought to play an important role in human cognition. Here we describe the in vivo effects of BMS-902483, a selective potent α7 nicotinic acetylcholine receptor partial agonist, in relationship to α7 nicotinic acetylcholine receptor occupancy. BMS-902483 has low nanomolar affinity for rat and human α7 nicotinic acetylcholine receptors and elicits currents in cells expressing human or rat α7 nicotinic acetylcholine receptors that are about 60% of the maximal acetylcholine response. BMS-902483 improved 24h novel object recognition memory in mice with a minimal effective dose (MED) of 0.1mg/kg and reversed MK-801-induced deficits in a rat attentional set-shifting model of executive function with an MED of 3mg/kg. Enhancement of novel object recognition was blocked by the silent α7 nicotinic acetylcholine receptor agonist, NS6740, demonstrating that activity of BMS-902483 was mediated by α7 nicotinic acetylcholine receptors. BMS-902483 also reversed ketamine-induced deficits in auditory gating in rats, and enhanced ex vivo hippocampal long-term potentiation examined 24h after dosing in mice. Results from an ex vivo brain homogenate binding assay showed that α7 receptor occupancy ranged from 64% (novel object recognition) to ~90% (set shift and gating) at the MED for behavioral and sensory processing effects of BMS-902483. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Interaction of alpha-conotoxin ImII and its analogs with nicotinic receptors and acetylcholine-binding proteins: additional binding sites on Torpedo receptor

    NARCIS (Netherlands)

    Kasheverov, I.E.; Zhmak, M.N.; Fish, A.; Rucktooa, P.; Khruschov, A.Y.; Osipov, A.V.; Ziganshin, R.H.; D'Hoedt, D.; Bertrand, D.; Sixma, T.K.; Smit, A.B.; Tsetlin, V.I.

    2009-01-01

    α-Conotoxins interact with nicotinic acetylcholine receptors (nAChRs) and acetylcholine-binding proteins (AChBPs) at the sites for agonists/competitive antagonists. α-Conotoxins blocking muscle-type or α7 nAChRs compete with α-bungarotoxin. However, α-conotoxin ImII, a close homolog of the α7

  5. Topological dispositions of lysine α380 and lysine γ486 in the acetylcholine receptor from Torpedo californica

    International Nuclear Information System (INIS)

    Dwyer, B.P.

    1991-01-01

    The locations have been determined, with respect to the plasma membrane, of lysine α380 and lysine γ486 in the α subunit and the γ subunit, respectively, of the nicotinic acetylcholine receptor from Torpedo californica. Immunoadsorbents were constructed that recognize the carboxy terminus of the peptide GVKYIAE released by proteolytic digestion from positions 378-384 in the amino acid sequence of the α subunit of the acetylcholine receptor and the carboxy terminus of the peptide KYVP released by proteolytic digestion from positions 486-489 in the amino acid sequence of the γ subunit. They were used to isolate these peptides from proteolytic digests of polypeptides from the acetylcholine receptor. Sealed vesicles containing the native acetylcholine receptor were labeled with pyridoxal phosphate and sodium [ 3 H]-borohydride. The effect of saponin on the incorporation of pyridoxamine phosphate into lysine α380 and lysine γ486 from the acetylcholine receptor in these vesicles was assessed with the immunoadsorbents. The conclusions that follow from these results are that lysine α380 is on the inside surface of a vesicle and lysine γ486 is on the outside surface. Because a majority (85%) of the total binding sites for α-bungarotoxin bind the toxin in the absence of saponin, the majority of the vesicles are right side out with the inside of the vesicle corresponding to the cytoplasmic surface and the outside of the vesicle corresponding to the extracytoplasmic, synaptic surface. Because lysine α380 and lysine γ486 lie on opposite sides of the membrane, a membrane-spanning segment must be located between the two positions occupied by these two amino acids in the common sequence of a polypeptide of the acetylcholine receptor

  6. Critical Evaluation of Acetylcholine Determination in Rat Brain Microdialysates using Ion-Pair Liquid Chromatography with Amperometric Detection

    Directory of Open Access Journals (Sweden)

    Yvette Michotte

    2008-08-01

    Full Text Available Liquid chromatography with amperometric detection remains the most widely used method for acetylcholine quantification in microdialysis samples. Separation of acetylcholine from choline and other matrix components on a microbore chromatographic column (1 mm internal diameter, conversion of acetylcholine in an immobilized enzyme reactor and detection of the produced hydrogen peroxide on a horseradish peroxidase redox polymer coated glassy carbon electrode, achieves sufficient sensitivity for acetylcholine quantification in rat brain microdialysates. However, a thourough validation within the concentration range required for this application has not been carried out before. Furthermore, a rapid degradation of the chromatographic columns and enzyme systems have been reported. In the present study an ion-pair liquid chromatography assay with amperometric detection was validated and its long-term stability evaluated. Working at pH 6.5 dramatically increased chromatographic stability without a loss in sensitivity compared to higher pH values. The lower limit of quantification of the method was 0.3 nM. At this concentration the repeatability was 15.7%, the inter-day precision 8.7% and the accuracy 103.6%. The chromatographic column was stable over 4 months, the immobilized enzyme reactor up to 2-3 months and the enzyme coating of the amperometric detector up to 1-2 months. The concentration of acetylcholine in 30 μl microdialysates obtained under basal conditions from the hippocampus of freely moving rats was 0.40 ± 0.12 nM (mean ± SD, n = 30. The present method is therefore suitable for acetylcholine determination in rat brain microdialysates.

  7. A synthetic combinatorial strategy for developing a-conotoxin analogs as potent a7 nicotinic acetylcholine receptor antagonists

    DEFF Research Database (Denmark)

    Armishaw, Christopher J; Singh, Narender; Medina-Franco, Jose L

    2010-01-01

    alpha-Conotoxins are peptide neurotoxins isolated from venomous cone snails that display exquisite selectivity for different subtypes of nicotinic acetylcholine receptors (nAChR). They are valuable research tools that have profound implications in the discovery of new drugs for a myriad of neurop......alpha-Conotoxins are peptide neurotoxins isolated from venomous cone snails that display exquisite selectivity for different subtypes of nicotinic acetylcholine receptors (nAChR). They are valuable research tools that have profound implications in the discovery of new drugs for a myriad...

  8. Differential Regulation of Receptor Activation and Agonist Selectivity by Highly Conserved Tryptophans in the Nicotinic Acetylcholine Receptor Binding Site

    OpenAIRE

    Williams, Dustin K.; Stokes, Clare; Horenstein, Nicole A.; Papke, Roger L.

    2009-01-01

    We have shown previously that a highly conserved Tyr in the nicotinic acetylcholine receptor (nAChR) ligand-binding domain (LBD) (α7 Tyr188 or α4 Tyr195) differentially regulates the activity of acetylcholine (ACh) and the α7-selective agonist 3-(4-hydroxy,2-methoxybenzylidene)anabaseine (4OH-GTS-21) in α4β2 and α7 nAChR. In this study, we mutated two highly conserved LBD Trp residues in human α7 and α4β2 and expressed the receptors in Xenopus laevis oocytes. α7 Re...

  9. Nicotinic acetylcholine receptor polymorphism, smoking behavior, and tobacco-related cancer and lung and cardiovascular diseases: a cohort study

    DEFF Research Database (Denmark)

    Kaur-Knudsen, Diljit; Bojesen, Stig E; Tybjærg-Hansen, Anne

    2011-01-01

    We examined the associations between the nicotinic acetylcholine receptor polymorphism (rs1051730) on chromosome 15q25 marking the gene cluster CHRNA3-CHRNB4-CHRNA5, smoking behavior, and tobacco-related cancer and lung and cardiovascular diseases in the general population.......We examined the associations between the nicotinic acetylcholine receptor polymorphism (rs1051730) on chromosome 15q25 marking the gene cluster CHRNA3-CHRNB4-CHRNA5, smoking behavior, and tobacco-related cancer and lung and cardiovascular diseases in the general population....

  10. Procaine rapidly inactivates acetylcholine receptors from Torpedo and competes with agonist for inhibition sites

    International Nuclear Information System (INIS)

    Forman, S.A.; Miller, K.W.

    1989-01-01

    The relationship between the high-affinity procaine channel inhibition site and the agonist self-inhibition site on acetylcholine receptors (AChRs) from Torpedo electroplaque was investigated by using rapid 86 Rb + quenched-flux assays at 4 degree C in native AChR-rich vesicles on which 50-60% of ACh activation sites were blocked with α-bungarotoxin (α-BTX). In the presence of channel-activating acetylcholine (ACh) concentrations alone, AChR undergoes one phase of inactivation in under a second. Addition of procaine produces two-phase inactivation similar to that seen with self-inhibiting ACh concentrations rapid inactivation complete in 30-75 ms is followed by fast desensitization at the same k d observed without procaine. The dependence of k r on [procaine] is consistent with a bimolecular association between procaine and its AChR site. Inhibition of AChR function by mixtures of procaine plus self-inhibiting concentrations of ACh or suberyldicholine was studied by reducing the level of α-BTX block in vesicles. The data support a mechanism where procaine binds preferentially to the open-channel AChR state, since no procaine-induced inactivation is observed without agonist and k r 's dependence on [ACh] in channel-activating range closely parallels that of 86 Rb + flux response to ACh

  11. cap alpha. -bungarotoxin binding properties of a central nervous system nicotinic acetylcholine receptor

    Energy Technology Data Exchange (ETDEWEB)

    Lukasiewicz, R J; Bennett, E L

    1978-01-01

    High-affinity, specific binding of radiolabeled ..cap alpha..-bungarotoxin to particulate fractions derived from rat brain shows saturability (B/sub max/ approx. = 37fmol/mg, K/sub D//sup app/ = 1.7 nM) and insensitivity to ionic strength, and is essentially irreversible (K/sub on/ = 5 x 10/sup 6/ min/sup -1/ x mol/sup -1/; K(displacement) = 1.9 x 10/sup -4/ min/sup -1/, tau/sub 1/2/ = 62 h). Subcellular distribution of specific sites is consistent with their location on synaptic junctional complex and post-synaptic membranes. These membrane-bound binding sites exhibit unique sensitivity to cholinergic ligands; pretreatment of membranes with cholinergic agonists (but not antagonists) induces transformation of ..cap alpha..-bungarotoxin binding sites to a high affinity form toward agonist. The effect is most marked for the natural agonist, acetylcholine. These results strongly support the notion that the entity under study is an authentic nicotinic acetylcholine receptor.

  12. Acetylcholine-induced inhibition of presynaptic calcium signals and transmitter release in the frog neuromuscular junction

    Directory of Open Access Journals (Sweden)

    Eduard Khaziev

    2016-12-01

    Full Text Available Acetylcholine (ACh, released from axonal terminals of motor neurones in neuromuscular junctions regulates the efficacy of neurotransmission through activation of presynaptic nicotinic and muscarinic autoreceptors. Receptor-mediated presynaptic regulation could reflect either direct action on exocytotic machinery or modulation of Ca2+ entry and resulting intra-terminal Ca2+ dynamics. We have measured free intra-terminal cytosolic Ca2+ ([Ca2+]i using Oregon-Green 488 microfluorimetry, in parallel with voltage-clamp recordings of spontaneous (mEPC and evoked (EPC postsynaptic currents in post-junctional skeletal muscle fibre. Activation of presynaptic muscarinic and nicotinic receptors with exogenous acetylcholine and its non-hydrolized analogue carbachol reduced amplitude of the intra-terminal [Ca2+]i transients and decreased quantal content (calculated by dividing the area under EPC curve by the area under mEPC curve. Pharmacological analysis revealed the role of muscarinic receptors of M2 subtype as well as d-tubocurarine-sensitive nicotinic receptor in presynaptic modulation of [Ca2+]i transients. Modulation of synaptic transmission efficacy by ACh receptors was completely eliminated by pharmacological inhibition of N-type Ca2+ channels. We conclude that ACh receptor-mediated reduction of Ca2+ entry into the nerve terminal through N-type Ca2+ channels represents one of possible mechanism of presynaptic modulation in frog neuromuscular junction.

  13. The Anti-Acetylcholine Receptor Antibody Test in Suspected Ocular Myasthenia Gravis

    Directory of Open Access Journals (Sweden)

    Jung Jin Lee

    2014-01-01

    Full Text Available Aim. To estimate the clinical significance of anti-acetylcholine receptor antibody (anti-AChR-Ab levels in suspected ocular myasthenia gravis. Methods. In total, 144 patients complaining of fluctuating diplopia and ptosis were evaluated for serum levels of anti-acetylcholine receptor antibody and their medical charts were retrospectively reviewed. Subjects were classified into three groups: variable diplopia only, ptosis only, and both variable diplopia and ptosis. We investigated serum anti-AChR-Ab titer levels and performed thyroid autoantibody tests. Results. Patients’ chief complaints were diplopia (N=103, ptosis (N=12, and their concurrence (N=29. Abnormal anti-AChR-Ab was observed in 21 of 144 patients (14.1%. Between the three groups, mean age, number of seropositive patients, and mean anti-AChR-Ab level were not significantly different (P=0.224, 0.073, and 0.062, resp.. Overall, 27.5% of patients had abnormal thyroid autoantibodies. Conclusion. The sensitivity of anti-AChR-Ab was 14.1% in suspected ocular myasthenia gravis and seropositivity in myasthenia gravis patients showed a high correlation with the presence of thyroid autoantibodies.

  14. Conformational changes in acetylcholine binding protein investigated by temperature accelerated molecular dynamics.

    Directory of Open Access Journals (Sweden)

    Zeynab Mohammad Hosseini Naveh

    Full Text Available Despite the large number of studies available on nicotinic acetylcholine receptors, a complete account of the mechanistic aspects of their gating transition in response to ligand binding still remains elusive. As a first step toward dissecting the transition mechanism by accelerated sampling techniques, we study the ligand-induced conformational changes of the acetylcholine binding protein (AChBP, a widely accepted model for the full receptor extracellular domain. Using unbiased Molecular Dynamics (MD and Temperature Accelerated Molecular Dynamics (TAMD simulations we investigate the AChBP transition between the apo and the agonist-bound state. In long standard MD simulations, both conformations of the native protein are stable, while the agonist-bound structure evolves toward the apo one if the orientation of few key sidechains in the orthosteric cavity is modified. Conversely, TAMD simulations initiated from the native conformations are able to produce the spontaneous transition. With respect to the modified conformations, TAMD accelerates the transition by at least a factor 10. The analysis of some specific residue-residue interactions points out that the transition mechanism is based on the disruption/formation of few key hydrogen bonds. Finally, while early events of ligand dissociation are observed already in standard MD, TAMD accelerates the ligand detachment and, at the highest TAMD effective temperature, it is able to produce a complete dissociation path in one AChBP subunit.

  15. some nutritional aspects of haemonchosis in experimentally infested ...

    African Journals Online (AJOL)

    nutritional aberration has been described as anorexia in both pure (Evans, Blunt & Southcott, 1963) and mixed infestations where Haemonchus contortus was prominent. (Clark, Ortlepp, Bosman, Laurence, Groenewald & Quin,. 1951; Shumard et al. 1957). Further observations on nutritional aspects of a pure infestation of ...

  16. Browse Title Index

    African Journals Online (AJOL)

    Items 51 - 100 of 238 ... Vol 3, No 1 (2013), Digit loss due to Demodex spp. infestation in a dog: ... Early detection of Haemonchus contortus infection in sheep using three ... of ensiled and dried pomegranate seeds for ruminants using in vitro gas ...

  17. Studies on the Leucocytic Response to Experimental Infection with ...

    African Journals Online (AJOL)

    Also, neutrophil numbers declined significantly (P < 0.05) in red fronted gazelles infected either singly with Trypanosoma brucei or concurrently with both parasites while those infected singly with Haemonchus contortus experienced a significant (P <0.05) rise in neutrophil counts which became evident from day 30 post ...

  18. In vitro anthelmintic effect of two medicinal plants (Anogeissus ...

    African Journals Online (AJOL)

    STORAGESEVER

    2009-09-15

    Sep 15, 2009 ... A study was conducted to evaluate Anogeissus leiocarpus leaf and Daniellia oliveri stem barks as effective remedy for gastrointestinal parasites. The anthelmintic activity of these extracts on eggs, first stage larvae and adults of Haemonchus contortus was examined by in vitro tests. The extracts were.

  19. Epidemiological study on gastrointestinal tract hel- minthosis of ...

    African Journals Online (AJOL)

    zone practice a crop - livestock mixed farming and keep combination of live- ... of animals (Goat and sheep), sex of animals (male and female), age of animals .... Mean worm burden of Haemonchus contortus was significantly high in older ..... high and mid land portion of the study site where sheep were forced to graze.

  20. Examination of commercially available copper oxide wire particles in combination with albendazole for control of gastrointestinal nematodes in lambs

    Science.gov (United States)

    Alternatives to synthetic anthelmintics remain critical due to the prevalence of anthelmintic resistance. The objective of the experiment was to determine the efficacy of copper oxide wire particles (COWP) from three commercial sources to control Haemonchus contortus in lambs. Naturally infected Ka...

  1. Molecular determinants of subtype-selective efficacies of cytisine and the novel compound NS3861 at heteromeric nicotinic acetylcholine receptors

    DEFF Research Database (Denmark)

    Harpsøe, Kasper; Hald, Helle; Timmermann, Daniel B

    2013-01-01

    Deciphering which specific agonist-receptor interactions affect efficacy levels is of high importance, because this will ultimately aid in designing selective drugs. The novel compound NS3861 and cytisine are agonists of nicotinic acetylcholine receptors (nAChRs) and both bind with high affinity...

  2. Hippocampal α7 nicotinic acetylcholine receptor levels in patients with schizophrenia, bipolar disorder, or major depressive disorder

    DEFF Research Database (Denmark)

    Thomsen, Morten Skøtt; Weyn, Annelies; Mikkelsen, Jens D

    2011-01-01

    The α7 nicotinic acetylcholine receptor (nAChR) is involved in cognitive function and synaptic plasticity. Consequently, changes in α7 nAChR function have been implicated in a variety of mental disorders, especially schizophrenia. However, there is little knowledge regarding the levels of the α7 n...

  3. Carbamoylcholine analogs as nicotinic acetylcholine receptor agonists--structural modifications of 3-(dimethylamino)butyl dimethylcarbamate (DMABC)

    DEFF Research Database (Denmark)

    Hansen, Camilla Petrycer; Jensen, Anders Asbjørn; Balle, Thomas

    2009-01-01

    Compounds based on the 3-(dimethylamino)butyl dimethylcarbamate (DMABC) scaffold were synthesized and pharmacologically characterized at the alpha(4)beta(2), alpha(3)beta(4,) alpha(4)beta(4) and alpha(7) neuronal nicotinic acetylcholine receptors (nAChRs). The carbamate functionality and a small...

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

    Directory of Open Access Journals (Sweden)

    Monica S Guzman

    2011-11-01

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

  5. The Actions of Piperidine Alkaloids at Fetal Muscle-Type and Autonomic-Type Nicotinic Acetylcholine Receptors

    Science.gov (United States)

    Piperidine alkaloids are found in many species of plants including Conium maculatum, Nicotiana spp., and Lupinus spp. A pharmacodynamic comparison was made of the alkaloids ammodendrine, anabasine, anabaseine, and coniine in; SH-SY5Y cells which express autonomic-type nicotinic acetylcholine recept...

  6. NMR Structure and Action on Nicotinic Acetylcholine Receptors of Water-soluble Domain of Human LYNX1

    Czech Academy of Sciences Publication Activity Database

    Lyukmanova, E. N.; Shenkarev, Z. O.; Shulepko, M. A.; Mineev, K. S.; D´Hoedt, D.; Kasheverov, I. E.; Filkin, S. Yu.; Krivolapova, A. P.; Janíčková, Helena; Doležal, Vladimír; Dolgikh, D. A.; Arseniev, A. S.; Bertrand, D.; Tsetlin, V.I.; Kirpichnikov, M. P.

    2011-01-01

    Roč. 286, č. 12 (2011), s. 10618-10627 ISSN 0021-9258 R&D Projects: GA ČR(CZ) GA305/09/0681 Institutional research plan: CEZ:AV0Z50110509 Keywords : NMR structure * nicotinic acetylcholine receptor * water-soluble domain Subject RIV: FH - Neurology Impact factor: 4.773, year: 2011

  7. Utrophin abundance is reduced at neuromuscular junctions of patients with both inherited and acquired acetylcholine receptor deficiencies

    NARCIS (Netherlands)

    Slater, CR; Young, C; Wood, SJ; Bewick, GS; Anderson, LVB; Baxter, P; Fawcett, PRW; Roberts, M; Jacobson, L; Kuks, J; Vincent, A; NewsomDavis, J

    Congenital myasthenic syndromes are a heterogenous group of conditions in which muscle weakness resulting from impaired neuromuscular transmission is often present from infancy. One form of congenital myasthenic syndrome is due to a reduction of the number of acetylcholine receptors (AChRs) at the

  8. An allosteric enhancer of M4muscarinic acetylcholine receptor function inhibits behavioral and neurochemical effects of cocaine

    DEFF Research Database (Denmark)

    Dencker, Ditte; Weikop, Pia; Sørensen, Gunnar

    2012-01-01

    The mesostriatal dopamine system plays a key role in mediating the reinforcing effects of psychostimulant drugs like cocaine. The muscarinic M4 acetylcholine receptor subtype is centrally involved in the regulation of dopamine release in striatal areas. Consequently, striatal M4 receptors could...

  9. Determination of anti-acetylcholine receptor antibodies in myasthenic patients by use of time-resolved fluorescence

    Czech Academy of Sciences Publication Activity Database

    Říčný, Jan; Šimková, L.; Vincent, A.

    2002-01-01

    Roč. 48, č. 3 (2002), s. 549-554 ISSN 0009-9147 R&D Projects: GA MZd NF4646 Institutional research plan: CEZ:AV0Z5011922 Keywords : nicotinic acetylcholine receptor * time-resolved fluorescence method * myasthenia gravis Subject RIV: FR - Pharmacology ; Medidal Chemistry Impact factor: 4.788, year: 2002

  10. Intravenously administered oxotremorine and atropine, in doses known to affect pain threshold, affect the intraspinal release of acetylcholine in rats

    DEFF Research Database (Denmark)

    Abelson, Klas S P; Höglund, A Urban

    2002-01-01

    muscarinic agonists and antagonists modify nociceptive threshold by affecting intraspinal release of acetylcholine (ACh). Catheters were inserted into the femoral vein in rats maintained on isoflurane anaesthesia for administration of oxotremorine (10-300 microg/kg) and atropine (0.1, 10, 5000 microg...

  11. Activation and desensitization of peripheral muscle and neuronal nicotinic acetylcholine receptors by selected, naturally-occurring pyridine alkaloids

    Science.gov (United States)

    Teratogenic alkaloids can cause developmental defects due to inhibition of fetal movement that results from desensitization of fetal muscletype nicotinic acetylcholine receptors (nAChRs). We investigated the ability of two known teratogens, the piperidinyl-pyridine anabasine and its 1,2-dehydropiper...

  12. Allosteric modulation by persistent binding of xanomeline of the interaction of competitive ligands with the M1 muscarinic acetylcholine receptor

    Czech Academy of Sciences Publication Activity Database

    Jakubík, Jan; Tuček, Stanislav; El-Fakahany, E. E.

    2002-01-01

    Roč. 301, č. 3 (2002), s. 1033-1041 ISSN 0022-3565 R&D Projects: GA ČR GP305/01/D119 Grant - others:NIH(US) NS25743 Institutional research plan: CEZ:AV0Z5011922 Keywords : xanomeline * M(1) muscarinic acetylcholine receptor Subject RIV: FH - Neurology Impact factor: 3.991, year: 2002

  13. Cholinesterase inhibition and acetylcholine accumulation following intracerebral administration of paraoxon in rats

    International Nuclear Information System (INIS)

    Ray, A.; Liu, J.; Karanth, S.; Gao, Y.; Brimijoin, S.; Pope, C.

    2009-01-01

    We evaluated the inhibition of striatal cholinesterase activity following intracerebral administration of paraoxon assaying activity either in tissue homogenates ex vivo or by substrate hydrolysis in situ. Artificial cerebrospinal fluid (aCSF) or paraoxon in aCSF was infused unilaterally (0.5 μl/min for 2 h) and ipsilateral and contralateral striata were harvested for ChE assay ex vivo. High paraoxon concentrations were needed to inhibit ipsilateral striatal cholinesterase activity (no inhibition at < 0.1 mM; 27% at 0.1 mM; 79% at 1 mM paraoxon). With 3 mM paraoxon infusion, substantial ChE inhibition was also noted in contralateral striatum. ChE histochemistry generally confirmed these concentration- and side-dependent effects. Microdialysates collected for up to 4 h after paraoxon infusion inhibited ChE activity when added to striatal homogenate, suggesting prolonged efflux of paraoxon. Since paraoxon efflux could complicate acetylcholine analysis, we evaluated the effects of paraoxon (0, 0.03, 0.1, 1, 10 or 100 μM, 1.5 μl/min for 45 min) administered by reverse dialysis through a microdialysis probe. ChE activity was then monitored in situ by perfusing the colorimetric substrate acetylthiocholine through the same probe and measuring product (thiocholine) in dialysates. Concentration-dependent inhibition was noted but reached a plateau of about 70% at 1 μM and higher concentrations. Striatal acetylcholine was below the detection limit at all times with 0.1 μM paraoxon but was transiently elevated (0.5-1.5 h) with 10 μM paraoxon. In vivo paraoxon (0.4 mg/kg, sc) in adult rats elicited about 90% striatal ChE inhibition measured ex vivo, but only about 10% inhibition measured in situ. Histochemical analyses revealed intense AChE and glial fibrillary acidic protein staining near the cannula track, suggesting proliferation of inflammatory cells/glia. The findings suggest that ex vivo and in situ cholinesterase assays can provide very different views into enzyme

  14. Acetylcholine-induced vasodilation in the uterine vascular bed of pregnant rats with adriamycin-induced nephrosis.

    Science.gov (United States)

    Yousif, Mariam H; Adeagbo, Ayotunde S; Kadavil, Elizabeth A; Chandrasekhar, Bindu; Oriowo, Mabayoje A

    2002-01-01

    This project was designed to study endothelium-dependent vasodilation in the uterine vascular bed during experimentally induced preeclampsia in rats. Uterine vascular beds were isolated from non-pregnant and pregnant rats with or without treatment with adriamycin (ADR) and perfused with physiological solution. Thereafter, vasodilator responses to acetylcholine were recorded. RECORDS: Pregnant ADR-treated rats displayed symptoms of preeclampsia including hypertension and proteinuria. Blood pressure was 110.0 +/- 4.7 mm Hg (n = 5) in control pregnant rats and 136.0 +/- 5.3 mm Hg (n = 5) in ADR-treated pregnant rats, and urinary protein concentrations were 0.35 mg/ml (n = 5) and 13.2 +/- 3.6 mg/ml (n = 9), respectively. Both blood pressure and proteinuria values were significantly (p acetylcholine-induced dose-dependent vasodilator responses in the vascular beds were not significantly different between the pregnant and nonpregnant rats. Although acetylcholine-induced vasodilation was significantly reduced by N omega-nitro-L-arginine methyl ester hydrochloride (L-NAME) in both groups, the residual response to acetylcholine was not affected by indomethacin, suggesting that prostanoids were not involved in this response. The L-NAME-resistant component, endothelium-derived hyperpolarizing factor (EDHF), was greater in ADR-treated uterine beds than in those of the controls, indicating a significant contribution from EDHF in these vessels. In the presence of an elevated external potassium ion concentration, acetylcholine produced similar vasodilator responses, indicating that the release of nitric oxide was not impaired. These results indicate that endothelium-dependent vasodilation was not impaired in this model of preeclampsia.

  15. Expression of nicotinic acetylcholine receptors on human B-lymphoma cells

    Directory of Open Access Journals (Sweden)

    Skok M. V.

    2009-12-01

    Full Text Available Aim. To find a correlation between the level of nicotinic acetylcholine receptor (nAChR expression and B lymphocyte differentiation or activation state. Methods. Expression of nAChRs in the REH, Ramos and Daudi cell lines was studied by flow cytometry using nAChR subunit-specific antibodies; cell proliferation was studied by MTT test. Results. It is shown that the level of 42/4 and 7 nAChRs expression increased along with B lymphocyte differentiation (Ramos > REH and activation (Daudi > > Ramos and depended on the antigen-specific receptor expression. The nAChR stimulation/blockade did not influence the intensity of cell proliferation.

  16. Steroids induce acetylcholine receptors on cultured human muscle: Implications for myasthenia gravis

    International Nuclear Information System (INIS)

    Kaplan, I.; Blakely, B.T.; Pavlath, G.K.; Travis, M.; Blau, H.M.

    1990-01-01

    Antibodies to the acetylcholine receptor (AChR), which are diagnostic of the human autoimmune disease myasthenia gravis, block AChR function and increase the rate of AChR degradation leading to impaired neuromuscular transmission. Steroids are frequently used to alleviate symptoms of muscle fatigue and weakness in patients with myasthenia gravis because of their well-documented immunosuppressive effects. The authors show here that the steroid dexamethasone significantly increases total surface AChRs on cultured human muscle exposed to myasthenia gravis sera. The results suggest that the clinical improvement observed in myasthenic patients treated with steroids is due not only to an effect on the immune system but also a direct effect on muscle. They propose that the identification and development of pharmacologic agents that augment receptors and other proteins that are reduced by human genetic or autoimmune disease will have broad therapeutic applications

  17. Differential expression of muscarinic acetylcholine receptor subtypes in Jurkat cells and their signaling.

    Science.gov (United States)

    Alea, Mileidys Perez; Borroto-Escuela, Dasiel O; Romero-Fernandez, Wilber; Fuxe, Kjell; Garriga, Pere

    2011-08-15

    Muscarinic acetylcholine receptors expression and signaling in the human Jurkat T cell line were investigated. Semiquantitative real-time PCR and radioligand binding studies, using a wide set of antagonist compounds, showed the co-existence of M(3), M(4), and M(5) subtypes. Stimulation of these subpopulations caused a concentration and time- dependent activation of second messengers and ERK signaling pathways, with a major contribution of the M(3) subtype in a G(q/11)-mediated response. In addition, we found that T-cell stimulation leads to increased expression of M(3) and M(5) both at transcriptional and protein levels in a PLC/PKCθ dependent manner. Our data clarifies the functional role of AChR subtypes in Jurkat cells and pave the way to future studies on the potential cross-talk among these subpopulations and their regulation of T lymphocytes immune function. Copyright © 2011 Elsevier B.V. All rights reserved.

  18. The Nicotinic Acetylcholine Receptor as a Target for Antidepressant Drug Development

    Directory of Open Access Journals (Sweden)

    Noah S. Philip

    2012-01-01

    Full Text Available An important new area of antidepressant drug development involves targeting the nicotinic acetylcholine receptor (nAChR. This receptor, which is distributed widely in regions of the brain associated with depression, is also implicated in other important processes that are relevant to depression, such as stress and inflammation. The two classes of drugs that target nAChRs can be broadly divided into mecamylamine- and cytisine-based compounds. These drugs probably exert their effects via antagonism at α4β2 nAChRs, and strong preclinical data support the antidepressant efficacy of both classes when used in conjunction with other primary antidepressants (e.g., monoamine reuptake inhibitors. Although clinical data remain limited, preliminary results in this area constitute a compelling argument for further evaluation of the nAChR as a target for future antidepressant drug development.

  19. Heteromeric α7β2 Nicotinic Acetylcholine Receptors in the Brain

    DEFF Research Database (Denmark)

    Wu, Jie; Liu, Qiang; Tang, Pei

    2016-01-01

    The α7 nicotinic acetylcholine receptor (α7 nAChR) is highly expressed in the brain, where it maintains various neuronal functions including (but not limited to) learning and memory. In addition, the protein expression levels of α7 nAChRs are altered in various brain disorders. The classic rule...... governing α7 nAChR assembly in the mammalian brain was that it was assembled from five α7 subunits to form a homomeric receptor pentamer. However, emerging evidence demonstrates the presence of heteromeric α7 nAChRs in heterologously expressed systems and naturally in brain neurons, where α7 subunits are co...... nAChR, which have provided new insights into the understanding of a novel target of cholinergic signaling....

  20. Molecular basis of the functional heterogeneity of the muscarinic acetylcholine receptor

    International Nuclear Information System (INIS)

    Numa, S.; Fukuda, K.; Kubo, T.; Maeda, A.; Akiba, I.; Bujo, H.; Nakai, J.; Mishina, M.; Higashida, H.

    1988-01-01

    The muscarinic acetylcholine receptor (mAChR) mediates a variety of cellular responses, including inhibition of adenylate cyclase, breakdown of phosphoinositides, and modulation of potassium channels, through the action of guanine-nucleotide-binding regulatory proteins (G proteins). The question then arises as to whether multiple mAChR species exist that are responsible for the various biochemical and physiological effects. In fact, pharmacologically distinguishable forms of the mAChR occur in different tissues and have been provisionally classified into M 1 (I), M 2 cardiac (II), and M 2 glandular (III) subtypes on the basis of their difference in apparent affinity for antagonists. Here, the authors have made attempts to understand the molecular basis of the functional heterogeneity of the mAChR, using recombinant DNA technology

  1. [3H]imidacloprid: synthesis of a candidate radioligand for the nicotinic acetylcholine receptor

    International Nuclear Information System (INIS)

    Latli, B.; Casida, J.E.

    1992-01-01

    Imidacloprid is an exceptionally potent insecticide known from physiological studies to act at the nicotinic acetylcholine receptor. To prepare [ 3 H]imidacloprid as a candidate radioligand, 6-chloronicotinoyl chloride was reduced with NaB 2 H 4 (in model studies) or NaB 3 H 4 in absolute ethanol to 2-chloro-5-pyridinylmethanol which was transformed to 2-chloro-5-chloromethylpyridine on refluxing with thionyl chloride. Coupling with 4,5-dihydro-N-nitro-1H-imidazol-2-amine then gave [ 2 H 2 ]imidacloprid incorporating about 95% of the deuterium or [ 3 H 2 ]imidacloprid (25 Ci/mmol) in 80% radiochemical yield. In studies not detailed here [ 3 H] imidacloprid was found to undergo high affinity, specific and saturable binding to a site in insect brain. (author)

  2. Design of ligands for the nicotinic acetylcholine receptors: the quest for selectivity.

    Science.gov (United States)

    Bunnelle, William H; Dart, Michael J; Schrimpf, Michael R

    2004-01-01

    In the last decade, nicotinic acetylcholine receptors (nAChRs) have emerged as important targets for drug discovery. The therapeutic potential of nicotinic agonists depends substantially on the ability to selectively activate certain receptor subtypes that mediate beneficial effects. The design of such compounds has proceeded in spite of a general shortage of data pertaining to subtype selectivity. Medicinal chemistry efforts have been guided principally by binding affinities to the alpha4beta2 and/or alpha7 subtypes, even though these are not predictive of agonist activity at either subtype. Nevertheless, a diverse family of nAChR ligands has been developed, and several analogs with promising therapeutic potential have now advanced to human clinical trials. This paper provides an overview of the structure-affinity relationships that continue to drive development of new nAChR ligands.

  3. Reactivity of Acetylcholine Esterase in inner Ear Maculae of Fish after Development at Hypergravity

    Science.gov (United States)

    Feucht, I.; Hilbig, R.; Anken, R.

    It has been shown earlier that the growth of inner ear otoliths of larval fish is (among other environmental factors) guided by the gravity vector. This guidance most probably is effected by the efferent vestibular system in the brainstem, because a transection of the nervus vestibularis has been shown to effect a cessation of the supply of calcium to the otoliths. The efferent innervation of fish inner ear maculae uses the synaptic transmitter acetylcholine (ACh). Therefore, we were - in order to further assess the role of the efferent system for otolith growth - prompted to determine ACh esterase-reactivity in the sensory epithelium of the utricle and the saccule (as well as in a non-gravity relevant brain region for control) in larval cichlid fish (Oreochromis mossambicus), which had been maintained at hypergravity during their development. The respective data will be communicated at the meeting. Acknowledgement: This work was financially supported by the German Aerospace Center (DLR) (FKZ: 50 WB 9997).

  4. Steroids induce acetylcholine receptors on cultured human muscle: Implications for myasthenia gravis

    Energy Technology Data Exchange (ETDEWEB)

    Kaplan, I.; Blakely, B.T.; Pavlath, G.K.; Travis, M.; Blau, H.M. (Stanford Univ. School of Medicine, CA (USA))

    1990-10-01

    Antibodies to the acetylcholine receptor (AChR), which are diagnostic of the human autoimmune disease myasthenia gravis, block AChR function and increase the rate of AChR degradation leading to impaired neuromuscular transmission. Steroids are frequently used to alleviate symptoms of muscle fatigue and weakness in patients with myasthenia gravis because of their well-documented immunosuppressive effects. The authors show here that the steroid dexamethasone significantly increases total surface AChRs on cultured human muscle exposed to myasthenia gravis sera. The results suggest that the clinical improvement observed in myasthenic patients treated with steroids is due not only to an effect on the immune system but also a direct effect on muscle. They propose that the identification and development of pharmacologic agents that augment receptors and other proteins that are reduced by human genetic or autoimmune disease will have broad therapeutic applications.

  5. Acetylcholine causes rooting in leaf explants of in vitro raised tomato (Lycopersicon esculentum Miller) seedlings.

    Science.gov (United States)

    Bamel, Kiran; Gupta, Shrish Chandra; Gupta, Rajendra

    2007-05-30

    The animal neurotransmitter acetylcholine (ACh) induces rooting and promotes secondary root formation in leaf explants of tomato (Lycopersicon esculentum Miller var. Pusa Ruby), cultured in vitro on Murashige and Skoog's medium. The roots originate from the midrib of leaf explants and resemble taproot. ACh at 10(-5) M was found to be the optimum over a wide range of effective concentrations between 10(-7) and 10(-3) M. The breakdown products, choline and acetate were ineffective even at 10(-3) M concentration. ACh appears to have a natural role in tomato rhizogenesis because exogenous application of neostigmine, an inhibitor of ACh hydrolysis, could mimic the effect of ACh. Neostigmine, if applied in combination with ACh, potentiated the ACh effect.

  6. Escherichia coli Protein Expression System for Acetylcholine Binding Proteins (AChBPs.

    Directory of Open Access Journals (Sweden)

    Nikita Abraham

    Full Text Available Nicotinic acetylcholine receptors (nAChR are ligand gated ion channels, identified as therapeutic targets for a range of human diseases. Drug design for nAChR related disorders is increasingly using structure-based approaches. Many of these structural insights for therapeutic lead development have been obtained from co-crystal structures of nAChR agonists and antagonists with the acetylcholine binding protein (AChBP. AChBP is a water soluble, structural and functional homolog of the extracellular, ligand-binding domain of nAChRs. Currently, AChBPs are recombinantly expressed in eukaryotic expression systems for structural and biophysical studies. Here, we report the establishment of an Escherichia coli (E. coli expression system that significantly reduces the cost and time of production compared to the existing expression systems. E. coli can efficiently express unglycosylated AChBP for crystallography and makes the expression of isotopically labelled forms feasible for NMR. We used a pHUE vector containing an N-terminal His-tagged ubiquitin fusion protein to facilitate AChBP expression in the soluble fractions, and thus avoid the need to recover protein from inclusion bodies. The purified protein yield obtained from the E. coli expression system is comparable to that obtained from existing AChBP expression systems. E. coli expressed AChBP bound nAChR agonists and antagonists with affinities matching those previously reported. Thus, the E. coli expression system significantly simplifies the expression and purification of functional AChBP for structural and biophysical studies.

  7. Characterization of prejunctional serotonin receptors modulating [3H]acetylcholine release in the human detrusor.

    Science.gov (United States)

    D'Agostino, Gianluigi; Condino, Anna M; Gallinari, Paola; Franceschetti, Gian P; Tonini, Marcello

    2006-01-01

    Bladder overactivity (OAB) is a chronic and debilitating lower urinary tract (LUT) disorder that affects millions of individuals worldwide. LUT symptoms associated with OAB, such as urgency and urinary incontinence, cause a hygienic and social concern to patients, but their current pharmacological treatment is largely inadequate due to the lack of uroselectivity. Although OAB etiology remains multifactorial and poorly understood, increasing evidence indicates that serotonin [5-hydroxytryptamine (5-HT)] is an endogenous substance involved in the control of micturition at central and peripheral sites. In this study, we demonstrated the presence of three distinct 5-HT receptors localized at parasympathetic nerve terminals of the human bladder by measuring electrically evoked tritiated acetylcholine release in isolated detrusor strips. These prejunctional receptors, involved in both positive and negative feedback mechanisms regulating cholinergic transmission, have been characterized by means of three highly selective 5-HT antagonists for 5-HT(4), 5-HT(7), and 5-HT(1A) receptors, namely GR113808A ([1-[2-[(-methylsulphonyl) amino] ethyl]4-piperinidyl]methyl1-methyl-1H-indole-3-carboxylate succinate), SB269970 [(R)-3-(2-(2-(4-methylpiperidin-1-yl)ethyl)pyrrolidine-1-sulfonyl)phenol hydrochloride], and WAY100635 [N-(2-(4-(2-methoxyphenyl)-1-piperazinyl)ethyl)-N-(2-pyridyl)-cyclohexane-carboxamide trichloride]. Under these conditions, we confirmed the facilitatory role of 5-HT(4) heteroreceptors on acetylcholine release and revealed for the first time the occurrence of 5-HT(7) and 5-HT(1A) heteroreceptors with a facilitatory and an inhibitory action, respectively. Our findings strengthen the novel concept for the use of recently patented selective 5-HT agonists and antagonists for the control of OAB dysfunctions associated with inflammatory conditions, although their therapeutic efficacy needs to be explored in the clinical setting.

  8. Acetylcholine muscarinic receptors and response to anti-cholinesterase therapy in patients with Alzheimer's disease

    International Nuclear Information System (INIS)

    Brown, Derek; Chisholm, Jennifer A.; Patterson, Jim; Wyper, David; Owens, Jonathan; Pimlott, Sally

    2003-01-01

    An acetylcholine deficit remains the most consistent neurotransmitter abnormality found in Alzheimer's disease and various therapeutic agents have been targeted at this. In this study we investigated the action of Donepezil, a cholinesterase inhibitor that has few side-effects. In particular we set out to investigate whether muscarinic acetylcholine receptor (mAChR) availability influences the response to this therapy. We used the novel single-photon emission tomography (SPET) tracer (R,R)[ 123 I]I-quinuclidinyl benzilate (R,R[ 123 I]I-QNB), which has high affinity for the M1 subtype of mAChR. Regional cerebral perfusion was also assessed using technetium-99m hexamethylpropylene amine oxime. We investigated 20 patients on Donepezil treatment and ten age-matched controls. The results showed a reduction in (R,R)[ 123 I]I-QNB binding in the caudal anterior cingulate in patients compared with controls and relatively high binding in the putamen and rostral anterior cingulate, suggesting a relative sparing of mAChR in these regions. The main finding of the study was that mAChR availability as assessed by (R,R)[ 123 I]I-QNB binding did not distinguish responders from non-responders. Interestingly, we found that the extent of cognitive improvement showed no positive correlation with (R,R)[ 123 I]I-QNB binding in any brain region but was inversely related to binding in the insular cortex. This suggests that, within the advised cognitive performance band for use of Donepezil, response is greater in those patients with evidence of a more marked cholinergic deficit. A larger study should investigate this. (orig.)

  9. Carbon monoxide-induced delayed amnesia, delayed neuronal death and change in acetylcholine concentration in mice

    International Nuclear Information System (INIS)

    Nabeshima, T.; Katoh, A.; Ishimaru, H.; Yoneda, Y.; Ogita, K.; Murase, K.; Ohtsuka, H.; Inari, K.; Fukuta, T.; Kameyama, T.

    1991-01-01

    We investigated the interrelationship of delayed amnesia, delayed neuronal death and changes in acetylcholine concentration induced by carbon monoxide (CO)-exposure in mice. In the test for retention of the passive avoidance task, amnesia was observed 5 and 7 days after CO-exposure when the mice were exposed to CO 1 day after training; in the case when the mice were exposed to CO 5 and 7 days before training, amnesia was also observed in a retention test given 1 day after training. The number of pyramidal cells in the hippocampal CA1 subfield was lower than that of the control 3, 5 and 7 days after CO-exposure. But the neurodegeneration in the parietal cortex, area 1, was not observed until 7 days after CO-exposure. The findings indicated that the amnesia and the neuronal death were produced after a delay when the mice were exposed to CO. In addition, the delayed amnesia was closely related to the delayed neuronal death in the hippocampal CA1 subfield. Moreover, [3H]glutamate and [3H]glycine binding sites did not change after CO-exposure but, 7 days after CO-exposure, the concentration of acetylcholine and the binding of [3H]quinuclidinyl benzilate in the frontal cortex and the striatum were found to have significantly changed, but those in the hippocampus did not show significant change. Therefore, we suggest that delayed amnesia induced by CO-exposure may result from delayed neuronal death in the hippocampal CA1 subfield and dysfunction in the acetylcholinergic neurons, in the frontal cortex, the striatum and/or the hippocampus

  10. Carbon monoxide-induced delayed amnesia, delayed neuronal death and change in acetylcholine concentration in mice

    Energy Technology Data Exchange (ETDEWEB)

    Nabeshima, T.; Katoh, A.; Ishimaru, H.; Yoneda, Y.; Ogita, K.; Murase, K.; Ohtsuka, H.; Inari, K.; Fukuta, T.; Kameyama, T. (Meijo Univ., Nagoya (Japan))

    1991-01-01

    We investigated the interrelationship of delayed amnesia, delayed neuronal death and changes in acetylcholine concentration induced by carbon monoxide (CO)-exposure in mice. In the test for retention of the passive avoidance task, amnesia was observed 5 and 7 days after CO-exposure when the mice were exposed to CO 1 day after training; in the case when the mice were exposed to CO 5 and 7 days before training, amnesia was also observed in a retention test given 1 day after training. The number of pyramidal cells in the hippocampal CA1 subfield was lower than that of the control 3, 5 and 7 days after CO-exposure. But the neurodegeneration in the parietal cortex, area 1, was not observed until 7 days after CO-exposure. The findings indicated that the amnesia and the neuronal death were produced after a delay when the mice were exposed to CO. In addition, the delayed amnesia was closely related to the delayed neuronal death in the hippocampal CA1 subfield. Moreover, (3H)glutamate and (3H)glycine binding sites did not change after CO-exposure but, 7 days after CO-exposure, the concentration of acetylcholine and the binding of (3H)quinuclidinyl benzilate in the frontal cortex and the striatum were found to have significantly changed, but those in the hippocampus did not show significant change. Therefore, we suggest that delayed amnesia induced by CO-exposure may result from delayed neuronal death in the hippocampal CA1 subfield and dysfunction in the acetylcholinergic neurons, in the frontal cortex, the striatum and/or the hippocampus.

  11. Effect of ethanol at clinically relevant concentrations on atrial inward rectifier potassium current sensitive to acetylcholine.

    Science.gov (United States)

    Bébarová, Markéta; Matejovič, Peter; Pásek, Michal; Hořáková, Zuzana; Hošek, Jan; Šimurdová, Milena; Šimurda, Jiří

    2016-10-01

    Alcohol intoxication tends to induce arrhythmias, most often the atrial fibrillation. To elucidate arrhythmogenic mechanisms related to alcohol consumption, the effect of ethanol on main components of the ionic membrane current is investigated step by step. Considering limited knowledge, we aimed to examine the effect of clinically relevant concentrations of ethanol (0.8-80 mM) on acetylcholine-sensitive inward rectifier potassium current I K(Ach). Experiments were performed by the whole-cell patch clamp technique at 23 ± 1 °C on isolated rat and guinea-pig atrial myocytes, and on expressed human Kir3.1/3.4 channels. Ethanol induced changes of I K(Ach) in the whole range of concentrations applied; the effect was not voltage dependent. The constitutively active component of I K(Ach) was significantly increased by ethanol with the maximum effect (an increase by ∼100 %) between 8 and 20 mM. The changes were comparable in rat and guinea-pig atrial myocytes and also in expressed human Kir3.1/3.4 channels (i.e., structural correlate of I K(Ach)). In the case of the acetylcholine-induced component of I K(Ach), a dual ethanol effect was apparent with a striking heterogeneity of changes in individual cells. The effect correlated with the current magnitude in control: the current was increased by eth-anol in the cells showing small current in control and vice versa. The average effect peaked at 20 mM ethanol (an increase of the current by ∼20 %). Observed changes of action potential duration agreed well with the voltage clamp data. Ethanol significantly affected both components of I K(Ach) even in concentrations corresponding to light alcohol consumption.

  12. Adenosine receptors and muscarinic receptors cooperate in acetylcholine release modulation in the neuromuscular synapse.

    Science.gov (United States)

    Santafe, M M; Priego, M; Obis, T; Garcia, N; Tomàs, M; Lanuza, M A; Tomàs, J

    2015-07-01

    Adenosine receptors (ARs) are present in the motor terminals at the mouse neuromuscular junction. ARs and the presynaptic muscarinic acetylcholine receptors (mAChRs) share the functional control of the neuromuscular junction. We analysed their mutual interaction in transmitter release modulation. In electrophysiological experiments with unaltered synaptic transmission (muscles paralysed by blocking the voltage-dependent sodium channel of the muscle cells with μ-conotoxin GIIIB), we found that: (i) a collaborative action between different AR subtypes reduced synaptic depression at a moderate activity level (40 Hz); (ii) at high activity levels (100 Hz), endogenous adenosine production in the synaptic cleft was sufficient to reduce depression through A1 -type receptors (A1 Rs) and A2 A-type receptors (A2 A Rs); (iii) when the non-metabolizable 2-chloroadenosine (CADO) agonist was used, both the quantal content and depression were reduced; (iv) the protective effect of CADO on depression was mediated by A1 Rs, whereas A2 A Rs seemed to modulate A1 Rs; (v) ARs and mAChRs absolutely depended upon each other for the modulation of evoked and spontaneous acetylcholine release in basal conditions and in experimental conditions with CADO stimulation; (vi) the purinergic and muscarinic mechanisms cooperated in the control of depression by sharing a common pathway although the purinergic control was more powerful than the muscarinic control; and (vii) the imbalance of the ARs created by using subtype-selective and non-selective inhibitory and stimulatory agents uncoupled protein kinase C from evoked transmitter release. In summary, ARs (A1 Rs, A2 A Rs) and mAChRs (M1 , M2 ) cooperated in the control of activity-dependent synaptic depression and may share a common protein kinase C pathway. © 2015 Federation of European Neuroscience Societies and John Wiley & Sons Ltd.

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

    Science.gov (United States)

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

    2004-07-13

    In fish, melanin pigment granules in the retinal pigment epithelium disperse into apical projections as part of the suite of responses the eye makes to bright light conditions. This pigment granule dispersion serves to reduce photobleaching and occurs in response to neurochemicals secreted by the retina. Previous work has shown that acetylcholine may be involved in inducing light-adaptive pigment dispersion. Acetylcholine receptors are of two main types, nicotinic and muscarinic. Muscarinic receptors are in the G-protein coupled receptor superfamily, and five different muscarinic receptors have been molecularly cloned in human. These receptors are coupled to adenylyl cyclase, calcium mobilization and ion channel activation. To determine the receptor pathway involved in eliciting pigment granule migration, we isolated retinal pigment epithelium from bluegill and subjected it to a battery of cholinergic agents. The general cholinergic agonist carbachol induces pigment granule dispersion in isolated retinal pigment epithelium. Carbachol-induced pigment granule dispersion is blocked by the muscarinic antagonist atropine, by the M1 antagonist pirenzepine, and by the M3 antagonist 4-DAMP. Pigment granule dispersion was also induced by the M1 agonist 4-[N-(4-chlorophenyl) carbamoyloxy]-4-pent-2-ammonium iodide. In contrast the M2 antagonist AF-DX 116 and the M4 antagonist tropicamide failed to block carbachol-induced dispersion, and the M2 agonist arecaidine but-2-ynyl ester tosylate failed to elicit dispersion. Our results suggest that carbachol-mediated pigment granule dispersion occurs through the activation of Modd muscarinic receptors, which in other systems couple to phosphoinositide hydrolysis and elevation of intracellular calcium. This conclusion must be corroborated by molecular studies, but suggests Ca2+-dependent pathways may be involved in light-adaptive pigment dispersion.

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

    Directory of Open Access Journals (Sweden)

    Crittenden Elizabeth L

    2004-07-01

    Full Text Available Abstract Background In fish, melanin pigment granules in the retinal pigment epithelium disperse into apical projections as part of the suite of responses the eye makes to bright light conditions. This pigment granule dispersion serves to reduce photobleaching and occurs in response to neurochemicals secreted by the retina. Previous work has shown that acetylcholine may be involved in inducing light-adaptive pigment dispersion. Acetylcholine receptors are of two main types, nicotinic and muscarinic. Muscarinic receptors are in the G-protein coupled receptor superfamily, and five different muscarinic receptors have been molecularly cloned in human. These receptors are coupled to adenylyl cyclase, calcium mobilization and ion channel activation. To determine the receptor pathway involved in eliciting pigment granule migration, we isolated retinal pigment epithelium from bluegill and subjected it to a battery of cholinergic agents. Results The general cholinergic agonist carbachol induces pigment granule dispersion in isolated retinal pigment epithelium. Carbachol-induced pigment granule dispersion is blocked by the muscarinic antagonist atropine, by the M1 antagonist pirenzepine, and by the M3 antagonist 4-DAMP. Pigment granule dispersion was also induced by the M1 agonist 4-[N-(4-chlorophenyl carbamoyloxy]-4-pent-2-ammonium iodide. In contrast the M2 antagonist AF-DX 116 and the M4 antagonist tropicamide failed to block carbachol-induced dispersion, and the M2 agonist arecaidine but-2-ynyl ester tosylate failed to elicit dispersion. Conclusions Our results suggest that carbachol-mediated pigment granule dispersion occurs through the activation of Modd muscarinic receptors, which in other systems couple to phosphoinositide hydrolysis and elevation of intracellular calcium. This conclusion must be corroborated by molecular studies, but suggests Ca2+-dependent pathways may be involved in light-adaptive pigment dispersion.

  15. Neuronal nicotinic acetylcholine receptors: Common molecular substrates of nicotine and alcohol dependence

    Directory of Open Access Journals (Sweden)

    Linzy M. Hendrickson

    2013-04-01

    Full Text Available Alcohol and nicotine are often co-abused. As many as 80-95% of alcoholics are also smokers, suggesting that ethanol and nicotine, the primary addictive component of tobacco smoke, may functionally interact in the central nervous system and/or share a common mechanism of action. While nicotine initiates dependence by binding to and activating neuronal nicotinic acetylcholine receptors (nAChRs, ligand-gated cation channels normally activated by endogenous acetylcholine (ACh, ethanol is much less specific with the ability to modulate multiple gene products including those encoding voltage-gated ion channels, and excitatory/inhibitory neurotransmitter receptors. However, emerging data indicate that ethanol interacts with nAChRs, both directly and indirectly, in the mesocorticolimbic dopaminergic (DAergic reward circuitry to affect brain reward systems. Like nicotine, ethanol activates DAergic neurons of the ventral tegmental area (VTA which project to the nucleus accumbens (NAc. Blockade of VTA nAChRs reduces ethanol-mediated activation of DAergic neurons, NAc DA release, consumption, and operant responding for ethanol in rodents. Thus, ethanol may increase ACh release into the VTA driving activation of DAergic neurons through nAChRs. In addition, ethanol potentiates distinct nAChR subtype responses to ACh and nicotine in vitro and in DAergic neurons. The smoking cessation therapeutic and nAChR partial agonist, varenicline, reduces alcohol consumption in heavy drinking smokers and rodent models of alcohol consumption. Finally, single nucleotide polymorphisms in nAChR subunit genes are associated with alcohol dependence phenotypes and smoking behaviors in human populations. Together, results from preclinical, clinical, and genetic studies indicate that nAChRs may have an inherent role in the abusive properties of ethanol, as well as in nicotine and alcohol co-dependence.

  16. Modulation of release of [3H]acetylcholine in the major pelvic ganglion of the rat.

    Science.gov (United States)

    Somogyi, G T; de Groat, W C

    1993-06-01

    Cholinergic modulation of [3H]acetylcholine release evoked by electrical stimulation was studied in the rat major pelvic ganglion, which was prelabeled with [3H]choline. Acetylcholine (ACh) release was independent of the frequency of stimulation; 0.3 Hz produced the same volley output as 10 Hz. Tetrodotoxin (1 microM) or omission of Ca2+ from the medium abolished ACh release. The M1 receptor agonist (4-hydroxy-2-butynyl)-1-trimethylammonium m-chlorocarbanilate chloride (McN-A 343, 50 microM) increased release (by 136%), whereas the M2 muscarinic agonist oxotremorine (1 microM) decreased ACh release (by 22%). The muscarinic antagonists, atropine (1 microM) or pirenzepine (M1 selective, 1 microM), did not change ACh release. However, pirenzepine (1 microM) blocked the facilitatory effect of McN-A 343, and atropine (1 microM) blocked the inhibitory effect of oxotremorine. The cholinesterase inhibitor physostigmine (1-5 microM), the nicotinic agonist 1,1-dimethyl-4-phenylpiperazinium (DMPP, 10 microM), and the nicotinic antagonist D-tubocurarine (50 microM) did not change ACh release. 4-Aminopyridine, a K+ channel blocker, significantly increased the release (by 146%). Seven days after decentralization of the major pelvic ganglion, the evoked release of ACh was abolished. It is concluded that release of ACh occurs from the preganglionic nerve terminals rather than from the cholinergic cell bodies and is not modulated by actions of endogenous ACh on either muscarinic or nicotinic autoreceptors. These data confirm and extend previous electrophysiological findings indicating that synapses in the major pelvic ganglion have primarily a relay function.

  17. Modes of Action, Resistance and Toxicity of Insecticides Targeting Nicotinic Acetylcholine Receptors.

    Science.gov (United States)

    Ihara, Makoto; Buckingham, Steven D; Matsuda, Kazuhiko; Sattelle, David B

    2017-01-01

    Nicotinic acetylcholine receptors (nAChRs) of insects play a key role in fast excitatory neurotransmission. Several classes of insecticides target insect nAChRs, which are composed of subunit members of a family of multiple subunit encoding genes. Alternative splicing and RNA A-to-I editing can add further to receptor diversity. Native and recombinant receptors have been explored as sites of insecticide action using radioligands, electrophysiology and site-directed mutagenesis. We have reviewed the properties of native and recombinant insect nAChRs, the challenges of functional recombinant insect nAChR expression, nAChR interactions with ligands acting at orthosteric and allosteric sites and in particular their interactions with insecticides. Actions on insect nAChRs of cartap, neonicotinoids, spinosyns, sulfoxamines, butenolides and mesoionic insecticides are reviewed and current knowledge of their modes of action are addressed. Mutations that add to our understanding of insecticide action and those leading to resistance are discussed. Co-crystallisation of neonicotinoids with the acetylcholine binding protein (AChBP), a surrogate for the nAChR ligand binding domain, has proved instructive. Toxicity issues relating to insecticides targeting nAChRs are also considered. An overview of insecticide classes targeting insect nAChRs has enhanced our understanding of these important receptors and their insecticide binding sites. However, the subunit composition of native nAChRs remains poorly understood and functional expression still presents difficulties. These topics together with improved understanding of the precise sites of insecticide actions on insect nAChRs will be the subject of future research. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  18. Super agonist actions of clothianidin and related compounds on the SAD beta 2 nicotinic acetylcholine receptor expressed in Xenopus laevis oocytes.

    Science.gov (United States)

    Ihara, Makoto; Matsuda, Kazuhiko; Shimomura, Masaru; Sattelle, David B; Komai, Koichiro

    2004-03-01

    To compare the actions of clothianidin, a neonicotinoid acting on insect nicotinic acetylcholine receptors, and related compounds with that of imidacloprid, the compounds were tested on the Drosophila SAD-chicken beta2 nicotinic acetylcholine receptor expressed in Xenopus laevis oocytes using two-electrode voltage-clamp electrophysiology. The maximum response of the SAD beta 2 nicotinic receptor to clothianidin was larger than that observed for acetylcholine. Ring breakage of the imidazolidine ring of imidacloprid resulting in the generation of a guanidine group was critical for this super agonist action.

  19. Neuroprotection of rat retinal ganglion cells mediated through alpha7 nicotinic acetylcholine receptors.

    Science.gov (United States)

    Iwamoto, K; Mata, D; Linn, D M; Linn, C L

    2013-05-01

    Glutamate-induced excitotoxicity is thought to play an important role in several neurodegenerative diseases in the central nervous system (CNS). In this study, neuroprotection against glutamate-induced excitotoxicity was analyzed using acetylcholine (ACh), nicotine and the α7 specific nicotinic acetylcholine receptor (α7 nAChR) agonist, N-[(3R)-1-azabicyclo[2.2.2]oct-3-yl]-4-chlorobenzamide hydrochloride (PNU-282987), in cultured adult rat retinal neurons. Adult Long Evans rat retinas were dissociated and retinal ganglion cells (RGCs) were isolated from all other retinal tissue using a two-step panning technique. Once isolated, RGCs were cultured under various pharmacological conditions to demonstrate excitotoxicity and neuroprotection against excitotoxicity. After 3 days, RGCs were immunostained with antibodies against the glycoprotein, Thy 1.1, counted and cell survival was assessed relative to control untreated conditions. 500 μM glutamate induced excitotoxicity in large and small RGCs in an adult rat dissociated culture. After 3 days in culture with glutamate, the cell survival of large RGCs decreased by an average of 48.16% while the cell survival of small RGCs decreased by an average of 42.03%. Using specific glutamate receptor agonists and antagonists, we provide evidence that the excitotoxic response was mediated through α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA)/kainic acid (KA) and N-methyl-d-aspartate (NMDA) glutamate receptors through an apoptotic mechanism. However, the excitotoxic effect of glutamate on all RGCs was eliminated if cells were cultured for an hour with 10 μM ACh, 100 μM nicotine or 100 nM of the α7 nAChR agonist, PNU-282987, before the glutamate insult. Inhibition studies using 10nM methyllycaconitine (MLA) or α-bungarotoxin (α-Bgt) supported the hypothesis that neuroprotection against glutamate-induced excitotoxicity on rat RGCs was mediated through α7 nAChRs. In immunocytochemical studies, double

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

    DEFF Research Database (Denmark)

    Wang, Jing; Novak, Ivana

    2013-01-01

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

  1. Simultaneous improvement in production of microalgal biodiesel and high-value alpha-linolenic acid by a single regulator acetylcholine

    OpenAIRE

    Parsaeimehr, Ali; Sun, Zhilan; Dou, Xiao; Chen, Yi-Feng

    2015-01-01

    Background Photoautotrophic microalgae are a promising avenue for sustained biodiesel production, but are compromised by low yields of biomass and lipids at present. We are developing a chemical approach to improve microalgal accumulation of feedstock lipids as well as high-value alpha-linolenic acid which in turn might provide a driving force for biodiesel production. Results We demonstrate the effectiveness of the small bioactive molecule ?acetylcholine? on accumulation of biomass, total li...

  2. Lipophilicity of acetylcholine and related ions examined by ion transfer voltammetry at a polarized room-temperature ionic liquid membrane

    Czech Academy of Sciences Publication Activity Database

    Langmaier, Jan; Záliš, Stanislav; Samec, Zdeněk

    2018-01-01

    Roč. 815, APR 2018 (2018), s. 183-188 ISSN 1572-6657 R&D Projects: GA ČR(CZ) GA15-03139S; GA ČR(CZ) GA17-05387S Institutional support: RVO:61388955 Keywords : acetylcholine * choline * ion transfer voltammetry Subject RIV: CG - Electrochemistry OBOR OECD: Electrochemistry (dry cells, batteries, fuel cells, corrosion metals, electrolysis) Impact factor: 3.012, year: 2016

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

    Directory of Open Access Journals (Sweden)

    Jake Bedore

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

  4. R+-methanandamide inhibits tracheal response to endogenously released acetylcholine via capsazepine-sensitive receptors.

    Science.gov (United States)

    Nieri, Paola; Martinotti, Enrica; Testai, Lara; Adinolfi, Barbara; Calderone, Vincenzo; Breschi, Maria Cristina

    2003-01-10

    The effects of cannabinoid drugs on the cholinergic response evoked by electrical field stimulation (0.2 ms pulse width, 20 V amplitude, 10 Hz, 7.5 s train duration) in guinea-pig tracheal preparations were investigated. The stable analogue of the endocannabinoid anandamide, R(+)-methanandamide (10(-7)-10(-4) M), produced a dose-dependent inhibition (up to 27+/-5% of control) of electrical field stimulation-mediated atropine-sensitive response. This effect was not blocked by the selective cannabinoid CB(1) receptor antagonist N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3 carboxamide hydrochloride (SR 141716A; 10(-6) M), and was not reproduced with the cannabinoid CB(1)/CB(2) receptor agonist R(+)-[2,3-dihydro-5-methyl-[(morpholinyl)methyl]pyrrolo [1,2,3-de]-1,4-benzoxazin-6-yl]-(1-naphthalenyl)methanone mesylate) (WIN 55,212-2; 10(-8)-10(-5) M) or the cannabinoid CB(2) receptor selective agonist 1-propyl-2-methyl-3-(1-naphthoyl)indole (JWH-015; 10(-8)-10(-5) M); it was, on the contrary, antagonized by the vanilloid antagonist 2-[2-(4-chlorophenyl)ethyl-amino-thiocarbonyl]-7,8-dihydroxy-2,3,4,5-tetrahydro-1H-2 benzazepine (capsazepine; 10(-6) M). At the postjunctional level, neither R(+)-methanandamide nor WIN 55,212-2 nor JWH-015 did affect tracheal contractions induced by exogenous acetylcholine (10(-6) M). An inhibitory vanilloid receptor-mediated effect on the cholinergic response evoked by electrical stimulation was confirmed with the vanilloid agonist capsaicin, at doses (3-6 x 10(-8) M) which poorly influenced the basal smooth muscle tone of trachea. In conclusion, our data indicate that in guinea-pig trachea (a) neither CB(1) nor CB(2) cannabinoid receptor-mediated modulation of acetylcholine release occurs; (b) vanilloid VR1-like receptors appear involved in R(+)-methanandamide inhibitory activity on the cholinergic response to electrical field stimulation.

  5. Distinctive Modulation of Dopamine Release in the Nucleus Accumbens Shell Mediated by Dopamine and Acetylcholine Receptors.

    Science.gov (United States)

    Shin, Jung Hoon; Adrover, Martin F; Alvarez, Veronica A

    2017-11-15

    Nucleus accumbens (NAc) shell shows unique dopamine (DA) signals in vivo and plays a unique role in DA-dependent behaviors such as reward-motivated learning and the response to drugs of abuse. A disynaptic mechanism for DA release was reported and shown to require synchronized firing of cholinergic interneurons (CINs) and activation of nicotinic acetylcholine (ACh) receptors (nAChRs) in DA neuron (DAN) axons. The properties of this disynaptic mechanism of DA transmission are not well understood in the NAc shell. In this study, in vitro fast-scan cyclic voltammetry was used to examine the modulation of DA transmission evoked by CINs firing in the shell of mice and compared with other striatal regions. We found that DA signals in the shell displayed significant degree of summation in response to train stimulation of CINs, contrary to core and dorsal striatum. The summation was amplified by a D2-like receptor antagonist and experiments with mice with targeted deletion of D2 receptors to DANs or CINs revealed that D2 receptors in CINs mediate a fast inhibition observed within 100 ms of the first pulse, whereas D2 autoreceptors in DAN terminals are engaged in a slower inhibition that peaks at ∼500 ms. ACh also contributes to the use-dependent inhibition of DA release through muscarinic receptors only in the shell, where higher activity of acetylcholinesterase minimizes nAChR desensitization and promotes summation. These findings show that DA signals are modulated differentially by endogenous DA and ACh in the shell, which may underlie the unique features of shell DA signals in vivo SIGNIFICANCE STATEMENT The present study reports that dopamine (DA) release evoked by activation of cholinergic interneurons displays a high degree of summation in the shell and shows unique modulation by endogenous DA and acetylcholine. Desensitization of nicotinic receptors, which is a prevailing mechanism for use-dependent inhibition in the nucleus accumbens core and dorsal striatum, is

  6. [Role of calcium ions in the mechanism of action of acetylcholine on energy metabolism in rat liver mitochondria].

    Science.gov (United States)

    Vatamaniuk, M Z; Artym, V V; Kuka, O B; Doliba, M M; Shostakovs'ka, I V

    1996-01-01

    It is shown that administration of acetylcholine to animals (50 micrograms per 100 g of body weight) leads to the activation of respiration and oxidative phosphorylation in the rat liver mitochondria under oxidation of alpha-ketoglutarate; this effect depends on the concentration of calcium ions in the incubation medium of mitochondria. The rate of ADP-stimulated respiration of mitochondria of experimental animals reaches its maximum level under lower concentrations of Ca2+ than in the control animals. The results of investigation of dependence of acetyl choline effect on respiration of mitochondria on the concentration of alpha-ketoglutarate in calcium and calcium-free incubation medium have shown that the half-maximum effect of acetylcholine is observed in calcium medium at lower concentration of the substrate than in calcium-free medium. The latter indicates to the increase of affinity of alpha-ketoglutarate dehydrogenase to alpha-ketoglutarate under these conditions. It is found out that acetylcholine (1.10(-8) M) increases the rate of ADP- and Ca(2+)-stimulated respiration of mitochondria of isolated perfused rat liver, while mutual effect of verapamyl and niphedipin removes this effect.

  7. Simultaneous improvement in production of microalgal biodiesel and high-value alpha-linolenic acid by a single regulator acetylcholine.

    Science.gov (United States)

    Parsaeimehr, Ali; Sun, Zhilan; Dou, Xiao; Chen, Yi-Feng

    2015-01-01

    Photoautotrophic microalgae are a promising avenue for sustained biodiesel production, but are compromised by low yields of biomass and lipids at present. We are developing a chemical approach to improve microalgal accumulation of feedstock lipids as well as high-value alpha-linolenic acid which in turn might provide a driving force for biodiesel production. We demonstrate the effectiveness of the small bioactive molecule "acetylcholine" on accumulation of biomass, total lipids, and alpha-linolenic acid in Chlorella sorokiniana. The effectiveness exists in different species of Chlorella. Moreover, the precursor and analogs of acetylcholine display increased effectiveness at higher applied doses, with maximal increases by 126, 80, and 60% over controls for biomass, total lipids, and alpha-linolenic acid, respectively. Production of calculated biodiesel was also improved by the precursor and analogs of acetylcholine. The biodiesel quality affected by changes in microalgal fatty acid composition was addressed. The chemical approach described here could improve the lipid yield and biodiesel production of photoautotrophic microalgae if combined with current genetic approaches.

  8. Quinuclidine compounds differently act as agonists of Kenyon cell nicotinic acetylcholine receptors and induced distinct effect on insect ganglionic depolarizations.

    Science.gov (United States)

    Mathé-Allainmat, Monique; Swale, Daniel; Leray, Xavier; Benzidane, Yassine; Lebreton, Jacques; Bloomquist, Jeffrey R; Thany, Steeve H

    2013-12-01

    We have recently demonstrated that a new quinuclidine benzamide compound named LMA10203 acted as an agonist of insect nicotinic acetylcholine receptors. Its specific pharmacological profile on cockroach dorsal unpaired median neurons (DUM) helped to identify alpha-bungarotoxin-insensitive nAChR2 receptors. In the present study, we tested its effect on cockroach Kenyon cells. We found that it induced an inward current demonstrating that it bounds to nicotinic acetylcholine receptors expressed on Kenyon cells. Interestingly, LMA10203-induced currents were completely blocked by the nicotinic antagonist α-bungarotoxin. We suggested that LMA10203 effect occurred through the activation of α-bungarotoxin-sensitive receptors and did not involve α-bungarotoxin-insensitive nAChR2, previously identified in DUM neurons. In addition, we have synthesized two new compounds, LMA10210 and LMA10211, and compared their effects on Kenyon cells. These compounds were members of the 3-quinuclidinyl benzamide or benzoate families. Interestingly, 1 mM LMA10210 was not able to induce an inward current on Kenyon cells compared to LMA10211. Similarly, we did not find any significant effect of LMA10210 on cockroach ganglionic depolarization, whereas these three compounds were able to induce an effect on the central nervous system of the third instar M. domestica larvae. Our data suggested that these three compounds could bind to distinct cockroach nicotinic acetylcholine receptors.

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

    International Nuclear Information System (INIS)

    Koenig, M.L.

    1985-01-01

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

  10. α7-Nicotinic acetylcholine receptor: role in early odor learning preference in mice.

    Directory of Open Access Journals (Sweden)

    Jennifer L Hellier

    Full Text Available Recently, we have shown that mice with decreased expression of α7-nicotinic acetylcholine receptors (α7 in the olfactory bulb were associated with a deficit in odor discrimination compared to wild-type mice. However, it is unknown if mice with decreased α7-receptor expression also show a deficit in early odor learning preference (ELP, an enhanced behavioral response to odors with attractive value observed in rats. In this study, we modified ELP methods performed in rats and implemented similar conditions in mice. From post-natal days 5-18, wild-type mice were stroked simultaneously with an odor presentation (conditioned odor for 90 s daily. Control mice were only stroked, exposed to odor, or neither. On the day of testing (P21, mice that were stroked in concert with a conditioned odor significantly investigated the conditioned odor compared to a novel odor, as observed similarly in rats. However, mice with a decrease in α7-receptor expression that were stroked during a conditioned odor did not show a behavioral response to that odorant. These results suggest that decreased α7-receptor expression has a role in associative learning, olfactory preference, and/or sensory processing deficits.

  11. Segregation of lipids near acetylcholine-receptor channels imaged by cryo-EM

    Directory of Open Access Journals (Sweden)

    Nigel Unwin

    2017-07-01

    Full Text Available Rapid communication at the chemical synapse depends on the action of ion channels residing in the postsynaptic membrane. The channels open transiently upon the binding of a neurotransmitter released from the presynaptic nerve terminal, eliciting an electrical response. Membrane lipids also play a vital but poorly understood role in this process of synaptic transmission. The present study examines the lipid distribution around nicotinic acetylcholine (ACh receptors in tubular vesicles made from postsynaptic membranes of the Torpedo ray, taking advantage of the recent advances in cryo-EM. A segregated distribution of lipid molecules is found in the outer leaflet of the bilayer. Apparent cholesterol-rich patches are located in specific annular regions next to the transmembrane helices and also in a more extended `microdomain' between the apposed δ subunits of neighbouring receptors. The particular lipid distribution can be interpreted straightforwardly in relation to the gating movements revealed by an earlier time-resolved cryo-EM study, in which the membranes were exposed briefly to ACh. The results suggest that in addition to stabilizing the protein, cholesterol may play a mechanical role by conferring local rigidity to the membrane so that there is productive coupling between the extracellular and membrane domains, leading to opening of the channel.

  12. Segregation of acetylcholine and GABA in the rat superior cervical ganglia: functional correlation.

    Directory of Open Access Journals (Sweden)

    Diana eElinos

    2016-04-01

    Full Text Available Sympathetic neurons have the capability to segregate their neurotransmitters (NTs and co-transmitters to separate varicosities of single axons; furthermore, in culture, these neurons can even segregate classical transmitters. In vivo sympathetic neurons employ acetylcholine (ACh and other classical NTs such as gamma aminobutyric acid (GABA. Herein, we explore whether these neurons in vivo segregate these classical NTs in the superior cervical ganglia of the rat. We determined the topographical distribution of GABAergic varicosities, somatic GABAA receptor, as well as the regional distribution of the segregation of ACh and GABA. We evaluated possible regional differences in efficacy of ganglionic synaptic transmission, in the sensitivity of GABAA receptor to GABA and to the competitive antagonist picrotoxin (PTX. We found that sympathetic preganglionic neurons in vivo do segregate ACh and GABA. GABAergic varicosities and GABAA receptor expression showed a rostro-caudal gradient along ganglia; in contrast, segregation exhibited a caudo-rostral gradient. These uneven regional distributions in expression of GABA, GABAA receptors, and level segregation correlate with stronger synaptic transmission found in the caudal region. Accordingly, GABAA receptors of rostral region show larger sensitivity to GABA and PTX. These results suggest the presence of different types of GABAA receptors in each region that result in a different regional levels of endogenous GABA inhibition. Finally, we discuss a possible correlation of these different levels of GABA modulation and the function of the target organs innervated by rostral and caudal ganglionic neurons.

  13. Metabolic stabilization of acetylcholine receptors in vertebrate neuromuscular junction by muscle activity

    International Nuclear Information System (INIS)

    Rotzler, S.; Brenner, H.R.

    1990-01-01

    The effects of muscle activity on the growth of synaptic acetylcholine receptor (AChR) accumulations and on the metabolic AChR stability were investigated in rat skeletal muscle. Ectopic end plates induced surgically in adult soleus muscle were denervated early during development when junctional AChR number and stability were still low and, subsequently, muscles were either left inactive or they were kept active by chronic exogenous stimulation. AChR numbers per ectopic AChR cluster and AChR stabilities were estimated from the radioactivity and its decay with time, respectively, of end plate sites whose AChRs had been labeled with 125 I-alpha-bungarotoxin (alpha-butx). The results show that the metabolic stability of the AChRs in ectopic clusters is reversibly increased by muscle activity even when innervation is eliminated very early in development. 1 d of stimulation is sufficient to stabilize the AChRs in ectopic AChR clusters. Muscle stimulation also produced an increase in the number of AChRs at early denervated end plates. Activity-induced cluster growth occurs mainly by an increase in area rather than in AChR density, and for at least 10 d after denervation is comparable to that in normally developing ectopic end plates. The possible involvement of AChR stabilization in end plate growth is discussed

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

    Directory of Open Access Journals (Sweden)

    Maria Di Bari

    2016-11-01

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

  15. Drugs Interfering with Muscarinic Acetylcholine Receptors and Their Effects on Place Navigation

    Directory of Open Access Journals (Sweden)

    Jan Svoboda

    2017-11-01

    Full Text Available Muscarinic acetylcholine receptors (mAChRs have been found to regulate many diverse functions, ranging from motivation and feeding to spatial navigation, an important and widely studied type of cognitive behavior. Systemic administration of non-selective antagonists of mAChRs, such as scopolamine or atropine, have been found to have adverse effects on a vast majority of place navigation tasks. However, many of these results may be potentially confounded by disruptions of functions other than spatial learning and memory. Although studies with selective antimuscarinics point to mutually opposite effects of M1 and M2 receptors, their particular contribution to spatial cognition is still poorly understood, partly due to a lack of truly selective agents. Furthermore, constitutive knock-outs do not always support results from selective antagonists. For modeling impaired spatial cognition, the scopolamine-induced amnesia model still maintains some limited validity, but there is an apparent need for more targeted approaches such as local intracerebral administration of antagonists, as well as novel techniques such as optogenetics focused on cholinergic neurons and chemogenetics aimed at cells expressing metabotropic mAChRs.

  16. Autoradiographic localization of nicotinic acetylcholine receptors in the brain of the zebra finch (Poephila guttata)

    International Nuclear Information System (INIS)

    Watson, J.T.; Adkins-Regan, E.; Whiting, P.; Lindstrom, J.M.; Podleski, T.R.

    1988-01-01

    We have localized nicotinic acetylcholine receptors in the zebra finch brain by using three 125I-labelled ligands: alpha bungarotoxin and two monoclonal antibodies to neuronal nicotinic receptors. Unfixed brains from intact adult male and female zebra finches were prepared for in vitro autoradiography. Low-resolution film autoradiograms and high-resolution emulsion autoradiograms were prepared for each of the three ligands. The major brain structures that bind all three of the ligands are hippocampus; hyperstriatum dorsalis; hyperstriatum ventralis; nucleus lentiformis mesencephali; nucleus pretectalis, some layers of the optic tectum; nucleus mesencephalicus lateralis; pars dorsalis; locus ceruleus; and all cranial motor nuclei except nucleus nervi hypoglossi. The major structures labelled only by [125I]-alpha bungarotoxin binding included hyperstriatum accessorium and the nuclei: preopticus medialis, medialis hypothalami posterioris, semilunaris, olivarius inferior, and the periventricular organ. Of the song control nuclei, nucleus magnocellularis of the anterior neostriatum; hyperstriatum ventralis, pars caudalis; nucleus intercollicularis; and nucleus hypoglossus were labelled. The binding patterns of the two antibodies were similar to one another but not identical. Both labelled nucleus spiriformis lateralis and nucleus geniculatus lateralis, pars ventralis especially heavily and also labelled the nucleus habenula medialis; nucleus subpretectalis; nucleus isthmi, pars magnocellularis; nucleus reticularis gigantocellularis; nucleus reticularis lateralis; nucleus tractus solitarii; nucleus vestibularis dorsolateralis; nucleus vestibularis lateralis; nucleus descendens nervi trigemini; and the deep cerebellar nuclei

  17. The effect of exogenous acetylcholine and other cholinergic agents on photoperiodic flower induction of Pharbitis nil

    Directory of Open Access Journals (Sweden)

    Hanna Łukasiewicz-Rutkowska

    2014-01-01

    Full Text Available Exogenous acetylcholine (ACh, acetylcholinesterase (AChE inhibitors, as well as agonists and antagonists of ACh receptors applied on the cotyledons of 5-day-old seedlings of Pharbitis nil during a 16-hour long inductive night or during a 12-hour long subinductive night, do not essentially influence the flower bud formation. Also the application of above mentioned substances to the seedlings growing under the conditions of 72 hours of darkness, 24 hours of light and then 24 hours of darkness does not influence in an essential way flowering of P. nil. On the other hand, applying these substances on the cotyledons of P. nil during 24-hour-long inductive night, preceded by 72 hours of darkness and then 24 hours of light of lowered intensity finished by 15-minute-long impulse of far red light which inhibit flowering, caused the reversion of the effect of far red light irradiation and stimulated the flowering. The obtained results suggest that endogenous system ACh/AChE could participate in the mechanism of a phytochrome controlled flowering of short day plants.

  18. A New Role for Attentional Corticopetal Acetylcholine in Cortical Memory Dynamics

    Science.gov (United States)

    Fujii, Hiroshi; Kanamaru, Takashi; Aihara, Kazuyuki; Tsuda, Ichiro

    2011-09-01

    Although the role of corticopetal acetylcholine (ACh) in higher cognitive functions is increasingly recognized, the questions as (1) how ACh works in attention(s), memory dynamics and cortical state transitions, and also (2) why and how loss of ACh is involved in dysfunctions such as visual hallucinations in dementia with Lewy bodies and deficit of attention(s), are not well understood. From the perspective of a dynamical systems viewpoint, we hypothesize that transient ACh released under top-down attention serves to temporarily invoke attractor-like memories, while a background level of ACh reverses this process returning the dynamical nature of the memory structure back to attractor ruins (quasi-attractors). In fact, transient ACh loosens inhibitions of py ramidal neurons (PYRs) by P V+ fas t spiking (FS) i nterneurons, while a baseline ACh recovers inhibitory actions of P V+ FS. Attentional A Ch thus dynamically modifies brain's connectivity. Th e core of this process is in the depression of GABAergic inhibitory currents in PYRs due to muscarinic (probably M2 subtype) presyn aptic effects on GABAergic synapses of PV+ FS neurons

  19. Functional interaction of nicotinic acetylcholine receptors and Na+/K+ ATPase from Locusta migratoria manilensis (Meyen).

    Science.gov (United States)

    Bao, Haibo; Sun, Huahua; Xiao, Youxin; Zhang, Yixi; Wang, Xin; Xu, Xiaoyong; Liu, Zewen; Fang, Jichao; Li, Zhong

    2015-03-06

    Associated proteins are important for the correct functioning of nicotinic acetylcholine receptors (nAChRs). In the present study, a neonicotinoid-agarose affinity column was used to isolate related proteins from a solubilized membrane preparation from the nervous system of Locusta migratoria manilensis (Meyen). 1530 peptides were identified and most of them were involved in the membranous structure, molecular interaction and cellular communication. Among these peptides, Na(+)/K(+) ATPase had the highest MASCOT score and were involved in the molecular interaction, which suggested that Na(+)/K(+) ATPase and nAChRs might have strong and stable interactions in insect central nervous system. In the present study, functional interactions between nAChRs and Na(+)/K(+) ATPase were examined by heterologous expression in Xenopus oocytes. The results showed that the activated nAChRs increased pump currents of Na(+)/K(+) ATPase, which did not require current flow through open nAChRs. In turn, Na(+)/K(+) ATPase significantly increased agonist sensitivities of nAChRs in a pump activity-independent manner and reduced the maximum current (Imax) of nAChRs. These findings provide novel insights concerning the functional interactions between insect nAChRs and Na(+)/K(+) ATPase.

  20. alpha7 Nicotinic acetylcholine receptor knockout selectively enhances ethanol-, but not beta-amyloid-induced neurotoxicity.

    Science.gov (United States)

    de Fiebre, Nancyellen C; de Fiebre, Christopher M

    2005-01-03

    The alpha7 subtype of nicotinic acetylcholine receptor (nAChR) has been implicated as a potential site of action for two neurotoxins, ethanol and the Alzheimer's disease related peptide, beta-amyloid. Here, we utilized primary neuronal cultures of cerebral cortex from alpha7 nAChR null mutant mice to examine the role of this receptor in modulating the neurotoxic properties of subchronic, "binge" ethanol and beta-amyloid. Knockout of the alpha7 nAChR gene selectively enhanced ethanol-induced neurotoxicity in a gene dosage-related fashion. Susceptibility of cultures to beta-amyloid induced toxicity, however, was unaffected by alpha7 nAChR gene null mutation. Further, beta-amyloid did not inhibit the binding of the highly alpha7-selective radioligand, [(125)I]alpha-bungarotoxin. On the other hand, in studies in Xenopus oocytes ethanol efficaciously inhibited alpha7 nAChR function. These data suggest that alpha7 nAChRs modulate the neurotoxic effects of binge ethanol, but not the neurotoxicity produced by beta-amyloid. It is hypothesized that inhibition of alpha7 nAChRs by ethanol provides partial protection against the neurotoxic properties of subchronic ethanol.

  1. Optical acetylcholine sensor based on free base porphyrin as a chromoionophore.

    Science.gov (United States)

    Mroczkiewicz, Monika; Pietrzak, Mariusz; Górski, Łukasz; Malinowska, Elżbieta

    2011-09-21

    In this work, the possibility of application of free base porphyrin as a lipophilic pH chromoionophore for the preparation of optical cation-selective sensors was investigated. The properties of polymeric membranes, containing porphyrins of different structures, namely tetraphenylporphyrin (TPP) and octaethylporphyrin (OEP), were compared. Changes in equilibrium between protonated and deprotonated form of porphyrin, resulting from variations in ACh concentration, were evaluated. The influence of various factors (kind and quantity of anionic additive and porphyrin in the membrane phase, pH of sample solution) on initial equilibrium was studied. The best membrane composition was chosen as: TPP 3 wt.%, KTFPB 175 mol.% relative to ionophore, PVC:o-NPOE (1 : 4) and measuring buffer solution: 0.05 M MES, pH 4.5. Selectivity, response stability, reversibility and repeatability tests were carried out for chosen sensor. Developed sensor allowed for the determination of a model analyte, acetylcholine, at the concentration range of 10(-5) to 10(-2) M, both in stationary and flow-injection system. Sensor response was reversible and repeatable in the mentioned concentration range.

  2. Selective decreases of nicotinic acetylcholine receptors in PC12 cells exposed to fluoride

    International Nuclear Information System (INIS)

    Chen Jia; Shan, K.-R.; Long, Y.-G.; Wang, Y.-N.; Nordberg, Agneta; Guan, Z.-Z.

    2003-01-01

    In an attempt to elucidate the mechanism by which excessive fluoride damages the central nervous system, the effects of exposure of PC12 cells to different concentrations of fluoride for 48 h on nicotinic acetylcholine receptors (nAChRs) were characterized here. Significant reductions in the number of binding sites for both [ 3 H]epibatidine and [ 125 I]α-bungarotoxin, as well as a significant decrease in the B max value for the high-affinity of epibatidine binding site were observed in PC12 cells subjected to high levels of fluoride. On the protein level, the α3 and α7 subunits of nAChRs were also significantly decreased in the cells exposed to high concentrations of fluoride. In contrast, such exposure had no significant effect on the level of the β2 subunit. These findings suggest that selective decreases in the number of nAChRs may play an important role in the mechanism(s) by which fluoride causes dysfunction of the central nervous system

  3. Acetylcholine release in the hippocampus during the operant conditioned reflex and the footshock stimulus in rats.

    Science.gov (United States)

    Dong, Yu; Mao, Jianjun; Shangguan, Dihua; Zhao, Rui; Liu, Guoquan

    2004-10-14

    The activity of the septo-hippocampal cholinergic pathway was investigated by measuring changes in the extracellular acetylcholine (ACh) levels in the hippocampus, by means of microdialysis, during the operant conditioned reflex and the repeated footshock stimulus. Microdialysis samplings were conducted in a Skinner box where lights were delivered as conditioned stimuli (CS) paired with footshocks as unconditioned stimuli (US). Two groups of rats were used. Extracellular ACh and choline (Ch) in samples collected at 6min intervals were assessed by high-performance liquid chromatography with electrochemical detection. The elevation of hippocampus ACh was observed in the two experimental groups. The increase in ACh during aversive stimulus (footshock) was significantly larger and was probably related to the number of footshocks. There might be moderate increase in the hippocampal ACh release during the retrieval of information. The concentration of choline showed no significant fluctuation in the two groups during the whole process. This experiment explored in more detail hippocampal cholinergic activity in relation to the two different procedures.

  4. Neonicotinoids target distinct nicotinic acetylcholine receptors and neurons, leading to differential risks to bumblebees

    Science.gov (United States)

    Moffat, Christopher; Buckland, Stephen T.; Samson, Andrew J.; McArthur, Robin; Chamosa Pino, Victor; Bollan, Karen A.; Huang, Jeffrey T.-J.; Connolly, Christopher N.

    2016-04-01

    There is growing concern over the risk to bee populations from neonicotinoid insecticides and the long-term consequences of reduced numbers of insect pollinators to essential ecosystem services and food security. Our knowledge of the risk of neonicotinoids to bees is based on studies of imidacloprid and thiamethoxam and these findings are extrapolated to clothianidin based on its higher potency at nicotinic acetylcholine receptors. This study addresses the specificity and consequences of all three neonicotinoids to determine their relative risk to bumblebees at field-relevant levels (2.5 ppb). We find compound-specific effects at all levels (individual cells, bees and whole colonies in semi-field conditions). Imidacloprid and clothianidin display distinct, overlapping, abilities to stimulate Kenyon cells, indicating the potential to differentially influence bumblebee behavior. Bee immobility was induced only by imidacloprid, and an increased vulnerability to clothianidin toxicity only occurred following chronic exposure to clothianidin or thiamethoxam. At the whole colony level, only thiamethoxam altered the sex ratio (more males present) and only clothianidin increased queen production. Finally, both imidacloprid and thiamethoxam caused deficits in colony strength, while no detrimental effects of clothianidin were observed. Given these findings, neonicotinoid risk needs to be considered independently for each compound and target species.

  5. Pathogenesis of Abdominal Aortic Aneurysms: Role of Nicotine and Nicotinic Acetylcholine Receptors

    Directory of Open Access Journals (Sweden)

    Zong-Zhuang Li

    2012-01-01

    Full Text Available Inflammation, proteolysis, smooth muscle cell apoptosis, and angiogenesis have been implicated in the pathogenesis of abdominal aortic aneurysms (AAAs, although the well-defined initiating mechanism is not fully understood. Matrix metalloproteinases (MMPs such as MMP-2 and -9 and other proteinases degrading elastin and extracellular matrix are the critical pathogenesis of AAAs. Among the risk factors of AAAs, cigarette smoking is an irrefutable one. Cigarette smoke is practically involved in various aspects of the AAA pathogenesis. Nicotine, a major alkaloid in tobacco leaves and a primary component in cigarette smoke, can stimulate the MMPs expression by vascular SMCs, endothelial cells, and inflammatory cells in vascular wall and induce angiogenesis in the aneurysmal tissues. However, for the inflammatory and apoptotic processes in the pathogenesis of AAAs, nicotine seems to be moving in just the opposite direction. Additionally, the effects of nicotine are probably dose dependent or associated with the exposure duration and may be partly exerted by its receptors—nicotinic acetylcholine receptors (nAChRs. In this paper, we will mainly discuss the pathogenesis of AAAs involving inflammation, proteolysis, smooth muscle cell apoptosis and angiogenesis, and the roles of nicotine and nAChRs.

  6. Gating of long-term potentiation by nicotinic acetylcholine receptors at the cerebellum input stage.

    Directory of Open Access Journals (Sweden)

    Francesca Prestori

    Full Text Available The brain needs mechanisms able to correlate plastic changes with local circuit activity and internal functional states. At the cerebellum input stage, uncontrolled induction of long-term potentiation or depression (LTP or LTD between mossy fibres and granule cells can saturate synaptic capacity and impair cerebellar functioning, which suggests that neuromodulators are required to gate plasticity processes. Cholinergic systems innervating the cerebellum are thought to enhance procedural learning and memory. Here we show that a specific subtype of acetylcholine receptors, the α7-nAChRs, are distributed both in cerebellar mossy fibre terminals and granule cell dendrites and contribute substantially to synaptic regulation. Selective α7-nAChR activation enhances the postsynaptic calcium increase, allowing weak mossy fibre bursts, which would otherwise cause LTD, to generate robust LTP. The local microperfusion of α7-nAChR agonists could also lead to in vivo switching of LTD to LTP following sensory stimulation of the whisker pad. In the cerebellar flocculus, α7-nAChR pharmacological activation impaired vestibulo-ocular-reflex adaptation, probably because LTP was saturated, preventing the fine adjustment of synaptic weights. These results show that gating mechanisms mediated by specific subtypes of nicotinic receptors are required to control the LTD/LTP balance at the mossy fibre-granule cell relay in order to regulate cerebellar plasticity and behavioural adaptation.

  7. Acetylcholine promotes the emergence and elongation of lateral roots of Raphanus sativus.

    Science.gov (United States)

    Sugiyama, Kou-ichi; Tezuka, Takafumi

    2011-10-01

    Radish (Raphanus sativus L.) was grown on four layers of paper towel moistened with distilled water with and without acetylcholine (ACh) for five days in the dark after sowing. ACh at 1 nM promoted the growth (emergence and elongation) of lateral roots of radish plants, but had no effect on the stems and main roots. Moreover, ACh enhanced the dry weight of roots [main (primary) + lateral roots]. Neostigmine, an inhibitor of acetylcholinesterase (AChE) also promoted the emergence and elongation of lateral roots, and atropine, a competitive inhibitor of ACh receptor, suppressed the emergence and elongation. ACh suppressed the activity of AChE and increased the amount of proteins and pyridine nucleotides (NAD and NADH) in the roots of the seedlings. It also increased the activities of NAD-forming enzymes [NAD synthetase and ATP-nicotinamide mononucleotide (ATP-NMN) adenyltransferase], and enhanced the amount of DNA in the roots of the seedlings. The relationship between ACh and the emergence and growth of lateral roots was discussed from a biochemical viewpoint.

  8. Integrated regulation of AMPA glutamate receptor phosphorylation in the striatum by dopamine and acetylcholine.

    Science.gov (United States)

    Xue, Bing; Chen, Elton C; He, Nan; Jin, Dao-Zhong; Mao, Li-Min; Wang, John Q

    2017-01-01

    Dopamine (DA) and acetylcholine (ACh) signals converge onto protein kinase A (PKA) in medium spiny neurons of the striatum to control cellular and synaptic activities of these neurons, although underlying molecular mechanisms are less clear. Here we measured phosphorylation of the α-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor (AMPAR) at a PKA site (S845) as an indicator of AMPAR responses in adult rat brains in vivo to explore how DA and ACh interact to modulate AMPARs. We found that subtype-selective activation of DA D1 receptors (D1Rs), D2 receptors (D2Rs), or muscarinic M4 receptors (M4Rs) induced specific patterns of GluA1 S845 responses in the striatum. These defined patterns support a local multitransmitter interaction model in which D2Rs inhibited an intrinsic inhibitory element mediated by M4Rs to enhance the D1R efficacy in modulating AMPARs. Consistent with this, selective enhancement of M4R activity by a positive allosteric modulator resumed the cholinergic inhibition of D1Rs. In addition, D1R and D2R coactivation recruited GluA1 and PKA preferentially to extrasynaptic sites. In sum, our in vivo data support an existence of a dynamic DA-ACh balance in the striatum which actively modulates GluA1 AMPAR phosphorylation and trafficking. This article is part of the Special Issue entitled 'Ionotropic glutamate receptors'. Copyright © 2016 Elsevier Ltd. All rights reserved.

  9. Synthesis of acetylcholine from choline derived from phosphatidylcholine in a human neuronal cell line

    International Nuclear Information System (INIS)

    Blusztajn, J.K.; Liscovitch, M.; Richardson, U.I.

    1987-01-01

    Cholinergic neurons are unique among cells since they alone utilize choline not only as a component of major membrane phospholipids, such as phosphatidylcholine (Ptd-Cho), but also as a precursor of their neurotransmitter acetylcholine (AcCho). It has been hypothesized that choline-phospholipids might serve as a storage pool of choline for AcCho synthesis. The selective vulnerability of cholinergic neurons in certain neurodegenerative diseases (e.g., Alzheimer disease, motor neuron disorders) might result from the abnormally accelerated liberation of choline (to be used a precursor of AcCho) from membrane phospholipids, resulting in altered membrane composition and function and compromised neuronal viability. However, the proposed metabolic link between membrane turnover and AcCho synthesis has been difficult to demonstrate because of the heterogeneity of the preparations used. Here the authors used a population of purely cholinergic cells (human neuroblastomas, LA-N-2), incubated in the presence of [methyl- 3 H]methionine to selectively label PtdCho synthesized by methylation of phosphatidylethanolamine, the only pathway of de novo choline synthesis. Three peaks of radioactive material that cochromatographed with authentic AcCho, choline, and phosphocholine were observed when the water-soluble metabolites of the [ 3 H]PtdCho were purified by high-performance liquid chromatography. The results demonstrate that AcCho can be synthesized from choline derived from the degradation of endogenous PtdCho formed de novo by methylation of phosphatidylethanolamine

  10. Effect of cobratoxin binding on the normal mode vibration within acetylcholine binding protein.

    Science.gov (United States)

    Bertaccini, Edward J; Lindahl, Erik; Sixma, Titia; Trudell, James R

    2008-04-01

    Recent crystal structures of the acetylcholine binding protein (AChBP) have revealed surprisingly small structural alterations upon ligand binding. Here we investigate the extent to which ligand binding may affect receptor dynamics. AChBP is a homologue of the extracellular component of ligand-gated ion channels (LGICs). We have previously used an elastic network normal-mode analysis to propose a gating mechanism for the LGICs and to suggest the effects of various ligands on such motions. However, the difficulties with elastic network methods lie in their inability to account for the modest effects of a small ligand or mutation on ion channel motion. Here, we report the successful application of an elastic network normal mode technique to measure the effects of large ligand binding on receptor dynamics. The present calculations demonstrate a clear alteration in the native symmetric motions of a protein due to the presence of large protein cobratoxin ligands. In particular, normal-mode analysis revealed that cobratoxin binding to this protein significantly dampened the axially symmetric motion of the AChBP that may be associated with channel gating in the full nAChR. The results suggest that alterations in receptor dynamics could be a general feature of ligand binding.

  11. Modulation of acetylcholine release from rat striatal slices by the GABA/benzodiazepine receptor complex

    Energy Technology Data Exchange (ETDEWEB)

    Supavilai, P.; Karobath, M.

    1985-02-04

    GABA, THIP and muscimol enhance spontaneous and inhibit electrically induced release of tritium labelled compounds from rat striatal slices which have been pre-labelled with /sup 3/H-choline. Baclofen is inactive in this model. Muscimol can inhibit electrically induced release of tritiated material by approximately 75% with half maximal effects at 2 ..mu..M. The response to muscimol can be blocked by the GABA antagonists bicuculline methobromide, picrotoxin, anisatin, R 5135 and CPTBO (cyclopentylbicyclophosphate). Drugs which act on the benzodiazepine receptor (BR) require the presence of muscimol to be effective and they modulate the effects of muscimol in a bidirectional manner. Thus BR agonists enhance and inverse BR agonists attenuate the inhibitory effects of muscimol on electrically induced release. Ro15-1788, a BR antagonist, does not modulate the inhibitory effects of muscimol but antagonizes the actions of clonazepam, a BR agonist, and of DMCM, an inverse BR agonist. These results demonstrate that a GABA/benzodiazepine receptor complex can modulate acetylcholine release from rat striatal slices in vitro. 24 references, 3 figures, 5 table.

  12. Bilirubin Modulates Acetylcholine Receptors In Rat Superior Cervical Ganglionic Neurons In a Bidirectional Manner

    Science.gov (United States)

    Zhang, Chengmi; Wang, Zhenmeng; Dong, Jing; Pan, Ruirui; Qiu, Haibo; Zhang, Jinmin; Zhang, Peng; Zheng, Jijian; Yu, Weifeng

    2014-01-01

    Autonomic dysfunction as a partial contributing factor to cardiovascular instability in jaundiced patients is often associated with increased serum bilirubin levels. Whether increased serum bilirubin levels could directly inhibit sympathetic ganglion transmission by blocking neuronal nicotinic acetylcholine receptors (nAChRs) remains to be elucidated. Conventional patch-clamp recordings were used to study the effect of bilirubin on nAChRs currents from enzymatically dissociated rat superior cervical ganglia (SCG) neurons. The results showed that low concnetrations (0.5 and 2 μM) of bilirubin enhanced the peak ACh-evoked currents, while high concentrations (3 to 5.5 µM) of bilirubin suppressed the currents with an IC50 of 4 ± 0.5 μM. In addition, bilirubin decreased the extent of desensitization of nAChRs in a concentration-dependent manner. This inhibitory effect of bilirubin on nAChRs channel currents was non-competitive and voltage independent. Bilirubin partly improved the inhibitory effect of forskolin on ACh-induced currents without affecting the action of H-89. These data suggest that the dual effects of enhancement and suppression of bilirubin on nAChR function may be ascribed to the action mechanism of positive allosteric modulation and direct blockade. Thus, suppression of sympathetic ganglionic transmission through postganglionic nAChRs inhibition may partially contribute to the adverse cardiovascular effects in jaundiced patients. PMID:25503810

  13. Recombinant human acetylcholine receptor alpha-subunit induces chronic experimental autoimmune myasthenia gravis.

    Science.gov (United States)

    Lennon, V A; Lambert, E H; Leiby, K R; Okarma, T B; Talib, S

    1991-04-01

    A synthetic gene encoding the 210 N-terminal residues of the alpha-subunit of the nicotinic acetylcholine receptor (AChR) of human skeletal muscle was cloned into an inducible expression plasmid to produce a fusion protein in high yield in Escherichia coli. Like native human AChR, the recombinant human alpha 1-210 protein induced AChR-binding, AChR-modulating, and AChR-blocking autoantibodies in rats when injected once intradermally as an emulsion in CFA, with Bordetella pertussis vaccine as supplementary adjuvant. The minimum dose of recombinant protein required to induce biochemical signs of experimental autoimmune myasthenia gravis (EAMG) with 100% incidence was 2.2 micrograms. With 6.6 to 22 micrograms, serum levels of autoantibodies were persistent, and clinically apparent EAMG lasted more than a month. Clinical, electrophysiological, and biochemical indices of EAMG induced by doses of 66 micrograms or more were more uniformly severe and persistent, with 33% fatality. Rats receiving a control extract of E. coli containing plasmid without the alpha 1-210 codon insert, with adjuvants, did not develop autoantibodies or signs of EAMG. This highly reproducible new model of EAMG induced by a recombinant human autoantigen should be valuable for testing Ag-specific immunotherapeutic strategies that might be applicable to treating acquired myasthenia gravis in humans.

  14. Immunological studies on the structure and function of the nicotinic acetylcholine receptor in mammalian muscle

    Energy Technology Data Exchange (ETDEWEB)

    Gu, Y.

    1989-01-01

    The specificity of the antibodies in the serum of a patient with myasthenia gravis for a the {alpha}-bungarotoxin binding sites of the acetylcholine receptor (AChR) was examined using AChRs in the C2 mouse muscle cell line as a model. The antibodies were shown to be specific for one of the two toxin-binding sites. The effect of the antibodies in this myasthenic serum on the functional response of the receptor to cholinergic agonists was also examined using carbamylcholine-induced {sup 22}Na uptake into C2 myotubes as a measured of the receptor function. Antibodies specific for the {gamma}, {delta}, and {epsilon} subunit, respectively, of mammalian muscle AChRs were developed using subunit-specific synthetic peptides as antigens. Using these antibodies and monoclonal antibodies for other subunits as probes, I have identified four ({alpha}, {beta}, {gamma}, and {delta}) subunits of mammalian muscle AChRs on immunoblots. When AChRs from embryonic, neonatal, normal and denervated adult muscles were compared on immunoblots, the {alpha}, {beta}, and {delta} subunits were identical in all four receptor preparations, with or without endoglycosidase digestion. The spatial and temporal distribution of the {gamma}- and {epsilon}- AChRs in developing and in denervated muscles corresponds to the distribution of AChRs with slow and fast channels, respectively, and that the development changes in the channel properties of the receptor arise from a change in the subunit composition of the receptor, in which the {gamma} is replaced by {epsilon}.

  15. Binding affinities of anti-acetylcholine receptor autoantibodies in myasthenia gravis

    Energy Technology Data Exchange (ETDEWEB)

    Bray, J.J.; Drachman, D.B.

    1982-01-01

    Antibodies directed against acetylcholine (ACh) receptors are present in the sera of nearly 90% of patients with myasthenia gravis (MG), and are involved in the pathogenesis of this autoimmune disease. However, the antibody titers measured by the standard radioimmunoassay correspond poorly with the clinical severity of the disease. To determine whether this disparity could be accounted for by differences in the binding affinities of anti-ACh receptor antibodies in different patients, we have measured the binding affinities of these autoantibodies in 15 sera from MG patients. The affinity constants (K/sub o/), as determined by Scatchard analysis, were all in the range of 10/sup 10/ M/sup -1/, comparable to the highest values reported in immunized animals. The affinity constants were truly representative of the population of autoantibodies detected by the radioimmunoassay, as shown by the remarkable linearity of the Scatchard plots (r/sup 2/>0.90) and the close correlation between the antibody titers determined by extrapolation of the Scatchard plots and by saturation analysis (r = 0.99; p < 0.001). There was only a 6-fold variation in affinity constants measured in this series of patients despite widely differing antibody titers and severity of the disease. Factors other than the titer and affinity of anti-ACh receptor antibodies may correlate better with the clinical manifestations of MG.

  16. Immunological studies on the structure and function of the nicotinic acetylcholine receptor in mammalian muscle

    International Nuclear Information System (INIS)

    Gu, Y.

    1989-01-01

    The specificity of the antibodies in the serum of a patient with myasthenia gravis for a the α-bungarotoxin binding sites of the acetylcholine receptor (AChR) was examined using AChRs in the C2 mouse muscle cell line as a model. The antibodies were shown to be specific for one of the two toxin-binding sites. The effect of the antibodies in this myasthenic serum on the functional response of the receptor to cholinergic agonists was also examined using carbamylcholine-induced 22 Na uptake into C2 myotubes as a measured of the receptor function. Antibodies specific for the γ, δ, and ε subunit, respectively, of mammalian muscle AChRs were developed using subunit-specific synthetic peptides as antigens. Using these antibodies and monoclonal antibodies for other subunits as probes, I have identified four (α, β, γ, and δ) subunits of mammalian muscle AChRs on immunoblots. When AChRs from embryonic, neonatal, normal and denervated adult muscles were compared on immunoblots, the α, β, and δ subunits were identical in all four receptor preparations, with or without endoglycosidase digestion. The spatial and temporal distribution of the γ- and ε- AChRs in developing and in denervated muscles corresponds to the distribution of AChRs with slow and fast channels, respectively, and that the development changes in the channel properties of the receptor arise from a change in the subunit composition of the receptor, in which the γ is replaced by ε

  17. Unraveling a molecular determinant for clathrin-independent internalization of the M2 muscarinic acetylcholine receptor

    Science.gov (United States)

    Wan, Min; Zhang, Wenhua; Tian, Yangli; Xu, Chanjuan; Xu, Tao; Liu, Jianfeng; Zhang, Rongying

    2015-01-01

    Endocytosis and postendocytic sorting of G-protein-coupled receptors (GPCRs) is important for the regulation of both their cell surface density and signaling profile. Unlike the mechanisms of clathrin-dependent endocytosis (CDE), the mechanisms underlying the control of GPCR signaling by clathrin-independent endocytosis (CIE) remain largely unknown. Among the muscarinic acetylcholine receptors (mAChRs), the M4 mAChR undergoes CDE and recycling, whereas the M2 mAChR is internalized through CIE and targeted to lysosomes. Here we investigated the endocytosis and postendocytic trafficking of M2 mAChR based on a comparative analysis of the third cytoplasmic domain in M2 and M4 mAChRs. For the first time, we identified that the sequence 374KKKPPPS380 servers as a sorting signal for the clathrin-independent internalization of M2 mAChR. Switching 374KKKPPPS380 to the i3 loop of the M4 mAChR shifted the receptor into lysosomes through the CIE pathway; and therefore away from CDE and recycling. We also found another previously unidentified sequence that guides CDE of the M2 mAChR, 361VARKIVKMTKQPA373, which is normally masked in the presence of the downstream sequence 374KKKPPPS380. Taken together, our data indicate that endocytosis and postendocytic sorting of GPCRs that undergo CIE could be sequence-dependent. PMID:26094760

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

    Energy Technology Data Exchange (ETDEWEB)

    Herholz, Karl [University of Manchester, Wolfson Molecular Imaging Centre, Clinical Neuroscience, Manchester (United Kingdom); University of Cologne, Cologne (Germany)

    2008-03-15

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

  19. Neuronal M3 muscarinic acetylcholine receptors are essential for somatotroph proliferation and normal somatic growth.

    Science.gov (United States)

    Gautam, Dinesh; Jeon, Jongrye; Starost, Matthew F; Han, Sung-Jun; Hamdan, Fadi F; Cui, Yinghong; Parlow, Albert F; Gavrilova, Oksana; Szalayova, Ildiko; Mezey, Eva; Wess, Jürgen

    2009-04-14

    The molecular pathways that promote the proliferation and maintenance of pituitary somatotrophs and other cell types of the anterior pituitary gland are not well understood at present. However, such knowledge is likely to lead to the development of novel drugs useful for the treatment of various human growth disorders. Although muscarinic cholinergic pathways have been implicated in regulating somatotroph function, the physiological relevance of this effect and the localization and nature of the receptor subtypes involved in this activity remain unclear. We report the surprising observation that mutant mice that selectively lack the M(3) muscarinic acetylcholine receptor subtype in the brain (neurons and glial cells; Br-M3-KO mice) showed a dwarf phenotype associated with a pronounced hypoplasia of the anterior pituitary gland and a marked decrease in pituitary and serum growth hormone (GH) and prolactin. Remarkably, treatment of Br-M3-KO mice with CJC-1295, a synthetic GH-releasing hormone (GHRH) analog, rescued the growth deficit displayed by Br-M3-KO mice by restoring normal pituitary size and normal serum GH and IGF-1 levels. These findings, together with results from M(3) receptor/GHRH colocalization studies and hypothalamic hormone measurements, support a model in which central (hypothalamic) M(3) receptors are required for the proper function of hypothalamic GHRH neurons. Our data reveal an unexpected and critical role for central M(3) receptors in regulating longitudinal growth by promoting the proliferation of pituitary somatotroph cells.

  20. Efficient Expression of Acetylcholine-Binding Protein from Aplysia californica in Bac-to-Bac System

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    Bo Lin

    2014-01-01

    Full Text Available The Bac-to-Bac baculovirus expression system can efficiently produce recombinant proteins, but the system may have to be optimized to achieve high-level expression for different candidate proteins. We reported here the efficient expression of acetylcholine-binding proteins from sea hares Aplysia californica (Ac-AChBP and a convenient method to monitor protein expression level in this expression system. Three key factors affecting expression of Ac-AChBP were optimized for maximizing the yield, which included the cell density, volume of the infecting baculovirus inoculums, and the culturing time of postinfection. We have found it to reach a high yield of ∼5 mg/L, which needs 55 h incubation after infection at the cell density of 2 × 106 cells/mL with an inoculum volume ratio of 1 : 100. The optimized expression system in this study was also applied for expressing another protein Ls-AChBP from Lymnaea stagnalis successfully. Therefore, this established method is helpful to produce high yields of AChBP proteins for X-ray crystallographic structural and functional studies.

  1. BDNF Up-Regulates α7 Nicotinic Acetylcholine Receptor Levels on Subpopulations of Hippocampal Interneurons

    Science.gov (United States)

    Massey, Kerri A.; Zago, Wagner M.; Berg, Darwin K.

    2006-01-01

    In the hippocampus, brain-derived neurotrophic factor (BDNF) regulates a number of synaptic components. Among these are nicotinic acetylcholine receptors containing α7 subunits (α7-nAChRs), which are interesting because of their relative abundance in the hippocampus and their high relative calcium permeability. We show here that BDNF elevates surface and intracellular pools of α7-nAChRs on cultured hippocampal neurons and that glutamatergic activity is both necessary and sufficient for the effect. Blocking transmission through NMDA receptors with APV blocked the BDNF effect; increasing spontaneous excitatory activity with the GABAA receptor antagonist bicuculline replicated the BDNF effect. BDNF antibodies blocked the BDNF-mediated increase but not the bicuculline one, consistent with enhanced glutamatergic activity acting downstream from BDNF. Increased α7-nAChR clusters were most prominent on interneuron subtypes known to innervate directly excitatory neurons. The results suggest that BDNF, acting through glutamatergic transmission, can modulate hippocampal output in part by controlling α7-nAChR levels. PMID:17029981

  2. Dopamine regulation of [3H]acetylcholine release from guinea-pig stomach

    International Nuclear Information System (INIS)

    Kusunoki, M.; Taniyama, K.; Tanaka, C.

    1985-01-01

    The involvement of dopamine receptors in cholinergic transmission of guinea-pig stomach was investigated by analyzing the effects of dopamine receptor agonists and antagonists on acetylcholine (ACh) release from this organ. Electrical stimulation (1-20 Hz) of strips of guinea-pig stomach preloaded with [ 3 H] choline induced a [ 3 H]ACh release that was calcium dependent and tetrodotoxin sensitive. Dopamine inhibited this transmural stimulation-induced [ 3 H]ACh release in a concentration-dependent manner (10(-8)-10(-4) M). This effect of dopamine was not altered by 10(-5) M hexamethonium, thereby suggesting that the major dopamine receptors are located on the postganglionic cholinergic neurons. Concentration-response curves for dopamine on [ 3 H]ACh release were inhibited by haloperidol, sulpiride and domperidone but not by prazosin, yohimbine, propranolol and ketanserin. LY 171555, an agonist for the D2 dopamine receptor, but not SKF 38-393, an agonist for the D1 dopamine receptor, to some extent decreased the release of [ 3 H]ACh induced by transmural stimulation. In view of the results, the release of ACh from postganglionic cholinergic neurons is probably required through dopamine receptors antagonized by D2 antagonists but not by adrenergic or serotonin receptor antagonists

  3. Action of hypo- and hyperthyroidism on the postmortal decay of acetylcholine in the rat spinal cord.

    Science.gov (United States)

    Molinengo, L; Cassone, M C; Oggero, L

    1986-01-01

    The postmortal decay of acetylcholine (Ach) was studied in the cervical spinal cords of rats in conditions of hyper- and hypothyroidism. The modifications of thyroid function were achieved either by chronic (20-25 days) administration of l-thyroxine or of methimazole. The basal metabolic rate and plasma T4 concentration were measured to estimate the degree of modification of thyroid activity. The levels of Ach at the start of postmortal decay were evaluated by extrapolation to time 0 of the curves of the postmortal decay of Ach and the levels of Ach at stabilization were estimated from the means of all the measures made at lapses of time over 100-200 s from death. In low and high hypothyroidism a reduction (53 and 72%, respectively) of the levels of Ach was found. A similar effect was found in hyperthyroidism: a 73 and 63% reduction of Ach levels in high and low hyperthyroidism, respectively. The level of Ach at stabilization of the postmortal decay increased only in hyperthyroid rats. The process by which Ach is destroyed is not modified in hyper- or hypothyroidism.

  4. Nicotinic Acetylcholine Receptor α4 Subunit Gene Variation Associated with Attention Deficit Hyperactivity Disorder

    Institute of Scientific and Technical Information of China (English)

    HUANG Xuezhu; XU Yong; LI Qianqian; LIU Pozi; YANG Yuan; ZHANG Fuquan; GUO Tianyou; YANG Chuang; GUO Lanting

    2009-01-01

    Previous pharmacological, human genetics, and animal models have implicated the nicotinic ace-tylcholine receptor a4 subunit (CHRNA4) gene in the pathogenesis of attention deficit/hyperactivity disorder (ADHD). The objective of this study is to examine the genetic association between single nucleotide poly-morphisms in the CHRNA4 gene (rs2273502, rs1044396, rs1044397, and rs3827020 loci) and ADHD. Both case-control and family-based designs are used. Children aged 6 to 16 years were interviewed and as-sessed with the children behavior checklist and the revised conner' parent rating scale to identify probands. No significant differences in the frequency distribution of genotypes or alleles were found between the case and control groups. However, further haplotype analyses showed the CCGG haplotype on dsk for ADHD in 164 case-control samples and the standard transmission disequilibrium test analyses suggest that the allele C of rs2273502 was over-transferred in 98 ADHD parent-offspring tdos. These findings suggest that the CHRNA4 gene may play a role in the pathogenesis of ADHD.

  5. The therapeutic potential of nicotinic acetylcholine receptor agonists for pain control.

    Science.gov (United States)

    Decker, M W; Meyer, M D; Sullivan, J P

    2001-10-01

    Due to the limitations of currently available analgesics, a number of novel alternatives are currently under investigation, including neuronal nicotinic acetylcholine receptor (nAChR) agonists. During the 1990s, the discovery of the antinociceptive properties of the potent nAChR agonist epibatidine in rodents sparked interest in the analgesic potential of this class of compounds. Although epibatidine also has several mechanism-related toxicities, the identification of considerable nAChR diversity suggested that the toxicities and therapeutic actions of the compound might be mediated by distinct receptor subtypes. Consistent with this view, a number of novel nAChR agonists with antinociceptive activity and improved safety profiles in preclinical models have now been identified, including A-85380, ABT-594, DBO-83, SIB-1663 and RJR-2403. Of these, ABT-594 is the most advanced and is currently in Phase II clinical evaluation. Nicotinically-mediated antinociception has been demonstrated in a variety of rodent pain models and is likely mediated by the activation of descending inhibitory pathways originating in the brainstem with the predominant high-affinity nicotine site in brain, the alpha4beta2 subtype, playing a critical role. Thus, preclinical findings suggest that nAChR agonists have the potential to be highly efficacious treatments in a variety of pain states. However, clinical proof-of-principle studies will be required to determine if nAChR agonists are active in pathological pain.

  6. Molecular Modeling of the M3 Acetylcholine Muscarinic Receptor and Its Binding Site

    Directory of Open Access Journals (Sweden)

    Marlet Martinez-Archundia

    2012-01-01

    Full Text Available The present study reports the results of a combined computational and site mutagenesis study designed to provide new insights into the orthosteric binding site of the human M3 muscarinic acetylcholine receptor. For this purpose a three-dimensional structure of the receptor at atomic resolution was built by homology modeling, using the crystallographic structure of bovine rhodopsin as a template. Then, the antagonist N-methylscopolamine was docked in the model and subsequently embedded in a lipid bilayer for its refinement using molecular dynamics simulations. Two different lipid bilayer compositions were studied: one component palmitoyl-oleyl phosphatidylcholine (POPC and two-component palmitoyl-oleyl phosphatidylcholine/palmitoyl-oleyl phosphatidylserine (POPC-POPS. Analysis of the results suggested that residues F222 and T235 may contribute to the ligand-receptor recognition. Accordingly, alanine mutants at positions 222 and 235 were constructed, expressed, and their binding properties determined. The results confirmed the role of these residues in modulating the binding affinity of the ligand.

  7. Action of nereistoxin on recombinant neuronal nicotinic acetylcholine receptors expressed in Xenopus laevis oocytes.

    Science.gov (United States)

    Raymond Delpech, Valérie; Ihara, Makoto; Coddou, Claudio; Matsuda, Kazuhiko; Sattelle, David B

    2003-11-01

    Nereistoxin (NTX), a natural neurotoxin from the salivary glands of the marine annelid worm Lumbriconereis heteropoda, is highly toxic to insects. Its synthetic analogue, Cartap, was the first commercial insecticide based on a natural product. We have used voltage-clamp electrophysiology to compare the actions of NTX on recombinant nicotinic acetylcholine receptors (nicotinic AChRs) expressed in Xenopus laevis oocytes following nuclear injection of cDNAs. The recombinant nicotinic AChRs investigated were chicken alpha7, chicken alpha4beta2 and the Drosophila melanogaster/chicken hybrid receptors SAD/beta2 and ALS/beta2. No agonist action of NTX (0.1-100 microM) was observed on chicken alpha7, chicken alpha4beta2 and the Drosophila/chicken hybrid nicotinic AChRs. Currents elicited by ACh were reduced in amplitude by NTX in a dose-dependent manner. The toxin was slightly more potent on recombinant Drosophila/vertebrate hybrid receptors than on vertebrate homomeric (alpha7) or heteromeric (alpha4beta2) nicotinic AChRs. Block by NTX of the chicken alpha7, chicken alpha4beta2 and the SAD/beta2 and ALS/beta2 Drosophila/chicken hybrid receptors is in all cases non-competitive. Thus, the site of action on nicotinic AChRs of NTX, to which the insecticide Cartap is metabolised in insects, differs from that of the major nicotinic AChR-active insecticide, imidacloprid.

  8. Synthesis and 125I labelling of a precursor for imaging nicotinic acetylcholine receptors

    International Nuclear Information System (INIS)

    Liu Yuxia; Liu Ning; Yang Yuanyou; Zan Liangbiao; Liao Jiali; Jin Jiannan; Sichuan Univ., Chengdu

    2006-01-01

    Nicotinic Acetylcholine Receptors (nAChRs) are involved in various pharmacological effects or diseases, such as Alzheimer's Disease, Parkinson's Disease and tobacco addiction. It will be very appealing to image nAChRs in vivo, diagnose and treat the above diseases, and probe the mechanism of nAChRs in tobacco addiction if the suitable radioactive labeled compound can be synthesized. In this study, (s)-5-(tri-butylstannyl)-3{[1-(tert-butoxycarbonyl)-2-azetidinyl]methoxy} pyridine, a precursor for imaging nAChRs, was synthesized with commercial 2-furfurylamine and (s)-2-azetidinecarboxylic acid as starting materials, and was further labeled with 125/123 I. The whole procedure for radiosynthesis needs 50-55 min and more than 30% of the 125 I are found in the purified 5-[ 125 I]-A-85380. Even staying for 3 days at room temperature in vitro, the purified 5-[ 125 I]-I-85380 can maintain its stability, with a radiochemical purity of more than 95%. (authors)

  9. [3H]acetylcholine synthesis in cultured ciliary ganglion neurons: effects of myotube membranes

    International Nuclear Information System (INIS)

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

    1987-01-01

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

  10. Stabilization of acetylcholine receptors at the neuromuscular synapse: the role of the nerve.

    Science.gov (United States)

    Ramsay, D A; Drachman, D B; Drachman, R J; Stanley, E F

    1992-05-29

    The majority of acetylcholine receptors (AChRs) at innervated neuromuscular junctions (NMJs) are stable, with half-lives averaging about 11 days in rodent muscles. In addition to the stable AChRs, approximately 18% of AChRs at these innervated junctions are rapidly turned over (RTOs), with half lives of less than 24 h. We have postulated that RTOs may be precursors of stable AChRs, and that the motor nerve may influence their stabilization. This hypothesis was tested by: (i) labeling AChRs in mouse sternomastoid (SM) muscles with 125I-alpha-BuTx; (ii) denervating one SM muscle in each mouse, and (iii) following the fate of the labeled AChRs through a 5-day period when RTOs were either stabilized or degraded. The hypothesis predicts that denervation should preclude stabilization of RTOs, resulting in a deficit of stable AChRs in denervated muscles. The results showed a highly significant (P less than 0.002) deficit of stable AChRs in denervated as compared with innervated muscles. Control experiments excluded the possibility that this deficit could be attributed to independent accelerated degradation of either RTOs or pre-existing stable AChRs. The observed deficit was quantitatively consistent with the deficit predicted by a mathematical model based on interruption of stabilization following denervation. We conclude that: (i) the observed deficit after denervation of NMJs is due to failure of stabilization of pre-existing RTOs; (ii) RTOs at normally innervated NMJs are precursors of stable AChRs; (iii) stabilization occurs after the insertion of AChRs at NMJs, and (iv) motor nerves play a key role in stabilization of RTOs. The concept of receptor stabilization has important implications for understanding the biology of the neuromuscular junction and post-synaptic plasticity.

  11. Fluoxetine Alleviates Behavioral Depression while Decreasing Acetylcholine Release in the Nucleus Accumbens Shell

    Science.gov (United States)

    Chau, David T; Rada, Pedro V; Kim, Kay; Kosloff, Rebecca A; Hoebel, Bartley G

    2011-01-01

    Selective serotonin reuptake inhibitors, such as fluoxetine, have demonstrated the ability to alleviate behavioral depression in the forced swim test; however, the sites and mechanisms of their actions remain to be further elucidated. Previous studies have suggested that behavioral depression in the swim test is mediated in part by acetylcholine (ACh) stimulating the cholinergic M1 receptors in the nucleus accumbens (NAc) shell. The current study tested whether acute, local, and chronic, subcutaneous fluoxetine treatments increase escape motivation during the swim test while simultaneously lowering extracellular ACh in the NAc shell. Experiment 1: Fluoxetine (1.0 mM) infused unilaterally in the NAc shell for 40 min reduced extracellular ACh while simultaneously increasing swimming time. Experiment 2: Fluoxetine (0.2, 0.5, and 0.75 mM) infused bilaterally in the NAc shell on day 3 dose-dependently decreased immobility and increased the total escape attempts (swimming and climbing) compared with Ringer given on day 2. Experiment 3: Fluoxetine (0.5 mM) infused bilaterally in the NAc for 40 min did not affect activities in an open field. Experiment 4: Chronic systemic fluoxetine treatment decreased immobility scores and increased total escape attempt scores compared with control saline treatment. In all, 14 days after the initial swim test, basal extracellular ACh in the shell was still elevated in the saline-treated group, but not in the fluoxetine-treated group. In summary, these data suggest that one of the potential mechanisms by which fluoxetine alleviates behavioral depression in the forced swim test may be to suppress cholinergic activities in the NAc shell. PMID:21525864

  12. Mechanism of acetylcholine receptor cluster formation induced by DC electric field.

    Directory of Open Access Journals (Sweden)

    Hailong Luke Zhang

    Full Text Available BACKGROUND: The formation of acetylcholine receptor (AChR cluster is a key event during the development of the neuromuscular junction. It is induced through the activation of muscle-specific kinase (MuSK by the heparan-sulfate proteoglycan agrin released from the motor axon. On the other hand, DC electric field, a non-neuronal stimulus, is also highly effective in causing AChRs to cluster along the cathode-facing edge of muscle cells. METHODOLOGY/PRINCIPAL FINDINGS: To understand its molecular mechanism, quantum dots (QDs were used to follow the movement of AChRs as they became clustered under the influence of electric field. From analyses of trajectories of AChR movement in the membrane, it was concluded that diffuse receptors underwent Brownian motion until they were immobilized at sites of cluster formation. This supports the diffusion-mediated trapping model in explaining AChR clustering under the influence of this stimulus. Disrupting F-actin cytoskeleton assembly and interfering with rapsyn-AChR interaction suppressed this phenomenon, suggesting that these are integral components of the trapping mechanism induced by the electric field. Consistent with the idea that signaling pathways are activated by this stimulus, the localization of tyrosine-phosphorylated forms of AChR β-subunit and Src was observed at cathodal AChR clusters. Furthermore, disrupting MuSK activity through the expression of a kinase-dead form of this enzyme abolished electric field-induced AChR clustering. CONCLUSIONS: These results suggest that DC electric field as a physical stimulus elicits molecular reactions in muscle cells in the form of cathodal MuSK activation in a ligand-free manner to trigger a signaling pathway that leads to cytoskeletal assembly and AChR clustering.

  13. Intermittent hypercapnic hypoxia effects on the nicotinic acetylcholine receptors in the developing piglet hippocampus and brainstem.

    Science.gov (United States)

    Vivekanandarajah, Arunnjah; Aishah, Atqiya; Waters, Karen A; Machaalani, Rita

    2017-05-01

    This study investigated the effects of acute (1 day) vs repeated (4 days) exposure to intermittent hypercapnic hypoxia (IHH) on the immunohistochemical expression of α2, α3, α5, α7, α9 and β2 nicotinic acetylcholine receptor (nAChR) subunits in the developing piglet hippocampus and brainstem medulla, and how prior nicotine exposure alters the response to acute IHH. Five piglet groups included: 1day IHH (1D IHH, n=9), 4days IHH (4D IHH, n=8), controls exposed only to air cycles for 1day (1D Air, n=6) or 4days (4D Air, n=5), and pre-exposed to nicotine for 13days prior to 1day IHH (Nic+1D IHH, n=7). The exposure period alternated 6min of HH (8%O 2 , 7%CO 2 , balance N 2 ) and 6min of air over 48min, while controls were switched from air-to-air. Results showed that: 1. repeated IHH induces more changes in nAChR subunit expression than acute IHH in both the hippocampus and brainstem medulla, 2. In the hippocampus, α2 and β2 changed the most (increased) following IHH and the CA3, CA2 and DG were mostly affected. In the brainstem medulla, α2, α5, α9 and β2 were changed (decreased) in most nuclei with the hypoglossal and nucleus of the solitary tract being mostly affected. 3. Pre-exposure to nicotine enhanced the changes in the hippocampus but dampened those in the brainstem medulla. These findings indicate that the nAChRs (predominantly with the α2/β2 complex) are affected by IHH in critical hippocampal and brainstem nuclei during early brain development, and that pre-exposure to nicotine alters the pattern of susceptibility to IHH. Copyright © 2017 Elsevier B.V. All rights reserved.

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

    Directory of Open Access Journals (Sweden)

    Bettina eNadorp

    2014-02-01

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

  15. A mechanism for acetylcholine receptor gating based on structure, coupling, phi, and flip.

    Science.gov (United States)

    Gupta, Shaweta; Chakraborty, Srirupa; Vij, Ridhima; Auerbach, Anthony

    2017-01-01

    Nicotinic acetylcholine receptors are allosteric proteins that generate membrane currents by isomerizing ("gating") between resting and active conformations under the influence of neurotransmitters. Here, to explore the mechanisms that link the transmitter-binding sites (TBSs) with the distant gate, we use mutant cycle analyses to measure coupling between residue pairs, phi value analyses to sequence domain rearrangements, and current simulations to reproduce a microsecond shut component ("flip") apparent in single-channel recordings. Significant interactions between amino acids separated by >15 Å are rare; an exception is between the αM2-M3 linkers and the TBSs that are ∼30 Å apart. Linker residues also make significant, local interactions within and between subunits. Phi value analyses indicate that without agonists, the linker is the first region in the protein to reach the gating transition state. Together, the phi pattern and flip component suggest that a complete, resting↔active allosteric transition involves passage through four brief intermediate states, with brief shut events arising from sojourns in all or a subset. We derive energy landscapes for gating with and without agonists, and propose a structure-based model in which resting→active starts with spontaneous rearrangements of the M2-M3 linkers and TBSs. These conformational changes stabilize a twisted extracellular domain to promote transmembrane helix tilting, gate dilation, and the formation of a "bubble" that collapses to initiate ion conduction. The energy landscapes suggest that twisting is the most energetically unfavorable step in the resting→active conformational change and that the rate-limiting step in the reverse process is bubble formation. © 2017 Gupta et al.

  16. Analysis of the negative inotropic effect of acetylcholine on frog atrial fibres.

    Science.gov (United States)

    Nargeot, J; Garnier, D; Rougier, O

    1981-03-01

    Voltage-clamp experiments have been performed on frog atrial preparations in order to study the mechanism of the inotropic effect of acetylcholine (ACh) at various concentrations. The amplitude of the slow inward current (Is) is reduced even at low ACh concentrations; such low concentrations have little or no effect on potassium permeability. Dose-effect relationships for Is inhibition (Is/Is max) by ACh show a half amplitude dose (K0.5 around 8 X 10(-8) M ACh. The reduction of Is is attributed largely to a decrease of the maximal conductance of the slow channel (gs). Steady-state activation and inactivation parameters are not affected by ACh. Experiments in a Na-free solution (Na replaced by Li ions) or in a Ca-free solution (with EGTA) indicate that the "slow sodium current" is more sensitive to ACh than the "slow Ca current", although these two currents both seem to flow through the slow channel. The decrease of the phasic component of contraction observed in the presence of ACh is very well correlated with the decrease of Is (K0.5 = 8 X 10(-8) M ACh), while the increase of the tonic tension may be related to the outward potassium current induced by high concentrations of ACh. The significant difference between the half amplitude dose (K0.5) observed in the dose effect curves with ACh for Is inhibition (K0.5 = 8 X 10(-8) M) and for ACh-induced extra-current (K0.5 - 10(-6) M) may indicate the presence of two muscarinic receptors.

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

    Science.gov (United States)

    Tuchek, S; Dolezhal, V; Richny, Ia

    1984-01-01

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

  18. Clinical significance of detection of antibodies to fetal and adult acetylcholine receptors in myasthenia gravis

    Institute of Scientific and Technical Information of China (English)

    Qi-Guang Shi; Zhi-Hong Wang; Xiao-Wei Ma; Da-Qi Zhang; Chun-Sheng Yang; Fu-Dong Shi; Li Yang

    2012-01-01

    Objective To evaluate the frequency,distribution and clinical significance of the antibodies to the fetal and/or adult acetylcholine receptor (AChR) in patients with myasthenia gravis (MG).Methods AChR antibodies were detected by cell-based assay in the serum of ocular MG (OMG) (n =90) and generalized MG (GMG) patients (n =110).The fetaltype (2α∶ β∶ γ∶ δ) and adult-type (2α∶ β∶ ε∶ δ) AChR were used as antigens,and their relevance to disease presentation was assessed.Results The overall frequencies of anti-adult and anti-fetal AChR antibodies were similar in all 200 patients examined,with 14 having serum specific to the AChR-γ subunit,and 22 to the AChR-ε subunit.The overall sensitivity when using the fetal and adult AChR antibodies was higher than that when using the fetal AChR antibody only (P =0.015).Compared with OMG patients,the mean age at disease onset and the positive ratio of antibodies to both isoforms of the AChR were significantly higher in patients who subsequently progressed to GMG.Older patients and patients with both anti-fetal and anti-adult AChR antibodies had a greater risk for developing generalized disease [odds ratio (OR),1.03;95% confidence interval (CI),1.01-1.06 and OR,5.09;95% CI,2.23-11.62].Conclusion Using both fetal-and adulttype AChRs as the antigens may be more sensitive than using either subtype.Patients with serum specific to both isoforms are at a greater risk of progressing to GMG.Patients with disease onset at an advanced age appear to have a higher frequency of GMG conversion.

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

    Science.gov (United States)

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

    2008-09-01

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

  20. Nicotine facilitates nicotinic acetylcholine receptor targeting to mitochondria but makes them less susceptible to selective ligands.

    Science.gov (United States)

    Uspenska, Kateryna; Lykhmus, Olena; Gergalova, Galyna; Chernyshov, Volodymyr; Arias, Hugo R; Komisarenko, Sergiy; Skok, Maryna

    2017-08-24

    Several nicotinic acetylcholine receptor (nAChR) subtypes are expressed in mitochondria to regulate the internal pathway of apoptosis in ion channel-independent manner. However, the mechanisms of nAChR activation in mitochondria and targeting to mitochondria are still unknown. Nicotine has been shown to favor nAChR pentamer assembly, folding, and maturation on the way of biosynthesis. The idea of the present work was to determine whether nicotine affects the content, glycosylation, and function of mitochondrial nAChRs. Experiments were performed in isolated liver mitochondria from mice, that either consumed or not nicotine with the drinking water (200μL/L) for 7days. Mitochondria detergent lysates were studied by sandwich or lectin ELISA for the presence and carbohydrate composition of different nAChR subunits. Intact mitochondria were examined by flow cytometry for the binding of fluorescently labeled α-cobratoxin and were tested in functional assay of cytochrome c release under the effect of either Ca 2+ or wortmannin in the presence or absence of nAChR-selective ligands, including PNU-282987 (1nM), dihydro-β-erythroidine (DhβE, 1μM), PNU-120596 (0.3, 3, or 10μM) and desformylflustrabromine hydrochloride (dFBr, 0.001, 0.3, or 1μM). It was found that nicotine consumption increased the ratio of mitochondrial vs non-mitochondrial nAChRs in the liver, enhanced fucosylation of mitochondrial nAChRs, but prevented the binding of α-cobratoxin and the cytochrome c release-attenuating effects of nAChR-specific agonists, antagonists, or positive allosteric modulators. It is concluded that nicotine consumption in vivo favors nAChR glycosylation and trafficking to mitochondria but makes them less susceptible to the effects of specific ligands. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Effect of donepezil hydrochloride (E2020) on extracellular acetylcholine concentration in the cerebral cortex of rats.

    Science.gov (United States)

    Kosasa, T; Kuriya, Y; Yamanishi, Y

    1999-10-01

    Donepezil hydrochloride (donepezil), a potent and selective acetylcholinesterase inhibitor, has been developed for the treatment of Alzheimer's disease. We studied the effect of oral administration of this drug on the extracellular acetylcholine (ACh) concentration in the cerebral cortex of rats using microdialysis. We also observed fasciculation, a peripheral cholinergic sign induced by activation of neuromuscular transmission, after oral administration of the drug as an index of peripheral cholinergic activation. Other cholinesterase inhibitors, tacrine, ENA-713 and TAK-147, were used as reference drugs. Donepezil significantly and dose-dependently increased the extracellular ACh concentration in the rat cerebral cortex within the dose range of 2.5-10 mg/kg. Tacrine, ENA-713 and TAK-147 also elevated the extracellular concentration of ACh. The minimum effective doses of donepezil, tacrine, ENA-713 and TAK-147 were (< or = 2.5, 10, 10 and < or = 10 mg/kg, respectively. Donepezil produced fasciculation at doses of 2.5 mg/kg and above, with a dose-dependent increase in incidence and intensity. The reference compounds also induced fasciculation in a dose-dependent manner. The threshold doses of tacrine, ENA-713 and TAK-147 for fasciculation were 5, 2.5 and 2.5 mg/kg, respectively. The values of the ratio of the minimum effective dose for the ACh-increasing action to that for the fasciculation-producing action were: donepezil, < or = 1; tacrine, 2; ENA-713, 4; TAK-147, < or = 4. These results indicate that orally administered donepezil has a potent and selective activity on the central cholinergic system.

  2. Nicotinic acetylcholine receptor availability in cigarette smokers: effect of heavy caffeine or marijuana use.

    Science.gov (United States)

    Brody, Arthur L; Hubert, Robert; Mamoun, Michael S; Enoki, Ryutaro; Garcia, Lizette Y; Abraham, Paul; Young, Paulina; Mandelkern, Mark A

    2016-09-01

    Upregulation of α4β2* nicotinic acetylcholine receptors (nAChRs) is one of the most well-established effects of chronic cigarette smoking on the brain. Prior research by our group gave a preliminary indication that cigarette smokers with concomitant use of caffeine or marijuana have altered nAChR availability. We sought to determine if smokers with heavy caffeine or marijuana use have different levels of α4β2* nAChRs than smokers without these drug usages. One hundred and one positron emission tomography (PET) scans, using the radiotracer 2-FA (a ligand for β2*-containing nAChRs), were obtained from four groups of males: non-smokers without heavy caffeine or marijuana use, smokers without heavy caffeine or marijuana use, smokers with heavy caffeine use (mean four coffee cups per day), and smokers with heavy marijuana use (mean 22 days of use per month). Total distribution volume (Vt/fp) was determined for the brainstem, prefrontal cortex, and thalamus, as a measure of nAChR availability. A significant between-group effect was found, resulting from the heavy caffeine and marijuana groups having the highest Vt/fp values (especially for the brainstem and prefrontal cortex), followed by smokers without such use, followed by non-smokers. Direct between-group comparisons revealed significant differences for Vt/fp values between the smoker groups with and without heavy caffeine or marijuana use. Smokers with heavy caffeine or marijuana use have higher α4β2* nAChR availability than smokers without these drug usages. These findings are likely due to increased nicotine exposure but could also be due to an interaction on a cellular/molecular level.

  3. Novel biomimetic composite material for potentiometric screening of acetylcholine, a neurotransmitter in Alzheimer's disease.

    Science.gov (United States)

    Sacramento, Ana S; Moreira, Felismina T C; Guerreiro, Joana L; Tavares, Ana P; Sales, M Goreti F

    2017-10-01

    This work describes a novel approach to produce an antibody-like biomimetic material. It includes preparing composite imprinted material never presented before, with highly conductive support nanostructures and assembling a high conductivity polymeric layer at low temperature. Overall, such highly conductive material may enhance the final features of electrically-based devices. Acetylcholine (ACh) was selected as target analyte, a neurotransmitter of importance in Alzheimer's disease. Potentiometric transduction was preferred, allowing quick responses and future adaptation to point-of-care requirements. The biomimetic material was obtained by bulk polymerization, where ACh was placed in a composite matrix of multiwalled carbon nanotubes (MWCNTs) and aniline (ANI). Subsequent polymerization, initiated by radical species, yielded a polymeric structure of polyaniline (PANI) acting as physical support of the composite. A non-imprinted material (NIM) having only PANI/MWCNT (without ACh) has been prepared for comparison of the biomimetic-imprinted material (BIM). RAMAN and Fourier Transform Infrared spectroscopy (FTIR), Transmission Electron microscopy (TEM), and Scanning Electron microscope (SEM) analysis characterized the structures of the materials. The ability of this biomaterial to rebind ACh was confirmed by including it as electroactive compound in a PVC/plasticizer mixture. The membranes with imprinted material and anionic additive presented the best analytical characteristics, with a sensitivity of 83.86mV decade -1 and limit of detection (LOD) of 3.45×10 -5 mol/L in HEPES buffer pH4.0. Good selectivity was observed against creatinine, creatine, glucose, cysteine and urea. The electrodes were also applied on synthetic serum samples and seemed a reliable tool for screening ACh in synthetic serum samples. The overall performance showed fast response, reusability, simplicity and low price. Copyright © 2017 Elsevier B.V. All rights reserved.

  4. Degradation rate of acetylcholine receptors inserted into denervated vertebrate neuromuscular junctions

    International Nuclear Information System (INIS)

    Shyng, S.L.; Salpeter, M.M.

    1989-01-01

    Many studies exist on the effect of denervation on the degradation of acetylcholine receptors (AChRs) at the vertebrate neuromuscular junction (nmj). These studies have described the behavior of either the total population of junctional receptors at different times after denervation, or of the receptors present at the time of denervation. No experimental studies yet exist on the degradation rate of the receptors newly inserted into denervated junctions. In the previous studies, the original receptors of mouse sternomastoid muscles were found to retain the slow degradation (t 1/2) of approximately 8-10 d of innervated junctional receptors for up to 10 d after denervation before accelerating to a t 1/2 of approximately 3 d. The total junctional receptors, on the other hand, showed a progressive increase in degradation rate from a t 1/2 of 8-10 d to a t 1/2 of 1 d. To reconcile these earlier observations, the present study examines the degradation of new receptors inserted into the nmj after denervation. To avoid possible contamination of the data with postdenervation extrajunctional receptors, we used transmission electron microscope autoradiography to study only receptors located at the postjunctional fold of the nmj. We established that the new receptors inserted into denervated junctions have a t 1/2 of approximately 1 d, considerably faster than that of the original receptors and equivalent to that of postdenervation extrajunctional receptors. Both original and new receptors are interspersed at the top of the junctional folds. Thus, until all the original receptors are degraded, the postjunctional membrane contains two populations of AChRs that maintain a total steady-state site density but degrade at different rates

  5. Blockade of central nicotine acetylcholine receptor signaling attenuate ghrelin-induced food intake in rodents.

    Science.gov (United States)

    Dickson, S L; Hrabovszky, E; Hansson, C; Jerlhag, E; Alvarez-Crespo, M; Skibicka, K P; Molnar, C S; Liposits, Z; Engel, J A; Egecioglu, E

    2010-12-29

    Here we sought to determine whether ghrelin's central effects on food intake can be interrupted by nicotine acetylcholine receptor (nAChR) blockade. Ghrelin regulates mesolimbic dopamine neurons projecting from the ventral tegmental area (VTA) to the nucleus accumbens, partly via cholinergic VTA afferents originating in the laterodorsal tegmental area (LDTg). Given that these cholinergic projections to the VTA have been implicated in natural as well as drug-induced reinforcement, we sought to investigate the role of cholinergic signaling in ghrelin-induced food intake as well as fasting-induced food intake, for which endogenous ghrelin has been implicated. We found that i.p. treatment with the non-selective centrally active nAChR antagonist, mecamylamine decreased fasting-induced food intake in both mice and rats. Moreover, central administration of mecamylamine decreased fasting-induced food intake in rats. I.c.v. ghrelin-induced food intake was suppressed by mecamylamine i.p. but not by hexamethonium i.p., a peripheral nAChR antagonist. Furthermore, mecamylamine i.p. blocked food intake following ghrelin injection into the VTA. Expression of the ghrelin receptor, the growth hormone secretagogue receptor 1A, was found to co-localize with choline acetyltransferase, a marker of cholinergic neurons, in the LDTg. Finally, mecamylamine treatment i.p. decreased the ability of palatable food to condition a place preference. These data suggest that ghrelin-induced food intake is partly mediated via nAChRs and that nicotinic blockade decreases the rewarding properties of food. Copyright © 2010 IBRO. Published by Elsevier Ltd. All rights reserved.

  6. The expression, localization and function of α7 nicotinic acetylcholine receptor in rat corpus cavernosum.

    Science.gov (United States)

    Faghir-Ghanesefat, Hedyeh; Rahimi, Nastaran; Yarmohammadi, Fatemeh; Mokhtari, Tahmineh; Abdollahi, Ali Reza; Ejtemaei Mehr, Shahram; Dehpour, Ahmad R

    2017-12-01

    Alpha7 nicotinic acetylcholine receptor (α7-nAChR), an emerging pharmacological target for a variety of medical conditions, is expressed in the most mammalian tissues with different effects. So, this study was designed to investigate the expression, localization and effect of α7-nAChR in rat corpus cavernosum (CC). Reverse transcription polymerase chain reaction (RT-PCR) revealed that α7-nAChR was expressed in rat CC and double immunofluorescence studies demonstrated the presence of α7-nAChR in corporal neurons. The rat CC segments were mounted in organ bath chambers and contracted with phenylephrine (0.1 μm -300 μm) to investigate the relaxation effect of electrical field stimulation (EFS,10 Hz) assessed in the presence of guanethidine (adrenergic blocker, 5 μm) and atropine (muscarinic cholinergic blocker, 1 μm) to obtain non-adrenergic non-cholinergic (NANC) response. Cumulative administration of nicotine significantly potentiated the EFS-induced NANC relaxation (-log EC50 = 7.5 ± 0.057). Whereas, the potentiated NANC relaxation of nicotine was significantly inhibited with different concentrations of methyllycaconitine citrate (α7-nAChR antagonist, P < 0.05) in preincubated strips. L-NAME (non-specific nitric oxide synthase inhibitor, 1 μm) completely blocked the neurogenic relaxation induced by EFS plus nicotine. To conclude α7-nAChR is expressed in rat CC and modulates the neurogenic relaxation response to nicotine. © 2017 Royal Pharmaceutical Society.

  7. Resistance to cycloxaprid in Laodelphax striatellus is associated with altered expression of nicotinic acetylcholine receptor subunits.

    Science.gov (United States)

    Zhang, Yueliang; Han, Yangchun; Yang, Qiong; Wang, Lihua; He, Peng; Liu, Zewen; Li, Zhong; Guo, Huifang; Fang, Jichao

    2018-04-01

    Cycloxaprid is a new oxabridged cis-configuration neonicotinoid insecticide, the resistance development potential and underlying resistance mechanism of which were investigated in the small brown planthopper, Laodelphax striatellus (Fallén), an important agricultural pest of rice. A cycloxaprid-resistant strain (YN-CPD) only achieved 10-fold higher resistance, in contrast to 106-fold higher resistance to buprofezin and 332-fold higher resistance to chlorpyrifos achieved after exposure to similar selection pressure, and the cycloxaprid selected line showed no cross-resistance to the buprofezin and chlorpyrifos-selected resistance strains. Moreover, we identified 10 nicotinic acetylcholine receptor (nAChR) subunits from the transcriptome of L. striatellus, and six segments had open reading frames (ORFs). While we did not find mutations in the nAChR genes of L. striatellus, subunits Lsα1 and Lsβ1 exhibited, respectively, 9.60-fold and 3.36-fold higher expression in the resistant strain, while Lsα8 exhibited 0.44-fold lower expression. Suppression of Lsα1 through ingestion of dsLsα1 led to an increase in susceptibility to cycloxaprid. The findings indicate that resistance to cycloxaprid develops slowly compared with resistance to other chemicals and without cross-resistance to chlorpyrifos or buprofezin; over-expressed Lsα1 is associated with low cycloxaprid resistance levels, but the importance of over-expressed Lsβ1 and reduced expression of Lsα8 could not be excluded. © 2017 Society of Chemical Industry. © 2017 Society of Chemical Industry.

  8. Selective expression of muscarinic acetylcholine receptor subtype M3 by mouse type III taste bud cells.

    Science.gov (United States)

    Mori, Yusuke; Eguchi, Kohgaku; Yoshii, Kiyonori; Ohtubo, Yoshitaka

    2016-11-01

    Each taste bud cell (TBC) type responds to a different taste. Previously, we showed that an unidentified cell type(s) functionally expresses a muscarinic acetylcholine (ACh) receptor subtype, M3, and we suggested the ACh-dependent modification of its taste responsiveness. In this study, we found that M3 is expressed by type III TBCs, which is the only cell type that possesses synaptic contacts with taste nerve fibers in taste buds. The application of ACh to the basolateral membrane of mouse fungiform TBCs in situ increased the intracellular Ca 2+ concentration in 2.4 ± 1.4 cells per taste bud (mean ± SD, n = 14). After Ca 2+ imaging, we supravitally labeled type II cells (phospholipase C β2 [PLCβ2]-immunoreactive cells) with Lucifer yellow CH (LY), a fluorescent dye and investigated the positional relationship between ACh-responding cells and LY-labeled cells. After fixation, the TBCs were immunohistostained to investigate the positional relationships between immunohistochemically classified cells and LY-labeled cells. The overlay of the two positional relationships obtained by superimposing the LY-labeled cells showed that all of the ACh-responding cells were type III cells (synaptosomal-associated protein 25 [SNAP-25]-immunoreactive cells). The ACh responses required no added Ca 2+ in the bathing solution. The addition of 1 μM U73122, a phospholipase C inhibitor, decreased the magnitude of the ACh response, whereas that of 1 μM U73343, a negative control, had no effect. These results suggest that type III cells respond to ACh and release Ca 2+ from intracellular stores. We also discuss the underlying mechanism of the Ca 2+ response and the role of M3 in type III cells.

  9. Elemental maps in human allantochorial placental vessels cells: 1. High K{sup +} and acetylcholine effects

    Energy Technology Data Exchange (ETDEWEB)

    Michelet-Habchi, C. E-mail: michelet@cenbg.in2p3.fr; Barberet, Ph.; Dutta, R.K.; Guiet-Bara, A.; Bara, M.; Moretto, Ph

    2003-09-01

    Regulation of vascular tone in the fetal extracorporeal circulation most likely depends on circulating hormones, local paracrine mechanisms and changes in membrane potential of vascular smooth muscle cells (VSMCs) and of vascular endothelial cells (VECs). The membrane potential is a function of the physiological activities of ionic channels (particularly, K{sup +} and Ca{sup 2+} channels in these cells). These channels regulate the ionic distribution into these cells. Micro-particle induced X-ray emission (PIXE) analysis was applied to determine the ionic composition of VSMC and of VEC in the placental human allantochorial vessels in a physiological survival medium (Hanks' solution) modified by the addition of acetylcholine (ACh: which opens the calcium-sensitive K{sup +} channels, K{sub Ca}) and of high concentration of K{sup +} (which blocks the voltage-sensitive K{sup +} channels, K{sub df}). In VSMC (media layer), the addition of ACh induced no modification of the Na, K, Cl, P, S, Mg and Ca concentrations and high K{sup +} medium increased significantly the Cl and K concentrations, the other ion concentrations remaining constant. In endothelium (VEC), ACh addition implicated a significant increase of Na and K concentration, and high K{sup +} medium, a significant increase in Cl and K concentration. These results indicated the importance of K{sub df}, K{sub Ca} and K{sub ATP} channels in the regulation of K{sup +} intracellular distribution in VSMC and VEC and the possible intervention of a Na-K-2Cl cotransport and corroborated the previous electrophysiological data.

  10. Pharmacological Characterisation of Nicotinic Acetylcholine Receptors Expressed in Human iPSC-Derived Neurons.

    Directory of Open Access Journals (Sweden)

    Anna Chatzidaki

    Full Text Available Neurons derived from human induced pluripotent stem cells (iPSCs represent a potentially valuable tool for the characterisation of neuronal receptors and ion channels. Previous studies on iPSC-derived neuronal cells have reported the functional characterisation of a variety of receptors and ion channels, including glutamate receptors, γ-aminobutyric acid (GABA receptors and several voltage-gated ion channels. In the present study we have examined the expression and functional properties of nicotinic acetylcholine receptors (nAChRs in human iPSC-derived neurons. Gene expression analysis indicated the presence of transcripts encoding several nAChR subunits, with highest levels detected for α3-α7, β1, β2 and β4 subunits (encoded by CHRNA3-CHRNA7, CHRNB1, CHRNB2 and CHRNB4 genes. In addition, similarly high transcript levels were detected for the truncated dupα7 subunit transcript, encoded by the partially duplicated gene CHRFAM7A, which has been associated with psychiatric disorders such as schizophrenia. The functional properties of these nAChRs have been examined by calcium fluorescence and by patch-clamp recordings. The data obtained suggest that the majority of functional nAChRs expressed in these cells have pharmacological properties typical of α7 receptors. Large responses were induced by a selective α7 agonist (compound B, in the presence of the α7-selective positive allosteric modulator (PAM PNU-120596, which were blocked by the α7-selective antagonist methyllycaconitine (MLA. In addition, a small proportion of the neurons express nAChRs with properties typical of heteromeric (non-α7 containing nAChR subtypes. These cells therefore represent a great tool to advance our understanding of the properties of native human nAChRs, α7 in particular.

  11. Orthosteric and Allosteric Ligands of Nicotinic Acetylcholine Receptors for Smoking Cessation

    Directory of Open Access Journals (Sweden)

    Tasnim S. Mohamed

    2015-11-01

    Full Text Available Nicotine addiction, the result of tobacco use, leads to over six million premature deaths world-wide, a number that is expected to increase by a third within the next two decades. While more than half of smokers want and attempt to quit, only a small percentage of smokers are able to quit without pharmacological interventions. Therefore, over the past decades, researchers in academia and the pharmaceutical industry have focused their attention on the development of more effective smoking cessation therapies, which is now a growing 1.9 billion dollar market. Because the role of neuronal nicotinic acetylcholine receptors (nAChR in nicotine addiction is well established, nAChR based therapeutics remain the leading strategy for smoking cessation. However, the development of neuronal nAChR drugs that are selective for a nAChR subpopulation is challenging, and only few neuronal nAChR drugs are clinically available. Among the many neuronal nAChR subtypes that have been identified in the brain, the α4β2 subtype is the most abundant and plays a critical role in nicotine addiction. Here, we review the role of neuronal nAChRs, especially the α4β2 subtype, in the development and treatment of nicotine addiction. We also compare available smoking cessation medications and other nAChR orthosteric and allosteric ligands that have been developed with emphasis on the difficulties faced in the development of clinically useful compounds with high nAChR subtype selectivity.

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

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    Lilia Zurkovsky

    2013-01-01

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

  13. The function of Shp2 tyrosine phosphatase in the dispersal of acetylcholine receptor clusters

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    Madhavan Raghavan

    2008-07-01

    Full Text Available Abstract Background A crucial event in the development of the vertebrate neuromuscular junction (NMJ is the postsynaptic enrichment of muscle acetylcholine (ACh receptors (AChRs. This process involves two distinct steps: the local clustering of AChRs at synapses, which depends on the activation of the muscle-specific receptor tyrosine kinase MuSK by neural agrin, and the global dispersal of aneural or "pre-patterned" AChR aggregates, which is triggered by ACh or by synaptogenic stimuli. We and others have previously shown that tyrosine phosphatases, such as the SH2 domain-containing phosphatase Shp2, regulate AChR cluster formation in muscle cells, and that tyrosine phosphatases also mediate the dispersal of pre-patterned AChR clusters by synaptogenic stimuli, although the specific phosphatases involved in this latter step remain unknown. Results Using an assay system that allows AChR cluster assembly and disassembly to be studied separately and quantitatively, we describe a previously unrecognized role of the tyrosine phosphatase Shp2 in AChR cluster disassembly. Shp2 was robustly expressed in embryonic Xenopus muscle in vivo and in cultured myotomal muscle cells, and treatment of the muscle cultures with an inhibitor of Shp2 (NSC-87877 blocked the dispersal of pre-patterned AChR clusters by synaptogenic stimuli. In contrast, over-expression in muscle cells of either wild-type or constitutively active Shp2 accelerated cluster dispersal. Significantly, forced expression in muscle of the Shp2-activator SIRPα1 (signal regulatory protein α1 also enhanced the disassembly of AChR clusters, whereas the expression of a truncated SIRPα1 mutant that suppresses Shp2 signaling inhibited cluster disassembly. Conclusion Our results suggest that Shp2 activation by synaptogenic stimuli, through signaling intermediates such as SIRPα1, promotes the dispersal of pre-patterned AChR clusters to facilitate the selective accumulation of AChRs at developing NMJs.

  14. Effects of Acetylcholine and Noradrenalin on Action Potentials of Isolated Rabbit Sinoatrial and Atrial Myocytes

    Science.gov (United States)

    Verkerk, Arie O.; Geuzebroek, Guillaume S. C.; Veldkamp, Marieke W.; Wilders, Ronald

    2012-01-01

    The autonomic nervous system controls heart rate and contractility through sympathetic and parasympathetic inputs to the cardiac tissue, with acetylcholine (ACh) and noradrenalin (NA) as the chemical transmitters. In recent years, it has become clear that specific Regulators of G protein Signaling proteins (RGS proteins) suppress muscarinic sensitivity and parasympathetic tone, identifying RGS proteins as intriguing potential therapeutic targets. In the present study, we have identified the effects of 1 μM ACh and 1 μM NA on the intrinsic action potentials of sinoatrial (SA) nodal and atrial myocytes. Single cells were enzymatically isolated from the SA node or from the left atrium of rabbit hearts. Action potentials were recorded using the amphotericin-perforated patch-clamp technique in the absence and presence of ACh, NA, or a combination of both. In SA nodal myocytes, ACh increased cycle length and decreased diastolic depolarization rate, whereas NA decreased cycle length and increased diastolic depolarization rate. Both ACh and NA increased maximum upstroke velocity. Furthermore, ACh hyperpolarized the maximum diastolic potential. In atrial myocytes stimulated at 2 Hz, both ACh and NA hyperpolarized the maximum diastolic potential, increased the action potential amplitude, and increased the maximum upstroke velocity. Action potential duration at 50 and 90% repolarization was decreased by ACh, but increased by NA. The effects of both ACh and NA on action potential duration showed a dose dependence in the range of 1–1000 nM, while a clear-cut frequency dependence in the range of 1–4 Hz was absent. Intermediate results were obtained in the combined presence of ACh and NA in both SA nodal and atrial myocytes. Our data uncover the extent to which SA nodal and atrial action potentials are intrinsically dependent on ACh, NA, or a combination of both and may thus guide further experiments with RGS proteins. PMID:22754533

  15. "Warming yang and invigorating qi" acupuncture alters acetylcholine receptor expression in the neuromuscular junction of rats with experimental autoimmune myasthenia gravis

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    Hai-peng Huang

    2016-01-01

    Full Text Available Myasthenia gravis is an autoimmune disorder in which antibodies have been shown to form against the nicotinic acetylcholine nicotinic postsynaptic receptors located at the neuromuscular junction. "Warming yang and invigorating qi" acupuncture treatment has been shown to reduce serum inflammatory cytokine expression and increase transforming growth factor beta expression in rats with experimental autoimmune myasthenia gravis. However, few studies have addressed the effects of this type of acupuncture on the acetylcholine receptors at the neuromuscular junction. Here, we used confocal laser scanning microscopy to examine the area and density of immunoreactivity for an antibody to the nicotinic acetylcholine receptor at the neuromuscular junction in the phrenic nerve of rats with experimental autoimmune myasthenia gravis following "warming yang and invigorating qi" acupuncture therapy. Needles were inserted at acupressure points Shousanli (LI10, Zusanli (ST36, Pishu (BL20, and Shenshu (BL23 once daily for 7 consecutive days. The treatment was repeated after 1 day of rest. We found that area and the integrated optical density of the immunoreactivity for the acetylcholine receptor at the neuromuscular junction of the phrenic nerve was significantly increased following acupuncture treatment. This outcome of the acupuncture therapy was similar to that of the cholinesterase inhibitor pyridostigmine bromide. These findings suggest that "warming yang and invigorating qi" acupuncture treatment increases acetylcholine receptor expression at the neuromuscular junction in a rat model of autoimmune myasthenia gravis.

  16. Aspects of dopamine and acetylcholine release induced by glutamate receptors; Aspectos das liberacoes de dopamina e acetilcolina mediadas por receptores de glutamato

    Energy Technology Data Exchange (ETDEWEB)

    Paes, Paulo Cesar de Arruda

    2002-07-01

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

  17. Detection of Ca2+-induced acetylcholine released from leukemic T-cells using an amperometric microfluidic sensor.

    Science.gov (United States)

    Akhtar, Mahmood H; Hussain, Khalil K; Gurudatt, N G; Shim, Yoon-Bo

    2017-12-15

    A microfluidic structured-dual electrodes sensor comprising of a pair of screen printed carbon electrodes was fabricated to detect acetylcholine, where one of them was used for an enzyme reaction and another for a detection electrode. The former was coated with gold nanoparticles and the latter with a porous gold layer, followed by electropolymerization of 2, 2:5,2-terthiophene-3-(p-benzoic acid) (pTTBA) on both the electrodes. Then, acetylcholinesterase was covalently attached onto the reaction electrode, and hydrazine and choline oxidase were co-immobilized on the detection electrode. The layers of both modified electrodes were characterized employing voltammetry, field emission scanning electron microscopy, X-ray photoelectron spectroscopy, and quartz crystal microscopy. After the modifications of both electrode surfaces, they were precisely faced each other to form a microfluidic channel structure, where H 2 O 2 produced from the sequential enzymatic reactions was reduced by hydrazine to obtain the analytical signal which was analyzed by the detection electrode. The microfluidic sensor at the optimized experimental conditions exhibited a wide dynamic range from 0.7nM to 1500μM with the detection limit of 0.6 ± 0.1nM based on 3s (S/N = 3). The biomedical application of the proposed sensor was evaluated by detecting acetylcholine in human plasma samples. Moreover, the Ca 2+ -induced acetylcholine released in leukemic T-cells was also investigated to show the in vitro detection ability of the designed microfluidic sensor. Interference due to the real component matrix were also studied and long term stability of the designed sensor was evaluated. The analytical performance of the designed sensor was also compared with commercially available ACh detection kit. Copyright © 2017 Elsevier B.V. All rights reserved.

  18. Prefrontal cortex, caloric restriction and stress during aging: studies on dopamine and acetylcholine release, BDNF and working memory.

    Science.gov (United States)

    Del Arco, Alberto; Segovia, Gregorio; de Blas, Marta; Garrido, Pedro; Acuña-Castroviejo, Dario; Pamplona, Reinald; Mora, Francisco

    2011-01-01

    This study was designed to investigate whether long-term caloric restriction during the life span of the rat changes the effects of an acute mild stress on the release of dopamine and acetylcholine in the prefrontal cortex (PFC) and on working memory performance. Spontaneous motor activity was also monitored and levels of BDNF measured in the prefrontal cortex, amygdala and hippocampus. Male Wistar rats (3 months of age) were housed during 3, 12, 21 and 27 months (6, 15, 24 and 30 months of age at the end of housing) in caloric restriction (CR; 40% food intake restriction) or control conditions. After behavioural testing, animals were further subdivided into two other groups. In one of the groups BDNF protein levels were determined. In the other group rats were implanted with guide cannulas into the PFC to perform microdialysis experiments. In CR rats the release of dopamine produced by handling stress did not differ from the response found in control rats of 6, 15 and 24 months of age. The release of acetylcholine was not changed at the ages of 6 and 15 months but reduced at the age of 24 months. Stress did not change dopamine or acetylcholine release in CR and control rats of 30 months of age. BDNF levels were increased in the hippocampus and amygdala, but not in the PFC, of 6 and 15 months CR rats. Spontaneous motor activity was increased in all groups of CR rats. Age, however, decreased motor activity in CR and control rats. Both experimental groups showed similar working memory performance in a delayed alternation task in basal conditions and after a situation of acute stress. These results suggest that CR does not modify the function of the PFC in response to an acute stress nor the changes found as a result of the normal process of aging. Copyright © 2010 Elsevier B.V. All rights reserved.

  19. Selective breeding of two lines of guinea pigs differing in bronchial sensitivity to acetylcholine and histamine exposure.

    Science.gov (United States)

    Mikami, H; Nishibata, R; Kawamoto, Y; Ino, T

    1991-04-01

    We developed two lines of guinea pigs, one as model animals for bronchial asthma with bronchial hypersensitivity and the other with hyposensitivity as a control. In the last four years, the bronchial hypersensitive line (BHS) and hyposensitive line (BHR), both derived from Hartley strain guinea pigs, have been selected by using bronchial reactivity to acetylcholine and to histamine as parameters. Both lines have reached the F6 generation. The following results were obtained with the two lines: 1) Sib and cous in matings, and mating of selected consanguineous individuals were adopted in breeding BHS and BHR. The breeding started with six families, each, but in the F6 generation the number of families decreased to two in each line. 2) Appearance rates of hyper- or hyposensitivity to acetylcholine and histamine increased with successive generations in both lines, which had been completely separated by the F6 generation. 3) Coefficients of inbreeding in BHS and BHR in the F6 generation ranged from 42% to 45% in the former and 42% in the latter. 4) Heritabilities (h2) of BHS and BHR for the appearance rates of sensitivity to acetylcholine were presumed to be 0.54 in the former and 0.69 in the latter. 5) No difference in the body weight of 0, 20, and 40 day-old BHS was observed in any generation. On the other hand, the body weight of 20 and 40 day-old BHR tended to decrease with successive generations. 6) Mean litter sizes of BHS and BHR in each of the generations ranged from 2.24 to 3.47 animals in the former and from 2.63 to 3.38 animals in the latter.(ABSTRACT TRUNCATED AT 250 WORDS)

  20. Study of acetylcholine and barium receptors in the rat duodeno-jejunum by means of labelled molecules

    International Nuclear Information System (INIS)

    Bataller, Georges

    1971-10-01

    The purpose of this work is the determination of the number and the localization of Acetylcholine and Barium receptors in the rat intestine. We used 'radioactive labelled' drugs to reach a high sensitiveness of detection. So we were able to point out the number of 'effective' molecules of drugs, that is to say the only ones combining with receptors. With the aid of some assumptions, we determine on the one hand the receptors localization by an assessment of the drug penetration depth before reaching their level and on the other hand the number of these receptors. (author) [fr

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

    International Nuclear Information System (INIS)

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

    1999-01-01

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

  2. Ablation of the Right Cardiac Vagus Nerve Reduces Acetylcholine Content without Changing the Inflammatory Response during Endotoxemia

    Directory of Open Access Journals (Sweden)

    Konstanze Plaschke

    2018-02-01

    Full Text Available Acetylcholine is the main transmitter of the parasympathetic vagus nerve. According to the cholinergic anti-inflammatory pathway (CAP concept, acetylcholine has been shown to be important for signal transmission within the immune system and also for a variety of other functions throughout the organism. The spleen is thought to play an important role in regulating the CAP. In contrast, the existence of a “non-neuronal cardiac cholinergic system” that influences cardiac innervation during inflammation has been hypothesized, with recent publications introducing the heart instead of the spleen as a possible interface between the immune and nervous systems. To prove this hypothesis, we investigated whether selectively disrupting vagal stimulation of the right ventricle plays an important role in rat CAP regulation during endotoxemia. We performed a selective resection of the right cardiac branch of the Nervus vagus (VGX with a corresponding sham resection in vehicle-injected and endotoxemic rats. Rats were injected with lipopolysaccharide (LPS, 1 mg/kg body weight, intravenously and observed for 4 h. Intraoperative blood gas analysis was performed, and hemodynamic parameters were assessed using a left ventricular pressure-volume catheter. Rat hearts and blood were collected, and the expression and concentration of proinflammatory cytokines using quantitative reverse transcription polymerase chain reaction and enzyme-linked immunosorbent assay were measured, respectively. Four hours after injection, LPS induced a marked deterioration in rat blood gas parameters such as pH value, potassium, base excess, glucose, and lactate. The mean arterial blood pressure and the end-diastolic volume had decreased significantly. Further, significant increases in blood cholinesterases and in proinflammatory (IL-1β, IL-6, TNF-α cytokine concentration and gene expression were obtained. Right cardiac vagus nerve resection (VGX led to a marked decrease in heart

  3. Ablation of the Right Cardiac Vagus Nerve Reduces Acetylcholine Content without Changing the Inflammatory Response during Endotoxemia.

    Science.gov (United States)

    Plaschke, Konstanze; Do, Thuc Quyen Monica; Uhle, Florian; Brenner, Thorsten; Weigand, Markus A; Kopitz, Jürgen

    2018-02-01

    Acetylcholine is the main transmitter of the parasympathetic vagus nerve. According to the cholinergic anti-inflammatory pathway (CAP) concept, acetylcholine has been shown to be important for signal transmission within the immune system and also for a variety of other functions throughout the organism. The spleen is thought to play an important role in regulating the CAP. In contrast, the existence of a "non-neuronal cardiac cholinergic system" that influences cardiac innervation during inflammation has been hypothesized, with recent publications introducing the heart instead of the spleen as a possible interface between the immune and nervous systems. To prove this hypothesis, we investigated whether selectively disrupting vagal stimulation of the right ventricle plays an important role in rat CAP regulation during endotoxemia. We performed a selective resection of the right cardiac branch of the Nervus vagus (VGX) with a corresponding sham resection in vehicle-injected and endotoxemic rats. Rats were injected with lipopolysaccharide (LPS, 1 mg/kg body weight, intravenously) and observed for 4 h. Intraoperative blood gas analysis was performed, and hemodynamic parameters were assessed using a left ventricular pressure-volume catheter. Rat hearts and blood were collected, and the expression and concentration of proinflammatory cytokines using quantitative reverse transcription polymerase chain reaction and enzyme-linked immunosorbent assay were measured, respectively. Four hours after injection, LPS induced a marked deterioration in rat blood gas parameters such as pH value, potassium, base excess, glucose, and lactate. The mean arterial blood pressure and the end-diastolic volume had decreased significantly. Further, significant increases in blood cholinesterases and in proinflammatory (IL-1β, IL-6, TNF-α) cytokine concentration and gene expression were obtained. Right cardiac vagus nerve resection (VGX) led to a marked decrease in heart acetylcholine

  4. Investigation of the presence and antinociceptive function of muscarinic acetylcholine receptors in the African naked mole-rat (Heterocephalus glaber)

    DEFF Research Database (Denmark)

    Jørgensen, Kristine B.; Krogh-Jensen, Karen; Pickering, Darryl S

    2016-01-01

    The present study investigated the cholinergic system in the African naked mole-rat (Heterocephalus glaber) with focus on the muscarinic acetylcholine receptor subtypes M1 and M4. The protein sequences for the subtypes m 1–5 of the naked mole-rat were compared to that of the house mouse (Mus...... musculus) using basic local alignment search tool (BLAST). The presence and function of M1 and M4 was investigated in vivo, using the formalin test with the muscarinic receptor agonists xanomeline and VU0152100. Spinal cord tissue from the naked mole-rat was used for receptor saturation binding studies...

  5. The main immunogenic region of acetylcholine receptors does not provoke the formation of antibodies of a predominant idiotype.

    Science.gov (United States)

    Killen, J A; Hochschwender, S M; Lindstrom, J M

    1985-08-01

    Anti-idiotype antibodies were induced in rats by immunization with rat monoclonal antibodies to the main immunogenic region of acetylcholine receptors. These anti-idiotype antibodies showed very little crossreaction with other rat monoclonal antibodies which bind to the same region of the receptor. When the rats producing these anti-idiotype antibodies were immunized with receptor, they showed no net decrease in anti-receptor antibody production. These data indicate that, although more than half of the antibodies produced by rats immunized with receptor are directed at a small region, many anti-receptor idiotypes are involved in this response and anti-idiotype therapy is not beneficial.

  6. The effect of acetylcholine, LatA and FAA on phloem assimilates translocation of Raphanus sativus L

    International Nuclear Information System (INIS)

    Yang Chongjun; Zhang Ping

    2005-01-01

    The petiole phloem of Raphanus sativus L. is treated with the medicaments of acetylcholine (Ach, the expansionist material of protoplasm), latrunculin A (LatA, the dissolubility of microfilament) and FAA (the regularization of phloem). The effects of treatments are measured by the accumulated content of dissoluble sugar and starch in the leaves, and 14 C-labelled assimilates. The regulating role of three medicaments on the translocation of assimilates in the phloem of Raphanus sativus L are investigated. The results indicate that low Ach improves assimilates translocation while LatA and FAA inhibit it in petiole phloem of Raphanus sativus L.. (authors)

  7. Myasthenic Crisis Complicated with Myxedema, Positive for Both Anti-acetylcholine Receptor and Anti-muscle-specific Tyrosine Kinase Antibodies.

    Science.gov (United States)

    Horiuchi, Kazuhiro; Nagai, Azusa; Wakita, Masahiro; Ito, Shotaro; Takamura, Kei; Houzen, Hideki

    2018-01-15

    We herein report the case of myasthenic crisis occurring in a 51-year-old man. He had experienced ptosis, increased body weight with edema, and fatigue with dyspnea. He presented at our emergency department with disturbed consciousness. He was originally diagnosed with myxedema coma, and he required artificial respiration. Because his weakness persisted and he was positive for anti-acetylcholine receptor antibodies and anti-muscle-specific tyrosine kinase antibodies, we diagnosed myasthenic crisis after various examinations. His clinical response to treatment was good and he was discharged in an ambulatory status 3 months after admission. This case demonstrates that myasthenic crisis may occur in association with myxedema.

  8. Effect of α-bungarotoxin and etorphine on acetylcholine-evoked release of endogenous and radiolabeled catecholamines from primary culture of adrenal chromaffin cells

    International Nuclear Information System (INIS)

    Kageyama, H.; Guidotti, A.

    1984-01-01

    Cell cultures of adrenal medulla have become an important research tool to study basic processes that regulate catecholamine storage, release and synthesis. Release has been studied either by labeling with [ 3 H]norepinephrine and measuring release of radioactivity or by measuring the endogenous catecholamines released with HPLC. Acetylcholine (5X10 -6 -10 -4 M) appears to release preferentially norepinephrine, although the cells store more epinephrine than norepinephrine. Etorphine and α-bungarotoxin antagonize the release of catecholamines elicited by acetylcholine. This inhibitory action appears to be greater when the measurement of endogenous catecholamines rather than radioactive norepinephrine is used to monitor the action of acetylcholine. The data suggest that the measurement of endogenous catecholamines by HPLC is preferable to the [ 3 H]NE loading and release technique, especially when analyzing the effects of low concentrations of drugs that are thought to affect nicotinic receptor function. (Auth.)

  9. Evidence that morphine and opioid peptides do not share a common pathway with adenosine in inhibiting acetylcholine release from isolated intestine.

    Science.gov (United States)

    Vizi, E S; Somogyi, G T; Magyar, K

    1981-12-01

    1 The release of acetylcholine from guinea-pig ileal isolated longitudinal muscle strip with intact Auerbach's plexus was measured by bioassay and by a radioisotope technique. 2 Normorphine (5 x 10(-7)M) and D-Met2, Pro5-enkephalinamide (D-Met, Pro-EA) reduced the release of acetylcholine. Theophylline, an adenosine antagonist, failed to prevent the inhibitory effect of normorphine or D-Met, Pro-EA. 3 Theophylline (1.7 x 10(-4)M) by itself enhanced the twitch responses to field stimulation (0.1 Hz) but did not prevent the inhibitory effect of normorphine and D-Met, Pro-EA. 4 From the results it can be concluded that morphine and opioid peptides do not share a common pathway with adenosine in inhibiting acetylcholine release from axon terminals of Auerbach's plexus.

  10. PI3K/Akt-independent NOS/HO activation accounts for the facilitatory effect of nicotine on acetylcholine renal vasodilations: modulation by ovarian hormones.

    Directory of Open Access Journals (Sweden)

    Eman Y Gohar

    Full Text Available We investigated the effect of chronic nicotine on cholinergically-mediated renal vasodilations in female rats and its modulation by the nitric oxide synthase (NOS/heme oxygenase (HO pathways. Dose-vasodilatory response curves of acetylcholine (0.01-2.43 nmol were established in isolated phenylephrine-preconstricted perfused kidneys obtained from rats treated with or without nicotine (0.5-4.0 mg/kg/day, 2 weeks. Acetylcholine vasodilations were potentiated by low nicotine doses (0.5 and 1 mg/kg/day in contrast to no effect for higher doses (2 and 4 mg/kg/day. The facilitatory effect of nicotine was acetylcholine specific because it was not observed with other vasodilators such as 5'-N-ethylcarboxamidoadenosine (NECA, adenosine receptor agonist or papaverine. Increases in NOS and HO-1 activities appear to mediate the nicotine-evoked enhancement of acetylcholine vasodilation because the latter was compromised after pharmacologic inhibition of NOS (L-NAME or HO-1 (zinc protoporphyrin, ZnPP. The renal protein expression of phosphorylated Akt was not affected by nicotine. We also show that the presence of the two ovarian hormones is necessary for the nicotine augmentation of acetylcholine vasodilations to manifest because nicotine facilitation was lost in kidneys of ovariectomized (OVX and restored after combined, but not individual, supplementation with medroxyprogesterone acetate (MPA and estrogen (E2. Together, the data suggests that chronic nicotine potentiates acetylcholine renal vasodilation in female rats via, at least partly, Akt-independent HO-1 upregulation. The facilitatory effect of nicotine is dose dependent and requires the presence of the two ovarian hormones.

  11. 11C-NS14492 as a novel PET radioligand for imaging cerebral alpha7 nicotinic acetylcholine receptors: in vivo evaluation and drug occupancy measurements

    DEFF Research Database (Denmark)

    Ettrup, Anders; Mikkelsen, Jens D; Lehel, Szabolcs

    2011-01-01

    Small-molecule α(7) nicotinic acetylcholine receptor (α(7)nAChR) agonists are currently validated for use as treatment for cognitive disturbances in schizophrenia and in Alzheimer disease. A suitable radiolabeled α(7)nAChR PET tracer would be important for in vivo quantification of α(7)nAChR bind......Small-molecule α(7) nicotinic acetylcholine receptor (α(7)nAChR) agonists are currently validated for use as treatment for cognitive disturbances in schizophrenia and in Alzheimer disease. A suitable radiolabeled α(7)nAChR PET tracer would be important for in vivo quantification of α(7)n...

  12. Dianicline, a novel α4β2 nicotinic acetylcholine receptor partial agonist, for smoking cessation: a randomized placebo-controlled clinical trial

    DEFF Research Database (Denmark)

    Tonstad, Serena; Holme, Ingar; Tønnesen, Philip

    2011-01-01

    Dianicline is a α4β2 nicotinic acetylcholine receptor partial agonist, a class of drugs that includes varenicline and cytisine. Varenicline is efficacious for smoking cessation, while cytisine has not been studied systematically. The efficacy of dianicline has not been previously tested in an ade......Dianicline is a α4β2 nicotinic acetylcholine receptor partial agonist, a class of drugs that includes varenicline and cytisine. Varenicline is efficacious for smoking cessation, while cytisine has not been studied systematically. The efficacy of dianicline has not been previously tested...

  13. 11C-NS14492 as a novel PET radioligand for imaging cerebral alpha7 nicotinic acetylcholine receptors: in vivo evaluation and drug occupancy measurements

    DEFF Research Database (Denmark)

    Ettrup, Anders; Mikkelsen, Jens D; Lehel, Szabolcs

    2011-01-01

    Small-molecule a(7) nicotinic acetylcholine receptor (a(7)nAChR) agonists are currently validated for use as treatment for cognitive disturbances in schizophrenia and in Alzheimer disease. A suitable radiolabeled a(7)nAChR PET tracer would be important for in vivo quantification of a(7)nAChR bind......Small-molecule a(7) nicotinic acetylcholine receptor (a(7)nAChR) agonists are currently validated for use as treatment for cognitive disturbances in schizophrenia and in Alzheimer disease. A suitable radiolabeled a(7)nAChR PET tracer would be important for in vivo quantification of a(7)n...

  14. Nicotine induces fibrogenic changes in human liver via nicotinic acetylcholine receptors expressed on hepatic stellate cells

    Energy Technology Data Exchange (ETDEWEB)

    Soeda, Junpei; Morgan, Maelle; McKee, Chad; Mouralidarane, Angelina; Lin, ChingI [University College London, Centre for Hepatology, Royal Free Hospital, London NW3 2PF (United Kingdom); Roskams, Tania [Department of Morphology and Molecular Pathology, University of Leuven (Belgium); Oben, Jude A., E-mail: j.oben@ucl.ac.uk [University College London, Centre for Hepatology, Royal Free Hospital, London NW3 2PF (United Kingdom); Department of Gastroenterology and Hepatology, Guy' s and St Thomas' Hospital, London SE1 7EH (United Kingdom)

    2012-01-06

    Highlights: Black-Right-Pointing-Pointer Cigarette smoke may induce liver fibrosis via nicotine receptors. Black-Right-Pointing-Pointer Nicotine induces proliferation of hepatic stellate cells (HSCs). Black-Right-Pointing-Pointer Nicotine activates hepatic fibrogenic pathways. Black-Right-Pointing-Pointer Nicotine receptor antagonists attenuate HSC proliferation. Black-Right-Pointing-Pointer Nicotinic receptor antagonists may have utility as novel anti-fibrotic agents. -- Abstract: Background and aims: Cigarette smoke (CS) may cause liver fibrosis but possible involved mechanisms are unclear. Among the many chemicals in CS is nicotine - which affects cells through nicotinic acetylcholine receptors (nAChR). We studied the effects of nicotine, and involved pathways, on human primary hepatic stellate cells (hHSCs), the principal fibrogenic cells in the liver. We then determined possible disease relevance by assaying nAChR in liver samples from human non-alcoholic steatohepatitis (NASH). Methods: hHSC were isolated from healthy human livers and nAChR expression analyzed - RT-PCR and Western blotting. Nicotine induction of hHSC proliferation, upregulation of collagen1-{alpha}2 and the pro-fibrogenic cytokine transforming growth factor beta 1 (TGF-{beta}1) was determined along with involved intracellular signaling pathways. nAChR mRNA expression was finally analyzed in whole liver biopsies obtained from patients diagnosed with non-alcoholic steatohepatitis (NASH). Results: hHSCs express muscle type ({alpha}1, {beta}1, delta and epsilon) and neuronal type ({alpha}3, {alpha}6, {alpha}7, {beta}2 and {beta}4) nAChR subunits at the mRNA level. Among these subunits, {alpha}3, {alpha}7, {beta}1 and {epsilon} were predominantly expressed as confirmed by Western blotting. Nicotine induced hHSC proliferation was attenuated by mecamylamine (p < 0.05). Additionally, collagen1-{alpha}2 and TGF-{beta}1 mRNA expression were significantly upregulated by nicotine and inhibited by

  15. Nicotine induces fibrogenic changes in human liver via nicotinic acetylcholine receptors expressed on hepatic stellate cells

    International Nuclear Information System (INIS)

    Soeda, Junpei; Morgan, Maelle; McKee, Chad; Mouralidarane, Angelina; Lin, ChingI; Roskams, Tania; Oben, Jude A.

    2012-01-01

    Highlights: ► Cigarette smoke may induce liver fibrosis via nicotine receptors. ► Nicotine induces proliferation of hepatic stellate cells (HSCs). ► Nicotine activates hepatic fibrogenic pathways. ► Nicotine receptor antagonists attenuate HSC proliferation. ► Nicotinic receptor antagonists may have utility as novel anti-fibrotic agents. -- Abstract: Background and aims: Cigarette smoke (CS) may cause liver fibrosis but possible involved mechanisms are unclear. Among the many chemicals in CS is nicotine – which affects cells through nicotinic acetylcholine receptors (nAChR). We studied the effects of nicotine, and involved pathways, on human primary hepatic stellate cells (hHSCs), the principal fibrogenic cells in the liver. We then determined possible disease relevance by assaying nAChR in liver samples from human non-alcoholic steatohepatitis (NASH). Methods: hHSC were isolated from healthy human livers and nAChR expression analyzed – RT-PCR and Western blotting. Nicotine induction of hHSC proliferation, upregulation of collagen1-α2 and the pro-fibrogenic cytokine transforming growth factor beta 1 (TGF-β1) was determined along with involved intracellular signaling pathways. nAChR mRNA expression was finally analyzed in whole liver biopsies obtained from patients diagnosed with non-alcoholic steatohepatitis (NASH). Results: hHSCs express muscle type (α1, β1, delta and epsilon) and neuronal type (α3, α6, α7, β2 and β4) nAChR subunits at the mRNA level. Among these subunits, α3, α7, β1 and ε were predominantly expressed as confirmed by Western blotting. Nicotine induced hHSC proliferation was attenuated by mecamylamine (p < 0.05). Additionally, collagen1-α2 and TGF-β1 mRNA expression were significantly upregulated by nicotine and inhibited by mecamylamine. α1 and α3-nAChR mRNA expression was significantly upregulated in NASH fibrosis compared to normal livers. Conclusion: Nicotine at levels in smokers’ blood is pro-fibrogenic, through

  16. Synthesis and evaluation of new imaging agent for central nicotinic acetylcholine receptor α7 subtype

    International Nuclear Information System (INIS)

    Ogawa, Mikako; Nishiyama, Shingo; Tsukada, Hideo; Hatano, Kentaro; Fuchigami, Takeshi; Yamaguchi, Hiroshi; Matsushima, Yoshitaka; Ito, Kengo; Magata, Yasuhiro

    2010-01-01

    Introduction: The nicotinic acetylcholine receptor (nAChR) α7 subtype (α 7 nAChR) is one of the major nAChR subtypes in the brain. We synthesized C-11 labeled α 7 nAChR ligands, (R)-2-[ 11 C]methylamino-benzoic acid 1-aza-bicyclo[2.2.2]oct-3-yl ester ([ 11 C](R)-MeQAA) and its isomer (S)-[ 11 C]MeQAA, for in vivo investigation with positron emission tomography (PET). Then, the potential of (R)- and (S)-[ 11 C]MeQAA for in vivo imaging of α 7 nAChR in the brain was evaluated in mice and monkeys. Methods: The binding affinity for α 7 nAChR was measured using rat brain. Biodistribution and in vivo receptor blocking studies were undertaken in mice. Dynamic PET scans were performed in conscious monkeys. Results: The affinity for α 7 nAChR was 41 and 182 nM for (R)- and (S)-MeQAA, respectively. The initial uptake in the mouse brain was high ([ 11 C](R)-MeQAA: 7.68 and [ 11 C](S)-MeQAA: 6.65 %dose/g at 5 min). The clearance of [ 11 C](R)-MeQAA was slow in the hippocampus (α 7 nAChR-rich region) but was rapid in the cerebellum (α 7 nAChR-poor region). On the other hand, the clearance was fast for [ 11 C](S)-MeQAA in all regions. The brain uptake of [ 11 C](R)-MeQAA was decreased by methyllycaconitine (α 7 nAChR antagonist) treatment. In monkeys, α 7 nAChRs were highly distributed in the thalamus and cortex but poorly distributed in the cerebellum. The high accumulation was observed in the cortex and thalamus for [ 11 C](R)-MeQAA, while the uptake was rather homogeneous for [ 11 C](S)-MeQAA. Conclusions: [ 11 C](R)-MeQAA was successfully synthesized and showed high uptake to the brain. However, since the in vivo selectivity for α 7 nAChR was not enough, further PET kinetic analysis or structure optimization is needed for specific visualization of brain α 7 nAChRs in vivo.

  17. The α7-nicotinic acetylcholine receptor mediates the sensitivity of gastric cancer cells to taxanes.

    Science.gov (United States)

    Tu, Chao-Chiang; Huang, Chien-Yu; Cheng, Wan-Li; Hung, Chin-Sheng; Uyanga, Batzorig; Wei, Po-Li; Chang, Yu-Jia

    2016-04-01

    Gastric cancer is difficult to cure because most patients are diagnosed at an advanced disease stage. Systemic chemotherapy remains an important therapy for gastric cancer, but both progression-free survival and disease-free survival associated with various combination regimens are limited because of refractoriness and chemoresistance. Accumulating evidence has revealed that the homomeric α7-nicotinic acetylcholine receptor (A7-nAChR) promotes human gastric cancer by driving cancer cell proliferation, migration, and metastasis. Therefore, A7-nAChR may serve as a potential therapeutic target for gastric cancer. However, the role of A7-nAChR in taxane therapy for gastric cancer was unclear. Cells were subjected to A7-nAChR knockdown (A7-nAChR KD) using short interfering RNA (siRNA). The anti-proliferative effects of taxane were assessed via 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide (MTT), terminal deoxynucleotidyl transferase-mediated nick-end labeling (TUNEL), and cell cycle distribution assays. A7-nAChR-KD cells exhibited low resistance to docetaxel and paclitaxel treatment, as measured by the MTT assay. Following paclitaxel treatment, the proportion of apoptotic cells was higher among A7-nAChR-KD cells than among scrambled control cells, as measured by cell cycle distribution and TUNEL assays. Further molecular analyses showed a reduction in the pAKT levels and a dramatic increase in the Bad levels in paclitaxel-treated A7-nAChR-KD cells but not in scrambled control cells. Following paclitaxel treatment, the level of Bax was slightly increased in both cell populations, whereas Poly (ADP-ribose) polymerase (PARP) cleavage was increased only in A7-nAChR-KD cells. These findings indicate that A7-nAChR-KD cells are more sensitive to paclitaxel treatment. We conclude that A7-nAChR may be a key biomarker for assessing the chemosensitivity of gastric cancer cells to taxane.

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

    International Nuclear Information System (INIS)

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

    2007-01-01

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

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

    Energy Technology Data Exchange (ETDEWEB)

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

    2007-11-15

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

  20. Neonicotinoid binding, toxicity and expression of nicotinic acetylcholine receptor subunits in the aphid Acyrthosiphon pisum.

    Directory of Open Access Journals (Sweden)

    Emiliane Taillebois

    Full Text Available Neonicotinoid insecticides act on nicotinic acetylcholine receptor and are particularly effective against sucking pests. They are widely used in crops protection to fight against aphids, which cause severe damage. In the present study we evaluated the susceptibility of the pea aphid Acyrthosiphon pisum to the commonly used neonicotinoid insecticides imidacloprid (IMI, thiamethoxam (TMX and clothianidin (CLT. Binding studies on aphid membrane preparations revealed the existence of high and low-affinity binding sites for [3H]-IMI (Kd of 0.16 ± 0.04 nM and 41.7 ± 5.9 nM and for the nicotinic antagonist [125I]-α-bungarotoxin (Kd of 0.008 ± 0.002 nM and 1.135 ± 0.213 nM. Competitive binding experiments demonstrated that TMX displayed a higher affinity than IMI for [125I]-α-bungarotoxin binding sites while CLT affinity was similar for both [125I]-α-bungarotoxin and [3H]-IMI binding sites. Interestingly, toxicological studies revealed that at 48 h, IMI (LC50 = 0.038 µg/ml and TMX (LC50 = 0.034 µg/ml were more toxic than CLT (LC50 = 0.118 µg/ml. The effect of TMX could be associated to its metabolite CLT as demonstrated by HPLC/MS analysis. In addition, we found that aphid larvae treated either with IMI, TMX or CLT showed a strong variation of nAChR subunit expression. Using semi-quantitative PCR experiments, we detected for all insecticides an increase of Apisumα10 and Apisumβ1 expressions levels, whereas Apisumβ2 expression decreased. Moreover, some other receptor subunits seemed to be differently regulated according to the insecticide used. Finally, we also demonstrated that nAChR subunit expression differed during pea aphid development. Altogether these results highlight species specificity that should be taken into account in pest management strategies.

  1. Neonicotinoid Binding, Toxicity and Expression of Nicotinic Acetylcholine Receptor Subunits in the Aphid Acyrthosiphon pisum

    Science.gov (United States)

    Taillebois, Emiliane; Beloula, Abdelhamid; Quinchard, Sophie; Jaubert-Possamai, Stéphanie; Daguin, Antoine; Servent, Denis; Tagu, Denis

    2014-01-01

    Neonicotinoid insecticides act on nicotinic acetylcholine receptor and are particularly effective against sucking pests. They are widely used in crops protection to fight against aphids, which cause severe damage. In the present study we evaluated the susceptibility of the pea aphid Acyrthosiphon pisum to the commonly used neonicotinoid insecticides imidacloprid (IMI), thiamethoxam (TMX) and clothianidin (CLT). Binding studies on aphid membrane preparations revealed the existence of high and low-affinity binding sites for [3H]-IMI (Kd of 0.16±0.04 nM and 41.7±5.9 nM) and for the nicotinic antagonist [125I]-α-bungarotoxin (Kd of 0.008±0.002 nM and 1.135±0.213 nM). Competitive binding experiments demonstrated that TMX displayed a higher affinity than IMI for [125I]-α-bungarotoxin binding sites while CLT affinity was similar for both [125I]-α-bungarotoxin and [3H]-IMI binding sites. Interestingly, toxicological studies revealed that at 48 h, IMI (LC50 = 0.038 µg/ml) and TMX (LC50 = 0.034 µg/ml) were more toxic than CLT (LC50 = 0.118 µg/ml). The effect of TMX could be associated to its metabolite CLT as demonstrated by HPLC/MS analysis. In addition, we found that aphid larvae treated either with IMI, TMX or CLT showed a strong variation of nAChR subunit expression. Using semi-quantitative PCR experiments, we detected for all insecticides an increase of Apisumα10 and Apisumβ1 expressions levels, whereas Apisumβ2 expression decreased. Moreover, some other receptor subunits seemed to be differently regulated according to the insecticide used. Finally, we also demonstrated that nAChR subunit expression differed during pea aphid development. Altogether these results highlight species specificity that should be taken into account in pest management strategies. PMID:24801634

  2. Honeybees Produce Millimolar Concentrations of Non-Neuronal Acetylcholine for Breeding: Possible Adverse Effects of Neonicotinoids.

    Directory of Open Access Journals (Sweden)

    Ignaz Wessler

    Full Text Available The worldwide use of neonicotinoid pesticides has caused concern on account of their involvement in the decline of bee populations, which are key pollinators in most ecosystems. Here we describe a role of non-neuronal acetylcholine (ACh for breeding of Apis mellifera carnica and a so far unknown effect of neonicotinoids on non-target insects. Royal jelly or larval food are produced by the hypopharyngeal gland of nursing bees and contain unusually high ACh concentrations (4-8 mM. ACh is extremely well conserved in royal jelly or brood food because of the acidic pH of 4.0. This condition protects ACh from degradation thus ensuring delivery of intact ACh to larvae. Raising the pH to ≥5.5 and applying cholinesterase reduced the content of ACh substantially (by 75-90% in larval food. When this manipulated brood was tested in artificial larval breeding experiments, the survival rate was higher with food supplemented by 100% with ACh (6 mM than with food not supplemented with ACh. ACh release from the hypopharyngeal gland and its content in brood food declined by 80%, when honeybee colonies were exposed for 4 weeks to high concentrations of the neonicotinoids clothianidin (100 parts per billion [ppb] or thiacloprid (8,800 ppb. Under these conditions the secretory cells of the gland were markedly damaged and brood development was severely compromised. Even field-relevant low concentrations of thiacloprid (200 ppb or clothianidin (1 and 10 ppb reduced ACh level in the brood food and showed initial adverse effects on brood development. Our findings indicate a hitherto unknown target of neonicotinoids to induce adverse effects on non-neuronal ACh which should be considered when re-assessing the environmental risks of these compounds. To our knowledge this is a new biological mechanism, and we suggest that, in addition to their well documented neurotoxic effects, neonicotinoids may contribute to honeybee colony losses consecutive to a reduction of the ACh

  3. Expression of five acetylcholine receptor subunit genes in Brugia malayi adult worms

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    Ben-Wen Li

    2015-12-01

    Full Text Available Acetylcholine receptors (AChRs are required for body movement in parasitic nematodes and are targets of “classical” anthelmintic drugs such as levamisole and pyrantel and of newer drugs such as tribendimidine and derquantel. While neurotransmission explains the effects of these drugs on nematode movement, their effects on parasite reproduction are unexplained. The levamisole AChR type (L-AChRs in Caenorhabditis elegans is comprised of five subunits: Cel-UNC-29, Cel-UNC-38, Cel-UNC-63, Cel-LEV-1 and Cel-LEV-8. The genome of the filarial parasite Brugia malayi contains nine AChRs subunits including orthologues of Cel-unc-29, Cel-unc-38, and Cel-unc-63. We performed in situ hybridization with RNA probes to localize the expression of five AChR genes (Bm1_35890-Bma-unc-29, Bm1_20330-Bma-unc-38, Bm1_38195-Bma-unc-63, Bm1_48815-Bma-acr-26 and Bm1_40515-Bma-acr-12 in B. malayi adult worms. Four of these genes had similar expression patterns with signals in body muscle, developing embryos, spermatogonia, uterine wall adjacent to stretched microfilariae, wall of Vas deferens, and lateral cord. Three L-AChR subunit genes (Bma-unc-29, Bma-unc-38 and Bma-unc-63 were expressed in body muscle, which is a known target of levamisole. Bma-acr-12 was co-expressed with these levamisole subunit genes in muscle, and this suggests that its protein product may form receptors with other alpha subunits. Bma-acr-26 was expressed in male muscle but not in female muscle. Strong expression signals of these genes in early embryos and gametes in uterus and testis suggest that AChRs may have a role in nervous system development of embryogenesis and spermatogenesis. This would be consistent with embryotoxic effects of drugs that target these receptors in filarial worms. Our data show that the expression of these receptor genes is tightly regulated with regard to localization in adult worms and developmental stage in embryos and gametes. These results may help to explain the

  4. Effect of purines on calcium-independent acetylcholine release at the mouse neuromuscular junction.

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    Veggetti, M; Muchnik, S; Losavio, A

    2008-07-17

    At the mouse neuromuscular junction, activation of adenosine A(1) and P2Y receptors inhibits acetylcholine release by an effect on voltage dependent calcium channels related to spontaneous and evoked secretion. However, an effect of purines upon the neurotransmitter-releasing machinery downstream of Ca(2+) influx cannot be ruled out. An excellent tool to study neurotransmitter exocytosis in a Ca(2+)-independent step is the hypertonic response. Intracellular recordings were performed on diaphragm fibers of CF1 mice to determine the action of the specific adenosine A(1) receptor agonist 2-chloro-N(6)-cyclopentyl-adenosine (CCPA) and the P2Y(12-13) agonist 2-methylthio-adenosine 5'-diphosphate (2-MeSADP) on the hypertonic response. Both purines significantly decreased such response (peak and area under the curve), and their effect was prevented by specific antagonists of A(1) and P2Y(12-13) receptors, 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) and N-[2-(methylthioethyl)]-2-[3,3,3-trifluoropropyl]thio-5'-adenylic acid, monoanhydride with dichloromethylenebiphosphonic acid, tetrasodium salt (AR-C69931MX), respectively. Moreover, incubation of preparations only with the antagonists induced a higher response compared with controls, suggesting that endogenous ATP/ADP and adenosine are able to modulate the hypertonic response by activating their specific receptors. To search for the intracellular pathways involved in this effect, we studied the action of CCPA and 2-MeSADP in hypertonicity in the presence of inhibitors of several pathways. We found that the effect of CPPA was prevented by the calmodulin antagonist N-(6-aminohexil)-5-chloro-1-naphthalenesulfonamide hydrochloride (W-7) while that of 2-MeSADP was occluded by the protein kinase C antagonist chelerythrine and W-7. On the other hand, the inhibitors of protein kinase A (N-(2[pbromocinnamylamino]-ethyl)-5-isoquinolinesulfonamide, H-89) and phosphoinositide-3 kinase (PI3K) (2-(4-morpholinyl)-8-phenyl-4H-1-benzopyran

  5. CB1 receptor antagonism increases hippocampal acetylcholine release: site and mechanism of action.

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    Degroot, Aldemar; Köfalvi, Attila; Wade, Mark R; Davis, Richard J; Rodrigues, Ricardo J; Rebola, Nelson; Cunha, Rodrigo A; Nomikos, George G

    2006-10-01

    Evidence indicates that blockade of cannabinoid receptors increases acetylcholine (ACh) release in brain cortical regions. Although it is assumed that this type of effect is mediated through CB1 receptor (CB1R) antagonism, several in vitro functional studies recently have suggested non-CB1R involvement. In addition, neither the precise neuroanatomical site nor the exact mechanisms underlying this effect are known. We thoroughly examined these issues using a combination of systemic and local administration of CB1R antagonists, different methods of in vivo microdialysis, CB1R knockout (KO) mice, tissue measurements of ACh, and immunochemistry. First, we showed that systemic injections of the CB1R antagonists N-(piperidin-1-yl)-5-(4-chlorophenyl)-1-(2,4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboximide hydrochloride (SR-141716A) and N-(piperidin-1-yl)-5-(4-iodophenyl)-1-(2, 4-dichlorophenyl)-4-methyl-1H-pyrazole-3-carboxamide (AM251) dose-dependently increased hippocampal ACh efflux. Likewise, local hippocampal, but not septal, infusions of SR141716A or AM251 increased hippocampal ACh release. It is noteworthy that the stimulatory effects of systemically administered CB1R antagonists on hippocampal ACh release were completely abolished in CB1R KO mice. CB1R KO mice had similar basal but higher stress-enhanced hippocampal ACh levels compared with wild-type controls. It is interesting that dopamine D1 receptor antagonism counteracted the stimulatory effect of CB1R blockade on hippocampal ACh levels. Finally, immunohistochemical methods revealed that a high proportion of CB1R-positive nerve terminals were found in hippocampus and confirmed the colocalization of CB1 receptors with cholinergic and dopaminergic nerve terminals. In conclusion, hippocampal ACh release may specifically be controlled through CB1Rs located on both cholinergic and dopaminergic neuronal projections, and CB1R antagonism increases hippocampal ACh release, probably through both a direct

  6. Acetylcholine versus cold pressor testing for evaluation of coronary endothelial function.

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    Ahmed AlBadri

    Full Text Available Assessment of coronary endothelial function with intracoronary acetylcholine (IC-Ach provides diagnostic and prognostic data in patients with suspected coronary microvascular dysfunction (CMD, but is often not feasible due in part to the time and expertise needed for pharmacologic mixing. Cold pressor testing (CPT is a simple and safe stimulus useful for either invasive or non-invasive endothelial function testing and myocardial perfusion imaging but has not been specifically evaluated among symptomatic women with signs of ischemic heart disease (IHD who have no obstructive coronary artery disease (CAD.163 women with signs and symptoms of IHD and no obstructive CAD from the NHLBI- Women's Ischemia Syndrome Evaluation-Coronary Vascular Dysfunction (WISE-CVD study underwent coronary reactivity testing with a Doppler flow wire (FloWire® Volcano, San Diego, CA in the proximal left anterior descending artery. Coronary artery diameter and coronary blood flow (CBF assessed by core lab using QCA before and after IC-Ach (18.2 μg/ml infused over 3 minutes and during CPT.Mean age was 55 ± 12 years. Rate pressure product (RPP in response to IC-Ach did not change (baseline to peak, P = 0.26, but increased during CPT (363±1457; P = 0.0028. CBF in response to CPT was poorly correlated to IC-Ach CBF. Change in coronary artery diameter after IC-Ach correlated with change after CPT (r = 0.59, P<0.001. The correlation coefficient was stronger in subjects with coronary dilation to IC-Ach (r = 0.628, P<0.001 versus those without dilation (r = 0.353, P = 0.002, suggesting that other factors may be important to this relationship when endothelium is abnormal.In women with no obstructive CAD and suspected CMD, coronary diameter changes with IC-Ach and CPT are moderately-well correlated suggesting that CPT testing may be of some use, particularly among patients with normal endothelial function, however, not an alternative to IC-Ach for diagnosis of coronary

  7. Effect of α7 nicotinic acetylcholine receptor agonists and antagonists on motor function in mice

    International Nuclear Information System (INIS)

    Welch, Kevin D.; Pfister, James A.; Lima, Flavia G.; Green, Benedict T.; Gardner, Dale R.

    2013-01-01

    Nicotinic acetylcholine receptors (nAChRs) are ligand-gated cation channels found throughout the body, and serve to mediate diverse physiological functions. Muscle-type nAChRs located in the motor endplate region of muscle fibers play an integral role in muscle contraction and thus motor function. The toxicity and teratogenicity of many plants (which results in millions of dollars in losses annually to the livestock industry) are due to various toxins that bind to nAChRs including deltaline and methyllycaconitine (MLA) from larkspur (Delphinium) species, and nicotine and anabasine from tobacco (Nicotiana) species. The primary result of the actions of these alkaloids at nAChRs is neuromuscular paralysis and respiratory failure. The objective of this study was to further characterize the motor coordination deficiencies that occur upon exposure to a non-lethal dose of nAChR antagonists MLA and deltaline as well as nAChR agonists nicotine and anabasine. We evaluated the effect of nAChR agonists and antagonists on the motor function and coordination in mice using a balance beam, grip strength meter, rotarod, open field analysis and tremor monitor. These analyses demonstrated that within seconds after treatment the mice had significant loss of motor function and coordination that lasted up to 1 min, followed by a short period of quiescence. Recovery to normal muscle coordination was rapid, typically within approximately 10 min post-dosing. However, mice treated with the nAChR agonist nicotine and anabasine required a slightly longer time to recover some aspects of normal muscle function in comparison to mice treated with the nAChR antagonist MLA or deltaline. -- Highlights: ► Mice treated with nAChR agonists and antagonists have a loss in motor function. ► These deficits are temporary as near normal motor function returns within 10 min. ► There are compound-specific differences in the effects on motor function.

  8. Clinical Utility of Acetylcholine Receptor Antibody Testing in Ocular Myasthenia Gravis.

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    Peeler, Crandall E; De Lott, Lindsey B; Nagia, Lina; Lemos, Joao; Eggenberger, Eric R; Cornblath, Wayne T

    2015-10-01

    The sensitivity of acetylcholine receptor (AChR) antibody testing is thought to be lower in ocular myasthenia gravis (OMG) compared with generalized disease, although estimates in small-scale studies vary. There is little information in the literature about the implications of AChR antibody levels and progression from OMG to generalized myasthenia gravis. To test the hypothesis that serum AChR antibody testing is more sensitive in OMG than previously reported and to examine the association between AChR antibody levels and progression from OMG to generalized myasthenia gravis. A retrospective, observational cohort study was conducted of 223 patients (mean [SD] age, 59.2 [16.4] years; 139 [62.3%] male) diagnosed with OMG between July 1, 1986, and May 31, 2013, at 2 large, academic medical centers. Baseline characteristics, OMG symptoms, results of AChR antibody testing, and progression time to generalized myasthenia gravis (if this occurred) were recorded for each patient. Multiple logistic regression was used to measure the association between all clinical variables and antibody result. Kaplan-Meier survival analysis was performed to examine time to generalization. Among the 223 participants, AChR antibody testing results were positive in 158 participants (70.9%). In an adjusted model, increased age at diagnosis (odds ratio [OR], 1.03; 95% CI, 1.01-1.04; P = .007) and progression to generalized myasthenia gravis (OR, 2.92; 95% CI, 1.18-7.26; P = .02) were significantly associated with positive antibody test results. Women were less likely to have a positive antibody test result (OR, 0.36; 95% CI, 0.19-0.68; P = .002). Patients who developed symptoms of generalized myasthenia gravis had a significantly higher mean (SD) antibody level than those who did not develop symptoms of generalized myasthenia gravis (12.7 [16.5] nmol/L vs 4.2 [7.9] nmol/L; P = .002). We demonstrate a higher sensitivity of AChR antibody testing than previously reported in the

  9. Extracellular matrix organization in developing muscle: correlation with acetylcholine receptor aggregates.

    Science.gov (United States)

    Bayne, E K; Anderson, M J; Fambrough, D M

    1984-10-01

    Monoclonal antibodies recognizing laminin, heparan sulfate proteoglycan, fibronectin, and two apparently novel connective tissue components have been used to examine the organization of extracellular matrix of skeletal muscle in vivo and in vitro. Four of the five monoclonal antibodies are described for the first time here. Immunocytochemical experiments with frozen-sectioned muscle demonstrated that both the heparan sulfate proteoglycan and laminin exhibited staining patterns identical to that expected for components of the basal lamina. In contrast, the remaining matrix constituents were detected in all regions of muscle connective tissue: the endomysium, perimysium, and epimysium. Embryonic muscle cells developing in culture elaborated an extracellular matrix, each antigen exhibiting a unique distribution. Of particular interest was the organization of extracellular matrix on myotubes: the build-up of matrix components was most apparent in plaques overlying clusters of an integral membrane protein, the acetylcholine receptor (AChR). The heparan sulfate proteoglycan was concentrated at virtually all AChR clusters and showed a remarkable level of congruence with receptor organization; laminin was detected at 70-95% of AChR clusters but often was not completely co-distributed with AChR within the cluster; fibronectin and the two other extracellular matrix antigens occurred at approximately 20, 8, and 2% of the AChR clusters, respectively, and showed little or no congruence with AChR. From observations on the distribution of extracellular matrix components in tissue cultured fibroblasts and myogenic cells, several ideas about the organization of extracellular matrix are suggested. (a) Congruence between AChR clusters and heparan sulfate proteoglycan suggests the existence of some linkage between the two molecules, possibly important for regulation of AChR distribution within the muscle membrane. (b) The qualitatively different patterns of extracellular matrix

  10. Distinct muscarinic acetylcholine receptor subtypes mediate pre- and postsynaptic effects in rat neocortex

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    Gigout Sylvain

    2012-04-01

    Full Text Available Abstract Background Cholinergic transmission has been implicated in learning, memory and cognition. However, the cellular effects induced by muscarinic acetylcholine receptors (mAChRs activation are poorly understood in the neocortex. We investigated the effects of the cholinergic agonist carbachol (CCh and various agonists and antagonists on neuronal activity in rat neocortical slices using intracellular (sharp microelectrode and field potential recordings. Results CCh increased neuronal firing but reduced synaptic transmission. The increase of neuronal firing was antagonized by pirenzepine (M1/M4 mAChRs antagonist but not by AF-DX 116 (M2/M4 mAChRs antagonist. Pirenzepine reversed the depressant effect of CCh on excitatory postsynaptic potential (EPSP but had marginal effects when applied before CCh. AF-DX 116 antagonized the depression of EPSP when applied before or during CCh. CCh also decreased the paired-pulse inhibition of field potentials and the inhibitory conductances mediated by GABAA and GABAB receptors. The depression of paired-pulse inhibition was antagonized or prevented by AF-DX 116 or atropine but only marginally by pirenzepine. The inhibitory conductances were unaltered by xanomeline (M1/M4 mAChRs agonist, yet the CCh-induced depression was antagonized by AF-DX 116. Linopirdine, a selective M-current blocker, mimicked the effect of CCh on neuronal firing. However, linopirdine had no effect on the amplitude of EPSP or on the paired-pulse inhibition, indicating that M-current is involved in the increase of neuronal excitability but neither in the depression of EPSP nor paired-pulse inhibition. Conclusions These data indicate that the three effects are mediated by different mAChRs, the increase in firing being mediated by M1 mAChR, decrease of inhibition by M2 mAChR and depression of excitatory transmission by M4 mAChR. The depression of EPSP and increase of neuronal firing might enhance the signal-to-noise ratio, whereas the

  11. Mice Lacking the Alpha9 Subunit of the Nicotinic Acetylcholine Receptor Exhibit Deficits in Frequency Difference Limens and Sound Localization

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    Amanda Clause

    2017-06-01

    Full Text Available Sound processing in the cochlea is modulated by cholinergic efferent axons arising from medial olivocochlear neurons in the brainstem. These axons contact outer hair cells in the mature cochlea and inner hair cells during development and activate nicotinic acetylcholine receptors composed of α9 and α10 subunits. The α9 subunit is necessary for mediating the effects of acetylcholine on hair cells as genetic deletion of the α9 subunit results in functional cholinergic de-efferentation of the cochlea. Cholinergic modulation of spontaneous cochlear activity before hearing onset is important for the maturation of central auditory circuits. In α9KO mice, the developmental refinement of inhibitory afferents to the lateral superior olive is disturbed, resulting in decreased tonotopic organization of this sound localization nucleus. In this study, we used behavioral tests to investigate whether the circuit anomalies in α9KO mice correlate with sound localization or sound frequency processing. Using a conditioned lick suppression task to measure sound localization, we found that three out of four α9KO mice showed impaired minimum audible angles. Using a prepulse inhibition of the acoustic startle response paradigm, we found that the ability of α9KO mice to detect sound frequency changes was impaired, whereas their ability to detect sound intensity changes was not. These results demonstrate that cholinergic, nicotinic α9 subunit mediated transmission in the developing cochlear plays an important role in the maturation of hearing.

  12. Gamma irradiation induces acetylcholine-evoked, endothelium-independent relaxation and activatesk-channels of isolated pulmonary artery of rats

    International Nuclear Information System (INIS)

    Eder, Veronique; Gautier, Mathieu; Boissiere, Julien; Girardin, Catherine; Rebocho, Manuel; Bonnet, Pierre

    2004-01-01

    Purpose: To test the effects of irradiation (R*) on the pulmonary artery (PA). Methods and materials: Isolated PA rings were submitted to gamma irradiation (cesium, 8 Gy/min -1 ) at doses of 20 Gy-140 Gy. Rings were placed in an organ chamber, contracted with serotonin (10 -4 M 5-hydroxytryptamine [5-HT]), then exposed to acetylcholine (ACh) in incremental concentrations. Smooth muscle cell (SMC) membrane potential was measured with microelectrodes. Results: A high dose of irradiation (60 Gy) increased 5HT contraction by 20%, whereas lower (20 Gy) doses slightly decreased it compared with control. In the absence of the endothelium, 5-HT precontracted rings exposed to 20 Gy irradiation developed a dose-dependent relaxation induced by acetylcholine (EI-ACh) with maximal relaxation of 60 ± 17% (n = 13). This was totally blocked by L-NAME (10 -4 M), partly by 7-nitro indazole; it was abolished by hypoxia and iberiotoxin, decreased by tetra-ethyl-ammonium, and not affected by free radical scavengers. In irradiated rings, hypoxia induced a slight contraction which was never observed in control rings. No differences in SMC membrane potential were observed between irradiated and nonirradiated PA rings. Conclusion: Irradiation mediates endothelium independent relaxation by a mechanism involving the nitric oxide pathway and K-channels

  13. Activation and modulation of human α4β2 nicotinic acetylcholine receptors by the neonicotinoids clothianidin and imidacloprid.

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    Li, Ping; Ann, Jason; Akk, Gustav

    2011-08-01

    Neonicotinoids are synthetic, nicotine-derived insecticides used for agricultural and household pest control. Though highly effective at activating insect nicotinic receptors, many neonicotinoids are also capable of directly activating and/or modulating the activation of vertebrate nicotinic receptors. In this study, we have investigated the actions of the neonicotinoids clothianidin (CTD) and imidacloprid (IMI) on human neuronal α4β2 nicotinic acetylcholine receptors. The data demonstrate that the compounds are weak agonists of the human receptors with relative peak currents of 1-4% of the response to 1 mM acetylcholine (ACh). Coapplication of IMI strongly inhibited currents elicited by ACh. From Schild plot analysis, we estimate that the affinity of IMI for the human α4β2 receptor is 18 μM. The application of low concentrations of CTD potentiated responses to low concentrations of ACh, suggesting that receptors occupied by one ACh and one CTD molecule have a higher gating efficacy than receptors with one ACh bound. Interestingly, subunit stoichiometry affected inhibition by CTD, with (α4)(2) (β2)(3) receptors significantly more strongly inhibited than the (α4)(3) (β2)(2) receptors. Copyright © 2011 Wiley-Liss, Inc.

  14. Fabrication of a Highly-sensitive Acetylcholine Sensor Based on AChOx Immobilized Smart-chips

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    M. M. RAHMAN

    2011-03-01

    Full Text Available Acetylcholine (ACh sensor based on acetylcholine oxidase (AChOx on EDC activated thioglycolic acid self-assembled monolayer (TGA-SAM using smart-chip has been developed. The simple cyclic voltammetry (CV, at 0.1 V/s technique is performed in total investigation, where 0.5M K3Fe(CN6 is utilized as a standard mediator in phosphate buffer solution (PBS, 0.1M. The ACh sensor exhibited a lower detection limit (DL, 0.1392 ± 0.005 nM, a wide linear dynamic range (LDR, 1.0 nM to 1.0 mM, good linearity (R=0.9951, and higher sensitivity (7.3543 ± 0.2 μAμM-1cm-2, and required small sample volume (70.0 μL as well as good stability and reproducibility. The smart-chip system employed a simple and efficient approach to the immobilization of enzymes onto active sensitive surface, which can enhance sensor performances to a large group of bio-molecules for wide range of biomedical applications in health care fields.

  15. Prediction of consensus binding mode geometries for related chemical series of positive allosteric modulators of adenosine and muscarinic acetylcholine receptors.

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    Sakkal, Leon A; Rajkowski, Kyle Z; Armen, Roger S

    2017-06-05

    Following insights from recent crystal structures of the muscarinic acetylcholine receptor, binding modes of Positive Allosteric Modulators (PAMs) were predicted under the assumption that PAMs should bind to the extracellular surface of the active state. A series of well-characterized PAMs for adenosine (A 1 R, A 2A R, A 3 R) and muscarinic acetylcholine (M 1 R, M 5 R) receptors were modeled using both rigid and flexible receptor CHARMM-based molecular docking. Studies of adenosine receptors investigated the molecular basis of the probe-dependence of PAM activity by modeling in complex with specific agonist radioligands. Consensus binding modes map common pharmacophore features of several chemical series to specific binding interactions. These models provide a rationalization of how PAM binding slows agonist radioligand dissociation kinetics. M 1 R PAMs were predicted to bind in the analogous M 2 R PAM LY2119620 binding site. The M 5 R NAM (ML-375) was predicted to bind in the PAM (ML-380) binding site with a unique induced-fit receptor conformation. © 2017 Wiley Periodicals, Inc. © 2017 Wiley Periodicals, Inc.

  16. In-vivo measurements of regional acetylcholine esterase activity in degenerative dementia: comparison with blood flow and glucose metabolism.

    Science.gov (United States)

    Herholz, K; Bauer, B; Wienhard, K; Kracht, L; Mielke, R; Lenz, M O; Strotmann, T; Heiss, W D

    2000-01-01

    Memory and attention are cognitive functions that depend heavily on the cholinergic system. Local activity of acetylcholine esterase (AChE) is an indicator of its integrity. Using a recently developed tracer for positron emission tomography (PET), C-11-labeled N-methyl-4-piperidyl-acetate (C11-MP4A), we measured regional AChE activity in 4 non-demented subjects, 4 patients with dementia of Alzheimer type (DAT) and 1 patient with senile dementia of Lewy body type (SDLT), and compared the findings with measurements of blood flow (CBF) and glucose metabolism (CMRGlc). Initial tracer extraction was closely related to CBF. AChE activity was reduced significantly in all brain regions in demented subjects, whereas reduction of CMRGlc and CBF was more limited to temporo-parietal association areas. AChE activity in SDLT was in the lower range of values in DAT. Our results indicate that, compared to non-demented controls, there is a global reduction of cortical AChE activity in dementia. Dementia, cholinergic system, acetylcholine esterase, positron emission tomography, cerebral blood flow, cerebral glucose metabolism.

  17. Isotopic rubidium ion efflux assay for the functional characterization of nicotinic acetylcholine receptors on clonal cell lines

    International Nuclear Information System (INIS)

    Lukas, R.J.; Cullen, M.J.

    1988-01-01

    An isotopic rubidium ion efflux assay has been developed for the functional characterization of nicotinic acetylcholine receptors on cultured neurons. This assay first involves the intracellular sequestration of isotopic potassium ion analog by the ouabain-sensitive action of a sodium-potassium ATPase. Subsequently, the release of isotopic rubidium ion through nicotinic acetylcholine receptor-coupled monovalent cation channels is activated by application of nicotinic agonists. Specificity of receptor-mediated efflux is demonstrated by its sensitivity to blockade by nicotinic, but not muscarinic, antagonists. The time course of agonist-mediated efflux, within the temporal limitations of the assay, indicates a slow inactivation of receptor function on prolonged exposure to agonist. Dose-response profiles (i) have characteristic shapes for different nicotinic agonists, (ii) are described by three operationally defined parameters, and (iii) reflect different affinities of agonists for binding sites that control receptor activation and functional inhibition. The rubidium ion efflux assay provides fewer hazards but greater sensitivity and resolution than isotopic sodium or rubidium ion influx assays for functional nicotinic receptors

  18. Thymus cells in myasthenia gravis selectively enhance production of anti-acetylcholine-receptor antibody by autologous blood lymphocytes

    International Nuclear Information System (INIS)

    Newsom-Davis, J.; Willcox, N.; Calder, L.

    1981-01-01

    We investigated the role of the thymus in 16 patients with myasthenia gravis without thymoma by studying the production of anti-acetylcholine-receptor antibody by thymic and blood lymphocytes cultured alone or together. In 10 responders (with the highest receptor-antibody titers in their plasma), cultured thymic cells spontaneously produced measurable receptor antibody. Receptor-antibody production by autologous blood lymphocytes was enhanced by the addition of responder's thymic cells, irradiated to abrogate antibody production and suppression (P<0.01). This enhancement was greater and more consistent than that by pokeweed mitogen; it depended on viable thymic cells, appeared to be selective for receptor antibody, and correlated with the ratio of thymic helper (OKT4-positive or OKT4+) to suppressor (OKT8+) T cells (P<0.01). These results suggest that myasthenic thymus contains cell-bound acetylcholine-receptor-like material or specific T cells (or both) that can aid receptor-antibody production. This may be relevant to the benefits of thymectomy in myasthenia and to the breakdown in self-tolerance in this and other autoimmune diseases

  19. Thymus cells in myasthenia gravis selectively enhance production of anti-acetylcholine-receptor antibody by autologous blood lymphocytes

    Energy Technology Data Exchange (ETDEWEB)

    Newsom-Davis, J.; Willcox, N.; Calder, L.

    1981-11-26

    We investigated the role of the thymus in 16 patients with myasthenia gravis without thymoma by studying the production of anti-acetylcholine-receptor antibody by thymic and blood lymphocytes cultured alone or together. In 10 responders (with the highest receptor-antibody titers in their plasma), cultured thymic cells spontaneously produced measurable receptor antibody. Receptor-antibody production by autologous blood lymphocytes was enhanced by the addition of responder's thymic cells, irradiated to abrogate antibody production and suppression (P<0.01). This enhancement was greater and more consistent than that by pokeweed mitogen; it depended on viable thymic cells, appeared to be selective for receptor antibody, and correlated with the ratio of thymic helper (OKT4-positive or OKT4+) to suppressor (OKT8+) T cells (P<0.01). These results suggest that myasthenic thymus contains cell-bound acetylcholine-receptor-like material or specific T cells (or both) that can aid receptor-antibody production. This may be relevant to the benefits of thymectomy in myasthenia and to the breakdown in self-tolerance in this and other autoimmune diseases.

  20. Identification, expression and functional characterization of M4L, a muscarinic acetylcholine M4 receptor splice variant.

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    Douglas A Schober

    Full Text Available Rodent genomic alignment sequences support a 2-exon model for muscarinic M4 receptor. Using this model a novel N-terminal extension was discovered in the human muscarinic acetylcholine M4 receptor. An open reading frame was discovered in the human, mouse and rat with a common ATG (methionine start codon that extended the N-terminus of the muscarinic acetylcholine M4 receptor subtype by 155 amino acids resulting in a longer variant. Transcriptional evidence for this splice variant was confirmed by RNA-Seq and RT-PCR experiments performed from human donor brain prefrontal cortices. We detected a human upstream exon indicating the translation of the mature longer M4 receptor transcript. The predicted size for the longer two-exon M4 receptor splice variant with the additional 155 amino acid N-terminal extension, designated M4L is 69.7 kDa compared to the 53 kDa canonical single exon M4 receptor (M4S. Western blot analysis from a mammalian overexpression system, and saturation radioligand binding with [3H]-NMS (N-methyl-scopolamine demonstrated the expression of this new splice variant. Comparative pharmacological characterization between the M4L and M4S receptors revealed that both the orthosteric and allosteric binding sites for both receptors were very similar despite the addition of an N-terminal extension.

  1. Identification, expression and functional characterization of M4L, a muscarinic acetylcholine M4 receptor splice variant.

    Science.gov (United States)

    Schober, Douglas A; Croy, Carrie H; Ruble, Cara L; Tao, Ran; Felder, Christian C

    2017-01-01

    Rodent genomic alignment sequences support a 2-exon model for muscarinic M4 receptor. Using this model a novel N-terminal extension was discovered in the human muscarinic acetylcholine M4 receptor. An open reading frame was discovered in the human, mouse and rat with a common ATG (methionine start codon) that extended the N-terminus of the muscarinic acetylcholine M4 receptor subtype by 155 amino acids resulting in a longer variant. Transcriptional evidence for this splice variant was confirmed by RNA-Seq and RT-PCR experiments performed from human donor brain prefrontal cortices. We detected a human upstream exon indicating the translation of the mature longer M4 receptor transcript. The predicted size for the longer two-exon M4 receptor splice variant with the additional 155 amino acid N-terminal extension, designated M4L is 69.7 kDa compared to the 53 kDa canonical single exon M4 receptor (M4S). Western blot analysis from a mammalian overexpression system, and saturation radioligand binding with [3H]-NMS (N-methyl-scopolamine) demonstrated the expression of this new splice variant. Comparative pharmacological characterization between the M4L and M4S receptors revealed that both the orthosteric and allosteric binding sites for both receptors were very similar despite the addition of an N-terminal extension.

  2. IN VITRO ANTIHELMINTIC ACTIVITY OF THREE PLANT SPECIES TRADITIONALLY USED IN TABASCO, MEXICO.

    Directory of Open Access Journals (Sweden)

    Judith Espinosa-Moreno

    2016-03-01

    Full Text Available Antihelmintic activity against Haemonchus contortus was searched as a new alternative for animal health improvement using three plants (Cydista aequinoctialis L., Heliotropium indicum L. and Momordica charantia L. of traditional usage in the rural area of Tabasco State, Mexico. Aqueous extracts were obtained of dried material. Each extract was diluted in distilled water to obtain a concentration of 20 mg ml-1. The antihelmintic activity was evaluated at 24, 48 y 72 h of exposition with 100 μL of each extract and 50 L3 unsheathed larvaes of H. contortus in a 96 well ELISA plaque.;/span;

  3. Prevalence of Ovine Haemonchosis in Wukro, Ethiopia

    Directory of Open Access Journals (Sweden)

    Lidya Gebresilassie

    2015-01-01

    Full Text Available Background. Haemonchosis caused by Haemonchus contortus is a predominant, highly pathogenic, and economically important disease of sheep and goats. Objective. Assessing the prevalence of Haemonchus parasite and its associated risk factors in sheep slaughtered at different restaurants of Wukro. Methods. Cross-sectional study using random sampling from November 2013 to April 2014 in a total of 384 sheep was conducted and SPSS version 20 software using descriptive statistics was used for data analysis and P0.05. Conclusion. The current finding revealed that significant numbers of sheep were affected by the parasites. Hence strategic deworming with good husbandry practice should be implemented.

  4. Design, synthesis, and pharmacological characterization of novel spirocyclic quinuclidinyl-Delta2 -isoxazoline derivatives as potent and selective agonists of alpha7 nicotinic acetylcholine receptors

    DEFF Research Database (Denmark)

    Dallanoce, Clelia; Magrone, Pietro; Matera, Carlo

    2011-01-01

    A set of racemic spirocyclic quinuclidinyl-¿(2) -isoxazoline derivatives was synthesized using a 1,3-dipolar cycloaddition-based approach. Target compounds were assayed for binding affinity toward rat neuronal homomeric (a7) and heteromeric (a4ß2) nicotinic acetylcholine receptors. ¿(2) -Isoxazol...

  5. The α7 nicotinic acetylcholine receptor ligands methyllycaconitine, NS6740 and GTS-21 reduce lipopolysaccharide-induced TNF-α release from microglia

    DEFF Research Database (Denmark)

    Thomsen, Morten Skøtt; Mikkelsen, Jens D

    2012-01-01

    The anti-inflammatory properties of, particularly the α7, nicotinic acetylcholine receptors (nAChRs) in the peripheral immune system are well documented. There are also reports of anti-inflammatory actions of nicotine in the CNS, but it is unclear, whether this is due to activation or inhibition...

  6. Positive allosteric modulators of the α7 nicotinic acetylcholine receptor potentiate glutamate release in the prefrontal cortex of freely-moving rats

    DEFF Research Database (Denmark)

    Bortz, D M; Upton, B A; Mikkelsen, J D

    2016-01-01

    Positive allosteric modulators (PAMs) of α7 nicotinic acetylcholine receptors (α7nAChRs) exhibit pro-cognitive effects in animal models of schizophrenia and are targets for the discovery of cognition-enhancing drugs. However, little is known about their in vivo mechanism of action because...

  7. Dual Nicotinic Acetylcholine Receptor α4β2 Antagonists/α7 Agonists: Synthesis, Docking Studies, and Pharmacological Evaluation of Tetrahydroisoquinolines and Tetrahydroisoquinolinium Salts

    DEFF Research Database (Denmark)

    Crestey, François; Jensen, Anders A; Soerensen, Christian

    2018-01-01

    We describe the synthesis of tetrahydroisoquinolines and tetrahydroisoquinolinium salts together with their pharmacological properties at various nicotinic acetylcholine receptors. In general, the compounds were α4β2 nAChR antagonists, with the tetrahydroisoquinolinium salts being more potent than...

  8. Structural differences in the two agonist binding sites of the Torpedo nicotinic acetylcholine receptor revealed by time-resolved fluorescence spectroscopy

    DEFF Research Database (Denmark)

    Martinez, K. L.; Corringer, P. J.; Edelstein, S. J.

    2000-01-01

    The nicotinic acetylcholine receptor (nAChR) from Torpedo marmorata carries two nonequivalent agonist binding sites at the αδ and αγ subunit interfaces. These sites have been characterized by time-resolved fluorescence with the partial nicotinic agonist dansyl-C6-choline (Dnscho). When bound...

  9. A combined α7 nicotinic acetylcholine receptor agonist and monoamine reuptake inhibitor, NS9775, represents a novel profile with potential benefits in emotional and cognitive disturbances

    DEFF Research Database (Denmark)

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

    2013-01-01

    also improve by nicotinic acetylcholine receptor (nAChR) activation. Preclinical studies have corroborated this and also demonstrated a synergistic antidepressant-like action when nAChR agonists and MRIs are combined. Here, we present the in vitro and in vivo profile of NS9775, a combined full α7 n...

  10. The nicotinic alpha7 acetylcholine receptor agonist ssr180711 is unable to activate limbic neurons in mice overexpressing human amyloid-beta1-42

    DEFF Research Database (Denmark)

    Soderman, A.; Spang-Thomsen, Mogens; Hansen, H.

    2008-01-01

    Recent studies have demonstrated that amyloid-beta1-42 (Abeta1-42) binds to the nicotinergic alpha7 acetylcholine receptor (alpha7 nAChR) and that the application of Abeta1-42 to cells inhibits the function of the alpha7 nAChR. The in vivo consequences of the pharmacological activation of the alp...

  11. Repeated potentiation of the metabotropic glutamate receptor 5 and the alpha 7 nicotinic acetylcholine receptor modulates behavioural and GABAergic deficits induced by early postnatal phencyclidine (PCP) treatment

    DEFF Research Database (Denmark)

    Kjaerby, Celia; Bundgaard, Christoffer; Fejgin, Kim

    2013-01-01

    GluR5), ADX47273, and the partial agonist of the α7 nicotinic acetylcholine receptor (α7 nAChR), SSR180711. Adolescent rats (4-5 weeks) subjected to PCP treatment during the second postnatal week displayed a consistent deficit in prepulse inhibition (PPI), which was reversed by a one-week treatment...

  12. Solid-phase synthesis and pharmacological evaluation of analogues of PhTX-12-A potent and selective nicotinic acetylcholine receptor antagonist

    DEFF Research Database (Denmark)

    Strømgaard, Kristian; Mellor, Ian R; Andersen, Kim

    2002-01-01

    Philanthotoxin-12 (PhTX-12) is a novel potent and selective, noncompetitive antagonist of nicotinic acetylcholine receptors (nAChRs). Homologues of PhTX-12 with 7-11 methylene groups between the primary amino group and the aromatic head-group were synthesized using solid-phase methodology. In vitro...

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

    DEFF Research Database (Denmark)

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

    2012-01-01

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

  14. Role of the cholinergic nervous system in rheumatoid arthritis: aggravation of arthritis in nicotinic acetylcholine receptor alpha7 subunit gene knockout mice

    NARCIS (Netherlands)

    van Maanen, Marjolein A.; Stoof, Susanne P.; Larosa, Gregory J.; Vervoordeldonk, Margriet J.; Tak, Paul P.

    2010-01-01

    BACKGROUND: The alpha7 subunit of nicotinic acetylcholine receptors (alpha7nAChR) can negatively regulate the synthesis and release of proinflammatory cytokines by macrophages and fibroblast-like synoviocytes in vitro. In addition, stimulation of the alpha7nAChR can reduce the severity of arthritis

  15. Early postdenervation depolarization develops faster at endplates of hibernating golden hamsters where spontaneous quantal and non-quantal acetylcholine release is very small

    Czech Academy of Sciences Publication Activity Database

    Moravec, J.; Vyskočil, František

    2005-01-01

    Roč. 51, č. 1 (2005), s. 25-29 ISSN 0168-0102 R&D Projects: GA AV ČR IAA5011411; GA ČR GA305/02/1333 Institutional research plan: CEZ:AV0Z5011922 Keywords : motor nerve ending * non-quantal * acetylcholine Subject RIV: ED - Physiology Impact factor: 2.184, year: 2005

  16. PASSIVE-AVOIDANCE TRAINING INDUCES ENHANCED LEVELS OF IMMUNOREACTIVITY FOR MUSCARINIC ACETYLCHOLINE-RECEPTOR AND COEXPRESSED PKC-GAMMA AND MAP-2 IN RAT CORTICAL-NEURONS

    NARCIS (Netherlands)

    VANDERZEE, EA; DOUMA, BRK; BOHUS, B; LUITEN, PGM

    1994-01-01

    Changes in neocortical immunoreactivity (ir) for muscarinic acetylcholine receptors (mAChRs), protein kinase C gamma (PKC gamma), microtubule-associated protein 2 (MAP-2), and the calcium-binding protein parvalbumin (PARV) induced by the performance of a one-trial passive shock avoidance (PSA) task

  17. Alpha-conotoxin analogs with additional positive charge show increased selectivity towards Torpedo californica and some neuronal subtypes of nicotinic acetylcholine receptors

    NARCIS (Netherlands)

    Kasheverov, I.E.; Zhmak, M.N.; Vulfius, C.A.; Corbacheva, E.V.; Mordvintsev, D.Y.; Utkin, Y.N.; van Elk, R.; Smit, A.B.; Tsetlin, V.I.

    2006-01-01

    α-Conotoxins from Conus snails are indispensable tools for distinguishing various subtypes of nicotinic acetylcholine receptors (nAChRs), and synthesis of α-conotoxin analogs may yield novel antagonists of higher potency and selectivity. We incorporated additional positive charges into α-conotoxins

  18. Presynaptic inhibition of spontaneous acetylcholine release induced by adenosine at the mouse neuromuscular junction.

    Science.gov (United States)

    De Lorenzo, Silvana; Veggetti, Mariela; Muchnik, Salomón; Losavio, Adriana

    2004-05-01

    1. At the mouse neuromuscular junction, adenosine (AD) and the A(1) agonist 2-chloro-N(6)-cyclopentyl-adenosine (CCPA) induce presynaptic inhibition of spontaneous acetylcholine (ACh) release by activation of A(1) AD receptors through a mechanism that is still unknown. To evaluate whether the inhibition is mediated by modulation of the voltage-dependent calcium channels (VDCCs) associated with tonic secretion (L- and N-type VDCCs), we measured the miniature end-plate potential (mepp) frequency in mouse diaphragm muscles. 2. Blockade of VDCCs by Cd(2+) prevented the effect of the CCPA. Nitrendipine (an L-type VDCC antagonist) but not omega-conotoxin GVIA (an N-type VDCC antagonist) blocked the action of CCPA, suggesting that the decrease in spontaneous mepp frequency by CCPA is associated with an action on L-type VDCCs only. 3. As A(1) receptors are coupled to a G(i/o) protein, we investigated whether the inhibition of PKA or the activation of PKC is involved in the presynaptic inhibition mechanism. Neither N-(2[p-bromocinnamylamino]-ethyl)-5-isoquinolinesulfonamide (H-89, a PKA inhibitor), nor 1-(5-isoquinolinesulfonyl)-2-methyl-piperazine (H-7, a PKC antagonist), nor phorbol 12-myristate 13-acetate (PHA, a PKC activator) modified CCPA-induced presynaptic inhibition, suggesting that these second messenger pathways are not involved. 4. The effect of CCPA was eliminated by the calmodulin antagonist N-(6-aminohexil)-5-chloro-1-naphthalenesulfonamide hydrochloride (W-7) and by ethylene glycol-bis(beta-aminoethyl ether)-N,N,N',N'-tetraacetic acid-acetoxymethyl ester epsilon6TDelta-BM, which suggests that the action of CCPA to modulate L-type VDCCs may involve Ca(2+)-calmodulin. 5. To investigate the action of CCPA on diverse degrees of nerve terminal depolarization, we studied its effect at different external K(+) concentrations. The effect of CCPA on ACh secretion evoked by 10 mm K(+) was prevented by the P/Q-type VDCC antagonist omega-agatoxin IVA. 6. CCPA failed to

  19. Cognitive correlates of α4β2 nicotinic acetylcholine receptors in mild Alzheimer's dementia.

    Science.gov (United States)

    Sabri, Osama; Meyer, Philipp M; Gräf, Susanne; Hesse, Swen; Wilke, Stephan; Becker, Georg-Alexander; Rullmann, Michael; Patt, Marianne; Luthardt, Julia; Wagenknecht, Gudrun; Hoepping, Alexander; Smits, Rene; Franke, Annegret; Sattler, Bernhard; Tiepolt, Solveig; Fischer, Steffen; Deuther-Conrad, Winnie; Hegerl, Ulrich; Barthel, Henryk; Schönknecht, Peter; Brust, Peter

    2018-06-01

    In early Alzheimer's dementia, there is a need for PET biomarkers of disease progression with close associations to cognitive dysfunction that may aid to predict further cognitive decline and neurodegeneration. Amyloid biomarkers are not suitable for that purpose. The α4β2 nicotinic acetylcholine receptors (α4β2-nAChRs) are widely abundant in the human brain. As neuromodulators they play an important role in cognitive functions such as attention, learning and memory. Post-mortem studies reported lower expression of α4β2-nAChRs in more advanced Alzheimer's dementia. However, there is ongoing controversy whether α4β2-nAChRs are reduced in early Alzheimer's dementia. Therefore, using the recently developed α4β2-nAChR-specific radioligand (-)-18F-flubatine and PET, we aimed to quantify the α4β2-nAChR availability and its relationship to specific cognitive dysfunction in mild Alzheimer's dementia. Fourteen non-smoking patients with mild Alzheimer's dementia, drug-naïve for cholinesterase therapy, were compared with 15 non-smoking healthy controls matched for age, sex and education by applying (-)-18F-flubatine PET together with a neuropsychological test battery. The one-tissue compartment model and Logan plot method with arterial input function were used for kinetic analysis to obtain the total distribution volume (VT) as the primary, and the specific binding part of the distribution volume (VS) as the secondary quantitative outcome measure of α4β2-nAChR availability. VS was determined by using a pseudo-reference region. Correlations between VT within relevant brain regions and Z-scores of five cognitive functions (episodic memory, executive function/working memory, attention, language, visuospatial function) were calculated. VT (and VS) were applied for between-group comparisons. Volume of interest and statistical parametric mapping analyses were carried out. Analyses revealed that in patients with mild Alzheimer's dementia compared to healthy controls

  20. Effect of donepezil hydrochloride (E2020) on basal concentration of extracellular acetylcholine in the hippocampus of rats.

    Science.gov (United States)

    Kosasa, T; Kuriya, Y; Matsui, K; Yamanishi, Y

    1999-09-10

    The effects of oral administration of the centrally acting acetylcholinesterase (AChE) inhibitors, donepezil hydrochloride (donepezil: E2020: (+/-)-2-[(1-benzylpiperidin-4-yl)methyl]-5,6-dimethoxy-indan-1-one monohydrochloride), tacrine (9-amino-1,2,3,4-tetrahydroacridine hydrochloride) and ENA-713 (rivastigmine: (S)-N-ethyl-3-[(1-dimethyl-amino)ethyl]-N-methyl-phenylcarbamate hydrogentartrate), which have been developed for the treatment of Alzheimer's disease, on the extracellular acetylcholine concentration in the hippocampus of rats were evaluated by using a microdialysis technique without adding cholinesterase inhibitor to the perfusion solution. We also compared the inhibition of brain AChE and the brain concentrations of these drugs. Donepezil at 2.5 mg/kg and tacrine at 5 mg/kg showed significant effects for more than 6 h. At these doses, the maximum increases were observed at about 1.5 h after administration of donepezil, and at about 2 h with tacrine, and were 499% and 422% of the pre-level, respectively. ENA-713 produced significant effects at doses of 0.625, 1.25 and 2.5 mg/kg, which lasted for about 1, 2 and 4 h, respectively. The maximum increases produced by these doses at about 0.5 h after administration were 190, 346 and 458% of the pre-level, respectively. The time courses of brain AChE inhibition with donepezil at 2.5 mg/kg, tacrine at 10 mg/kg and ENA-713 at 2.5 mg/kg were mirror images of the extracellular acetylcholine-increasing action at the same doses. The time courses of the brain concentrations of drugs after oral administration of donepezil at 2.5 mg/kg and tacrine at 10 mg/kg were consistent with those of brain AChE inhibition at the same doses, and there was a linear relation between these parameters. Brain concentration of ENA-713 at 2.5 mg/kg was below the limit of quantification at all time points measured. These results suggest that oral administration of donepezil, tacrine and ENA-713 increases acetylcholine concentration in the